XXII. BIOCHEMICKÝ ZJAZD - Jesseniova lekárska fakulta

Slovenská spoločnosť 

pre biochémiu a molekulárnu biológiu, člen IUBMB a FEBS 

Česká společnost 

pro biochemii a molekulární biologii, člen IUBMB a FEBS 

Ústav lekárskej biochémie JLF UK 

XXII. BIOCHEMICKÝ ZJAZD 

8. – 12. septembra 2010 

Jesseniova lekárska fakulta Univerzity Komenského 

v Martine

Proceedings 

from XXII. Biochemistry Congress 

held in Martin September 8 – 12, 2010 

Chairman: 

Vicechairman: 

Members: 

Honorary advisory board 

J. Turňa 

V. Pačes 

D. Dobrota, A. Hrnčiar, D. Mištuna, J. Pastorek 

Program Committee 

I. Barák A. Breier A. Drgová Z. Ďuračková 

P. Griač P. Kaplán J. Korduláková J. Kormanec 

O. Križanová Ľ. Lacinová J. Lehotský M. Mareková 

K. Mikušová P. Račay S. Stuchlík Z. Sulová 

Ľ. Škultéty J. Turňa Ľ. Varečka 

Organizing Committee 

E. Babušíková M. Bittšanský D. Dobrota A. Drgová 

A. Evinová J. Hatok M. Chomová J. Jurečeková 

P. Kaplán R. Kirschnerová M. Kovalská S. Kuka 

J. Lehotský L. Letková T. Matáková M. Pavlíková 

P. Račay M. K. Sivoňová A. Štefaníková Z. Tatarková 

Edited by: E. Babušíková, D. Dobrota, J. Hatok, J. Lehotský 

ISBN 978-80-88866-83-1 

© Comenius University in Bratislava, Jessenius Faculty of Medicine in Martin, 

Department of Medical Biochemistry 

XXII. Biochemistry Congress, Martin 

1

XXII. Biochemistry Congress is supported by: 

MaIN SPONSOrs 

BIOMEDICA SLOVAKIA s.r.o. 

BIOTECH s.r.o. 

ROCHE SLOVENSKO s.r.o. 

KRD molecular technologies s.r.o. 

K-TRADE 

LAMBDA LIFE a.s. 

MERCK spol. s r.o. 

SHIMADZU SLOVAKIA o.z. 

SIEMENS s.r.o. 

SIGMA-ALDRICH® 

TRIGON s.r.o. 

SPONSOrs 

Beckman Coulter 

Bio-Rad 

E-Colli s.r.o. 

Ecomed 

Eppendorf Czech & Slovakia s.r.o. 

Fermentas 

GeneTiCA s.r.o. 

Hermes LabSystems Ltd. 

Merci Slovakia s.r.o. 

Mettler-Toledo s.r.o. 

MGP spol s r.o. 

Millipore 

Randox s.r.o. 

Scintila s.r.o. 

THANK YOU. 

2 XXII. Biochemistry Congress, Martin

TOPICS 

I. BIOCHEMISTRy AND MOLECULAR BIOLOGy OF NERVOUS sySTEM 

II. 

III. 

IV. 

BIOTECHNOLOGY 

BIOINFORMATICS 

GENOMICS 

V. CELL REGULATIONS aND SIGNAL TRANSDUCTION 

VI. 

VII. 

GLYCOMICS 

mEMBRANE BIOCHEMISTRY AND BIOENERGETICS 

VIII. NEW METHODOLOGIC PROCEDURES 

IX. 

PATHOBIOCHEMISTRY AND TRANSLATIONAL MEDICINE 

X. PROTEOMICS AND ENZYMOLOGY 

XI. 

XII. 

REACTIVE SPECIES IN BIOMEDICINE 

TEACHING OF BIOCHEMISTRY AND MOLECULAR BIOLOGY 

XIII. XENOBIOCHEMISTRY 


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Wednesday, 8 September 2010 

14:00 - 18:00 Registration 

CC 

18:00 - 23:00 

WELCOME RECEPTION 

CC 

Thursday, 9 September 2010 

8:15 - 8:45 OPENING CEREMONY 

CC 

8:45 - 9:00 WINNER'S CEREMONY CC 

9:00 - 9:45 

PLENARY LECTURE I. 

CC 

9:45 - 10:15 

COFFEE BREAK 

10:15 - 11:55 

11:55 - 12:15 

12:15 - 12:40 

12:40 - 13:45 

13:45 - 14:10 

14:10 - 15:25 

15:25 - 15:45 

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Cell regulations and signal transduction 

CC 

Beckman lecture – cell biology 

Sigma lecture – biochemistry/biology 

Commercial lecture - Biotech 

CC 

CC 

CC 

Reactive species in biomedicine 

LUNCH & POSTER VIEWING V, VI, VII, XI 

Membrane biochemistry and bioenergetics I. 

CC 

PLENARY LECTURE II. 

COFFEE BREAK 

Glycomics 

A 

A 

15:45 - 16:35 

16:35 - 17:00 

CC 

Membrane biochemistry and bioenergetics I. 

CC 

Glycomics 

A 

17:00 - 21:15 

19:00 - 21:00 

Museum 

Slovak chamber theatre 

Aquapark Turčianské Teplice 

Friday, 10 September 2010 

8:30 - 9:15 PLENARY LECTURE III. 

CC 

9:15 - 9:35 

9:35 - 11:15 

11:15 - 11:40 

11:40 - 12:00 

12:00 - 13:00 

13:00 - 13:45 

13:45 - 14:10 

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Pathobiochemistry and translational 

medicine I. 

Commercial lecture - KRD 

LUNCH 

COFFEE BREAK 

CC 

CC 

POSTER VIEWING II, III, IV, IX, XIII 

PLENARY LECTURE IV. 

Applied molecular biology 

Beckman lecture - Genomics A 

A 

KRD workshop A 

14:10 - 15:10 

15:10 - 15:30 

Xenobiochemistry 

Membrane biochemistry and bioenergetics II. 

CC 

A 

COFFEE BREAK 

15:30 - 16:30 

CC 

Xenobiochemistry 

CC 

Membrane biochemistry and bioenergetics II. 

A 

17:00 - 18:00 

18:00 - 19:00 

19:00 - 20:00 

Concert: Cantica Collegium Musicum 


10:15 - 10:35 

10:35 9:15 --10:15 

11:35 

10:15 11:35 - 10:35 12:00 

10:35 11:35 

12:00 9:15 --10:15 

13:00 

10:15 11:35 - 10:35 12:00 

13:00 - 14:00 

10:35 

12:00 

- 11:35 

13:00 

14:00 - 14:55 

11:35 - 12:00 

13:00 14:00 

14:55 - 15:15 

12:00 13:00 

14:00 - 14:55 

13:00 15:15 - 14:00 15:55 

14:55 - 15:15 

14:00 15:55 - 14:55 16:35 

15:15 - 15:55 

14:55 16:45 - 15:15 17:00 

15:55 - 16:35 

15:15 

17:00 - 

15:55 

18:00 

16:45 17:00 

15:55 16:35 

17:00 

18:00 - 

18:00 

22:00 

16:45 - 17:00 

18:00 - 22:00 

17:00 - 18:00 

18:00 - 22:00 

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Saturday, 11 September COFFEE 2010 BREAK 

Proteomics 

Proteomics 

and 

and 

enzymology 

enzymology 

CC 

Teaching of biochemistry and 

Beckman lecture Saturday, - proteomics 

11 September COFFEE 2010 BREAK molecular biology 

CC 

A 

Proteomics 

Proteomics 

and 

and 

enzymology 

enzymology LUNCH 

CC Teaching of biochemistry and 

Beckman lecture - proteomics COFFEE molecular biology 

CC BREAK 

A 

POSTER VIEWING I, VIII, X, XII 

Proteomics and enzymology 

LUNCH 

CC 

Pathobiochemistry and translational Biochemistry Teaching of and biochemistry molecular biology and 

Beckman lecture medicine - II. proteomics CC 

molecular of nervous biology system A 

POSTER VIEWING CC I, VIII, X, XII 

A 

COFFEE 

Pathobiochemistry and translational LUNCH 

BREAK 

Biochemistry and molecular biology 

medicine II. 

CC 

of nervous system A 


medicine POSTER II. VIEWING CC I, VIII, X, XII 

COFFEE BREAK 


Pathobiochemistry and translational Biochemistry of nervous and molecular system biology 

Pathobiochemistry medicine and II. translational CC 

of nervous system 

A 

medicine II. 

CC Biochemistry and molecular biology 

Closing COFFEE ceremony BREAK of nervous system CC 

A 


medicine II. 

CC 

Closing ceremony 


CC 

of nervous system 

Farewell party 

A 

Teaching of biochemistry and molecular biology 

A 

Teaching of biochemistry and molecular biology 

A 

Closing ceremony 

CC 



Sunday, 12 September 2010 

8:00 - 12:00 

8:00 - 12:00 

13:00 

13:00 

8:00 - 12:00 

13:00 

Sunday, 12 September 2010 

Orava Castle 

Orava Castle 

Sunday, 12 September Departure 2010 

Departure 

Orava Castle 

Departure 

Šútovo waterfall 




5

Program in details: Wednesday 

PROGraM IN DETAILS 

WeDNESDAY, 8 September 2010 

Jessenius Faculty of Medicine – Convention Centre 

14.00 - 18.00 REGISTRATION 

18.00 - 22.00 WELCOME rECEPTION 


Program in details: Thursday 

ThURSDAy, 9 September 2010 


8.00 - 17.00 REGISTRATION 

8.15 - 8.45 Opening Ceremony 

8.45 - 9.00 WINNER´S CEREMONY 

9.00 - 9.45 PLENarY LECTURE I. 

Chairs: 

Ján Turňa, Dušan Dobrota 

Mathias Sprinzl: ELECTROCHEMICAL DETECTION OF NUCLEIC ACIDS 

ON BIOSENSORS 

9.45 - 10.15 CoffeE BREAK 

10.15 - 12.40 CELL rEGULaTIONS aND SIGNal traNSDUCTION 

Chairs: 

Ján Kormanec, Imrich Barák 

10.15 - 10.35 Ján Kormanec, Renáta Nováková, Ľubica Fecková, Peter Kutaš, 

Alena Reháková: REGULATION OF AURICIN BIOSYNTHESIS IN 

STREPTOMYCES AUREFACIENS CCM 3239 

10.35 - 10.55 Imrich Barák, Katarína Muchová, Naďa Pavlendová, Ján Jamroškovič: 

LIPID HELICES FORMATION IN BACILLUS SUBTILIS CELL MEMBRANE 

10.55 - 11.15 Veronika Benson, Valeria Grobárová, Katarína Hulíková, Jan 

Svoboda, Daniel Rozbeský, Daniel Kavan, Alan Kádek, Karel Křenek, 

Anna Fišerová, Vladimír Křen, Karel Bezouška: HIGH AFFINITY 

CARBOHYDRATE AND NON-CARBOHYDRATE LIGANDS FOR LECTIN- 

TYPE ACTIVATION RECEPTORS OF NATURAL KILLER CELLS REGULATE 

EFFECTOR FUNCTION THROUGH PI3K PATHWAY, AND GENERATE 

PERMANENT IMMUNE PROTECTION AGAINST MELANOMAS 

11.15 - 11.35 Radim Černý, Elerin Kärner, Christian Unger, Mikael Wendel: OSTERIX 

OVER-EXPRESSION IN HUMAN EMBRYONIC STEM CELLS AND ITS 

EFFECT ON CELL DIFFERENCIATION 


7


11.35 - 11.55 Katarína Kršková, Daniela Ježová, Lucia Gajdošechová, Miroslava 

Eckertová, Štefan Zorad: THE ROLE OF ANGIOTENSIN II AND OXYTOCIN 

IN REGULATION OF ADIPOCYTE CELL SIZE 

LECTUre - BECKMan CZE: CELL BIOLOGY 

11.55 - 12.15 Jana Morová, Radim Osička, Jiří Mašín, Peter Šebo: RTX CYTOTOXINS 

RECOGNIZE β2 INTEGRIN RECEPTORS THROUGH N-LINKED 

OLIGOSACHARIDES 

LECTUre - SIGMA ALDRICH CZE 

12.15 - 12.30 Lucie Piterková, Jana Kučerová, Karel Indrák, Vladimír Divoký: 

INTRONIC LINE-1 INSERTION IN β–GLOBIN GENE CAUSE β–TALASEMIA 

DUE TO ABERRANT SPLICING, NONSENSE-MEDIATED DECAY AND 

DECREASED RATE OF β–GLOBIN L1 

ALLELE TRANSCRIPTION 

COMMERCIAL LECTURE - BIOTECH 

12.30 - 12.40 Martin Meluzín: MODERNÍ TRENDY ZOBRAZOVACÍCH METOD 

Jessenius Faculty of Medicine – Lecture Hall A 

10.15 - 12.40 rEaCTIve SPECIES IN BIOMEDICINE 

Chairs: 

Zdeňka Ďuračková, Peter Kaplán 

10.15 - 10.50 Karl-Heinz Wagner, Oliver Neubauer: HOW CAN OXIDATIVE STRESS 

AND DNA STABILITY BE INFLUENCED BY DIET AND PHYSICAL ACTIVITY 

10.50 - 11.15 Oľga Pecháňová, Andrej Barta, Stanislava Vranková, Jana Parohová, 

Mária Kovácsová: THE CROSS-TALK OF NITRIC OXIDE AND NUCLEAR 

factor kappa B in EXPERIMENTAL hypertension 

11.15 - 11.40 Jan Borovanský, Adéla Lipšová, Jiří Vachtenheim: FREE RADICAL 

SITUATION IN PIGMENT CELLS 

11.40 - 11.55 Jan Pláteník, Juraj Gáll, Jan Škrha, Jr., Richard Buchal, Eva Sedláčková, 

Karina Verébová: INDUCTION OF MITOCHONDRIAL PERMEABILITY 

TRANSITION BY MICROMOLAR IRON 



11.55 - 12.10 Zuzana Tatarková, Eva Babušíková, Stanislav Kuka, Ján Lehotský, 

Peter Račay, Dušan Dobrota, Peter Kaplán: OXIDATIVE STRESS AND 

HEART AGING 

12.10 - 12.25 Eva Babušíková, Miloš Jeseňák, Peter Bánovčin, Dušan Dobrota: 

BRONCHIAL ASTHMA AND EFFECT OF OXIDATIVE STRESS ON ITS 

DEVELOPMENT 

12.25 - 12.40 Jana Muchová, Zuzana Nagyová, Iveta Ondrejovičová, Zdeňka 

Ďuračková: oxidative risk in atherosclerosis 

12.40 - 13.30 LUNCH 

13.00 - 13.45 POSTER VIEWING, Section V, VI, VII, XI (Convention Centre) 


13.45 - 14.10 PLENarY LECTURE II: WINNEr of „DrOBNICOv MEMOriÁl“ 

Chair: 

Albert Breier 

Mária Balážová, Peter Griač: IDENTIFICATION OF 

PHOSPHATIDYLGLYCEROL SPECIFIC PHOSPHOLIPASE C IN YEAST 

SACCHAROMYCES CEREVISIAE 

14.10 - 16.35 MEMBraNE BIOCHEMISTry aND BIOENErGETICS I. 

Chairs: 

Peter Griač, Ľubica Lacinová 

14.10 - 14.50 Anton Horváth, Ingrid Škodová, Anna Gnipová, Alena Zíková, Vladislava 

Benkovičová, Zdeněk Verner, Zdeněk Paris, Július Lukeš: SPECIALITIES 

OF OXIDATIVE PHOSPHORYLATION OF TRYPANOSOMATIDS AND 

EUGLENAS 

14.50 - 15.25 Peter Šmigáň, Monika Vidová, Janette Bobalova, Zuzana Nováková: 

ENERGETIC ASPECTS OF A MODIFICATION OF THE Na + /H + 

ANTIPORTER ACTIVITY IN A HARMALINE RESISTANT MUTANT OF 

METHANOTHERMOBACTER THERMAUTOTROPICUS 

15.25 - 15.45 Coffee BREAK 


9


15.45 - 16.00 Katarína Poloncová, Roman Holič, Peter Griač: IS 

PHOSPHATIDYLINOSITOL TRANSFER ACTIVITY ESSENTIAL FOR THE 

FUNCTION OF Pdr16p? 

16.00 - 16.15 Andrea Faltinová, Jana Gaburjáková, Ľubica Urbániková, Matúš 

Hajduk, Nataša Tomášková, Marián Antalík, Alexandra Zahradníková: 

EFFECT OF DOMAIN PEPTIDES OF THE CARDIAC ryANODINE RECEPtor 

ON THE STABILITy OF BILAYER LIPID MEMBRANES AND ON RyR2 

ACTIVITY 

16.15 - 16.35 Iveta Waczulíková, Oľga Uličná, Oľga Vančová, Jarmila Kucharská, 

Veronika Ilovská, Libuša Šikurová: ATORVASTATIN CHANGES 

MEMBRANE LIPID FLUIDITY IN MITOCHONDRIA ISOLATED FROM 

VARIOUS TISSUES OF RATS 

16.35 - 16.50 Roman Oros: ADVANCED TECHNOLOGY FOR METABOLIC 

INVESTIGATIONS 


14.10 - 17.00 GLYCOMICS 

Chairs: 

Zdenka Sulová, Katarína Mikušová 

14.10 - 14.35 Igor Tvaroška: MOLECULAR MODELING INSIGHT INTO CATALYTIC 

MECHANISMS OF GLYCOSYLTRANSFERASES 

14.35 - 15.00 Marek Baráth, Igor Tvaroška, Ján Hirsch: SYNTHESIS OF GlcNAc-TS 

MIMETICS AS A POTENT INHIBITORS OF GLYCOSYLTRANSFERASES 

15.00 - 15.25 Michaela Wimmerová: LECTINS FROM PATHOGENS: MYSTERY OF 

LIFE 


15.45 - 16.10 Vladimír Farkaš: TRANSGLYCOSYLATION - A UNIVERSAL PRINCIPLE 

IN TAILORING THE PLANT AND FUNGAL CELL WALLS 

16.10 - 16.35 Mária Vršanská, Katarína Šuchová, Vladimír Puchart, Peter Biely: 

DIFFERENCIES BETWEEN TWO ENDOXYLANASES FROM GH5 


16.35 - 17.00 Zuzana Svetlíková, Marcelo E. Guerin, Mary Jackson, Jana Korduláková, 

Katarína Mikušová: MyCOBACTERIAL MANNOSyl TRANSFERASE PimA 

as A TARGET FOR the DEVELOPMENT of NEW ANTITUBERCULAR 

DRUGS 

16.45 - 21.15 SOCIal evENTS 


17.30 - 18.30 Slovak National Museum 

16.45 - 21.15 Aquapark Turčianske Teplice 

19.00 - 21.00 Slovak Chamber Theatre, Martin 


11

Program in details: Friday 

friday, 10 September 2010 


8.30 - 9.15 PLENarY LECTURE III. 

Chairs: 

Ján Lehotský 

Peter Biely: MICROBIAL XYLANASES: PROPERTIES AND APPLICATIONS 


9.35 - 11.15 PaTHOBIOCHEMISTry aND traNSLaTIONal MEDICINE I. 

Chairs: 

Peter Račay, Oľga Križanová 

09.35 - 10.00 Juraj Kopáček, Jaromír Pastorek, Silvia Pastoreková: Molecular 

mechanisms involved in response to hypoxia 

10.00 - 10.25 Ľubomíra Lenčešová, Marta Sírová, Lucia Csáderová, Marcela Lauková, 

Zdena Sulová, Richard Kvetňanský, Oľga Križanová: CHANGES AND 

ROLE OF ADRENOCEPTORS IN PC12 CELLS AFTER PHENYLEPHRINE 

ADMINISTRATION AND APOPTOSIS INDUCTION 

10.25 - 10.50 Miroslav Barančík, Petra Šimončíková, Monika Ivanová: EFFECTS OF 

doxorubicin TREATMENT on MATRIX METALLOPROTEINASES IN 

RATS 

10.50 - 11.15 Attila Ziegelhöffer, Jana Mujkošová, Oľga Uličná, Iveta Waczulíková, 

Miroslav Ferko, Norbert Vrbjar, Štefan Polák, Tanya Ravingerová, 

Adriana Adameová: FUNCTION AND PROPERTIES OF HEART AND 

KIDNEY MITOCHONDRIA (MIT) IN SPONTANEOUSLY HYPERTENSIVE 

(HYP) RATS: INFLUENCE OF CAPTOPRIL AND NIFEDIPINE 

LECTUre - KRD 

11.40 - 11.50 Jiří Vašák: LIVING COLOURS: ILLUMINATE yOUR ASSAys WITH 

Clontech 




9.35 - 12.00 aPPLIED MOLECULar BIOLOGY 

Chairs: 

Stanislav Stuchlík, Ján Turňa 

09.35 - 10.00 Peter Májek, Vladimír Špitalský, Gabriel Minárik, Tomáš Szemes: 

A METHOD FOR AUTOMATED DETECTION OF HETEROZYGOUS 

INSERTION-DELETION MUTATIONS 

10.00 - 10.25 Anna Hrabovska, Veronique Bernard, Eric Krejci: PROTEIN 

IMMUNIZATION OF MUTANT MOUSE – AN EFFICIENT WAY TO 

GENERATE SELECTIVE AND SENSITIVE ANTIBODIES 

10.25 - 10.50 Michal Kaliňák, Tibor Liptaj: NEW POSSIBILITIES FOR THE STUDY OF 

METABOLISM IN SLOVAKIA 

10.50 - 11.15 Martin Benej, Martina Poturnajová: TOXCAT METHOD: APPLICATION 

IN MOLECULAR ONCOLOGY 

11.15 - 11.40 Laco Kačáni: FROM BASIC BIOMEDICAL RESEARCH TO BIOTECHNOLOGy 

LECTUre - BECKMan CZE: GENOMICS 

11.40 - 12.00 Ondřej Mihola, Zdeněk Trachtulec, Jiří Forejt: The IDENTIFICATION 

and CHARACTERIZATION OF THE FIRST VERTEBRATE hyBRID STERILity 

gene (HST1/PRDM9) 

WorKSHOP 

12.00 - 13.00 KRD Molecular Technologies, s.r.o. 

12.00 - 13.00 LUNCH 

13.00 - 13.45 POSTER VIEWING, Sections II, III, IV,IX, XIII (Convention Centre) 


13



13.45 - 14.10 PLENaRY LECTURE Iv: WINNEr of „KoŠTířova CENA“ 

Chair: 

Václav Pačes 

Dana Douděrová, Pavel Martásek: STRUCTURAL-FUNCTIONAL 

CORRELATIONS OF HYDROXYMETHYLBILANE SYNTHASE 

14.10 - 16.30 XENOBIOCHEMISTry 

Chairs: 

Albert Breier, Ľudovít Varečka 

14.10 - 14.30 Zdena Sulová, Mário Šereš, Miroslav Barančík, Lenka Gibalová, 

Branislav Uhrík, Lenka Poleková, Albert Breier: DOES EXISTS ANY 

RELATION BETWEEN P-GLYCOPROTEIN MEDIATED MULTIDRUG 

RESISTANCE AND INTRACELLULAR CALCIUM HOMEOSTASIS 

14.30 - 14.50 Pavlína Janů, Markéta Thimová, Petra Lovecká, Martina Macková, 

Kateřina Demnerová: EVALUATION OF TOXICITY AND GENOTOXICITY 

OF ORGANOHALOGEN PESTICIDES 

14.50 - 15.10 Monika Kmeťová Sivoňová, Dušan Dobrota, Tatiana Matáková, Zuzana 

Tatarková, Mária Kovalská, Martina Pavlíková, Róbert Dušenka, Ján 

Kliment: XENOBIOTIC-METABOLIZING ENZYMES POLYMORPHISMS 

AND CANCER RISK 

15.10 - 15.30 COFFEE BREAK 

15.30 - 15.50 Helena Mertlíková-Kaiserová, Antonín Holý, Ivan Votruba: Origin 

of ACqUIRED RESISTANCE TO cyTOTOXIC ACyCLIC NUCLEOSIDE 

phosphonates 

15.50 - 16.10 Marie Stiborová, Eva Frei: CYTOCHROME P450- AND PEROXIDASE- 

MEDIATED OXIDATION OF ELLIPTICINE DICTATES ITS ANTI-TUMOR 

EFFICIENCY 

16.10 - 16.30 Vladimíra Tomečková: SYNTHETIC CYCLIC CHALCONE ANALOGUES 

AS NOVEL BIOLOGICALLY ACTIVE DYES 




14.10 - 16.10 MEMBraNE BIOCHEMISTry aND BIOENErGETICS II. 

Chairs: 

Peter Griač, Ľubica Lacinová 

14.10 - 14.50 Milan Štengl, František Barták, Roman Sýkora, Jiří Chvojka, Jan 

Beneš, Aleš Kroužecký, Ivan Novák, Jitka Švíglerová, Jitka Kuncová, 

Martin Matějovič: CARDIAC L-TYPE CALCIUM CURRENT IN SEPSIS 

14.50 - 15.05 Viera Kominková, Zuzana Tomášková, Ľubica Máleková, Karol 

Ondriaš: EFFECT OF ADENINE NUCLEOTIDES AND Mg 2+ IONS ON 

mitochondrial chloride channels 


15.30 - 16.00 Ľubica Lacinová, Mária Karmažínová: GATING OF THE T-TYPE CALCIUM 

CHANNELS 

16.00 - 16.15 Mária Karmažínová, Edward Perez-Reyes, Ľubica Lacinová: GATING 

OF THE NEURONAL CA V 

3.3 CHANNEL 

19.00 - 20.00 CONCERT: CanTICa COLLEGIUM MUSICUM (EVANGELIC CHURCH) 

20.30 - 21.30 Biznis meeting: Members of committees Slovak and Czech biochemical 

societes 


15

Program in details: Saturday 

SATURDAY, 11 September 2010 


9.15 - 12.00 PrOTEOMICS aND ENZYMOLOGY 

Chairs: 

Ľudovít Škultéty, Peter Račay 

09.15 - 09.35 Daniela Krajčíková, Denisa Mullerová, Wan Qiang, Per Bullogh, Jilin 

Tang, Imrich Barák: ASSEMBLY OF BACILLUS SUBTILIS SPORE COAT: 

INVESTIGATION OF PROTEIN-PROTEIN INTERACTIONS AMONG THE 

SPORE COAT PROTEINS OF BACILLUS SUBTILIS 

09.35 - 09.55 Katarína Bíliková, Jozef Šimúth: PROTEOMICS OF MULTIFUNCTIONAL 

ROYAL JELLY PROTEINS 

09.55 - 10.15 Andrea Antošová, Katarína Šipošová, Martina Koneracká, Vlasta 

Závišová, Peter Kopčanský, Zuzana Gažová: INHIBITION OF INSULIN 

AMYLOID AGGREGATION WITH ALBUMIN FUNCTIONALIZED 

MAGNETIC FLUID 


10.35 - 10.55 H. Mrazek, L. Weignerova, D. Manglova, D. Kavan , , V. Kren, K. Bezouska: 

FUNGAL α–N-ACETYLGALACTOSAMINIDASE FROM ASPERGILLUS 

NIGER: CLONING AND EXPRESSION IN YEAST 

10.55 - 11.15 Gabriela Flores Ramírez, Zuzana Bílková, Pavol Vadovič, Ľudovít 

Škultéty: IDENTIFICATION OF Surface PROTEINS OF THE OBLIGATE 

INTRACELLULAR bacterium CoxIELLA BURNETII 

11.15 - 11.35 Martin Hajduch, Katarína Klubicová, Maksym Danchenko, Ľudovít 

Škultéty, Namik Rashydov, Anna Preťová: TWENTy FOUR yEARS 

since Chernobyl DISASTER: WHAT seed PROTEIN can tell us? 

LECTUre - BECKMan CZE: prOTEOMICS 

11.35 - 12.00 Pavel Bouchal, Monika Mudrochová, Eva Budinská, Zbyněk Bortlíček, 

Iva Struhárová, Lenka Hernychová, Theodoros Roumeliotis, Spiros D. 

Garbis, Roman Hrstka, Petr Müller, Rudolf Nenutil, Bořivoj Vojtěšek: 

BIOMARKERS OF lyMPH NODE METASTASIS IN LOW-GRADE BREAST 

cancer: An integrated, proteomics-based approach 




9.15 - 12.00 TeaCHING of BIOCHEMISTry aND MOLECULar BIOLOGY 

Chairs: 

Anna Drgová, Mária Mareková, Jana Korduláková 

09.15 - 09.35 Ľubomír Tomáška: WHEN MORE IS LESS: A DILEMMA OF A BIOMEDIcal 

educator 

09.35 - 09.55 Zuzana Kostecká: TEACHING BIOCHEMISTRY AT THE UNIVERSITY OF 

VETERINARY MEDICINE AND PHARMACY IN KOŠICE 

09.55 - 10.15 Jiří Hudeček: DO WE TEACH BIOCHEMISTRY IN A LOGICAL WAY? 

REMARKS CONCERNING THE CONTENTS AND LEARNING APPROACH 


10.35 - 10.55 Mária Kožurková, Marián Antalík, Dušan Podhradský: TEACHING 

BIOCHEMISTRY AT THE FACULTY OF SCIENCE IN KOŠICE 

10.55 - 11.15 Daniel Rajdl, Jaroslav Racek, Marie Šolcová: E-LEARNING – FRIEND 

OF FOE? 

11.15 - 11.35 Zdeňka Ďuračková, Zuzana Országhová: HISTORY AND PRESENT OF 

SCIENTIFIC AND PEDAGOGIC CONFERENCES OF TEACHERS FROM 

CHEMICAL INSTITUTES AND DEPARTMENTS OF SLOVAK AND CZECH 

MEDICAL FACULTIES 

11.35 - 11.55 Mária Mareková, Jana Mašlanková, Peter Urban, Juraj Guzy: 

BIOCHEMISTRY IN THE PICTURES – INTERACTIVE BIOCHEMISTRY 

12.00 - 13.00 LUNCH 

13.00 - 14.00 POSTER vIEWING, Sections: I, VIII, X (Convention Centre) 


17



14.00 - 16.00 PaTHOBIOCHEMISTry aND traNSLaTIONal MEDICINE II. 

Chairs: 

Peter Račay, Oľga Križanová 

14.00 - 14.35 Nadežda Lukáčová, Alexandra Dávidová, Ľudmila Capková, Andrea 

Kucharíková: SPINAL CORD INJURY: PATHOGENESIS AND TREATMENT 

14.35 - 14.55 Peter Račay, Jozef Hatok, Mária Chomová, Jana Jurečeková, Peter 

Chudý, Juraj Chudej, Andrea Štefániková, Dušan Dobrota: APOPTOSIS 

– DOUBLE EDGED SWORD 


15.15 - 15.35 Jozef Hatok, Jana Jurečeková, Peter Chudý, Pavol Hollý, Anton 

Dzian, Eduard Huľo, Eva Fabianová, Tatiana Matáková, Peter Račay: 

APOPTOSIS IN RELATION TO the DEVELOPMENT of CANCER AND 

RESISTANCE of cancer cells to cytostatics 

15.35 - 15.55 Lenka Surdeníková, Fei Ru, Marian Kollárik: UTILITY OF SINGLE 

CELL RT-PCR (scRT-PCR) FOR THE STUDY OF PRIMARY AFFERENT 

NEURONS - PRELIMINARY VALIDATION 


14.00 - 16.00 BIOCHEMISTry aND MOLECULar BIOLOGY of NErvOUS 

system 

Chairs: 

Ján Lehotský, Dušan Dobrota 

14.00 - 14.35 Jozef Michalik, Egon Kurča: BIOCHEMICAL MARKERS OF MULTIPLE 

SCLEROSIS 

14.35 - 14.55 Eva Babušíková, Dušan Dobrota, Anthony J. Turner, Natalia N. 

Nalivaeva: PROCESSING OF AMYLOID PRECURSOR PROTEIN AFTER 

IN VIVO INDUCED ISCHEMIA 




15.15 - 15.35 Dušan Dobrota, Michal Bittšanský: MAGNETIC RESONANCE 

SPECTROSCOPY IN DIAGNOSTIC PROTOCOL OF THE BRAIN DISEASES 

15.35 - 15.55 Michal Bittšanský, Veronika Husárová, Igor Ondrejka, Valéria 

Kerná, Pavol Adamík, Huber Poláček, Dušan Dobrota: CHANGES 

IN NEURONAL METABOLITES MEASURED BY PROTON MAGNETIC 

RESONANCE SPECTROSCOPY IN DEPRESSED PATIENTS DURING 

TREATMENT 

15.55 - 16.15 Ján Lehotský, Mária Chomová, Andrea Evinová, Mária Kovalská, 

Martina Pavlíková, Zuzana Tatarková, Peter Kaplán, Peter Račay: 

INDUCTION OF ISCHEMIC TOLERANCE IN SENSITIVE NEURONS: 

coordinated role of multiple mechanisms 

16.15 - 16.35 Rostislav Skrabana, Radovan Dvorsky, Branislav Kovacech, Jozef 

Sevcik,Michal Novak: STRUCTURAL ANALySIS OF TAU PROTEIN, THE 

constituent of NEUROFIBRILLARy PATHOLOGy IN ALZHEIMER›s 

DISEASE 

16.45 - 17.00 CLOSING CEREMONY 

18.00 - 22.00 FAREWELL PARTy (Open-air Museum of Slovak Village, Martin) 


19

Program in details: Sunday 

SUNDAY, 12 September 2010 

08.00 - 12.00 GUIDED TOUrs 

Orava Castle 


13:00 DEPARTURE 


POSTER VIEWING 


21

Program in details: Poster viewing 

V. 

POSTER vIEWING 1., SECTION V, VI, VII, XI 

THUrSDay, 9 SEPTEMBEr 2010, 

13.00 - 13.45 

29. Lucia Gajdošechová, Miroslava Eckertová, Katarína Kršková, Štefan Zorad: THE ROLE 

OF 14-3-3 PROTEIN IN REGULATION OF GLUCOSE TRANSPORTER GLUT4 TRANSLOCATION 

TO ADIPOCyTE PLASMA MEMBRANE 

30. Dana Grebeňová, Michaela Pluskalová, Zbyněk Hrkal, Kateřina Kuželová: CHANGES 

in COFILIN PHOSPHORyLATION DURING THE APOPTOSIS OF LEUKEMIC JURL-MK1 CELLS 

31. Dagmar Homerová, Bronislava Řežuchová, Henrieta Škovierová, Ján Kormanec: 

CHARACTERIZATION OF A GENE ENCODING A SMALL REGULATORy RPOE – DEPENDENT 

RNA IN SALMONELLA ENTERICA SEROVAR tyPHIMURIUM 

32. Iva Jelínková, Olga Vondálová Blanářová, Jiřina Hofmanová, Petr Sova, Alois Kozubík, 

Alena Vaculová: MOLECULAR MECHANISMS INVOLVED IN POTENT PLATINUM (IV) COMPLEXmediated 

COLON CANCER CELL SENSITIZATION TO TRAIL-INDUCED APOPTOSIS 

33. Lenka Kočí, Martina Hýžďalová, Alena Vaculová, Jiřina Hofmanová, Alois Kozubík: 

TRAIL-INDUCED APOPTOSIS CAUSES ACTIVATION OF PRO-SURVIVAL PATHWAys IN NON- 

ADHERENTLy GROWING COLON CANCER CELLS 

34. Gabriel Kollárovič, Miroslava Kretová, Lucia Lichá, Peter Baráth, Katarína Luciaková: 

PREPARATION AND FUNCTIONAL ANALySIS OF PHOSPHORyLATION MUTANT FORMS OF 

THE TRANSCRIPTION FACTOR NFI 

35. Soňa Kontseková, Anna Repič, Monika Baráthová, Katarína Polčicová, Jaromír Pastorek: 

GENERATION AND CHARACTERIZATION MONOCLONAL ANTIBODIES AGAINST ENDOSIALIN, 

the POTENTIAL MARKER OF TUMOR ANGIOGENESIS 

36. Miroslava Kretová, Ľudmila Šabová,Katarína Luciaková: THE ROLE OF NF1 IN p21 GENE 

expression 

37. Peter Kutaš, Ľubomíra Fecková, Alena Reháková, Renáta Nováková, Ján Kormanec: 

STRICT CONTROL OF AURICIN PRODUCTION IN STREPTOMyCES AUREOFACIES CCM 3239 

INVOLVES A FEEDBACK MECHANISM 

38. Ingrid Lajdová, Viera Spustová, Adrián Okša, Dušan Chorvát Jr.: ALTERED CALCIUM 

signaling IN PERIPHERAL BLOOD MONONUCLEAR CELLS OF CHRONIC KIDNEy DISEASE 

PATIENTS 

39. Ľubica Ondrušová, Jiří Vachtenheim: EXPRESSION OF MICROPHTHALMIA-ASSOCIATED 

TRANSCRIPTION FACTOR CRITICALLy REqUIRES ACTIVE SWI/SNF CHROMATIN REMODEL- 

ING COMPLEX 



40. Barbora Brodská, Petra Otevřelová, Aleš Holoubek: MCL-1 AS REGULATOR OF APOPTOSIS 

in CML CELL LINE AND PERIPHERAL BLOOD MONONUCLEAR CELLS 

41. Jana Plšíková, Ján Kovaľ, Jaromír Mikeš, Mária Kožurková, Peter Fedoročko, Ladislav 

Janovec, Ján Ungvarský, Danica Sabolová: NOVEL GUANIDINE DERIVATIVES AND EVALUA- 

TION OF THEIR DNA BINDING AFFINITIES AND POSSIBLE ANTICANCER EFFECT 

42. Michaela Pluskalová, Dana Grebeňová, Zbyněk Hrkal, Kateřina Kuželová: HISTONE 

acetyLATION OF LEUKEMIC JURL-MK1 CELLS WITHIN SAHA TREATMENT 

43. Jiřina Procházková, Lenka Umannová, Alois Kozubík, Miroslav Machala, Jan Vondráček: 

REGULATION OF PLAKOGLOBIN EXPRESSION, A KEy DESMOSOMAL CONSTITUENT, by ARyl 

hyDROCARBON RECEPTOR AND cAMP SIGNALING 

44. Alena Reháková, Renáta Nováková, Ľubomíra Fecková, Peter Kutaš, Ján Kormanec: 

CHARAKTERIZATION OF SARP REGULATORy GENE INVOLVED IN POSITIVE REGULATION OF AN 

angucyCLINE-LIKE POLyKETIDE ANTIBIOTICS AURICIN GENE CLUSTER IN STREPTOMYCES 

aureofaciens CCM 3239 

45. Bronislava Řežuchová, Beatrica Ševčíková, Dagmar Homerová, Ján Kormanec: THE COMplex 

NETWORK REGULATORy CIRCUITS IN THE REGULATION OF SIGMA FACTORS INVOLVED 

in DIFFERENTIATION AND STRESS RESPONSE IN STREPTOMYCES COELICOLOR A3(2) 

VI. 

46. Tatiana Kurucová, Helena Kavcová, Kristína Rogozanová, Lucia Messingerová, Danica 

Mislovčová, Albert Breier, Zdena Sulová: DIFFERENCES IN INTERACTION OF LECTINS SPEcifically 

RECOGNIZING SIALIC ACID RESIDUES WITH SURFACE OF P-GP NEGATIVE OR 

positive L1210 CELLS 

47. Zdena Sulová, Peter Ditte, Tatiana Kurucová, Eva Poláková, Kristína Rogozanová Lucia 

Škvarková, Ján Sedlák, Jaromír Pastorek, Albert Breier: THE PRESENCE OF P-GLyCOPROTEIN 

in L1210 CELLS DIRECTLy INDUCES DOWN-REGULATION OF CELL SURFACE SACCHARIDE- 

TARGETS OF CONCANAVALIN A 

VII. 

48. Jana Antalíková, Jana Jankovičová, Katarína Michalková, Michal Simon, Ľubica Horovská: 

EPITOP OF IVA-520 MONOCLONAL ANTIBODy ON THE BOVINE SPERM CD46 MOLECULE 

49. Cagala M. , Lencesova L., Hudecova S., Csaderova L., Sirova M., Cholujova D., Kopacek 

J., Pastorekova S., Krizanova O.: DIMETHyLOXALLyl gyCINE MODULATES GENE EXPRESION 

and PROTEIN LEVELS OF THE SODIUM CALCIUM EXCHANGER IN HEK 293 CELL LINE 


23


50. Peter Kohút, Martin Valachovič, Lucia Hronská, Ivan Hapala: DEHyDROERGOSTEROL 

ELUCIDATES STEROL UPTAKE PROCESS IN yEAST S. CEREVISIAE 

51. Jan Madacki, Katarína Mikušová, Mary Jackson, Jana Korduláková: MyCOBACTERIAL 

EPOXIDEHyDROLASE EPHD IS INVOLVED IN FATTy ACID METABOLISM 

52. Boris Lakatoš, Lucia Bialešová, Eva Harnišová: ISOFORMS OF AMP-ACTIVATED PROTEIN 

kinase SUBUNITS IN lyMPHOCyTES AND OBESITy 

53. Katarína Michalková, Michal Simon, Jana Antalíková, Ľubica Horovská: DISTRIBUTION 

AND BIOCHEMICAL CHARACTERIZATION OF CD52-LIKE MOLECULE IN BULL EPIDIDyMIS 

54. Hana Rauchová, Martina Vokurková, , 

Tomáš Soukup: IDEBENONE ACTIVATION OF 

glyCEROL-3-PHOSPHATE OXIDATION IN LIVER MITOCHONDRIA FROM CONTROL AND 

hyPERTHyROID RATS 

55. Oľga Uličná, Oľga Vančová, Jarmila Kucharská, Peter Božek, Iveta Waczulíková, Libuša 

Šikurová: EFFECT OF ATORVASTATIN ON BIOENERGETICS OF THE LIVER MITOCHONDRIA 

ON A HIGH LIPID DIET 

56. Oľga Vančová, Oľga Uličná, Katarína Šebeková, Magdalena Labieniec, Cezary Watala: 

EFFECT OF PAMAM G4 DENDRIMER ON LIVER MITOCHONDRIA OXIDATIVE PHOSPHORy- 

LATION AND LONG-TERM MARKERS OF hyPERGLyCAEMIA IN EXPERIMENTAL DIABETES 

57. Monika Vidová, Zuzana Nováková, Peter Šmigáň: BIOCHEMICAL AND MOLECULAR ANALySIS 

of NITRATE-RESISTANT MUTANT OF METHANOTHERMOBACTER THERMAUTOTROPHICUS 

XI. 

90. Lucia Andrezálová, Zuzana Országhová, Jana Muchová, Zdeňka Ďuračková: EFFECT OF 

OMEGA-3 FATTy ACIDS ON PON 1 ARyLESTERASE AND LACTONASE ACTIVITy IN CHILDREN 

SUFFERING FROM hyPERCHOLESTEROLEMIA 

91. Ima Dovinová, Zuzana Pakanová, Stanislava Vranková, Oľga Pecháňová, Soňa Čačányiová, 

František Kristek, Helena Paulíková: ANTIOXIDANT RESPONSE TO TREATMENT WITH NATUral 

COMPOUNDS AND AN NO DONOR IN EXPERIMENTAL hyPERTENSION 

92. Marián Koláček, Jana Muchová, Eva Uhlíková, Viera Kupčová, Ladislav Turecký: WILSON´s 

disease AND OXIDATIVE STRESS 

93. Stanislav Kuka, Zuzana Tatarková, Peter Račay, Ján Lehotský, Dušan Dobrota, Peter 

Kaplán: AGE-RELATED CHANGES IN ACTIVITIES OF MITOCHONDRIAL ELECTRON TRANSPORT 

chain COMPLEXES IN THE RAT HEART 

94. Daniela Mokrá, Anna Drgová, Rudolf Pullmann st., Andrea Čalkovská: SELECTIVE PHOSphodiesterase-3 

INHIBITOR OLPRINONE ALLEVIATES OXIDATIVE LUNG INJURy INDUCED 

by MECONIUM 



95. Zuzana Országhová, Zuzana Paduchová, Ingrid Žitňanová, Cezary Watala, Jana Muchová, 

Zdeňka Ďuračková: METyLNICOTINAMIDE AND DNA OXIDATION DAMAGE IN RATS WITH 

STREPTOZOTOCINE INDUCED DIABETES MELLITUS 

96. Eliška Procházková, Petr Jansa, Lucie Čechová, Ivan Votruba, Helena Mertlíková-Kaiserová: 

ANTIOXIDANT CAPACITy OF SELECTED ANALOGS OF NUCLEIC ACID COMPONENTS: COM- 

PARISON OF CELL-FREE AND CELL-BASED ASSAys 

97. Beáta Veliká, Ivan Kron: EFFECTIVENESS OF PHENOLS AS ANTIOXIDANTS AGAINST 

superoxide RADICAL 

98. Martina Vokurková, Hana Rauchová, Stanislav Pavelka, Tomáš Soukup, Narcis Tribulová: 

EFFECT OF n-3 POLyUNSATURATED FATTy ACIDS SUPPLEMENTATION ON RAT LIVER IN 

different THyROID STATUS 

99. Adéla Zdařilová, Alena Rajnochová Svobodová, Jana Zapletalová, Pavel Štrebl, Josef 

Zadražil, Jitka Vostálová: EFFECT OF RENAL TRANSPLANTATION ON OXIDATIVE STRESSrelated 

BIOMARKERS 

POSTER VIEWING 2., SECTION II, III, IV,IX, XIII 

FrIDay, 10 SEPTEMBEr 2010, 

13.00 - 13.45 

II. 

8. Hind Al Alami, Ľubomíra Tóthová, Michal Kajsík, Jana Gajdošová, Hana Drahovská, Ján 

Turňa: CHARACTERIZATION OF BACTERIOPHAGES INFECTING CRONOBACTER STRAINS 

9. Andrea Balažová, Víťazoslava Blanáriková, Jindra Valentová, František Bilka, Ivana 

Holková: EFFECT OF METHyl JASMONATE ON THE PRODUCTION OF SANGUINARINE AND 

ITS PRECURSORS IN OPIUM POPPy SUSPENSION CULTURES 

10. František Bilka, Andrea Balažová, Andrea Bilková, Víťazoslava Blanáriková, Ivana Holková, 

Marián Vanko: COMPARISON OF SANGUINARINE PRODUCTION OF PAPAVERACEAE FAMILy 

plants IN VITRO CULTURES 

11. Michaela Kandričáková , 

Stanislav Stuchlík, Ján Turňa: DESIGN OF AN EXPRESSION system 

FOR THE PRODUCTION OF RECOMBINANT HUMAN THROMBIN IN ESCHERICHIA COLI 

12. Tatiana Kraková, Jozef Šimúth, Katarína Bíliková: HETEROLOGOUS EXPRESSION OF 

royAL JELLy APALBUMINS IN E. COLI 

13. Mahesh Madyagol, Stanislav Stuchlík, Ján Turňa: EXPRESSION, PURIFICATION AND 

functional CHARACTERIZATION OF TWO FORMS OF AGROBACTERIUM SP. STRAIN CP4 

EPSPS GENE IN ESCHERICHIA COLI FOR HORIZONTAL GENE TRANSFER STUDIES 


25


14. Jozef Parnica, Lukáš Kandráč, Marián Antalík: CONFORMATION TRANSITIONS OF cy- 

TOCHROME c IN DEEP EUTECTIC SOLVENTS 

15. Lucia Pánčiová, Zdenko Levarski, Pavol Utekal, Stanislav Stuchlík, Ján Turňa: PRODUCTION 

of RECOMBINANT PROTEINS USING HIGH CELL DENSITy CULTURES OF ESCHERICHIA COLI 

in BIOREACTOR 

16. Michaela Šimšíková, Marián Antalík: SyNTHESIS AND SURFACE MODIFICATION OF ZINC 

oxide NANOPARTICLES 

17. Csaba Tóth, Roland Pálffy, Juraj Gašperík, Stanislav Stuchlík, Ján Turňa: CLONING, EX- 

PRESSION AND ANTIMICROBIAL ACTIVITy OF THE HUMAN CATHELICIDIN LL-37 

18. Ľubomíra Tóthová, Hind Al Alami, Jana Lintnerová, Hana Drahovská, Ján Turňa: 

CHARACTERIZATION OF BACTERIOPHAGES INFECTING SALMONELLA ENTERICA 

19. Pavol Utekal, Lucia Pánčiová, Stanislav Stuchlík, Ján Turňa: PRODUCTION OF TWO 

recombinant ALCOHOLDEHyDROGENASES SUITABLE FOR BIOTRANSFORMATION OF C-6 

aldehyDES INTO CORESPONDING ALCOHOLS 

20. Ondřej Vaněk, Petra Celadová, Jan Bláha, Daniel Kavan, Petr Pompach, Karel Bezouška: 

EUKARyOTIC EXPRESSION AS AN INDISPENSABLE TOOL FOR PREPARATION OF NATIVE 

dimeric FORMS OF NK CELL C-tyPE LECTIN-LIKE RECEPTORS 

III. 

21. Matej Stano, Ľuboš Kľučár: phiGENOME - A WEB-BASED GENOME BROWSER INTENDED 

for DISPLAy OF PHAGE GENOMES 

IV. 

22. Jarmila Farkašovská, Andrej Godány: SITE-SPECIFIC INTEGRATION OF BACTERIOPHAGE 

µ1/6 INTO THE STREPTOMYCES AUREOFACIENS CHROMOSOME 

23. Jana Gajdošová, Natália Kamodyová, Kristína Benedikovičová, Hana Drahovská, 

Eva Kaclíková, Ján Turňa: STUDy OF THERMOTOLERANCE ISLAND IN CRONOBACTER 

spp. 

24. Lucia Letková, Tatiana Matáková, Erika Halašová, Anna Drgová, Dušan Dobrota: DNA 

POLyMORPHISMS OF SELECTED REPAIR GENES AND RISK OF LUNG CANCER 

25. Eva Lincová/Slabáková, Zuzana Pernicová, Eva Slavíčková, Alois Kozubík, Karel Souček: 

EXPRESSION OF TRANSCRIPTION FACTORS AND microRNAs IN TGF-β1-INDUCED EMT OF 

BENIGN pROSTATE EPITHELIAL CELLS 



26. Silvia Mahmood, Tatiana Matáková, Lucia Letková, Monika Kmeťová Sivoňová, Jozef 

Hatok, Dušan Dobrota: THE ASSOCIATION BETWEEN EGF 61 G/A POLYMORPHISM AND 

COLORECTAL CANCER DEVELOPMENT 

27. Milena Matejovičová, Michaela Králíková, Jitka Melounová, Martina Vodová, Jana Žáková, 

Igor Crha: SPERM DNA INTEGRITy ASSESMENT USING DIFFERENT COMET ASSAy PROTOCOLS 

28. Marika Matoušová, Ivan Votruba, Miroslav Otmar, Helena Mertlíková-Kaiserová: 

COMPARATIVE STUDy OF hyPOMETHyLATING ACTIVITIES OF 5-AZACyTIDINE CONGENERS 

IX. 

60. Soňa Bálentová, Eva Hajtmanová, Yvetta Mellová, Ivana Kinclová, Marián Adamkov: 

EFFECT OF FRACTIONATED DOSES OF GAMA RAys ON THE ROSTRAL MIGRATORy STREAM 

OF ADULTS RATS 

61. Ľudmila Capková, Alexandra Dávidová, Andrea Kucháriková, Nadežda Lukáčová: Is 

respiratory PATHWAy ACTING THROUGH NO-sGC? 

62. Monika Dvořáková, Jana Muchová, Branislav Trebatický, Ján Breza, Zdeňka Ďuračková: 

THE EFFECT OF NATURAL POLyPHENOLS ON ADIPONECTINE LEVEL IN PATIENTS SUFFERING 

from ERECTILE dySFUNCTION 

63. Jana Jurečeková, Jozef Hatok, Andrea Štefániková, Dušan Dobrota, Peter Račay: STUDy 

of ANTIAPOPTOTIC PROTEINS RESPONSIBLE FOR DEVELOPMENT OF DRUG RESISTANCE 

IN ACUTE LEUKEMIA 

64. Vlastimil Kulda, Martin Pešta, Ondřej Topolčan, Lukáš Řehoř, Martin Svatoň, Václav 

Liška, Václav Babuška, Luboš Holubec, Radim Černý: PROGNOSTIC SIGNIFICANCE OF MIR- 

21 AND MIR-143 EXPRESSION IN TISSUE SAMPLES OF COLORECTAL CARCINOMA AND 

colorectal LIVER METASTASES 

65. Erika Moravčíková, Evžen Křepela, Jan Prochádzka, Jan Čermák, Kamila Benková: THE 

functionality OF APOPTOSOME APPARATUS AND THE EXPRESSION OF ITS REGULATORS 

in NON-SMALL CELL LUNG CARCINOMA 

66. Iveta Ondrejovičová, Jana Muchová, Zuzana Paduchová, Zuzana Nagyová, Zdeňka 

Ďuračková: EFFECT OF OMEGA-3 PUFA ON LIPID PROFILE AND OXIDATIVE STRESS IN hy- 

PERCHOLESTEROLEMIC CHILDREN 

67. Blanka Stibůrková, Makoto Hosoyamada, Kimiyoshi Ichida, Ivan Šebesta: ANALySIS OF 

urate TRANSPORTERS SLC22A12 AND SLC2A9 IN PATIENTS WITH RENAL hyPOURICEMIA 

in CZECH POPULATION 

68. Andrea Štefániková, Jozef Hatok, Jana Jurečeková, Ivana Plameňová, Dušan Dobrota, 

Peter Račay: STUDy OF THE EFFECT OF HISTONE DEACETyLASE INHIBITOR ON THE SENSI- 

TIVITy OF LEUKAEMIC CELLS TO THE cyTOSTATICS 


27


69. Ladislav Vaško, Janka Vašková: CONTENT OF FATTy ACIDS IN FOOD AND HEALTH STATUS 

70. Janka Vašková, Ladislav Vaško: Effect of humic acids in vivo 

XIII. 

100. Hana Bártíková, Jana Firbasová, Ivan Vokřál, Lenka Skálová, Jiří Lamka, Vladimír 

Kubíček, Barbora Szotáková: BIOTRANSFORMATION OF SELECTED ANTHELMINTICS IN RAT 

tapeworm hyMENOLEPIS DIMINUTA 

101. Iva Boušová, Zuzana Průchová, Lucie Trnková, Jaroslav Dršata: INHIBITORy EFFECT OF 

natural FLAVONOIDS ON eqUINE LIVER GLUTATHIONE S-TRANSFERASE 

102. Lenka Gibalová, Ján Sedlák, Alena Reháková, Martina Labudová, Zdena Sulová, 

Albert Breier: MULTIDRUG RESISTANT P-GLyCOPROTEIN POSITIVE CELLS ARE ALSO CROSS- 

RESISTANT TO CISPLATIN 

103. Veronika Hanušová, Lenka Vildová, Věra Králová, Ladislava Schröterová, Lenka 

Trilecová, Alena Pakostová, Lenka Skálová: THE EFFECTIVENESS OF ORACIN IN ENHANC- 

ING THE cyTOTOXICITy OF DOXORUBICIN THROUGH THE INHIBITION OF DOXORUBICIN 

DEACTIVATION IN BREAST CANCER CELL LINE MCF-7 

104. Věra Kotrbová, Barbora Mrázová, Eva Frei, Marie Stiborová: CyTOCHROME b 5 

POTENtiates 

ACTIVITIES OF cyTOCHROMES P450 1A1 AND 1A2 TO OXIDIZE ANTICANCER DRUG 

ELLIPTICINE TO PHARMACOLOGICALLy EFFICIENT METABOLITES 

105. Věra Králová, Emil Rudolf: SELENITE-INDUCED CELL DEATH IN HUMAN COLON CAN- 

CER CELLS 

106. Jitka Křížková, Kamila Burdová, Petr Hodek, Marie Stiborová: EFFECTS OF FLAVONOIDS 

on cyTOCHROMES P450 AFTER PERORAL SINGLE DOSE ADMINISTRATION TO MALE RATS 

107. Tamara Lasotová, Hana Bártíková, Ivan Vokřál, Barbora Szotáková, Vladimír Kubíček, 

Jiří Lamka, Marián Várady, Lenka Skálová: ACTIVITIES OF DRUG-METABOLIZING AND AN- 

TIOXIDANT ENZyMES IN HEAMONCHUS CONTORTUS STRAINS RESISTANT OR SENSITIVE 

TO ANTHELMINTICS 

108. Kateřina Levová, Jana Šístková, Eva Frei, Volker M. Arlt, David H. Phillips, Heinz H. 

Schmeiser, Marie Stiborová: CyTOCHROMES P450 1A1/2 OXIDIZE CARCINOGENIC ARIStolochic 

ACID I FORMING ITS DETOXICATION METABOLITE AND DECREASING LEVELS OF 

AA-DNA ADDUCTS IN VIVO 

109. Anna Sobeková, Ľuboslava Lohajová, Peter Javorský: THE EFFECT OF BENDIOCARB ON 

antioxidant PARAMETERS IN MALE AND FEMALE ORGANS OF RABBIT 

110. Miroslava Štefanišinová, Mária Kožurková, Vladimíra Tomečková, Mária Mareková: 

INTERACTION OF PLASMID DNA AND MITOCHONDRIA WITH cyCLIC CHALCONE ANALOGUES 



111. Tatiana Matáková, Erika Halašová, Lucia Letková, Anton Dzian, Dušan Dobrota: 

ASSOCIATION POLyMORPHISMS OF GST, HOGG1 AND XRCC1 GENES WITH LUNG 

adenocarcinoma 

112. Jana Mizerovská, Helena Dračínská, Volker M. Arlt, Heinz H. Schmeiser, Eva Frei, Marie 

Stiborová: OXIDATION OF 3-AMINOBENZANTRONE, A HUMAN METABOLITE OF CARCINO- 

GENIC 3-NITROBENZANTHRONE, by HUMAN AND RAT cyTOCHROMES P450 

113. Michaela Moserová, Miroslav Šulc, Volker M. Arlt, David H. Phillips, Eva Frei, Marie 

Stiborová: METABOLIC ACTIVATION OF CARCINOGENIC BENZO[a]pyRENE by cyTOCHROME 

P450 1A1 IS DICTATED by COMPOSITION OF THE MIXED-FUNCTION-MONOOXyGENASE sySTEM 

114. Barbora Mrázová, Eva Martínková, Radek Indra, Eva Frei, Marie Stiborová: THE 

study ON THE cyTOCHROME b 5 

–MEDIATED STIMULATION OF ELLIPTICINE OXIDATION 

by cyTOCHROME P450 3A4 TO ITS PHARMACOLOGICALLy MORE EFFICIENT METABOLITES 

115. Miloslava Netopilová, Libuše Černá, Lucie Škarydová, Vladimír Wsol: IMMUNODETECTION 

of 11β-hyDROXySTEROID DEHyDROGENASE DURING PURIFICATION OF A NEW HUMAN 

MEMBRANE-BOUND CARBONyl REDUCING ENZyME 

116. Radka Podlipná, Petra Šídlová, Kotyza Jan, Tomáš Vaněk: PhyTOREMEDIATION – 

the PROMISING METHOD FOR THE REMOVAL OF PHARMACEUTICAL RESIDUES FROM 

wastewater 

117. Jitka Poljaková, Tomáš Eckschlager, Eva Frei, Marie Stiborová: ELLIPTICINE cyTOTOXICity 

TO HUMAN THyROID CANCER CELL LINES 

118. Alena Rajnochová Svobodová, Adéla Zdařilová, Dana Kylarová, Bohumil Zálešák, Jitka 

Vostálová: qUALITy AND TIME-STABILITy OF HUMAN SKIN EXPLANTS 

119. Anna Sobeková, Katarína Holovská, Peter Javorský: OXIDATIVE STRESS IN TURKEys 

CAUSED by CHRONIC CADMIUM EXPOSURE 

120. Martina Svobodová, Markéta Martínková, Helena Dračínská, Marie Stiborová: 

SIMILARITy BETWEEN RAT AND HUMAN ENZyMES INVOLVED IN OXIDATION 2-NITROPHE- 

NOL, A METABOLITE OF CARCINOGENIC 2-NITROANISOLE 

121. Mário Šereš, Eva Poláková, Oľga Križanová, Zdena Sulová, Albert Breier: OVEREXPRESSION 

of P-GLyCOPROTEIN IN L1210/VCR CELLS IS ASSOCIATED WITH CHANGES IN SEVERAL EN- 

DOPLASMIC RETICULUM PROTEINS 

122. Lenka Umannová, Miroslav Machala, Alois Kozubík, Jan Vondráček: ENVIRONMENTAL 

pollutants AS FACTOR MODULATING THE INFLAMMATORy RESPONSE AND FUNCTIONS 

of LUNG CELLS 

123. Zuzana Vantová, Helena Paulíková, Mária Kožurková, Danica Sabolová, Ima Dovinová, 

Pavol Kristián, Ján Imrich, Ján Ungvarský, Ladislav Janovec: THE MECHANISM OF cyTOTOXIC 

effect OF NOVEL ACRIDINE INTERCALATORS 


29


124. Jiří Vrba, Jitka Ulrichová: RETINOIC ACID-INDUCED DIFFERENTIATION MODULATES 

the APOPTOTIC EFFECT OF SODIUM VALPROATE IN HL-60 CELLS 

POSTER VIEWING 3., SECTION I, VIII, X 

SaTUrday, 11 SEPTEMBEr 2010, 

13.00 - 14.00 

I. 

1. Daniel Čierny, Stanislav Celec, Mária Kovalská, Peter Kaplán, Ivan Ondrejka, Egon Kurča, 

Ján Lehotský: LABORATORy BIOMARKERS IN ISCHEMIC STROKE AND DEPRESSION IN HU- 

MAN PATIENTS 

2. Monika Ďurfinová, Marta Brechtlová, Ľubica Procházková, Peter Kukumberg, Ľubomír 

Kuračka, Branislav Líška: Is IT POSSIBLE TO IMPROVE DEMyELINATION DISEASES MONItoring 

by DETERMINATION OF SOME ENZyME ACTIVITIES CHARACTERISTIC FOR THE 

central NERVOUS sySTEM? 

3. Andrea Evinová, Eva Babušíková, Pavol Adamík, Ivan Ondrejka, Egon Kurča, Milan Grófik, 

Ján Lehotský: SELECTED GENE POLyMORPHISMS IN ISCHEMIC STROKE AND DEPRESSED 

HUMAN PATIENTS FROM CENTRAL SLOVAKIA 

4. Mária Chomová, Peter Račay: An ANALySIS OF THE IMPACT OF CNS ISCHEMIA ON MI- 

TOCHONDRIAL RESPIRATORy COMPLEXES 

5. Mária Kovalská, Martina Pavlíková, Zuzana Tatarková, Peter Kaplán, Dušan Dobrota, 

Marián Adamkov, Ján Lehotský: THE ROLE OF MAP-KINASE PATHWAy IN GLOBAL ISCHEMIA/ 

reperfusion INJURy OF RAT BRAIN AFTER INDUCED hyPERHOMOCySTEINEMIA 

6. Marcela Martončíková, Kamila Lievajová, Juraj Blaško, Judita Orendáčová, Enikő 

Račeková: ANATOMICAL DISTRIBUTION OF dyING CELLS WITHIN ADULT RATS ROSTRAL 

migratory STREAM 

7. Martina Pavlíková, Mária Kovalská, Monika Sivoňová, Zuzana Tatarková, Ján Lehotský: 

SECRETORy PATHWAys SPCA1-Ca 2+ PUMP EXPRESSION AS A PART OF ISCHEMIC PRECON- 

DITIONING IN RAT FOREBRAIN 

VIII. 

58. Katarína Mrvová, Anna Hrabovská: DEVELOPMENT OF A DETECTION TOOL TO FOLLOW 

the SPECIFIC ACTIVITy OF BUTyryLCHOLINESTERASE IN HUMAN PATIENTS 

59. Dominika Neuschlová, Anna Hrabovská: OPTIMALIZATION OF ELLMAN´s ASSAy TO 

study THE KINETICS OF CHOLINESTERASES 


X. 


71. Vladimír Pevala, Jacob A. Bauer, Javier García-Nafría, Gabriela Ondrovičová, Ľuboš 

Ambro, Elena Blagova, Vladimir M. Levdikov, Anthony J. Wilkinson, Keith S. Wilson, Eva 

Kutejová: HEXAMER FORMATION TRIGGERS A SWITCH FROM AN INACTIVE TO AN ACTIVE 

CONFORMATION IN HUMAN MITOCHONDRIAL LON PROTEASE 

72. Milo Bystrický, Martina Beláňová, Mary Jackson, Katarína Mikušová, Jana Korduláková: 

BIOCHEMICAL CHARACTERIZATION OF RV1459C PROTEIN – PUTATIVE GT-C GLyCOSyltransferase 

FROM myCOBACTERIA 

73. Ľubomír Borko, Vladena Bauerová-Hlinková, Eva Hostinová, Juraj Gašperík, Jozef Ševčík: 

THE STUDy OF RyANODINE RECEPTOR 2 N-TERMINAL REGION RESPONSIBLE FOR HEART 

ARRyTHMIAS AND HEART FALIURE 

74. Petronela Dianišková, Jana Korduláková, Henrieta Škovierová, Devinder Kaur, Mary 

Jackson, Patrick J. Brennan, Katarína Mikušová: THE FUNCTIONAL CHARACTERIZATION 

of THE PUTATIVE myCOBACTERIAL ABC TRANSPORTER MSMEG_6366 - MSMEG_6369 

75. Veronika Doubnerová, Lucia Miedzińska, Jana Dobrá, Radomíra Vaňková, Helena Ryšlavá: 

EFFECT OF DROUGHT ON THE METABOLISM OF TOBACCO PLANTS (NICOTIANA TABACUM L.) 

76. Diana Fedunová, Zuzana Flachbartová, Jaroslava Bágeľová, Zuzana Gažová, Marián 

Antalík: THERMAL STABILITy OF cyTOCHROME c AND α-LACTALBUMIN COMPLEXES 

77. Peter Grones, Zuzana Odnogová, Jozef Grones: REP 34 

PROTEIN ENCODE by PLASMID 

pGP2 FROM ACETOBACTER 

78. Hana Kiňová Sepová, Andrea Bilková, František Bilka, Lýdia Bezáková: PRODUCTION OF 

3-hyDROXyPROPIONALDEHyd by THE STRAINS OF LACTOBACILLUS REUTERI 

79. Michaela Koháryová, Marta Kollárová: THIOREDOXIN sySTEM OF STREPTOMyCETES 

80. Mária Kožurková, Danica Sabolová, Slávka Hamuľáková, Jana Janočková, Jana Plšíková, 

Pavol Kristian, Ján Imrich, Ondrej Holas, Miroslav Pohanka, Kamil Kuča: STUDIES OF NOVEL 

bivalent TACRINE DERIVATIVES TARGETING CHOLINESTERASES 

81. Lucia Lichardusová, Jaroslav Kušnír, Mária Mareková: APPLICATION OF CONCENTRATION 

fluorescence MATRICES TO THE DETECTION OF FLUORESCENCE METABOLITES IN URINE 

82. Marián Mazáň, Noelia Blanco, Javier Arroyo, Vladimír Farkaš: CRH TRANSGLyCOSyLASES 

catalyZE INTER-POLyMERIC LINKAGES IN FUNGAL CELL WALL 

83. Ľuboš Nižnanský, Svetlana Kryštofová, Ľudovít Varečka: DELETION OF GLUTAMATE 

decarboxyLASE GENE FROM TRICHODERMA VIRIDE F-534 STRAIN 

84. Helena Ryšlavá, Veronika Doubnerová, Robert Valenta, Kateřina Kloudová, Jana Trefancová, 

Helena Synková, Noemi Čeřovská: CHARACTERIZATION OF β-N-ACETyLHEXOSAMINIDASE 

IN LEAVES OF TOBACCO PLANTS 


31


85. Danica Sabolová, Lucia Krajňáková, Jana Plšíková, Mária Kožurková: DNA BINDING 

study OF 9-OXO-9,10-DIHyDROACRIDINCARBOXyhyDRAZIDES AS A POTENT TOPOISOM- 

ERASE I INHIBITORS 

86. Jana Schubertová Aradská, Dušan Blaškovič, Ján Turňa: IN VIVO cross-LINKING FOR 

IDENTIFICATION OF TELLURITE RESISTANCE-ASSOCIATED PROTEINS 

87. Martin Šimkovič, Anita Gdovinová, Zuzka Zemková, Ľudovít Varečka: MULTIPLE PRO- 

TEASES ARE SECRETED by VEGETATIVE TRICHODERMA VIRIDE myCELIUM CULTIVATED 

WITH PROTEIN INDUCER 

88. Katarína Šipošová, Andrea Antošová, Peter Kutschy, Zuzana Daxnerová, Zuzana Gažová: 

PhyTOALEXINS REDUCE INSULIN AMyLOID AGGREGATION 

89. Barbora Vidová, Michal Chotár, Andrej Godány: THE LysM DOMAIN IN SURFACE IM- 

MUNOGENIC PROTEIN (SIP) AND ITS INFLUENCE ON ELICITATION OF IMMUNITy AGAINST 

STREPTOCOCCUS AGALACTIAE 


BOOK OF ABSTraCTS 


33


PLENarY LECTURES 


35

Plenary lectures 

IDENTIFICATION OF PHOSPHATIDYLGLYCEROL SPECIFIC PHOSPHOLIPASE 

C IN YEAST SACCHarOMYCES CEREVISIAE 

Mária Balážová and Peter Griač 

Institute of Animal Biochemistry and Genetics SAV, Ivanka pri Dunaji 

Phosphatidylglycerol (PG) is a metabolic precursor to the unique anionic mitochondrial 

phospholipid, cardiolipin (CL). CL and PG are phospholipids with important functions in 

promoting cell growth, anaerobic metabolism, mitochondrial functions and biogenesis. 

Considering their important role in eukaryotic cell physiology, little is known about the 

mechanisms by which PG membrane composition is controlled. 

Product of the open reading frame YPL206c, Pgc1p, of the yeast Saccharomyces cerevisiae 

is homologous to bacterial and mammalian glycerophosphodiester phosphodiesterases. 

Deletion of PGC1 causes accumulation of PG, which was evident especially under the 

conditions of inositol limitation. To test if the product of PGC1 has an effect on degradation 

of PG, an in vitro assay was devised. Data obtained from this assay indicated that 

in the strain without Pgc1p, production of NBD-diacylglycerol (NBD-DAG) is significantly 

decreased compared to the wild type strain. In addition, NBD-DAG production was highly 

increased in the strain with overexpression of the Pgc1p. Two localizations of GST tagged- 

Pgs1p were observed: mitochondrion and lipid particles. However, in vitro phospholipase 

C activity of Pgc1 protein was detected only using mitochondrial protein extract. Based 

on these results we suggest that the product of YPL206c encodes mitochondrial PG 

specific phospholipase C (Pgc1p) involved in regulation of PG levels. 

Acknowledgement: Work was supported by LPP-0291-09 and VVCE-0064-07 grants. 



MICROBIAL XYLANASES: PROPERTIES AND APPLICATIONS 

Peter Biely 

Institute of Chemistry, Center of Glycomics, Slovak Academy of Sciences, Bratislava 

Considerable attention of current research is devoted to development of environmentally 

friendly processes for utilization of renewable resources. This effort includes also 

bioconversion of the major plant hemicellulose, xylan, after cellulose, the second most 

abundant polysaccharide in nature. Xylan is a heteropolysaccharide with a main chain 

built of b-1,4-liked xylopyranosyl residues. Depending on a plant source the main chain is 

decorated with uronic acids, arabinofuranose or esterified with acetic acid. Decomposition 

of xylan in nature by microorganisms is a part of the carbon cycle and involves concerted 

action of several enzymes. The enzymes attacking the xylan main chain are the depolymerizing 

endo-b-1,4-xylanases and xylose-releasing b-xylosidases. The acetyl groups and 

carbohydrate substituents of the main chain and are liberated with so called accessory 

enzymes. The group led by the author contributed significantly to current knowledge 

on the production of xylanolytic enzymes, mode of their action, substrate structure 

requirements and diversity of endoxylanases and xylosidases. Important impact had 

the discovery of hemicellulolytic deacetylases and introduction of efficient assays of 

xylanolytic enzymes. Partial amino acid sequences of novel accessory enzymes enabled 

isolation of the corresponding genes, their expression and the search for homologous 

sequences in known microbial genomes. This work resulted in establishment of new 

glycoside hydrolase and carbohydrate esterase families (http://www.cazy.org) with important 

synthetic and biotechnological potential. Microbial enzymes hydrolyzing xylan 

to oligosaccharides and fermentable sugars, and decreasing viscosity of xylan solutions 

became important industrial enzymes. They found applications in the pulp and paper 

industry, food industry and animal feed. 


37


STRUCTUraL-FUNCTIONAL COrrELATIONS OF HYDROXYMETHYLBILANE 

SYNTHASE 

Dana Douděrová and Pavel Martásek 

Department of Pediatrics, 1 st Faculty of Medicine, Charles University in Prague 

Acute intermittent porphyria (AIP) is an autosomal dominantly inherited disorder, classified 

as acute hepatic porphyria. It is characterized by a deficiency of hydroxymethylbilane 

synthase (HMBS, EC 4.3.1.8), the third enzyme in heme biosynthesis. Clinical features 

include gastrointestinal, neurologic and cardiovascular symptoms, but the most common 

clinical presentation is abdominal pain caused by neurovisceral crises. 

The purpose of this study was first to perform molecular analysis of the AIP patients. In 

each affected family, this becomes an important tool for individualised medicine, allowing 

for careful drug prescription; in addition, it is very important for the asymptomatic 

carriers to be warned of precipitating factors, thus avoiding an acute attack. 

The proper DNA diagnostics can be achieved by a combination of a robust and effective 

pre-screening method and a confirmatory DNA sequencing step. We decided to 

establish a new generation pre-screening method, which will be highly sensitive and 

relatively time- and cost-effective. Our method of choice was high-resolution melting 

(HRM) analysis using the LightScanner instrument. 

Another important aspect of this project was to study the molecular heterogeneity of 

AIP in relation to the HMBS protein. We aimed at characterisation of the impact of the 

HMBS gene mutation on the structure and function of the enzyme, and demonstration 

of how this aids the interpretation of clinical, biochemical and genetic data in establishing 

an AIP diagnosis. To demonstrate this, we used expression and characterisation of 

mutant HMBS enzymes in the prokaryotic system together with the use of predictive 

computer-assisted structure-function correlation studies. 



“BIOMacrOMOLECULar INTEraCTIONS ON ELECTrICaLY 

readaBLE MICrOCHIPS” 

Mathias Sprinzl 

Laboratorium für Biochemie, Universität Bayreuth, Germany 

Transduction of specific biochemical interactions to electrically readable signals is the 

main objective of the investigations. The aim is to develop analytical devices (biochips) 

for use in diagnostics, biotechnology and environmental analysis. 

Specific biomacromolecular interactions direct a reporter enzyme (1) for binding to gold 

electrodes where an electrochemically detectable molecule is enzymatically synthetized. 

Biomacromolecular reactions used for electrically readable biochips can work on different 

principles e.g. DNA/DNA, DNA/protein, protein/protein or RNA/protein interactions. 

RNA-aptamers, ribozymes and riboswitches can be also used for construction of biochips 

and used for sensitive and simple identification of whole cells, proteins, metabolites and 

small organic molecules with hand-hold, electrically readable, instruments. Examples 

for detection of bacteria by hybridisation with 16S RNA, amino acid analysis in physiological 

fluids, detection of toxic substances in water and analysis of micro RNA (2,3) 

will be presented. 

Supported by the Deutsche Forschungsgemeinschaft Sp 243/1-1/2, Forschungskreis der 

Ernährungsindustrie AiF 230 ZN and Siemens AG-CT, Erlangen, Germany 

1) Wang Y, Stanzel M, Gumbrecht W, Humenik M, Sprinzl M. (2007)Esterase 

2-oligodeoxynucleotide conjugates as sensitive reporter for electrochemical detection of nucleic 

acid hybridization. Biosens Bioelectron. 15, 1798-806. 

2) Pöhlmann C, Wang Y, Humenik M, Heidenreich B, Gareis M, Sprinzl M. (2009)Rapid, specific 

and sensitive electrochemical detection of foodborne bacteria. Biosens Bioelectron. 15, 2766-71. 

3) Humenik M, Pöhlmann C, Wang Y, Sprinzl M. (2008) Enhancement of electrochemical signal on 

gold electrodes by polyvalent esterase-dendrimer clusters. Bioconjug Chem. 19, 2456-61. 


39

LECTURES 


Lectures 

PROCESSING OF AMYLOID PRECURSOR PROTEIN afTER 

IN VIVO INDUCED ISCHEMIA 

Eva Babušíková 1 , Dušan Dobrota 1 , Anthony J. Turner 2 and Natalia N. Nalivaeva 2 

1 

Department of Medical Biochemistry, Comenius University in Bratislava, Jessenius 

Faculty of Medicine in Martin, Martin, Slovakia, 2 Institute of Molecular and Cellular 

Biology, University of Leeds, Leeds, United Kingdom 

Ischemia stroke results from a transient or permanent reduction in cerebral blood flow. 

In recent years it has been suggested that neurological disorder in elderly human population 

as Alzheimer’s disease (AD) is linked to certain brain pathologies, which promote its 

development and progression via accumulation of toxic amyloid peptide (Aβ) deposits 

in the brain. In the present study we determined the effect of the global ischemia (the 

four-vessel occlusion model) on the amount of amyloid precursor protein (APP) and 

some amyloid peptide degrading metalloproteinases. We observed that ischemia result 

in increased amyloidogenic processing of APP in hippocampus and cortex as well. Levels 

of APP increased significantly after ischemia as well as the amount of sAPPPβ soluble 

fragment produced by APP cleavage by β-secretase (BACE). Levels of BACE were significant 

increase. Amounts of Aβ degrading enzymes neprilysin and endothelin-converting enzyme 

decreased significantly after ischemia. We observed oxidative damage after ischemia. 

Oxidative modifications of proteins were demonstrated by significant accumulation of 

dityrosines and formation of lysine conjugates with the lipid peroxidation end products. 

After ischemia levels of conjugated dienes increased significantly. Concentrations of free 

sulfhydryl groups and thiobarbituric acid-reactive substances did not change during 

ischemic insult. Our results suggest that global ischemia may lead to amyloid peptide 

deposits accumulation and promote Alzheimer’s disease, which in turn might induce 

protein and lipid oxidation and reactive oxygen species formation. 


41

Lectures 

LIPID HELICES formaTION IN BaCILLUS SUBTILIS CELL MEMBraNE 

Imrich Barák, Katarína Muchová, Nada Pavlendová and Ján Jamroškovič 

Institute of Molecular Biology, Slovak Academy of Sciences, Dúbravská cesta 21, 

845 51 Bratislava, Slovakia 

The domains of different lipid composition are present in eukaryotic and prokaryotic cell 

membranes. Using membrane binding fluorescent dyes, we demonstrate previously, the 

presence of lipid spirals extending along the long-axis of cells of the rod-shaped bacterium 

B. subtilis. These data indicate a higher level of membrane lipid organization than 

previously observed. Little is known however of the origin of these helical structures. 

Principally, there are at least three main specifically localized molecular structures in 

the membrane or close proximity to it what can help to form or influence the formation 

of lipid helixes. In our work we have focused on analyzing these lipid structures in correlation 

with other above mentioned helical structures in the cell membrane or its close 

proximity. We were analyzing lipid domains by using lipid specific dyes in protoplasted 

cells, in Mbl, MreB and MreBH mutant strains. We have used FRAP and FRET experiments 

to determine dynamics of lipid domains and co-localization of lipid dyes with GFP fused 

proteins, respectively. 

We have also studied the role of lipid helices in cell division by directing the Min system 

to the helices from pole to pole. We inspected cell division when E. coli Min-system was 

introduced into B. subtilis cells. We show that MinD Ec 

can partially substitute function of 

its B. subtilis protein counterpart. Additionally, we observed dynamic behavior of MinD Ec 

and MinE in B. subtilis when expressed together. All these findings indicate that these 

two Min systems resemble each other more than was thought previously 


Lectures 

effECTS of DOXOrUBICIN treaTMENT ON MatrIX 

METaLLOPrOTEINaSES IN raTS 

Miroslav Barančík, Petra Šimončíková and Monika Ivanová 

Institute for Heart Research CEKVY SAS, Bratislava 

The anthracycline doxorubicin (DOX) is an effective chemotherapeutic agent which is 

frequently used in the treatment of many types of malignancies. Limitation of its use is 

a cardiotoxicity associated with the development of cardiomyopathy and chronic heart 

failure. Matrix metalloproteinases (MMPs) are enzymes that play an important role in 

degradation and remodeling of extracellular matrix under physiological and pathological 

conditions. Especially MMP-2 and MMP-9 are suggested to play an important role also in 

pathogenesis of several cardiovascular diseases. The aim of the study was to investigate 

the involvement of MMPs in the responses of rats to prolonged doxorubicin treatment. 

In the study, male Wistar rats were used. DOX was administered to rats by intraperitoneal 

injections of 7 doses in 3-day’s intervals (total cumulative dose of DOX was 15 mg 

per kg of body weight). The control animals were treated with saline. The samples of 

tissue or plasma were collected 4, 8 and 12 weeks after application of last dose of DOX. 

The protein levels were determined by immunoblot assay and MMPs activities were 

measured by gelatin zymography. Determined were also blood pressure, body weight 

and weight of several organs (heart, brain, liver, kidney) and the parameters obtained 

in DOX-treated rats were compared with parameters of control animals. The investigation 

of changes associated with action of DOX revealed that prolonged exposure of rats 

to DOX led to changes in MMPs activation in heart tissue. Moreover, the effects of DOX 

were connected with time-dependent changes in plasma MMPs activities. Our results 

suggest that MMPs are involved in the responses of rat hearts to chronic DOX treatment. 

Acknowledgement: Supported by VEGA SR 2/0205/09, APVV 51-027404 


43

Lectures 

SYNTHESIS OF GLCNaC-TS MIMETICS AS a POTENT INHIBITORS OF 

GLYCOSYLTraNSFEraSES 

Marek Baráth, Igor Tvaroška and Ján Hirsch 

Institute of Chemistry, Slovak Academy of Sciences, Dúbravská cesta 9, 845 38 

Bratislava, Slovakia 

Glysosyltransferases are enzymes that catalyst transfer of monosacharidic unit from an 

activated sugar phosphate to an acceptor molecule, usually an alcohol. The result of 

glycosyl transfer can be a monosaccharide glycoside, an oligosaccharide or polysaccharide. 

Many functions have been implicated for protein glycosylation, including promoting 

protein folding or stabilizing cell-surface glycoproteins. 

Different strategies have been used in order to identify potent inhibitors of glycosyltransferases. 

The main goal of this contribution is to synthesized of the transition state 

(TS) analogs starting from the donor UDP-GlcNAc. A leading idea of all these TS analogs 

is a „ 1-thio“ linker between a mimetic of GlcNAc in TS geometry and a mimetic of the 

acceptor bearing the β-D-psicofuranose, β-D-tagatofuranose and backbones with the 

key substitution on position C-1 by the phosphate group and on position C-2 by the 

thiophenyl group, which has been designed. 

Couple of potential inhibitors have been synthesized bearing β-D-psico and β-D-tagato 

configuration using a multi step synthesis. The β-D-psico analogs were as well as utilized 

for the synthesis of N-acetylated β-D-fructo analogs as their epimers in three more steps 

using a Walden inversion at C-3 position. 

Acknowledgement: This work was partially supported by the grants: VEGA 2/6129/27, 

2/0128/08 and Centre of Excelence- GLYCOMED. 


Lectures 

CHANGES IN NEURONAL METABOLITES MEASURED BY PROTON 

MAGNETIC RESONANCE SPECTROSCOPY IN DEPRESSED PATIENTS 

DURING TREATMENT 

Michal Bittšanský, Veronika Husárová, Igor Ondrejka, Valéria Kerná, Pavol Adamík, 

Hubert Poláček and Dušan Dobrota 

Jessenius Faculty of Medicine, Comenius University, Martin, Slovakia 

Previous works have shown that the symptoms of depressive disorder can be correlated 

to the concentrations of proton MR metabolites. In our study, 20 depressive patients 

were at the admission to the hospital and after the hospital treatment clinically examined 

using MADRS scale (The Montgomery-Åsberg Depression Rating Scale) and using 1 H 

MR-spectroscopy (1.5 Tesla, single-voxel spectroscopy) in both hippocampi. We evaluated 

the absolute and relative signals of N-acetyl aspartate (NAA), total creatine (Cre), 

total cholines (Cho), myo-inositol (mI) measured with short (30 ms) and long (135 ms) 

time echo using calibrated LCModel. We observed statistically significant correlations of 

MRS-observable metabolites to the clinical MADRS score, and also significant differences 

between the patient groups before and after treatment. Some of the changes seem to 

reflect changes in the relaxation times of the metabolites. 

Acknowledgement: This work was supported by the grant 2007/57-UK-17 of Slovak 

Ministry of Health and by project “CREATING A NEW DIAGNOSTIC ALGORITHM FOR 

SELECTED CANCER DISEASES” co-financed from EU sources and European Regional 

Development Fund. 


45

Lectures 

TOXCAT METHOD: APPLICATION IN MOLECULar ONCOLOGY 

Martin Benej 1 and Martina Poturnajová 2 

1 

Department of Molecular Biology UK Bratislava, 

2 

Cancer Research Institute SAS Bratislava 

Understanding the molecular mechanisms of cancer onset is one of the goals of contemporary 

cancer research. Individual approach to each disease is of crucial importance 

in this issue. Moreover, not only each disease, but also each patient carrying causative 

mutation in his/her genome requires individual approach. Thus, a need of a whole set 

of methods for characterization of causative mutations has arisen. ToxCAT is a method 

simulating the natural lipid bilayer environment, enabling the study of transmembrane 

(TM) domain interactions in vitro. The method is based on introduction of chimaeric constructs 

containing a TM domain of interest into periplasmic region of E.coli MM39 strain. 

Interactions of the chimerae result in expression of chloramphenicol acetyltransferase 

(CAT) reporter gene. The quantity of CAT expression corresponds with the strength of TM 

domain association. We illustrate the ToxCAT method application on the specific model 

of our interest – Medullary thyroid carcinoma (MTC). MTC is caused predominantly by 

single point mutations in six RET proto-oncogene exons. RET protein, the product of the 

RET gene is a tyrosine kinase cell-surface receptor. Mutations in extracellular domain of 

the receptor result in dimerization of the RET protein with other mutant RET molecule, 

thus enabling ligand-independent permanent activation of the RET receptor kinase. For 

this interaction, the strength of transmembrane domain oligomerization of the two RET 

molecules is responsible. We focus on the impact of RET TM domain mutations of Slovak 

MTC patients on the strength of TM domain oligomerization, to investigate a possible 

correlation with the age of onset and aggressiveness of the disease. 


Lectures 

HIGH affINITY carBOHYDraTE aND NON-carBOHYDraTE LIGaNDS 

for LECTIN-TYPE aCTIvaTION rECEPTOrs of naTUral KILLEr CELLS 

rEGULaTE effECTOr fUNCTION THrOUGH PI3K paTHWay, aND 

GENEraTE PErmaNENT IMMUNE prOTECTION agaINST MELaNOMas 

Veronika Benson 1 , Valeria Grobárová, 1 Katarína Hulíková 1 Jan Svoboda 1 , 

Daniel Rozbeský 1,2 , Daniel Kavan 1,2 , Alan Kádek 1,2 , Karel Křenek 1 , Anna Fišerová 1 , 

Vladimír Křen 1 and Karel Bezouška 1,2 

1 

Institute of Microbiology v.v.i., Academy of Sciences of Czech Republic and 

2 

Department of Biochemistry, Faculty of Science, Charles University Prague, Praha, 

Czech Republic 

Our laboratories are interested in understanding of complex interactions between activation 

receptors of natural killer (NK) cells, their target structures at tumor cell surface, and 

intracellular activation pathways resulting in the activation of NK cell effector functions at 

molecular and cellular level. To identify high afinity ligands, we produce stable recombinant 

soluble forms of NK cell receptors such as NKR-P1, CD69, and NKG2D and use them 

in binding, inhibition and precipitation studies based on standard biochemical assays and 

oligosaccharide arrays. High affinity ligand mimetics are constructed by attachment of the 

active compounds to polyamidoamine or calix arene cores, or by dimerization of the ligand 

through a defined chemical linker. GlcNAc-coated polyamidoamine dendrimers induce 

upregulation of antibody formation that triggered by their interaction with mNKR-P1C. 

GlcNAc-coated calyx arene downregulated the expression of GlcNAc transferases MGAT3 

and MGAT 5, increased the susceptibility of tumor cells to natural killing, and increased 

the expression of mNKG2D through the activation of PI3K–ERK but not phospholipase C-γ- 

JNK pathway. GlcNAc dimers can provide permanent protection in 70 % of mice bearing 

syngeneic B16S melanomas. This is due to activation of NKT lymphocytes, and subsequent 

infiltration of tumors by CD8 + cytotoxic lymphocytes. The exceptional signaling efficiency 

of GlcNAc dimers is explaned by sequential cooperative engagement of mNKR-P1A leading 

to the formation of large signaling complexes of about 20 MDa containing G proteins, ß- 

arrestin, phosphorylated dynamin, Src kinase, Vav, Rac1, Grb2, and Ras. Use of combined 

ligand mimetics results in engagement of several target receptors, and efficient activation 

of NK cell effector functions due to effective receptor cross-talk. 

Supported by grants by Ministry of Education of Czech Republic (MSM_21620808 

and 1M0505), by the Institutional Research Concept for the Institute of Microbiology 

(AVOZ50200510), by Czech Science Foundation (303/09/0477 and 305/09/H008), Grant 

Agency of Academy of Sciences of the Czech Republic nebo (ASCR) IAA500200620, and by 

the European Commission (Project Spine 2 Complexes, contract LSHG-CT-2006-031220). 


47

Lectures 

PROTEOMICS OF MULTIFUNCTIONAL ROYAL JELLY PROTEINS 

Katarína Bíliková and Jozef Šimúth 

Department of Molecular Apidology, Institute of Molecular Biology, 

SAS, Bratislava 

Presented study demonstrated how neofunctionalization results from various posttranslational 

modifications of maternal proteins of honeybee royal jelly (RJ). We have purified 

a minority protein of RJ, named apalbumin2a. Characterization of apalbumin2a by LC-MALDI 

TOF/TOF MS revealed it as a homologue of major basic royal jelly protein apalbumin2, 

carrying two fully occupied N-glycosylation sites, one with high-mannose structure, 

HexNAc2Hex9, and other carrying complex type antennary structures, HexNAc4Hex3 and 

HexNAc5Hex4, while the maternal protein, apalbumin2, contained only high-mannose 

N-linked glycans. We have found that apalbumin2a inhibit growth of Paenibacillus 

larvae, the primary honeybee pathogen of American foulbrood disease, similarly to RJ 

peptide royalisin. In spite of a single gene in honeybee genome for apalbumin2, presence 

of various forms of the protein, having different N-terminal sequences could be 

a result of specific proteolytic degradation of mature protein, alternative splicing or 

heterogonous transcription start sites by „leakage“ of the RNA transcription machinery. 

Obtained data call attention for functional plasticity of RJ proteins with potential impact 

on fundamental research, namely studies of novel mechanisms of action of antibacterial 

proteins, as well as on the field of drug development and therapeutic application of RJ 

proteins as antibiotics. 

Acknowledgments: This work was supported by Max-Planck Society for Partner Group 

of Slovak Academy of Sciences and by 6RP EU-BeeShop No.: 022568. 


Lectures 

FREE raDICAL SITUATION IN PIGMENT CELLS 

Jan Borovanský, Adéla Lipšová and Jiří Vachtenheim 

Institute of Biochemistry & Experimental Oncology, 1 st Faculty of Medicine, 

Charles University, Prague 

Biochemical specificity of pigment cells consists in their capacity to synthesize specific 

metabolic products – cytoprotective eumelanins and cytotoxic phaeomelanins in the 

process of melanogenesis. Melanogenesis represents a potential threat for the pigment 

cell because the intermediates belong to cytotoxic species – quinones, semiquinones and 

the synthesis of melanins is accompanied by the production of superoxide anions and 

H 2 

O 2 

. For that reason melanogenesis is strictly compartmentalized to melanosomes. The 

free radical situation is quite complex because superoxide anions are tyrosinase substrate 

and melanin polymer behaves as a pseudosuperoxide dismutase producing H 2 

O 2 

. In 1991 

we demonstrated the presence of aberrant melanosomes with membrane defects (with 

subsequent leakage of cytotoxic species) as a common phenomenon in melanoma cells. 

Pigment cells are protected by scavenging mechanisms (a) intramelanosomal binding of 

cytotoxic species to proteins which can be illustrated by our finding of protein-bound dopa 

in melanosomal proteins; b) conversion of quinones into adducts with cysteine and GSH 

into cysteinyldopa which is excreted via urine; c) prevention of diphenol conversion into 

quinones by COMT. If the capacity of scavenging mechanisms is overcome, pathological 

reactions ensue which can be exploited in melanoma therapy: a) Trojan horse approach = 

administration of tyrosine and DOPA analogues that are converted by tyrosinase specifically 

in pigment cells to cytotoxic molecules; b) inhibition of scavenging mechanisms = using 

COMT inhibitors we were able to inhibit proliferation of melanoma cells in vitro but not 

in vivo. Tumour proliferation is often free radical burden for the host. To our surprise the 

growth of B16 and S91 melanomas in mice and MeLiM melanoma in minipigs was not accompanied 

with signs of free radical damage. For that reason we compared the activities 

of antioxidant enzymes in tumour cells between 7 human melanoma cell lines and human 

osteosarcoma, glioma, colorectal carcinoma, lung carcinoma and neuroblastoma cell lines. 

The comparison showed significantly higher (p=0,004) catalase activity in melanoma lines 

compared to nonmelanoma lines, whereas there were no significant differences as for 

glutathione peroxidase, SOD and γ-glutamyltransferase. The total antioxidant status (TAS) 

of melanoma cells was also significantly higher than in nonmelanoma cells (p=0.004). 

Correlation between catalase activity and TAS (R=0.909, p level = 0,00004) confirmed that 

the defense of melanoma was based on catalase activity. Taking into account the role of H 2 

O 2 

in cell proliferation, angiogenesis, invasion and metastasizing, apoptosis, the manipulation 

of catalase can be promising tool in experimental melanoma therapy. 

Ackowledgement: Supported by VZ MSM 21620808. 


49

Lectures 

BIOMarKErs of LYMPH NODE METaSTaSIS IN LOW-graDE breaST 

caNCEr: aN INTEGraTED, prOTEOMICS-baSED aPProaCH 

Pavel Bouchal 1,2 , Monika Mudrochová 1,2 , Eva Budinská 3 , Zbyněk Bortlíček 3 , 

Iva Struhárová 1,2 , Lenka Hernychová 4 , Theodoros Roumeliotis 5 , Spiros D. Garbis 5 , 

Roman Hrstka 1 , Petr Müller 1 , Rudolf Nenutil 1 and Bořivoj Vojtěšek 1 

1 

Masaryk Memorial Cancer Institute, Brno; 2 Masaryk University, Faculty of Science, Brno; 

3 

Masaryk University, Institute of Biostatistics and Analyses, Brno; 

4 

University of Defence, Faculty of Military Health Sciences, Hradec Králové, 

5 

Academy of Athens, Greece 

Our effort has been focused on biomarker discovery in the set of 96 breast cancer tumors 

divided into groups according to grade and presence or absence of lymph node metastases. 

Three proteomics approaches were involved in complex protein analysis of tissue 

lysates: (i) SELDI-TOF MS enabled us to quantify 130 intact protein peaks. One protein 

peak correlating with lymph node metastases was detected and identified. (ii) Selected 

set of tumors was analyzed using iTRAQ-2DLC-MS/MS approach. This approach provides 

a simultaneous identification and quantitation of more than 600 proteins. Differentially 

expressed proteins belong to several functional groups which will be described in the 

presentation. (iii) Set of selected tumor lysates was analyzed also using 2-D SDS-PAGE. 

Additionally, the expression variability of 26 selected gene products was evaluated using 

qRT-PCR. 

Generally, our results indicate the differential expression of (i) cytoskeletal proteins, (ii) 

anterior gradient protein family members and (iii) proteins involved in heme biosynthesis 

between the two studied groups. 

Acknowledgments: This work was supported by Czech Science Foundation (Project No. 

304/10/0868), by Czech Ministry of Health (Project No. MZMOU2005) and by Czech 

Ministry of Education (MSM0021622413). 


Lectures 

DOES EXISTS ANY RELATION BETWEEN P-GLYCOPROTEIN MEDIATED 

MULTIDRUG RESISTANCE AND INTraCELLULar CALCIUM HOMEOSTASIS 

Zdena Sulová 1 , Mário Šereš 1 , Miroslav Barančík 2 , Lenka Gibalová 1 , Branislav Uhrík 1 , 

Lenka Poleková 1 and Albert Breier 1 

1 

Institute of Molecular Physiology and Genetics, Centre of Excellence of the Slovak 

Research and Development Agency „BIOMEMBRANES2008”, Slovakia, 2 Institute 

for Heart Research, SAV, Bratislava 

Multidrug resistance (MDR) of neoplastic tissue represents real obstacle in effective 

chemotherapy of cancer. Several mechanisms of MDR were identified, from which 

over-expression and efflux activity of P-glycoprotein (P-gp) – plasma membrane ATPase 

(ABCB1 member of ABC transporter family) – represent most common observed reason 

of neoplastic diseases chemotherapy malfunction. Process of P-gp mediated MDR 

seems to be related to intracellular calcium homeostasis at least indirectly because: i. 

substances blocking calcium influx through L-type of calcium channels like verapamil 

were often found to antagonize P-gp mediated MDR; ii. calcium signal abnormalities 

were observed in cells over-expressing P-gp; iii. cells with P-gp mediated MDR were often 

resistant to thapsigargin; iv. several differences in intracellular calcium localization were 

observed when P-gp negative and P-gp positive cells were compared; v. differences in 

contents of several proteins of endoplasmic reticulum involved in calcium homeostasis 

were observed to be associated with P-gp over-expression. Current study represents an 

attempt to summarize knowledge about possible relations between P-gp mediated MRD 

and intracellular calcium homeostasis. 

Acknowledgments: This work was supported by: APVV-0084-07, VVCE-0064-07, VEGA- 

2/0123/10, VEGA-2/0155/09 


51

Lectures 

OSTERIX OVER-EXPRESSION IN HUMAN EMBRYONIC STEM CELLS AND 

ITS EffECT ON CELL DIffERENCIATION 

Radim Černý 1 , Elerin Kärner 2 , Christian Unger 3 and Mikael Wendel 2 

1 

Department of Biochemistry, LFUK Plzeň, 2 Center for Oral Biology and 3 Department 

of Medicine, Karolinska Institutet, Stockholm, Sweden 

Osterix (Osx) is a recently identified zinc finger-containing transcription factor encoded 

by the Sp7 gene, which regulates gene expression in committed osteoblastic precursor 

cells, acting downstream of Runx2 (Nakashima et al.: Cell 108, 143, 2002). We have overexpressed 

Osx after lentiviral transfer of Osx cDNA recombined with enhanced green 

fluorescent protein (EGFP) into the genome of human embryonic stem cells (HESC) line 

H9. We obtained two HESC subpopulations expressing two significantly different levels 

of Osx. Both subpopulations exhibited spontaneous differentiation and reduced expression 

of markers characteristic of the pluripotent phenotype, such as SSEA3, Tra I-60, and 

Nanog. The high level of Osx expression, compared to endogenous levels found in primary 

human osteoblasts, did not enhance osteogenic differentiation, and did not up-regulate 

collagen I expression. Instead, the high Osx levels induced the commitment towards 

the hematopoietic-endothelial lineage by up-regulating the expression of CD34 and 

Gata I. However, low levels of Osx expression up-regulated collagen I, bone sialoprotein 

and osteocalcin production. Conversely, forced high level expression of the homeobox 

transcription factor HoxB4, a known regulator for early hematopoiesis, promoted osteogenesis 

in HESCs, while low levels of HoxB4 lead to hematopoietic gene expression. We 

conclude that for an enhanced osteogenesis originating from in vitro cultured HESCs, 

the correct levels of ectopic transcription factors need to be established. Our data also 

highlight the notion of close relationship between early blood and bone development. 


Lectures 

HISTORY AND PRESENT OF SCIENTIFIC AND PEDAGOGIC CONFERENCES 

OF TEACHERS frOM CHEMICAL INSTITUTES AND DEParTMENTS OF 

SLOvaK AND CZECH MEDICAL faCULTIES 

Zdeňka Ďuračková and Zuzana Országhová 

Institute of Medical Chemistry, Biochemistry and Clinical Biochemistry, 

Medical School, Comenius University, Bratislava, Slovakia; 

The regular meetings of Slovak and Czech teachers from institutes and departments 

providing education of chemical and biochemical disciplines at the Medical faculties 

are organized more than 50 years. The first meeting of teachers assembled by Prof. A.F. 

Richter in Prague was already in 1952. 

Primarily these meetings were focused only to the problems of education of chemistry and 

biochemistry. The way of interviewing and entrance tests, level of candidates, postgradual 

study as well as contemporary methods of education (e-learning) are also discussed. 

In the last years our conferences are also place for presentation of research activities 

of institutes. The discussions at the meetings and reciprocal deal with practical experiences 

are the big contribution to the work of teachers, scientists and doctorands of 

participated institutes. 

From 20 th to 21 st May 2010 pedagogical conference of Slovak and Czech chemical and biochemical 

teachers was organized in Modra by Institute of Medical Chemistry, Biochemistry 

and Clinical biochemistry, Medical School, Comenius University in Bratislava. This conference 

gave the chance to young colleagues to present the scientific programmes of 

individual institutes as well as parts of their own research. 


53

Lectures 

MAGNETIC RESONANCE SPECTROSCOPY IN DIAGNOSTIC PROTOCOL OF 

THE BraIN DISEASES 

Dušan Dobrota and Michal Bittšanský 

Jessenius Faculty of Medicine in Martin, Comenius University in Bratislava, 

Slovak Republic 

Magnetic resonance spectroscopy (MRS allowed study of some biochemical changes, 

and metabolic pathway in vitro and in vivo. MRS is a physical technique that has been 

used as an analytical method, predominantly by chemists to describe the structure of 

molecules in a specific solution. In biological application the method allowed to study 

low molecular compounds containing atoms that have magnetic properties (e.g. 1 H, 31 P, 

13 

C, 19 F etc..Proton magnetic resonance spectroscopy ( 1 H MRS) can measure levels of 

cerebral metabolites with the low molecular weight such as N-acetylaspartate (NAA), 

choline (Cho), creatine (Cre), lactate (Lac) and some others. 1 H MRS application in vivo in 

the diagnostic protocol some brain diseases (brain tumors, epilepsy, neurodegenerative 

diseases and schizophrenia) was the study’s focus. In vivo magnetic resonance spectra 

were obtained from the different parts of the brain using clinical scanner Siemens 

Symphony (1,5T) and standard protocols. 1 H magnetic resonance spectroscopy (MRS) 

provides valuable information about the changes in the concentration of above mentioned 

metabolites and their ratios, which are typical for each brain diseases. The great 

advantage of MRS in vivo study is that we are allowed study of these biochemical events 

in real time without any disturbance of the tissue. We may conclude, that in vitro 1 H MRS 

provides good information about biochemical differences in various type of human brain 

pathologies and could be used in the standard diagnostic protocol. 

Acknowledgement: This work was supported by the Ministry of Education Grant 048- 

010UK-8/2008 and by project “CENTER OF TRANSLATIONAL MEDICINE” co-financed from 

EU sources and European Regional Development Fund. 


Lectures 

EffECT of DOMaIN PEPTIDES OF THE carDIac ryaNODINE rECEPTOr 

ON THE STaBILITY of BILAYER LIPID MEMBraNES 

AND ON rYr2 ACTIVITY 

Andrea Faltinová 1 , Jana Gaburjáková 1 , Ľubica Urbániková 2 , Matúš Hajduk 2 , 

Nataša Tomášková 3 , Marián Antalík 3 and Alexandra Zahradníková 1 

1 

Institute of Molecular Physiology and Genetics SAS, Bratislava, Slovakia, 

2 

Institute of Molecular Biology SAS, Bratislava, Slovakia, 

3 

Institute of Experimental Physics SAS, Košice, Slovakia 

The cardiac ryanodine receptor (RyR2) contains one N-terminal, one central and two 

C-terminal domains where mutations related to the cardiac arrhythmia, CPVT, tend to be 

clustered. It is assumed that interaction between the N-terminal and the central domain 

plays a role in forming the “domain switch” that regulates the stability of the resting 

(closed) state of the RyR2. The aim of our study was to test whether mutation-prone 

regions of the RyR2 suppress the stability of the closed conformation. 

We constructed two peptides, DPcpvtN2 and DPcpvtC, corresponding to the N-terminal 

and central part of the RyR2 with the highest occurrence of CPVT mutations. We examined 

their effect on the resting activity of the RyR2. DPcpvtC (20 – 30 M) moderately 

increased the RyR2 open probability, in accordance with the hypothesis. However, before 

an effect on the RyR2 activity could be observed, DPcpvtN2 interacted with the BLM. In 

the concentration range of 0.5 – 2.0 M the peptide perforated the BLM regardless of 

the presence of the RyR2. Secondary structure analysis of DPcpvtN2 using bioinformatics, 

CD-spectroscopy and mapping on the known tertiary structure of the IP3R ligand-binding 

domain that is homological with the distal part of the N-terminal domain has shown 

a high incidence of αhelix (45 - 76 %) as well as ascending hydrophobicity gradient in 

the DPcpvtN2. These properties might explain the observed effect of DPcpvtN2 on the 

stability of BLM. 

Supported by grants APVV-0139-06, APVV-0441-09, VEGA 02/0190/10 and by the European 

Union Contract No. LSHM-CT-2005-018833/EUGeneHeart. 


55

Lectures 

TraNSGLYCOSYLATION - a UNIVERSAL PRINCIPLE IN TAILORING 

THE PLANT AND FUNGAL CELL WALLS 

Vladimír Farkaš 

Institute of Chemistry, Center for Glycomics, Slovak Academy of Sciences, Department 

of Glycobiology, Dúbravská cesta 9, 84538 Bratislava, Slovakia 

Plant and fungal cell walls are composite structures composed of polysaccharides and 

protein-polysaccharides mutually cross-linked by non-covalent interactions and covalent 

bonds. Individual wall polymers are being synthesized separately, either intracellularly 

or at the plasma membrane and exported into the cell wall. The final stage of cell wall 

formation involves the formation of cross-links between the individual polymer molecules, 

either of the same or of the diverse types. The enzymes catalyzing the latter 

type of reactions are transglycosylases. They are either GPI-anchored to the plasma 

membrane or embedded in the cell wall. As an example from the plant kingdom, the 

enzyme xyloglucan endotransglycosylase (XET) will be presented. The enzyme catalyzes 

cleavage of xyloglucan molecules and transferring the cleaved fragments to other xyloglucan 

molecules in the plant cell walls. Transglycosylases operate also in the fungal 

cell walls. As the examples can serve the β-1,3-glucan elongases of the Gas family or 

the chitin endotransglycosylases of the Crh family from yeast. Biochemical properties 

of these enzymes heterologously expressed in Pichia were determined in vitro using 

specially devised assays. In these assays, soluble polysaccharide derivatives were used as 

the glycosyl donors and diverse fluorescently labeled oligosaccharides as the acceptors. 

As the measure of enzyme activity served the amount of the fluorescence incorporated 

into the polymer. 


Lectures 

INHIBITION OF INSULIN AMYLOID AGGREGATION WITH ALBUMIN 

FUNCTIONALIZED MAGNETIC FLUID 

Andrea Antošová 1 , Katarína Šipošová 2 , Martina Koneracká 1 , Vlasta Závišová 1 , 

Peter Kopčanský 1 and Zuzana Gažová 1 

1 

Department of Biophysics, Department of Magnetism, Institute of Experimental 

Physics SAS, Kosice, Slovakia, 2 Department of Biochemistry, Faculty of Science, 

P. J. Safarik University, Kosice, Slovakia 

Amyloid-related diseases, such as Alzheimer’s disease or diabetes type II, are associated 

with self assembly of protein into amyloid aggregates. Recently, there are only few reports 

dealing with the effect of nanomaterials on the amyloid aggregation. We investigated 

effect of magnetic fluid consists of Fe 3 

O 4 

nanoparticles sterically stabilized by sodium 

oleate with adsorbed BSA (MF-BSA) on amyloid aggregation of insulin. The ability of MF- 

BSA to inhibit formation of insulin amyloid aggregates (Iagg) in vitro was studied by ThT 

assay and TEM. We have found that MF-BSA is able to prevent formation of aggregates, 

the extent of amyloid formation depends on MF-BSA concentration with extensive 70% 

inhibiting activity for ratio Iagg:MF-BSA = 1:7. The obtained results indicate that presence 

of MF-BSA led to the inhibition of insulin amyloid aggregation. Our findings make 

MF-BSA of potential interest as therapeutic agents against amyloid-related diseases. 

Ackowledgement: This work was supported within the projects Nos. 26220220005, 

26220120033 and 26220120021 in frame of SF EU, Centre of Excellence of SAS Nanofluid, 

VEGA 0079, 0056, 0077 and VVGS PF 13/2010/ Ch. 


57

Lectures 

TWENTY fOUr YEars SINCE CHErNOBYL DISaSTEr: 

What SEED prOTEIN can TELL US? 

Martin Hajduch 1 , Katarína Klubicová 1 , Maksym Danchenko 1, 3 , Ludovit Škultéty 2 , 

Namik Rashydov 3 and Anna Preťová 1 

1 

Department of Reproduction and Developmental Biology, Institute of Plant Genetics 

and Biotechnology, Slovak Academy of Sciences, Nitra, Slovakia 

2 

Center for Molecular Medicine, BITCET, Institute of Virology, Slovak Academy 

of Sciences, Bratislava, Slovakia, 3 Department of Biophysics and Radiobiology, 

Institute of Cell Biology and Genetic Engineering, National Academy of Sciences 

of Ukraine, Kyiv, Ukraine 

The explosion of one of the four reactors of Chernobyl nuclear power plant (CNPP) on 

26 April 1986 caused the worst environmental nuclear disaster in the history. Huge 

amounts of radioactivity were released not only to the close surroundings of the power 

plant but also to large parts of Europe. Despite the fact that since 1986 radiation levels 

in the affected environment have declined several hundred folds, dangerous long-living 

isotopes such as 137 Cs and 90 Sr remains as main contaminants. Now, 24 years after the 

accident, the question how plants in radio-contaminated Chernobyl were able to adapt 

is still open, and needs to be fully answered. Plants are stationary and thus must adapt 

to extreme conditions in order to survive. The main objective of our research is to 

characterize quantitative differences on protein levels between soybean (Glycine max) 

and flax (Linum usitatissimum) grown in contaminated (~5 km from CNPP) and control 

(~10 km from CNPP) experimental fields in order to elucidate molecular mechanisms 

plants used for adaptation. To acquire complex proteome information seed proteins 

are quantitatively analyzed using two-dimensional gel electrophoresis and identified by 

tandem mass spectrometry. 


Lectures 

Apoptosis IN rELaTION TO THE DEvELOPMENT of caNCEr 

aND rESISTaNCE of caNCEr CELLS TO CYTOSTaTICS 

Jozef Hatok 1 , Jana Jurečeková 1 , Peter Chudý 2 , Pavol Hollý 2 , Anton Dzian 3 , 

Eduard Huľo 3 , Eva Fabianová 1 , Tatiana Matáková 1 and Peter Račay 1 

1 

Department of Medical Biochemistry, 2 Clinic of Hematology and Transfusiology, 

3 

Clinic of Surgery – JFM and MFH in Martin, CU in Bratislava, Slovakia 

Apoptosis plays an important role in development and homeostasis of the multicellular 

organisms and its deregulation may result in many serious diseases, including cancer. 

In addition, dysregulation of apoptosis is associated with resistance of cancer cells to 

cytostatics. We studied the expression of mRNA of apoptotic proteins (p53, Bax, Bcl-2 

and Bcl-X L 

) in samples from cancer patients. In addition, we focused on their correlation 

with the results of chemoresistance testing. We examined 102 samples from patients 

with haematological malignancies (60) and solid tumors (42). To determine the levels of 

mRNA we used the RT-PCR. The in vitro chemoresistance of leukaemic cells was evaluated 

by MTT assay. Statistically significant differences of mRNA expression of all investigated 

proteins between the group of leukaemia samples and leukocytes from healthy volunteers 

were determined (p

Lectures 

SPECIALITIES OF OXIDATIVE PHOSPHORYLATION OF TRYPANOSOMATIDS 

AND EUGLENAS 

Anton Horváth 1 , Ingrid Škodová 1 , Anna Gnipová 1 , Alena Zíková 2 , 

Vladislava Benkovičová 1 , Zdeněk Verner 2 , Zdeněk Paris 2 and Július Lukeš 2 

1 

Department of Biochemistry FNS UK, Bratislava, Slovak republic, 2 Institute 

of Parasitology, Czech Academy of Sciences, České Budějovice, Czech Republic 

Trypanosomatids and euglenas are protists that belong to common phylum Euglenozoa. 

Despite their relatively close relations they differ quite strongly. Euglenas usually live in 

freshwater environments and contain chloroplasts with functional photosynthetic apparatus. 

In contrast with that all trypanosomatids are obligatory parasites and they are 

the only known eukaryotes with glycolysis separated from cytosol to special organelles 

- glycosomes. Euglenas and trypanosomatids are able suppress and again activate their 

oxidative phosphorylation depending on changing living conditions and they both have 

some alternative pathways to classical respiratory chain. So they are good models for 

study individual enzyme of oxidative phosphorylation and their importance for metabolism 

in different living conditions. In our lab we work with 4 different trypanosomatids and 

with Euglena gracilis and its mutants that are not able photosynthesis. Here we report 

about our two lines of our research: 

1. Localization and activity of two different FAD dependent glycerol-3-phosphate dehydrogenases 

and functions of several polypeptides associated with cytochrome C oxidase 

in Trypanosoma brucei 

2. General characterization of enzymes of respiratory chain in Euglena gracilis and their 

significance in cells grown on light and dark 


Lectures 

PROTEIN IMMUNIZATION OF MUTANT MOUSE – AN EffICIENT WAY 

TO GENEraTE SELECTIVE AND SENSITIVE ANTIBODIES 

Anna Hrabovska 1,2 , Veronique Bernard 1 and Eric Krejci 1 

1 

Centre d’Etude de la Sensori-Motricité, Université Paris Descartes- CNRS- UMR8194, 

45 rue des Saints Pères, 75006 Paris, France 

2 

Dpt. of pharmacology and toxicology, Faculty of Pharmacy, Comenius University, 

Odbojarov 10, 832 32 Bratislava 

Despite a long history, the successful generation of specific and selective antibodies is 

still a task fraught with considerable uncertainty. 

We examine a simple, fast, and highly efficient strategy to produce an antibody, which 

utilizes immunization of mutant mouse strains with antigens that the host strains themselves 

have been genetically targeted to be deficient for. To test this strategy we choose 

butyrylcholinesterase, an antigen that has been considered to be difficult to generate 

antibody against. Antigens of different origins all provided a strong immune response, 

while the characteristic of the resulting antibodies depended on the preparation for the 

antigen prior to the immunization. 

This method, introduced previously but since neglected until now due probably to the 

lack of specific resources, should at this time, based on our data presented here, be 

considered a reasonable and reliable choice for antibody production. 

Acknowledgement: Work was supported by grants AFM; grant ANR Neuroscience and 

APVV grants (SK-FR-0031-09 a SK-CZ-0028-09). 


61

Lectures 

DO WE TEACH BIOCHEMISTRY IN a LOGICAL WAY? REMarKS 

CONCERNING THE CONTENTS AND LEarNING APPROACH 

Jiří Hudeček 

Department of Biochemistry, Faculty of Science, Charles University 

Hlavova 2030/8, 128 40 Praha 2, Czech Republic 

I was always puzzled with the fact that among many of the most motivated and gifted 

students of “pure” chemical disciplines, there was a sort of depreciatory attitude towards 

biochemistry. (Occasionally, I could feel a similar attitude also among teachers.) At the 

same time, biochemical problems are often “invading” the field of the “pure” chemical 

disciplines, and one can say that at present a considerable percentage of all research in 

our chemical Departments has a strong connection with biochemistry. Certainly, part of 

the reasons for this situation might be historical, but there is a tendency for reproduction 

of these feelings. After conducting some interviews with students, I understand now that 

for the more logically thinking students, biochemistry is often a discipline too “biological” 

(in the sense “historical”), just describing things and showing some a posteriori explanations. 

Additionally, the traditional approach to teaching (I call it “synthetical”) forces 

students to learn a lot of facts, for long time seemingly without much logical coherence. 

They sometimes have a difficulty to understand the reasons why to learn, say, details of 

the citrate cycle. As the overall view of the intermediate metabolism comes only later, 

they may lose the enthusiasm long before coming close to get any sense of the beauty 

of the discipline. In the present contribution, I would like to discuss several possibilities 

how the situation might be improved: (1) using the “analytical” concept (an overall 

view later completed with details), (2) stressing the “chemical logic” in certain parts of 

biochemistry (structure and properties of biopolymers), (3) strong connection between 

the description (results) and experimental background for it, etc. 


Lectures 

EvaLUATION OF TOXICITY AND GENOTOXICITY Of ORGANOHALOGEN 

PESTICIDES 

Pavlína Janů 1 , Markéta Thimová 1 , Petra Lovecká 1 , 

Martina Macková 1 and Kateřina Demnerová 1 

Department of Biochemistry and Microbiology, Faculty of Food and Biochemical 

Technology, ICT Prague, Technická 5, 166 28 Prague, Czech Republic 

This work is focused on toxicity and genotoxicity analysis of the worldwide commonly 

used benzonitrile herbicides dichlobenil, chloroxynil, bromoxynil and ioxynil and their 

metabolites 2,6-dichlorobenzamid, 2,6-dichlorobenzoic acid, 3,5-dichloro-4-hydroxybenzoic 

acid, 3,5-dibromo-4-hydroxybenzoic acid, 3,5-diiodo-4-hydroxybenzoic acid. The toxicity 

was also determined for restricted organochlorine pesticides (fungicide HCB, insecticide 

γ-HCH, 4,4’-DDT and its metabolites 4,4’-DDA and 4,4’-DDE). 

Sea luminescent bacteria Vibrio fischeri, gramnegative bacteria Escherichia coli, grampositive 

sporulating bacteria Bacillus subtilis and microorganism isolated from contaminated 

soil (Burkholderia glathei) were chosen as the prokaryotic model systems for investigation 

of the acute toxicity. The eukaryotic model systems were represented by the seeds 

of Lactuca sativa, var. capitata and Avena sativa, by the „hairy root“ culture of Solanum 

nigrum and by the human cell line HEK 293T. Genotoxicity was determinated using 

the Ames test with bacteria Salmonella typhimurium His - TA98 and TA100 and the comet 

assay with the HEK 293T cell line. 

Our data point to that toxicity of pesticides is higher than their corresponding metabolites 

practically in all used systems. The toxic effect of studied compounds was similar in the 

context of used model system. The comet assay confirmed genotoxicity of pesticides 

ioxynil, bromoxynil, hexachlorobenzene and DDE, the metabolite of DDT. 

Acknowledgements: The authors thank for the support of grants Tandem FT –TA4/101, 

FT-TA5/043 a MSM 6046137305. 


63

Lectures 

BRONCHIAL ASTHMA AND EffECT OF OXIDATIVE STRESS ON ITS 

DEVELOPMENT 

Eva Babušíková 1 , Miloš Jeseňák 2 , Peter Bánovčin 2 and Dušan Dobrota 1 

1 

Department of Medical Biochemistry, 2 Institute of Children and Adolescents, 

Comenius University in Bratislava, Jessenius Faculty of Medicine in Martin, 

Martin, Slovakia 

Bronchial asthma (BA) is associated with increased oxidative stress. Oxidative stress is 

increasing due to the shift of the balance between pro-oxidant and antioxidant to side of 

pro-oxidant. Bronchial asthma is a complex chronic inflammatory disorder of the airways 

and as a complex disease is multifactorial, with many candidate genes suspected as being 

important in its development. In our study we analyzed oxidative damage of proteins 

and lipids and polymorphism of glutathione-S-transferase (GST: GSTT1, GSTM1) which 

is a BA candidate gene due to its role in protection against oxidative stress. The total 

content of sulfhydryl groups was decreased significantly (p < 0.001) in asthmatic patients 

compared to healthy children. Concentrations of thiobarbituric acid-reactive substances 

were increased significantly (p < 0.001) in asthma patients. The GSTT1 and GSTM1 null 

genotypes were more frequent (OR 1.63, respectively OR 1.18) among the asthmatic 

patients. The null genotype for both representatives of glutathione-S-transferase family 

represented higher risk for development of BA (OR 2.23) than present of one of them. 

Asthmatic children with null genotypes had higher oxidative damage as well. These results 

suggested that increased oxidative stress may play role in the asthma pathogenesis. 

Oxidative stress and changed antioxidant defense are included in the asthma pathology 

and therefore elimination of oxidative stress could be potentially an appropriate strategy 

for treatment of bronchial asthma. 

Acknowledgements: This work was supported by Ministry of Health of the Slovak Republic 

2007/47-UK-12. 


Lectures 

FrOM baSIC BIOMEDICal rESEarCH TO BIOTECHNOLOGY 

Laco Kačáni 

CEMIT – Center of Excellence in Medicine &IT GmbH, Innsbruck, Austria 

Recent business success of the biotech and pharma industry is mainly based on the commercialization 

of basic research discoveries done by researchers at public universities. 

As a result of these new economic developments, the strict division between basic and 

applied research in life sciences has weakened, thereby linking the research capacities 

of universities with the commercial capabilities of industry. The protection of research 

results in the life sciences by means of intellectual property rights is a prerequisite for 

commercial exploitation of research results in biotechnology. However, increased awareness 

among academic researchers for the commercialization of their research is even 

more important for the transfer of new biotechnologies to the business sector. 

The way, in which academic researchers cooperate and build partnerships with businesses, 

as well as the importance of academic research for biotech and pharma industry, 

changed dramatically in recent years. Consequently, new models of technology transfer 

and collaboration with biotech industry emerged in the last decade, some stimulated by 

state interventions and others formed directly between universities and biotech companies. 

These models enable a rapid translation of new knowledge into technologies, 

reduce the overall costs of R&D and facilitate the launching of new products or services 

onto the market place. 

In this talk, various models of technology transfer and collaboration with biotech industry 

will be analyzed and exemplified from different points of view, using examples of successful 

commercialization of academic biomedical research in biotechnology. 


65

Lectures 

NEW POSSIBILITIES FOR THE STUDY OF METABOLISM IN SLOvaKIA 

Michal Kaliňák 1 and Tibor Liptaj 2 

1 

Department of Biochemistry and Microbiology, 2 Department of NMR and MS, 

Faculty of Chemical and Food Technology Slovak University of Technology, 

Radlinského 9, 812 37 Bratislava 

NMR solves chemical structures for decades, but only after some laboratories in Slovakia 

were equipped with new NMR spectrometers and appropriate software it is possible to 

use metabonomic approach to tackle various biological questions. We have summarized 

various aspects of sample preparation, measurement setup and post-acquisition processing 

needed to be followed to obtain results giving biologically relevant information. 

Restrictions and disadvantages of NMR in terms of sensitivity, signal overlap and sample 

volume are presented. 

The metabonomic 1 H NMR data of germination, growth and conidiation of filamentous 

fungus Trichoderma viride are shown as an example of a study in fungal microbiology. 

The onset of metabolic activity can be observed during germination. Different cultivation 

conditions can be discerned on the basis of multivariate statistical analysis without the 

need to identify individual metabolites. 

Acknowledgement: This work was supported by Slovak Grant Agency VEGA 1/0462/08 

and APVV 0642-07. NMR experimental part of this work was facilitated by the support 

of Slovak National Research and Development Program No. 2003SP200280203. 


Lectures 

GATING OF THE NEURONAL CA V 

3.3 CHANNEL 

Mária Karmažínová 1 , Edward Perez-Reyes 2 and Ľubica Lacinová 1 

1 

Institute of Molecular Physiology and Genetics, Slovak Academy of Sciences, 

Bratislava, Slovakia, 2 Department of Pharmacology, University of Virginia, 

Charlottesville, Virginia 22908, USA 

Low-voltage activated Ca V 

3 Ca 2+ channels have activation threshold about -60 mV. Kinetics 

of their activation at membrane voltages just above activation threshold is much slower 

that the activation kinetics of other VDCC. It was demonstrated that intracellular loop 

connecting repeats I and II of all three Ca V 

3 channels contains so-called “gating brake”. 

Disruption of this brake yielded channels that activated at even more hyperpolarized 

potentials with significantly accelerated kinetics. We have compared gating of a wild type 

Ca V 

3.3 channel and a mutated ID12 channel, in which putative gating brake at proximal 

part of the I-II loop was removed. The whole cell Ca 2+ current was measured using the 

HEKA-10 patch clamp amplifier. Holding potential (HP) in all experiments was -100 mV. 

Gating currents were measured by 50 ms long depolarizing pulses to membrane potentials 

between -90 mV and +70 mV. 

Voltage dependence of gating current activation was shifted by 18.5 mV towards more 

hyperpolarized potentials in ID12 channel. Kinetics of the on-charge activation was 

significantly accelerated. Kinetics of the off-charge was not altered. Value of maximal 

on-charge normalized in respect to maximal inward current amplitude was doubled. 

We concluded that the putative gating brake in I-II loop hinders not only opening of 

the conducting pore but also the activating movement of voltage sensing S4 segments 

stabilizing the channel in its closed state. 

Acknowledgements: Supported by VVCE-0064-07 and VEGA 2/0195/10. 


67

Lectures 

MOLECULar MECHaNISMS INvOLvED IN rESPONSE TO HYPOXIa 

Juraj Kopáček, Jaromír Pastorek and Silvia Pastoreková 

Institute of Virology, Slovak Academy of Sciences, Dúbravská cesta 9, 

845 05 Bratislava 

Cellular responses to diminished supply of oxygen include growth arrest, apoptosis, 

anaerobic glycolysis, angiogenesis etc. The primary response to the lack of oxygen at 

the molecular level is the stabilization of a subunit of HIF-1 transcriptional complex, 

a key regulator of the genes involved in adaptation to the hypoxic stress. In normoxia, 

HIF-1a undergoes hydroxylation that is required for its interaction with the product of 

the with type von Hippel-Lindau (VHL) tumor suppressor gene. This interaction results in 

fast ubiquitilation and proteasome degradation of HIF-1a. Loss or mutation in VHL, the 

main negative regulator of the hypoxic pathway, leads to development of the hypoxic 

phenotype also under normoxic conditions. In addition, stabilization of HIF-1a could be 

achieved by signal transduction through the pathways regulated by activated oncogenes 

that can contribute to or amplify the effects of of HIF-1 transcriptional complex. HIF-1 is 

a key regulator of a broad range of cellular and systemic responses to hypoxia and acts 

in all mammalian cells. HIF-1 activity is dependent upon the availability of the HIF-1α 

subunit, which is in turn regulated by cellular oxygen levels. 

This work was supported by the Research & Development Operational Programme funded 

by the ERDF „TRANSMED” and by VEGA 2/0194/09. 


Lectures 

rEGULaTION of aurICIN BIOSYNTHESIS IN 

StrePTomyces aureofaCIens CCM 3239 

Ján Kormanec, Renáta Nováková, Ľubica Fecková, Peter Kutaš and Alena Reháková 

Institute of Molecular Biology, Slovak Academy of Sciences, Dubravska cesta 21, 


Gram-positive bacteria Streptomycetes are the main producers of bioactive natural products 

including many antibiotics. The polyketides, which are synthesized by multifunctional 

enzymes called polyketide synthase (PKS), belong to the most important classes of antibiotics. 

We previously identified a type II polyketide synthase (PKS) gene cluster, aur1, in 

Streptomyces aureofaciens CCM3239 with the highest similarity to angucycline polyketide 

subgroup of PKS clusters. Deletion of two critical biosynthetic genes resulted in a lack of 

antibiotic that was named auricin. However, its purification has been hampered by very 

low yields. This cryptic phenotype has been recently described for several homologous 

angucycline gene clusters, indicating their tight regulation. Sequence analysis of the 

whole aur1 cluster revealed a huge number of regulatory genes, including six genes for 

homologues of TetR family repressors, five genes for homologues of specific Streptomyces 

Antibiotic Regulatory Proteins (SARP family), a single gene for homologue of AraC/ 

XylS-family regulators, a single genes for homologue of response regulators of bacterial 

two-component signal transduction systems, and two genes for gama-butyrolactone 

autoregulator-receptor system. Their partial characterization indicated that several of 

them play a positive or negative role in auricin production in a cascade scheme. 

Acknowledgements: This work was supported by the Slovak Research and Development 

Agency under the contract No. APVV-0017-07. 


69

Lectures 

TEACHING BIOCHEMISTRY AT THE UNIVERSITY OF VETERINarY 

MEDICINE AND PHarMACY IN KOŠICE 

Zuzana Kostecká 

Institute of Biochemistry, Department of Chemistry, Biochemistry and Biophysics, 

The University of Veterinary Medicine and Pharmacy in Košice 

Implementation of credit system in the study at the University of Veterinary Medicine 

and Pharmacy in Košice was coupled with innovation of curricula of all obligatory study 

subjects, creating of new compulsory optional and optional subjects including biochemistry. 

Nowadays the teachers of biochemistry institute participate in teaching of obligatory 

subjects: Biochemistry for the study pogramme General veterinary medicine (SP GVM) and 

SP Food hygiene (SP FH), General Biochemistry for SP Pharmacy, Aplied Biochemistry for 

SP Safety of food and feed and Biochemistry for SP GVM in English language; compulsory 

optional study subjects: Molecular mechanisms of metabolic and inherent diseases for 

SP GVM and SP FH, Molecular mechanisms of metabolic and inherent diseases for SP 

GVM in English, Clinical biochemistry for SP GVM and SP FH and Clinical biochemistry 

for SP GVM in English. The obligatory subject Clinical and pathological biochemistry for 

SP Pharmacy is provided by external teachers (teachers of Faculty of Medicine of Pavol 

Jozef Šafárik University in Košice). 

The aim of this presentation is to inform about changes in teaching biochemistry and 

to evaluate the positives and negatives according to our experiences. Lecture shows 

the advantages and disadvantages of credit system of study, including the questions of 

actual problems in field of biochemistry teaching at our university, e.g. organisation of 

teaching, position of study subjects in curriculum, continuity of study subjects, using of 

multimedia technology, etc. In conclusion, considering the possible solutions and seeking 

answers to questions will be resulted in successful educational process. 


Lectures 

TEACHING BIOCHEMISTRY AT THE faCULTY OF SCIENCE IN KOŠICE 

Mária Kožurková, Marián Antalík and Dušan Podhradský 

Department of Biochemistry, Institute of Chemistry, Faculty of Science, 

P. J. Šafárik University, Košice, Slovakia 

We have been teaching biochemistry for 20 year. Our department provides an education 

of Biochemistry for students of the bachelor and master degree, according to the credit 

system of study at Faculty of Science. 

The subject Biochemistry was broken up into part I and II. Biochemistry I taught for 2 hour 

per week (for students of bachelor study) and will give a survey of new technologies and 

materials. The lectures are suitable for both scientific and pedagogically oriented students. 

Biochemistry II (for students of master study) will focus of structures and functions of 

saccharides and lipids, metabolism, regulation of metabolic pathways, basic metabolic 

processes and principle of bioenergetic. The subject is taught for 2 hours per week and 

is finished by final written test. Biochemistry III - Modern trends in biochemistry (for 

students of master study) provide biochemical view into the modern trends: evolution 

of energetic metabolism, protein splicing, protein sequencing, protein - DNA interaction, 

problems of DNA replication, cell division, viruses, and apoptosis. Students are eligible 

to graduate with a title of Master of Science upon passing of a state examination in the 

areas of Biochemistry, Molecular Biology, Biophysical Chemistry, Bioorganic Chemistry, 

Clinical Biochemistry and Biotechnology. 

The department is also an educational workplace for post-graduate students in the field 

of the structure and functions of biomolecules. 

Acknowledgement. This work was supported by the Grant Agency VEGA 1/0053/08 and 

KEGA 3/6301/08. 


71

Lectures 

Assembly of BaCILLus suBTILIS SPOre COat: INvESTIGaTION of 

prOTEIN-prOTEIN INTEraCTIONS aMONG THE SPOre COat prOTEINS 

of BaCILLLUS SUBTILIS 

Daniela Krajčíková 1 , Denisa Mullerová 1 , Wan Qiang 2 , Per Bullogh 2 , 

Jilin Tang 3 and Imrich Barák 1 

1 

Institute of Molecular Biology, Slovak Academy of Sciences, Bratislava, Slovakia; 

2 

Krebs Institute for Biomolecular Research, Department of Molecular Biology 

and Biotechnology, University of Sheffield, United Kingdom; 

3 

State Key Laboratory of Electroanaytical Chemistry, Changchun Institute of Applied 

Chemistry Chinese Academy of Sciences, Changchun, P. R. China 

Spores, dormant cell types of Bacillus subtilis, are incased in thick proteinaceous multilayered 

shell, called the coat, with significant protective role from the environment. While 

providing high level of resistance, the coat allows the spore to respond to the renewed 

presence of nutrients and start the cell growth in the process called germination. The 

unique properties of the coat are determined by the architecture of complex spore coat 

structure. Being formed by more than 70 different proteins, two main layer of coat are 

easily distinguished – the lamellar inner coat and thick striated outer coat. The order of 

assembly and final destination of the coat structural components rely mainly on specific 

protein-protein interactions and on the action of small group of morphogenetic proteins 

which guide the deposition of rest of the coat components onto the spore surface. Since 

the process of assembly is still poorly understood, by searching for direct protein-protein 

interactions we want to gradually generate the data to obtain the entire picture of whole 

coat formation event. 

In our studies we implemented yeast two hybrid system and other genetic and biochemical 

methods to examine protein interactions among a group of morphogenetic coat 

proteins and proteins of coat insoluble fraction. Investigation of properties of individual 

recombinant coat proteins showed, that they frequently form high molecular weight 

oligomeric structures in vitro. We employed AFM and EM microscopy to analyze these 

structures in detail. 


Lectures 

CHANGES AND ROLE OF ADRENOCEPTORS IN PC12 CELLS afTER 

PHENYLEPHRINE ADMINISTraTION AND APOPTOSIS INDUCTION 

Ľubomíra Lenčešová 1,2 , Marta Sírová 1 , Lucia Csáderová 2 , Marcela Lauková 3 , 

Zdena Sulová 1 , Richard Kvetňanský 3 and Oľga Križanová 1 

1 

Institute of Molecular Physiology and Genetics, Center of Excellence 

for Cardiovascular Research, Slovak Academy of Sciences, Bratislava, Slovak Republic, 

2 

Molecular Medicine Center, Slovak Academy of Sciences, Bratislava, Slovak Republic, 

3 

Institute of Experimental Endocrinology, Slovak Academy of Sciences, 

Bratislava, Slovak Republic 

Adrenergic regulation might modulate the effect of apoptosis. Therefore we studied, 

whether a1-adrenergic receptor’s agonist phenylephrine (PE) can affect or induce apoptosis 

in rat pheochromocytoma (PC12) cells. We have shown that PE treatment did not 

increase level of the apoptosis, or level of the caspase 3 mRNA. When apoptosis was 

induced in the presence of PE, caspase 3 mRNA was significantly increased, while the 

percentage of apoptotic cells remained unchanged compared to apoptotic group without 

PE. During this process, a1D-, b2- and b3-adrenergic receptors (ARs) were upregulated. 

Since all these three types of ARs are differently localized in the cell, we assume that 

mutual communication of all three ARs is crucial to participate in this signaling and during 

development of apoptosis, some of these systems might translocate. Another inportant 

system in handling noradrenaline during apoptosis might be noradrenaline transporter 

(NET), since it was downregulated in apoptotic cells treated with PE, compared to untreated 

apoptotic cells. However, precise mechanism of mutual communication among 

all these systems remains to be elucidated. 

This work was supported with scientific grants APVV 51/0397 and VEGA 2/0049/10. 


73

Lectures 

GATING OF THE T-TYPE CALCIUM CHANNELS 

Ľubica Lacinová and Mária Karmažínová 

Institute of Molecular Physiology and Genetics, SAV, Bratislava, Slovak Republic 

T-type calcium channels are distinguished by relatively low voltage threshold for an activation 

and steep voltage dependence of activation and inactivation kinetics just above 

the activation threshold. Further, while macroscopic current kinetics of Ca V 

3.1 and Ca V 

3.2 

channels are virtually identical, kinetics of the Ca V 

3.3 channel is almost one order more 

slow. Kinetics and voltage dependence of macroscopic inward calcium current through 

Ca V 

3 channels was described in a detail. In contrast, very little information is available 

on gating current of these channels. Therefore we compared gating currents measured 

from all three Ca V 

3.1, Ca V 

3.2 and Ca V 

3.3 channels. 

Voltage dependencies of macroscopic current activation are similar for all three Ca V 

3 

channels. While gating kinetics of macroscopic calcium current is virtually identical for 

Ca V 

3.1 and Ca V 

3.2 channels it is about one order slower for the Ca V 

3.3 channel. Voltage 

dependencies of charge movement differ dramatically from those for macroscopic current. 

First, their slope factors are several-fold bigger that slope factors of macroscopic 

current activation. Second, activation mid-point for Ca V 

3.3 channels on-gating is shifted 

to more positive membrane potentials by about 20 mV compare to Ca V 

3.1 and Ca V 

3.2 

channels, whose activation mid-points are similar. The same is truth for off-gating voltage 

dependences. Kinetics of both on- and off-gating is remarkably faster for Ca V 

3.1 and 

Ca V 

3.2 channels compare to Ca V 

3.3 channels. Further, more charge is moved per unit of 

macroscopic current amplitude in Ca V 

3.3 channels compare to Ca V 

3.1 and Ca V 

3.2 channels. 

Acknowledgement: Supported by VVCE-0064-07 and VEGA 2/0195/10. 


Lectures 

INDUCTION of ISCHEMIC TOLEraNCE IN SENSITIve NEUrONS: 

COOrDINaTED rOLE of MULTIPLE MECHaNISMS 

Ján Lehotský, Mária Chomová, Andrea Evinová, Mária Kovalská, Martina Pavlíková, 

Zuzana Tatarková, Peter Kaplán and Peter Račay 

Comenius University, Jessenius Faculty of Medicine, Department of Medical 

Biochemistry, Martin, Slovakia 

Ischemic brain injuries are among the most common and important causes of disability 

and death worldwide. Sublethal ischemia, ischemic preconditioning (IPC), triggers 

endogenous responses that protect the brain against a subsequent severe ischemic 

insult, a phenomenon known as tolerance. These treatments can initiate and amplify 

the endogenous adaptive/restorative processes in brain. The aim of this study was to 

determine whether altered interplay between intracellular Ca2 + stores resulting in 

the apoptotic and unfolded protein response is linked with the neuronal adapation 

induced by ischemic preconditioning. We refer here that ischemic/reperfusion injury 

(IRI) is manifested by: i) functional mitochondrial ganges, and ii) altered gene expression 

and translation of endoplasmic reticular (ER) key UPR proteins. Tissue response to 

ischemic preconditioning includes changes in the: i) level of initiation and execution of 

apoptosis ii) activation of inhibition of p53 translocation to mitochondria, iii) changes 

of endoplasmic reticular (ER) expression of Ca 2+ binding GRP78 and ATF6 proteins, and 

iv) effects on Secretory Pathways Calcium Pump (SPCA) gene expression and partial recovery 

of depressed SPCA activity. The results suggest that adaptation process /ischemic 

tolerance induced by preischemic challenge includes interplay between intracellular 

Ca 2+ stores, mitochondria, ER and Golgi apparatus, and have potential to translate into 

novel strategies for the treatment of ischemic stroke. In addition, we present here an 

overview of pathways which eventually maturates in tolerant phenotype of neuronal 

cells in affected brain areas. 

Acknowledgements: This work was supported by the VEGA grant No. 49/09, VVCE 64/07, 

55 UK-16/2007 and by project “Center of translational medicine” co-financed from EC 

sources and European regional development fund. 


75

Lectures 

SPINAL CORD INJURY: PATHOGENESIS AND TREATMENT 

Nadežda Lukáčová, Alexandra Dávidová, Ľudmila Capková and Andrea Kucharíková 

Institute of Neurobiology, Slovak Academy of Sciences, Košice 

Transversal spinal cord lesions interrupt a neuronal pathways which provide an inhibitory 

effect on reflex activity. Subsequently, spasticity develops below the injury site. The aim of 

this study was to find out whether neuronal degeneration correlates with up-regulation 

of nitric oxide synthase (NOS) forming nitric oxide, and whether these neurons are protected 

by parvalbumin (PV), buffering free intracellular calcium. Fluoro-Jade B was used 

to detect dying neurons. 7 and 14 days after spinal cord injury both the level of nNOS 

protein and nNOS mRNA level were significantly increased in segments below the site of 

injury. We noted strong nNOS upregulation in motoneurons and in neurons of laminae 

VII. However, a-motoneurons were not Fluoro-Jade B positive. While PV-IR was increased 

in a number of small neurons in rat, rabbit’s a-motoneurons exhibited very strong PV 

fluorescent staining. The results indicate the participation of PV in motor control. After 

spinal injury the animals were treated with GABA B 

receptor agonist Baclofen (3 µg/2 x 

daily from 7th day), or with NNLA (an inhibitor of neuronal NOS dosed at 20 mg/b.w.) 

independently, or combined together. Intrathecal treatment with Baclofen for three days 

restored both the NO synthase and PV levels almost to control value. NNLA or Baclofen 

decreased the level of nNOS mRNA in spinal cord, but combined use of both drugs was 

not effective. 

Acknowledgements: Supported by APVV 0314-06 and by VEGA 2/0015/08. 


Lectures 

A method for aUTOMaTED DETECTION of HETErOZYGOUS 

INSErTION-DELETION MUTaTIONS 

Peter Májek 1 , Vladimír Špitalský 1 , Gabriel Minárik 2 and Tomáš Szemes 2 

1 

ADINIS s.r.o., Bratislava 

2 

Department of Molecular Biology, FNS, Comenius University in Bratislava 

Structural and insertion-deletion (indel) variants are of considerable importance, mostly 

because of their phenotypic consequences. Indels shorter than 30 bp currently constitute 

about 24 % of all disease causing mutations reported in Human Gene Mutation Database. 

However, typical tools for analysis of sequence traces with indels obtained from diploid 

samples do not have sufficient accuracy and therefore extensive manual review of such 

samples is needed. 

We present here a new algorithm, implemented in software ADINIS IndelFinder, for 

automated detection of indel mutations from diploid sequence traces. The algorithm 

identifies 95% of indel mutations selected by a human expert with 5% false positives 

rate on a set of 39 sequence traces of exon 11 of KIT gene of subjects diagnosed with 

gastrointestinal cancer. 

The algorithm is based on a parametric digitalization of the electropherogram signal to 

a discrete set of nucleotide peaks followed by three rounds of the Needleman-Wunsch 

(NW) algorithm. The parameters of electropherogram digitalization as well as the scoring 

matrices used in the NW are optimized by a Monte-Carlo sampling to automatically find 

the best performing set of parameters. 


77

Lectures 

BIOCHEMISTry IN THE PICTUrES - INTEraCTIve BIOCHEMISTry 

Mária Mareková, Jana Mašlanková, Peter Urban and Juraj Guzy 

Department of Medical Chemistry, Biochemistry and Clinical Biochemistry, 

UPJŠ - Faculty of Medicine, Tr. SNP 1, 040 01 Košice, Slovak Republic 

Modern informatics technologies (IT) including internet, essentially change the classical 

teaching methods. Traditional book illustrations cannot compete to electronic assigns, 

which provides except high-quality image documentation with zoom support also video 

recordings, animations or comments. Electronic form provides quick searching and intermediate 

updates. Our aim is to prepare interactive atlas of biochemistry – biochemistry 

in the pictures, with pictures, schemes, brief texts and the testing system for student`s 

own control. This upcoming atlas will be adjusted also for multimedia support of teaching, 

f.e. for it`s using as e-learning component, which leads students to more active form of 

studying. For the development of interactive atlas is also necessary cooperation between 

teachers and IT professionals. We are searching for help with preparation and servicing 

of internet electronical atlas on internet web servers, using administrator’s rights with 

possibility to manage the personal user accounts and with creation of copyright protection 

for multimedia content. 

Acknowledgments: This work was supported by grant project KEGA3/7130/09. 


Lectures 

OrIGIN of aCQUIrED rESISTaNCE TO CYTOTOXIC aCYCLIC 

NUCLEOSIDE PHOSPHONaTES 

Helena Mertlíková-Kaiserová, Antonín Holý and Ivan Votruba 

Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech 

Republic, Gilead Sciences & IOCB Research Center, 166 10 Prague, Czech Republic 

Acquired resistance to chemotherapy upon repeated administration of the cytotoxic drugs 

remains a serious clinical issue. Disclosing the mechanisms that lead to its development is 

necessary for introduction of successful prevention/reversal strategies. In our laboratory, 

we have prepared CCRF-CEM leukemic cells resistant to high concentrations of cytotoxic 

nucleotide analogs PMEG and PMEDAP. Employing [8- 3 H] radiolabeled compounds we 

aimed to describe changes in membrane transport and/or intracellular metabolism of 

the compounds. We found that the uptake of both PMEG and PMEDAP was unaffected in 

resistant cells. The resistance is therefore not due to decreased intracellular concentrations 

of the parent compounds. However, their metabolites PMEG/PMEDAP phosphate 

and PMEG/PMEDAP diphosphate were only present in sensitive but not resistant cells. 

As only the latter two can be incorporated into the DNA and act as chain terminators it 

is clear that the origin of resistance lies in the phosphorylation step catalyzed by relevant 

nucleoside monophosphate kinases - guanylate kinase (GUK) for PMEG and mitochondrial 

adenylate kinase (AK2) for PMEDAP. Expression of these proteins was indeed decreased 

in resistant cells. It is possible that apart from lower amount of GUK and AK2 protein, 

catalytic activity of these enzymes might also be affected. This will further be explored. 

Acknowledgments: Research project of the IOCB #OZ40550506; Project #1M0508 by the 

Ministry of Education, Youth and Sports of the Czech Republic. 


79

Lectures 

THE IDENTIfICaTION aND CHaraCTErIZaTION of THE firST 

verTEBraTE HYBrID STErILITY GENE (HST1/PrDM9) 

Ondřej Mihola, Zdeněk Trachtulec and Jiří Forejt 

Department of Mouse Molecular Genetics, Institute of Molecular Genetics, Academy 

of Sciences of the Czech Republic, Videnska 1083, 142 20 Prague, Czech Republic 

The mouse Hybrid sterility 1 (Hst1) gene participates in a breakdown of spermatogenesis 

in the F1 offspring of crosses between some laboratory strains predominantly of Mus m. 

domesticus origin (e.g., B6 or B10) and certain mice of Mus m. musculus (sub)species, 

such as of the PWD strain. Other hybrid males, e.g. (PWD x C3H), are fertile. The Hst1 gene 

has been mapped on a high-resolution ((B10 xC3H) x B10) backcross and by transgenesis. 

The Hst1 candidate region was narrowed down to a single gene, PR-domain 9 (Prdm9) 

or Meisetz, encoding a histone 3 K4 trimethyltransferase. The gene was confirmed as 

Hst1 by comparing the phenotypes of its null allele with the phenotypes of the sterile 

hybrids. To learn about the mechanisms regulating the germ-cell development through 

PRDM9, the expression profile of fertile and sterile hybrid testes differing only in the 

allele of Prdm9 were analysed by microarrays and real-time qRT-PCR. Furthemore, the 

localization of PRDM9 protein in germ cells was studied by indirect immunofluorescence. 

Our data suggest that Prdm9 is one of the master transactivators regulating meiotic 

epigenetic events. 


Lectures 

BIOCHEMICAL MarKERS OF MULTIPLE SCLEROSIS 

Jozef Michalik and Egon Kurča 

Clinic of Neurology, Jessenius Faculty of Medicine and Faculty Hospital in Martin 

Multiple sclerosis is an autoimmune, inflammatory, demyelinating disease of the central 

nervous system. Several pathophysiological mechanisms such as alteration of the 

immune system, disruption of blood-brain barrier, inflammation, oxidative stress and 

excitotoxicity, demyelination, axonal, neuronal damage, gliosis, remyelination and repair, 

cortical reorganisation are involved in the pathogenesis of the disease. Because of the 

heterogenity in clinical presentations and courses, a subtyping of patients by clinical, 

neuroradiological, genetical, neuroimmunological, biochemical parameters is necessary 

at present. 

This paper discusses the potential applicability of some biological markers for the diagnosis, 

disease activity, prediction of clinical courses and response to disease modifying therapies. 

There are some immunological markers, biomarkers of neuronal and axonal damage, 

biomarkers of demyelination, oxidative stress and excitotoxicity, gliosis (cytokines and 

their receptors, adhesion molecules, matrix metalloproteinases, 24S-hydroxycholesterol, 

antibodies to myelin oligodendrocyte glycoprotein, axonal antigens, amyloid precursor 

protein, 14-3–3 protein, MxA protein, B cell activating factor of the TNF family - BAFF). 


81

Lectures 

RTX CYTOTOXINS rECOGNIZE β 2 

INTEGrIN rECEPTOrs THrOUGH 

N-LINKED OLIGOSaCCHarIDES 

Jana Morová, Radim Osička, Jiří Mašín and Peter Šebo 

Institute of Microbiology of the Academy of Sciences of the Czech Republic, 


Bordetella pertussis Adenylate cyclase toxin-hemolysin (CyaA, ACT, or AC-Hly) is a bifunctional 

protein belonging to the RTX (Repeat in ToXin) family of bacterial cytotoxins. 

CyaA delivers into target cells an adenylate cyclase domain, which catalyzes uncontrolled 

conversion of ATP to cAMP, a key signaling molecule subverting cell functions. The toxin 

utilizes as a specific cellular receptor, the CD11b/CD18 integrin (α M 

β 2 

, Mac-1, or CR3) 

that is heavily Nglycosylated. Here, we demonstrate that deglycosylation of cell surface 

proteins by glycosidases completely abolished CyaA binding to CD11b-expressing cells. 

Moreover, cAMP intoxication of the deglycosylated cells exposed to the toxin was significantly 

reduced, suggesting a requirement of CD11b/CD18 glycosylation. Similar results 

were obtained, when N-glycosylation of de novo synthesized cellular proteins was inhibited 

by the antibiotic tunicamycin. Moreover, binding of CyaA to CD11b-expressing cells 

was significantly inhibited in the presence of excess of free saccharides found in building 

units of the oligosaccharide complex of the integrin. On the other hand, saccharides 

not occurring in integrin oligosaccharide chains were unable to inhibit CyaA binding to 

CD11b/CD18 to any significant extent, showing that CyaA selectively recognizes the sugar 

residues of N-linked oligosaccharides of the integrin. Furthermore, the requirement for 

integrin glycosylation could be demonstrated also for binding of another RTX protein, the 

leukotoxin of Aggregatibacter actinomycetemcomitans (LtxA), which specifically binds 

to target cells via another receptor of the β 2 

integrin family, CD11a/CD18. These results 

demonstrate that glycosylation of β 2 

integrin receptors facilitates CyaA and LtxA binding 

to and killing of various target cells and set a new paradigm for action of RTX cytotoxins. 


Lectures 

Fungal ɑ-N-aCETYLGalaCTOSaMINIDaSE frOM aSPergILLus niger: 

CLONING aND EXPrESSION IN YEaST 

H.Mrázek 1,2 , L. Weignerová 2 , D. Manglová 1,2 , D. Kavan 1,2 , V. Křen 2 and K. Bezouška 2 

1 

Department of Biochemistry, Faculty of Science, Charles University in Prague, Prague, 

Czech Republic, 2 Institute of Microbiology v.v.i., Academy of Sciences 

of Czech Republic, Prague, Czech Republic 

Alpha-N-acetylgalactosaminidase is an exoglycosidase specific for the hydrolysis of terminal 

ɑ-linked N-acetylgalactosamine in various sugar chain. A large screening study of extracellular 

ɑ-N-acetylgalactosaminidase activity of a library of filamentous fungi (42 strains), led 

to the identification of the best constitutive producer Aspergillus niger CCIM K2.They occur 

widely in microorganisms, plants and animals, and have considerable potential in various 

industrial application. Stability and activity at high temperatures are important properties 

of ɑ-N-acetylgalactosaminidases. This enzyme from Aspergillus niger has certain unique 

properties, and it was thus interesting to clone it for further structural investigations. 

Alpha-N-acetylgalactosaminidase from Aspergillus niger CCIM K2 was partially sequenced 

by Edman degradation and MALDI MS. A gene fragment encoding a putative part of the 

ɑ-N-acetylgalactosaminidase was amplified using cDNA prepared from Aspergillus niger 

CCIM K2. Degenerated PCR primers were designed according to amino acids found in ɑ-Nacetylgalactosaminidase 

from Aspergillus niger CCIM K2. The full-length coding sequence 

of ɑ-N-acetylgalactosaminidase was cloned into pPICZɑ and the recombinant protein was 

expressed in yeast. The cloned DNA consists of 1450 base pair, and the deduced amino acid 

sequence (480 amino acid residues with molecular mass 54,814kDa) is almost identical to that 

of purified enzymes (as determined by SDS-PAGE). Comparison of this amino acid sequence 

with the GenBank database revealed significant homology (96% identity) with sequence of 

ɑ-galactosidase (EC3.2.1.22) from Aspergillus niger. Analysis of the identified sequences 

shows that ɑ-N-acetylgalactosaminidase is composed of three domains. 

The ɑ-N-acetylgalactosaminidase from Aspergillus niger CCIM K2 was expressed in yeast. In 

further experiments we would like to express the individual domains of this complex enzyme, 

and to evaluate, if there is an active carbohydrate-binding (lectin) domain within this enzyme. 

Acknowledgements: This work was supported, by Grant agency of Charles University in 

Prague (19309) and the Institutional Research Concept for the Institute of Microbiology 

(AVOZ5020051), and grants from the Czech Science Foundations 

(203/05/0172, 204/06/0771, I) 


83

Lectures 

oxidaTIve rISK IN aTHErOSCLErOSIS 

Jana Muchová 1 , Zuzana Nagyová 2 , Iveta Ondrejovičová 1 and Zdeňka Ďuračková 1 

1 



2 

Juvenalia, Paediatric Center, Dunajská Streda, Slovakia; 

Atherosclerosis is the most common pathological process that leads to cardiovascular 

diseases and is known to be associated with inflammation, oxidative stress and endothelial 

dysfunction. In the vasculature reactive oxidant species may oxidatively modify 

lipids and proteins with deleterious consequences for vascular function. The ROS are 

common by-products of many oxidative biochemical and physiological processes. They 

can be released by increased activation of xanthine oxidase, NAD(P)H oxidase, lipoxygenases, 

mitochondria, or the uncoupling of nitric oxide synthase in vascular cells, as 

well as decreased cellular antioxidant capacity. ROS mediate various signaling pathways 

that underlie vascular inflammation in atherogenesis. The dysfunctional vasculature is 

characterized by lipid peroxidation and aberrant lipid deposition, inflammation, immune 

cells cell activation, platelet activation, thrombus formation, and disturbed hemodynamic 

flow. Each of these pathological states is associated with increasing of free radical 

species-derived oxidation products and, thereby, implicates increased oxidant stress in 

the pathogenesis of vascular diseases. 


Lectures 

AdvaNCED TECHNOLOGY for METaBOLIC INvESTIGaTIONS 

Roman Oros 

Shimadzu Austria HmbH, Korneuburg, Laaer Strasse 7-9, A-2100, Austria 

This lecture presents the use of MS n measurment with prediction software tool to identify 

the formulas and structures in fields such as impurity analysis of pharmacological active 

substances, metabolic profiling and biomarker research. 

Discerning the chemical formula or structure of unknowns is a difficult task that can be 

partially alleviated by acquiring high mass accuracy data, however, data interpretation 

is tedious and time consuming. By using fragmentation spectra collected from LCMS-IT- 

TOF(a hybrid ion-trap time-of-flight mass spectrometer) along with enhanced formula 

prediction software, samples are rapidly analyzed to identify chemical formulas and 

structures. 

The composition prediction is based on three main steps:a) composition calculated from 

mass, b)using isotopic pattern, c)MS n spectral filtering 

The LCMS n data for metabolite structural detection in a metabolomic field will be presented. 


85

Lectures 

AcrIDINES – USEfUL MUTaGENS 

Helena Paulíková 

Department of Biochemistry and Microbiology, Faculty of Chemical and Food 

Technology, Slovak Technical University, SK-81237 Bratislava 

Acridine based compounds comprise of an important class of DNA-intercalating anticancer 

drugs, and are structurally characterised by the presence of a planar chromophore 

capable of intercalation into DNA. However acridines are a double-edged sword, they 

are known as substances cause mutation and cancer. Most notable among their mutagenic 

effects is the induction of frameshift mutations. In spite of their mutagenicity, 

DNA affinity makes them attractive for the design of antitumor drug targeting DNA. Four 

new classes of acridine derivatives have been investigated by our research group and 

their anticancer potential has been assessed. Intercalation into DNA was confirmed by 

a variety of spectroscopic and electrophoresis techniques and anti-proliferative activity 

against three tumor cell lines (L1210, HL-60; A2780) was observed. The mechanism of 

antitumor activity of acridines involves intercalator-dependent formation of irreparable 

DNA double or single strand breaks arising from the inhibiting of DNA topoisomerases. 

Although acridines are also known as mutagens, the data obtained from the literature 

and our results showed that not all acridine intercalators are mutagens and it seems that 

the presence of electrophilic functional groups is nescessary for mutagenicity. 

Acknowledgements: Supported by Slovak Grant Agency, grants No 1/0097/10 and 

1/0053/08. 


Lectures 

THE CROSS-TALK OF NITRIC OXIDE AND NUCLEar faCTOr kaPPa B 

IN EXPERIMENTAL HYPErTENSION 

Olga Pecháňová, Andrej Barta, Stanislava Vranková, 

Jana Parohová and Mária Kovácsová 

Institute of Normal and Pathological Physiology and Centre of Excellence 

for Cardiovascular Research, Slovak Academy of Sciences, Bratislava, Slovak Republic 

Recently we have demonstrated involvement of NF-κB in the upregulation of endothelial 

nitric oxide synthase (eNOS) in hypertension induced by N G -nitro-L-arginine methyl 

ester (L-NAME). Thus, the goal of our study was to analyze an effect of NF-κB inhibitor, 

lactacystin, in L-NAME-induced hypertension. Adult 12-week-old male Wistar rats were 

subjected to treatment with L-NAME (40 mg/kg/day) for seven weeks (n=14). Half of 

the rats received lactacystin together with L-NAME for last three weeks. Next 16-weekold 

male Wistar rats received lactacystin only for 3 weeks (n=7). Blood pressure was 

measured by tail-cuff plethysmography every week. Total NOS activity was determined 

by measuring the formation of L-[ 3 H] citrulline from L-[ 3 H] arginine. Endothelial NOS 

and NF-κB (p65) protein expressions were determined immunohistochemically and by 

Western blot analysis. Membrane oxidative damage was analyzed by conjugated diene 

(CD) level determination. Lactacystin treatment did not affect the blood pressure (103±6 

mmHg), while 7-week-L-NAME treatment increased blood pressure (141±3 mmHg) by 38% 

comparing the age-matched untreated animals (104±5 mmHg). Addition of lactacystin to 

the L-NAME increased blood pressure significantly (159±4 mmHg) by 54% comparing the 

untreated control group and by 12% comparing the L-NAME group. L-NAME treatment 

led to increased NF-κB expression followed by elevation of both eNOS protein expression 

and total NOS activity in the aorta, heart and kidney. Addition of lactacystin blocked, 

however, elevated eNOS protein expression in all tissues investigated. CD concentration 

was increased by lactacystin treatment. 

We hypothesized that NF-κB is responsible for upregulation of eNOS protein expression 

which may represent one of the counterregulatory mechanisms activated to compensate 

decreased NO production and increased blood pressure after long-term L-NAME treatment. 

Acknowledgements: This study was supported by the grants VEGA 2/0178/09 and 

APVV-0538-07. 


87

Lectures 

INTrONIC LINE-1 INSErTION IN THE β-gloBIn GENE caUSES 

β-THalaSSEMIa DUE TO aBErraNT SPLICING, NONSENSE-MEDIaTED 

DECay aND DECreaSED raTE of β-gloBIn L1 

aLLELE traNSCrIPTION 

Lucie Piterková 1 , Jana Kučerová 1 , Karel Indrák 1,2 and Vladimír Divoký 1,2 

1 

Department of Biology, Faculty of Medicine, UP Olomouc 

2 

Department of Hemato-Oncology, University Hospital Olomouc 

ß-thalassemia is a common hereditary hemoglobin disorder characterized by quantitative 

reduction of functional ß globin chains. The patients, mother and daughter of Ukrainian 

descent exhibited typical laboratory features of ß -thalassemia trait. Molecular analyses 

revealed that the full-length (6 kb) retrotransposon L1 was inserted in the antisense 

orientation into the intron-2 of the ß-globin gene (collaboration with Dr. J. Prchal, Salt 

Lake City, UT). The total level of expression of the affected ß-globin gene transcript was 

reduced to 10-15% of the total ß-globin mRNA and thus leading to ß + -thalassemia. Based 

on recently published data we hypothesize that RNA production of mutated gene was 

affected by the combination of several events. We demonstrated that the observed 

reduction in steady-state level of ß-globin mRNA is partially caused by aberrant splicing 

followed by activation of nonsense-mediated decay (NMD) pathway, leading to increased 

degradation of aberrant ß-globin mRNA variants. Reduction in expression of ß-globin 

mRNA from ß -globin L1 

allele comes also from altered rate of transcription. We performed 

PCR-based nuclear run-on assay and forty minutes of in vitro transcription revealed 30% 

decrease in ß-globin L1 

allele transcription rate compared to wild-type ß-globin allele. 

We also observed the ß-globin L1 

3’ enhancer sequence was fully methylated. However, 

treatment with a demethylating agent did not increase the expression of the ß-globin 

transcript of the ß-globin L1 

gene. Therefore the methylation of the ß-globin L1 

3’ enhancer 

sequence is only a secondary event probably associated with enhancer displacement by 

L1 insertion. The other known mechanisms of intronic L1-mediated gene disruption as 

premature polyadenylation and gene breaking were not detected in our case. 


Lectures 

INDUCTION OF MITOCHONDRIAL PERMEABILITY 

TraNSITION BY MICROMOLar IRON 

Jan Pláteník, Juraj Gáll, Jan Škrha, Jr., Richard Buchal, 

Eva Sedláčková and Karina Verébová 

Institute of Medical Biochemistry, First Faculty of Medicine, 

Charles University in Prague 

Mitochondrial permeability transition (MPT) plays an important role in necrotic and 

apoptotic cell death. MPT is induced by calcium and promoted by oxidative stress. In 

vivo the oxidative stress is often catalyzed by iron. In this study we investigated ability of 

micromolar iron to induce MPT in isolated mitochondria. According to literary data, Fe(II) 

over-loads antioxidant defense that shifts NAD(P)H/NAD(P) to oxidation, and the loss of 

reduced NAD(P)H promotes MPT opening. Iron can also be imported to mitochondrial 

matrix by calcium uniporter. In this study, isolated rat liver mitochondria were initially 

stabilized with EDTA and bovine serum albumin. They were energized by succinate or 

malate/pyruvate, and for MPT induction stimulated by addition of Ca or Fe(II). We measured 

mitochondrial swelling (light scatter), the inner membrane potential (fluorescent 

probe JC-1) and NAD(P)H oxidation (autofluorescence 340/465 nm). Both Ca and Fe(II) 

could induce cyclosporin A-inhibitable depolarization and swelling (MPT). Fe(II) induced 

MPT only in the presence of some residual EDTA that formed an iron complex catalyzing 

rapid oxidation of NAD(P)H. Effect of iron also required membrane potential and 

could be prevented by post-addition of membrane permeant, but not impermeant iron 

chelators. Iron was apparently needed only for induction of MPT, while its propagation 

continued through calcium release/reuptake. We conclude that both iron import and 

NAD(P)H oxidation must occur simultaneously for MPT to occur. This observation can 

help to elucidate mechanism of iron toxicity, which may be involved in pathogenesis of 

several liver diseases where cytosolic iron sequestration fails, e.g. hemochromatosis 

and alcoholic liver disease. 

Acknowledgements: Supported by GAUK 43/2006/C/1.LF and MSM0021620807. 


89

Lectures 

Is PHOSPHaTIDYLINOSITOL traNSfer aCTIvITY ESSENTIal for 

THE fUNCTION of Pdr16p? 

Katarína Poloncová, Roman Holič and Peter Griač 

Institute of Animal Biochemistry and Genetics SAV, Ivanka pri Dunaji 

A common feature of phosphatidylinositol transfer proteins (PITPs) is their ability to 

transport phosphatidylinositol (PI) between membranes in vitro. The main PITP of the 

yeast Saccharomyces cerevisiae is Sec14p. It is an essential protein that participates in 

the vesicular transport from the Golgi membranes. Two point mutations in sites invariably 

conserved among all Sec14p homologues present in yeast led to the loss of PI transfer 

activity of the Sec14p (Phillips et al. 1999, Molecular Cell 4, p. 187). 

Pdr16p is one of the Sec14p homologues with 24% homology of the primary amino acid 

sequence to Sec14p. Though the exact function of this protein is yet to be described, it is 

known that deletion of the PDR16 gene leads to increased sensitivity to azole antifungals. 

Importantly, clinical isolates of Candida albicans and Candida lusitanie resistant to azoles 

with increased expression of Pdr16p were identified. 

To determine whether the PI transfer activity is essential for the Pdr16p function, we 

prepared a mutant that is predicted to be deficient in PI transfer activity of the Pdr16p. 

Yeast cells with PI transfer deficient Pdr16p as a sole Pdr16p showed increased sensitivity 

to azole antifungals similar to the pdr16Δ cells. Sterol composition of this mutant 

was also changed compared to the wt and resembles sterol composition of the pdr16Δ 

cells. These data indicate that PI transfer activity of the Pdr16p has to be present for this 

protein to fulfill its cellular function. 

Acknowledgements: This work was supported by VEGA 2/0077/10 and VVCE-0064-07 

grants. 


Lectures 

Apoptosis – DOUBLE EDGED SWORD 

Peter Račay 1 , Jozef Hatok 1 , Mária Chomová 1 , Jana Jurečeková 1 , Peter Chudý 2 , 

Juraj Chudej 2 , Andrea Štefániková 1 and Dušan Dobrota 1 

1 

Department of Medical Biochemistry, 

2 

Clinic of Haematology and Transfusiology, JLF UK Martin 

Apoptosis is an evolutionarily conserved process that is crucial for development and homeostasis 

of tissues of multicellular organisms. Its deregulation can elicit inappropriate 

cell death associated with different diseases (e. g. neurodegenerative diseases, diabetes, 

etc.) or is associated with survival of undifferentiated or transformed cells and can lead 

to development of cancer. 

Here we present results documenting initiation of apoptosis after global brain ischemia as 

well as dysfunction of apoptosis execution associated with acute myeloblastic leukaemia. 

We have documented that global brain ischemia induces transcription independent p53- 

mediated mitochondrial apoptosis. Execution of apoptosis was associated with genomic 

DNA fragmentation and death of pyramidal neurones in CA1 layer of rat hippocampus. 

On the other hand, RT-PCR analysis documented significant increase of mRNA level of 

apoptotic protein Bax in leukaemic cells in comparison to normal leukocytes, indicating 

transcription dependent initiation of p53-mediated apoptosis. Despite apoptosis initiation, 

survival of leukaemic cells is probably associated with inhibition of apoptosis execution 

mediated by increased transcription of both bcl-X and bcl-2 genes. 

Our results showed that detailed study of mechanism of apoptosis might by useful in 

search for new effective treatment of diseases associated with either cell death or cell 

survival due to deregulation of apoptosis. 

Acknowledgement: This work was supported by grant VVCE 0064-07 


91

Lectures 

E-LEarNING – FrIEND of fOE? 

Daniel Rajdl, Jaroslav Racek and Marie Šolcová 

Institution of Clinical Biochemistry and Hematology, Medical Faculty, 

Charles University in Pilsen and Charles University Hospital in Pilsen 

E-learning as a way of teaching and learning is still encountering controversial reactions. 

Opponents argue that authoring e-learning materials is very complicated and dedicated 

to computer specialists. Furthermore, depersonalization, lack of dialogue among students 

plus teachers and poor quality and usability of many published e-learning materials add 

further strong arguments to opponents. On the other hand, e-learning fans appreciate 

its time flexibility, versatility, better control of outputs and emphasize enhanced possibilities 

of individual approach even in the large groups of students. 

In the presentation, we will focus on our personal experience with authoring e-learning 

materials and tutoring e-learning courses. We will emphasize Learning Mangement System 

Moodle and its usage especially in blended-learning (e-learning support for presence 

classes) and testing of students (exam tests). Furthermore, basic examples of Adobe 

Captivate (interactive presentations with multimedia), GoogleDocs (on-line office suite) 

and Adobe Connect Pro (with emphasis on videoconferencing) utilization in teaching will 

also be discussed. Moreover, some tips for technical equipment useful for interactive 

teaching (voting systems, interactive whiteboards) will be presented. 


Lectures 

IDENTIfICaTION of SurfaCE PrOTEINS of THE OBLIGaTE 

INTraCELLULar baCTErIUM CoXIeLLa burneTII 

Gabriela Flores Ramírez 1 , Zuzana Bílková 2 , Pavol Vadovič 1 and Ľudovít Škultéty 1,3 

1 

Department of Rickettsiology. Institute of Virology, SAS, 845 05 Bratislava, Slovakia. 

2 

Department of Biological and Biochemical Sciences, Faculty of Chemical Technology, 

University of Pardubice, Pardubice, Czech Republic, 3 Centre of Molecular Medicine, 

Slovak Academy of Sciences, Bratislava, Slovakia. 

Coxiella burnetti is an obligatory, intracellular bacterium that causes Q fever in humans. 

Due to the important role of surface proteins in adhesion, invasion, and intracellular 

survival of pathogens in the host, these proteins represent the crucial bacterial antigens, 

drug targets or biomarkers for diagnostic purposes. The aim of our investigation was to 

extract and identify the surface proteins of C. burnetii in virulent phase I. 

Two different strategies were applied. The first is based on protein extraction by detergent 

Triton X-114 followed by separation of the extracted proteins by SDS-PAGE and tryptic 

digestion. The second one is the surface shaving procedure employing magnetic beads 

with immobilized trypsin. The tryptic peptides were then separated by nanocapillary liquid 

chromatography and the surface proteins were identified by tandem mass spectrometry. 

Each of the presented methods has some advantages over those conventional ones and 

they allowed us to identify high numbers of proteins predicted to be surface localized. In 

addition to those lipoproteins, ribosomal proteins and metabolic enzymes were identified. 

Acknowledgements: This contribution/publication is the result of the project implementation: 

„TRANSMED“ supported by the Research & Development Operational Programme 

funded by the ERDF. 


93

Lectures 

XENOBIOTIC-METABOLIZING ENZYMES POLYMORPHISMS 

AND CANCER RISK 

Monika Kmeťová Sivoňová 1 , Dušan Dobrota 1 , Tatiana Matáková 1 , Zuzana Tatarková 1 , 

Mária Kovalská 1 , Martina Pavlíková 1 , Róbert Dušenka 2 and Ján Kliment 2 

1 

Department of Medical Biochemistry, Comenius University, Jessenius Faculty 

of Medicine, Martin, 2 Department of Urology, Comenius University, Jessenius Faculty 

of Medicine and University Hospital, Martin 

Research in the past few years has shown that genetic, socioeconomic and environmental 

factors, particularly diet and lifestyle can affect the process of carcinogenesis. 

It is assumed that increased exposure to procarcinogens and carcinogens contained in 

tobacco smoke, debris, fermented food, polluted water, air etc., is implicated in multistage 

carcinogenesis. Xenobiotic-metabolizing enzymes play a key role in the metabolism of 

drugs and environmental chemicals and in the metabolic activation and detoxification 

of procarcinogens. Phenotyping analyses have revealed an association between these 

enzymes activities and the risk of developing several forms of cancer. The gene polymorphism 

of xenobiotic-metabolizing enzymes can cause interindividual differences in 

the activation/inactivation of anticancer agents/carcinogens and understanding of the 

contribution of these enzymes gene polymorphisms and their interactions with other 

relevant factors may improve screening diagnostic assays for cancer. 

Acknowledgements: This work was supported by grants MH SR 2007/45-UK-10, MH SR 

2007/57-UK-17, MVTS Bil/ČR/SR/UK/06, AV 4/0013/05 and project “Center of Translational 

Medicine” co-financed from EC sources and European Regional Development Fund. 


Lectures 

StrUCTUral anaLYSIS of tau prOTEIN, THE CONSTITUENT of 

NEUrofIBrILLary paTHOLOGY IN aLZHEIMEr’s DISEaSE 

Rostislav Skrabana 1,2 , Radovan Dvorsky 3 , Branislav Kovacech 1,2 , 

Zuzana Flachbartova 1 , Ondrej Cehlar 1 , Jozef Sevcik 4 and Michal Novak 1,2 

1 

Institute of Neuroimmunology, Slovak Academy of Sciences, Bratislava, Slovakia; 

2 

Axon Neuroscience GmbH, Vienna, Austria; 3 Max Planck Institute for Molecular 

Physiology, Dortmund, Germany; 4 Institute of Molecular Biology, Slovak Academy 

of Sciences, Bratislava, Slovakia 

Neuronal protein tau is one of the prominent microtubule-associated proteins in the 

brain. Tau shows typical features of an intrinsically disordered protein including low sequence 

complexity and large proportion of polar and charged amino acids. Tau protein 

amino acid composition implicates a high net charge at physiologic pH and a low mean 

hydrophobicity, increased hydrodynamic volume and a negligible content of secondary 

structure. Tau protein has an important role for the development of axons; on the other 

hand it is implicated in misfolded paired helical filaments (PHF) occurring in Alzheimer’s 

disease and other tauopathies. Traditionally, tau protein flexibility posed substantial 

limitations for assessing its structure using spectroscopic and/or diffraction techniques. 

However, recent technology advancements allowed deeper insight into tau protein 

conformational freedom. Using monoclonal antibodies as surrogate tau binding partners 

it was possible to stabilize a distinct tau protein conformation which can be studied by 

X-ray crystallography. Monoclonal antibody MN423, which recognizes a conformation 

of tau protein assembled into Alzheimer PHF core could be used as a molecular mould 

for studying PHF fold using disordered molecule of recombinant tau and adopting X-ray 

crystallography, biophysical techniques and molecular dynamics methods. Similarly, 

other antibodies directed against distinct parts of tau protein molecule, provided data 

about conformational space of tau. 

Acknowledgement: This work was supported by the Slovak Research and Development 

Agency under the contracts Nos. APVV-0471-06, APVV-0634-07, LPP-0038-09, by the 

Slovak Grant Agency VEGA grants Nos. 2/6172/26, 2/0162/10, 2/0217/10 and by the 

International Centre for Genetic Engineering and Biotechnology grant No. CRP/SVK05-01. 


95

Lectures 

ENErGETIC aSPECTS of a MODIfICaTION of THE Na + /H + 

aNTIPOrTEr aCTIvITY IN a HarmaLINE rESISTaNT MUTaNT of 

MethanothermobaCTer thermautotrophICus 

Peter Šmigáň 1 , Monika Vidová 1 , Janette Bobalová 2 and Zuzana Nováková 1 

1 

Institute of Animal Biochemistry and Genetics, SAS, Ivanka pri Dunaji, 

2 

Institute of Analytical Chemistry, Academy of Sciences of the Czech Republic, Brno 

One of the most remarkable features of methanogenic archaea is the coexistence of two 

+ 

primary ion gradients, Δμ~ H+ 

and Δμ~ Na 

participating in ATP synthesis. Na + /H + antiport is 

uniquely able to balance these electrochemical gradients. However, physiological functions 

and protein components responsible for Na + /H + remain hypothetical. In this study 

a spontaneous mutant of M. thermautotrophicus resistant to the Na + /H + antiporter inhibitor 

harmaline was isolated. The Na + /H + exchanger activity in the mutant cells was remarkably 

decreased in comparison with wild -type cells. ATP synthesis driven by methanogenic 

electron transport was significantly enhanced in the mutant cells. To define the protein 

basis of harmaline resistance, the composition of membrane-associated proteins was 

partially characterized and compared with that of the wild- type strain. The experimental 

data revealed the differential expression in this mutant of A flavoprotein and Molybdenum 

–containing formylmethanofuran dehydrogenase 1 subunit C which play a direct role in 

flavin based electron bifurcation. The overexpression of these proteins might contribute 

to harmaline resistance. Taken together the results indicate that harmaline resistance 

in this mutant has arisen as a consequence of mutation(s) in an antiporter gene(s) or 

a protein(s) that links or influences an antiporter activity. 

Acknowledgements: This investigation was supported in part by the Science and Technology 

Assistance Agency (Slovak Republic) APVT-51- 024904, APVV-VVCE 0064-07 and by Research 

Grants VEGA 2/0015/09 from the Slovak Academy of Sciences and the Institutional 

Research Plan AV0Z40310501 of the Institute of Analytical Chemistry of the ASCR, v.v.i 


Lectures 

CarDIAC L-TYPE CALCIUM CUrrENT IN SEPSIS 

Milan Štengl 1 , František Barták 1 , Roman Sýkora 2 , Jiří Chvojka 2 , Jan Beneš 3 , 

Aleš Kroužecký 2 , Ivan Novák 2 , Jitka Švíglerová 1 , Jitka Kuncová 1 and Martin Matějovič 2 

1 

Department of Physiology, 2 First Medical Department, 3 Department of Anesthesiology 

and Critical Care Medicine, Charles University in Prague, Faculty of Medicine 

and Teaching Hospital in Plzen, Plzen, Czech Republic 

Myocardial depression is a well recognized manifestation of sepsis and septic shock. We 

hypothesize that reduced L-type calcium current (ICaL) with consequent shortening of 

cardiac repolarization contributes to the myocardial depression in a clinically relevant 

porcine model of hyperdynamic septic shock. In anesthetized, mechanically ventilated 

and instrumented pigs (n=22), sepsis was induced by bacteremia (central venous infusion 

of live P. aeruginosa) and continued for 22 hours. Electrocardiogram was recorded before 

and 22 hours after induction of bacteremia. In vitro, action potentials were recorded 

in right ventricular trabeculae. ICaL was measured in isolated ventricular myocytes. RR, 

QT and QTc intervals were significantly shortened by sepsis. Action potential durations 

(APD) were shortened in septic preparations. TNF-α did not influence APD. Peak ICaL 

density was reduced in myocytes from septic animals (8.3±0.4 pA/pF vs. 11.2±0.6 pA/pF 

in control). The voltage dependence of both ICaL activation and inactivation was shifted 

to more negative potentials in myocytes from septic animals. Action potential-clamp 

experiments revealed that the contribution of ICaL to the septic action potential was 

significantly diminished. In cardiac myocytes incubated with TNF-α ICaL was not further 

affected. We conclude that in a clinically relevant porcine model, hyperdynamic septic 

shock induced reduction of ICaL and shortening of ventricular repolarization. 

Acknowledgement: Supported by the Research Project MSM 0021620819, Replacement 

of and support to some vital organs, from the Ministry of Education, Czech Republic. 


97

Lectures 

CYTOCHrOME P450- aND PErOXIDaSE-MEDIaTED OXIDaTION of 

ELLIPTICINE DICTaTES ITS aNTI-TUMOr effICIENCY 

Marie Stiborová 1 and Eva Frei 2 

1 

Department of Biochemistry, Faculty of Science, Charles University in Prague, 

2 

Division of Molecular Toxicology, German Cancer Research Center, 

Heidelberg, Germany 

An antineoplastic alkaloid ellipticine is a prodrug, whose pharmacological efficiency is 

dependent on its cytochrome P450 (CYP)- and/or peroxidase-mediated activation in target 

tissues. The CYP-mediated ellipticine metabolites 9-hydroxy- and 7-hydroxyellipticine 

and the product of ellipticine oxidation by peroxidases, the ellipticine dimer, are the 

detoxication metabolites of this compound. Two carbenium ions, ellipticine-13-ylium and 

ellipticine-12-ylium, derived from two activation ellipticine metabolites, 13-hydroxyellipticine 

and 12-hydroxyellipticine, generate two major deoxyguanosine adducts in DNA 

found in the human breast adenocarcinoma MCF-7 cells, leukemia HL-60 and CCRF-CEM 

cells, neuroblastoma IMR-32, UKF-NB-3 and UKF-NB-4 cells and glioblastoma U87MG 

cells in vitro and in rat breast carcinoma in vivo. Formation of these covalent DNA adducts 

by ellipticine is the predominant mechanism of its cytotoxicity and anti-tumor activity 

to these cancer cell lines. Ellipticine is also an inducer of CYP1A, 1B1 and 3A4 enzymes 

in the cancer cells and/or in vivo in rats exposed to this compound, thus modulating 

its own pharmacological efficiencies. The study forms the basis to further predict the 

susceptibility of human cancers to ellipticine and suggests this alkaloid for treatment 

in combination with CYP and/or peroxidase gene transfer increasing the anticancer 

potential of this prodrug. 

Acknowledgements: Supported by GACR (P301/10/0356) and Czech Ministry of Education 

(MSM0021620813 and 1M0505). 


Lectures 

UTILITY OF SINGLE CELL RT-PCR (SCrT-PCR) FOR THE STUDY OF 

PRIMarY affERENT NEURONS - PRELIMINarY vaLIDATION 

Lenka Surdeníková 1 , Fei Ru 2 and Marian Kollárik 1,2# 

1 

Pathophysiology, Jessenius Medical School, 2 Medicine, Johns Hopkins School 

of Medicine, # correspondence: kollarik@jhmi.edu 

We addressed the hypothesis that scRT-PCR detection of selected receptors in primary 

sensory neurons provides information predictive of their functional response mediated 

by these receptors. scRT-PCR was performed on individual mouse or guinea pig primary 

sensory neurons. Functional response of the neurons or their putative peripheral nerve 

terminals was evaluated by calcium imaging, whole cell patch clamp or single nerve fiber 

recordings. scRT-PCR detection of MrgPrA 3 

receptor in neurons correlated with their 

responsiveness to the MrgPrA 3 

selective agonist chloroquine (1mM) in calcium imaging 

studies. MrgPrA 3 

mRNA was detected in 8 of 9 chloroquine-responsive neurons but not 

in 11 chloroquine -unresponsive neurons. Similarly, detection of TRPV1 receptor correlated 

with the responsiveness to the TRPV1 agonist capsaicin (1microM) in patch clamp 

studies (n=9). Detection of the purinergic receptors P2X 2 

/P2X 3 

in the nodose nociceptive 

neurons correlated with the P2X 2 

/P2X 3 

current signature in these neurons in patch 

clamp studies. scRT-PCR detection of the adenosine A 1 

and A 2A 

receptors, and the TRPA1 

receptor in the vagal nodose nociceptive neurons correlated with the responsiveness of 

their putative nerve terminals to the selective adenosine A 1 

and A 2A 

receptor agonists 

and TRPA1 agonists, respectively, in single fiber recording studies. We conclude that 

scRT-PCR detection of all receptors evaluated thus far in our studies in primary sensory 

neurons correlated with the functional response mediated by these receptors. Our data 

indicate that scRT-PCR on the individual primary sensory neurons provides information 

predictive of their functional response and is a suitable complementary tool for the 

study of these neurons. 


99

Lectures 

MYCOBaCTErial maNNOSYL traNSferaSE PIMa as a TarGET for 

THE DEvELOPMENT of NEW aNTITUBErCULar drUGS 

Zuzana Svetlíková 1 , Marcelo E. Guerin 2 , Mary Jackson 3 , 

Jana Korduláková 1 and Katarína Mikušová 1 

1 

Comenius University, Faculty of Natural Sciences, Bratislava, Slovakia; 

2 

University of the Basque Country, Unit of Biophysics, Bilbao, Spain; 

3 

Colorado State University, Department of Microbiology, Immunology and Pathology, 

Fort Collins, USA 

Tuberculosis still remains one of the most serious infectious diseases in the world with 

several million new cases each year. At present there are more than 2 billion people 

infected with Mycobacterium tuberculosis, the causative agent of tuberculosis, with 

a chance of one in ten for the development of the active disease. Increased prevalence 

of multidrug-resistant and extensively drug-resistant strains of M. tuberculosis diminishes 

the number of effective drugs available for the treatment of infections they cause. For 

this reason there is a need for drugs with new mode of action. The unique mycobacterial 

cell envelope provides an array of novel drug targets since its integrity is necessary 

for bacterial viability. Phosphatidylinositol mannosides (PIM) are important part of this 

crucial structure. They appear to be involved in host-pathogen interactions and serve as 

a basis for the biosynthesis of other key molecules - mycobacterial lipopolysaccharides 

lipoarabinomannan and lipomannan. Build up of PIM is initiated by the action of mannosyl 

transferase PimA catalyzing the transfer of the mannose residue from GDP-mannose to 

phosphatidylinositol. Since the reaction is essential for survival of M. tuberculosis, PimA 

represents an attractive target for the drug development. 

Acknowledgment: This work was supported by European Comission under contract 

LSHP-CT-2005-018923”NM4TB“ 


Lectures 

oxidaTIve STrESS aND HEart aGING. 

Zuzana Tatarková, Eva Babušíková, Stanislav Kuka, Ján Lehotský, Peter Račay, 

Dušan Dobrota and Peter Kaplán 

Department of medical biochemistry JLF UK Martin 

Aging plays in our life uniquely role and oxidative stress (OS) is considered by many 

authors as a major factor in this process. The causes and consequences of aging involve 

the interaction of many processes. Mitochondria are the main sources of ROS (reactive 

oxygen species) during normal metabolism. ROS production by mitochondria is important 

because it underlies oxidative damage in a lot of pathologies and is characterized 

by destruction of structural integrity of membrane, leading to a decrease in membrane 

fluidity and enzyme activities. Therefore, the objective of our study was to determine 

the specific relationship between heart aging and changes in the level of OS, lipid peroxidation 

and OS-sensitive enzymes activities. We used three different age groups (6-, 

15- and 26-months old rats) which represented adult, old and senescence. The activities 

of all respiratory enzymes significantly decreased with the highest activity losses (37%) 

in complex IV. Inhibition of citric acid cycle enzymes, a-KGDH and SDH in the hearts of 

senescent rats, is also due to reaction of ROS with the thiol groups of these proteins. For 

this reason we studied total thiol group content, which decreases by 30%. The whole aging 

process was associated with elevation in basal levels of lipid peroxidation-end products, 

MDA and HNE. Cells are continually challenged by conditions that cause acute or chronic 

stress and there are many antioxidant mechanisms that cope with this situation. Activity 

of the Mn-SOD was together with cytosolic CuZn-SOD continuously depressed with age, 

but mitochondria creates higher level of MnSOD protein, which possibly could help to 

another antioxidant enzymes fight against OS. In all this reasons, OS in mitochondria 

may drive myocardial aging. But there are still many processes that we need investigate. 

Acknowledgements: This work was supported by grants VEGA 1/0027/08, VEGA 1/0049/09, 

VVCE-0064-07. 


101

Lectures 

WHEN MOre IS LESS: a DILEMMa of a BIOMEDICal EDUCaTOr 

Ľubomír Tomáška 

Department of Genetics, Comenius University, Bratislava 

The exponential growth of data due to a dramatic increase in the number of active 

researchers as well as development of advanced and powerful technologies results in 

a general, yet difficult question related to science education: How and what to teach the 

future generation of biomedical scientists? Whereas 30 years ago the classical textbooks 

covered essentially the whole fields by means of their presentation in a form of highly 

readable and entertaining recapitulation of major discoveries, the modern textbooks, due 

to their factual nature, do not provide the reader with an opportunity to „participate“ in 

the exciting experiments. It seems logical that in 1976 the key experiment in molecular 

genetics describing the fine structure of a gene [1] is elaborated on 15 pages [2], while 

the most recent edition of the same textbook barely mentions its main message [3]. The 

changes of textbooks in both the richness of their content and style should reflect the 

content of undergraduate curricula. Or should they, really? Should the student have an 

overview of everything considered important by the major textbooks and thus naturally 

gain a shallow and superficial knowledge? Or should we employ key, yet technologically 

outdated experiments to catalyze creative thinking risking that the student will not have 

comprehensive information about the state-of-the-art in the corresponding discipline? 

We need to look for means to dissect this dilemma and think about the changes in the 

curricula [4] and ultimately in re-definition of goals of higher biomedical education [5]. 

[1] Benzer, S. (1955). Proc. Natl. Acad. Sci. USA 41: 344-354. 

[2] Watson, J.D. (1976). Molecular biology of the gene. 3 rd Edition, W.A. Benjamin. 

[3] Watson, J.D. et al. (2008). Molecular biology of the gene. 6 th Edition, CSHL Press. 

[4] http://www.asbmb.org/CareersAndEducation.aspx?id=432 

[5] Connelly. T. et al. (2008). A new biology for the 21 st century. The NAS Press. 


Lectures 

EffECT of aDENINE NUCLEOTIDES aND Mg 2+ IONS ON 

MITOCHONDrial CHLOrIDE CHaNNELS 

Viera Kominková, Zuzana Tomášková, Ľubica Máleková and Karol Ondriaš 

Institute of molecular physiology and genetics, Slovak Academy of Sciences, 

Bratislava, Slovak Republic 

Chloride channels are involved in many physiological and pathological processes such 

as the apoptosis, stress, reperfusion injury or cardioprotection. The cells in ischemic 

tissues are depleted of oxygen and lose their capacity for ATP production. Due to this 

energy deprivation, destructive processes are activated which lead to cell injury or death. 

Molecular mechanisms of these processes are still not fully understood. The aim of our 

work was to study the possible role of mitochondrial chloride channels in these processes. 

We report the effect of adenine nucleotides (ATP, ADP and AMP-PNP) and Mg 2+ on the 

activity of mitochondrial chloride channels. Submitochondrial vesicles isolated from 

rat heart were incorporated into bilayer lipid membrane and single chloride channel 

currents were measured. We found that ATP inhibited chloride channels and affected 

both kinetics and current amplitude. This inhibitory effect was not dependent on phosphorylation 

of the channel. Furthermore, ADP did not affect the channel activity but 

only decreased the current amplitude. When the effect of ATP was studied in Mg 2+ free 

solutions, a hyperbolic current-voltage relationship was observed after addition of ATP. 

Addition of MgCl 2 

(up to 5mmol/L) partially relieved this effect of ATP. 

ATP, within the range of physiological concentrations, inhibits the mitochondrial chloride 

channels, which invokes a hypothesis that these channels have a role during pathological 

processes connected with ATP depletion. 

Acknowledgements: This work was supported by VEGA 2/0150/10 and VVCE-0064-07. 


103

Lectures 

SYNTHETIC CYCLIC CHALCONE ANALOGUES AS NOVEL BIOLOGICALLY 

ACTIVE DYES 

Vladimíra Tomečková 

Department of Medical Chemistry, Biochemistry, Clinical Biochemistry 

and LABMED a.s. UPJŠ Košice 

This study demonstrates the behaviour of synthetic fluorescent substituted chalcone 

(1) and its cyclic chalcone analogues: indanone (2), tetralone (3), benzosuberone (4) 

with dimethylamino substituent in para position in the presence of BSA proteins, egg 

yolk lecithine vesicules and cells studied by fluorescence spectroscopy. These dyes with 

anticancer properties are yellow and have protein, lipid and cell staining properties. 

From the spectral results we can assume that all studied compounds interacted with 

lipids and proteins by hydrophobic interactions. Tetralone and benzosuberone showed 

the best hydrophobic interaction with lipids and proteins. In the presence of proteins 

all compounds showed batochromic shift of the spectra and increase of fluorescence 

intensity. Biologically the most active dye (4) was the least fluorescent and the least biologically 

active dye (1) was the most fluorescent. Interaction of these unpolar substances 

with proteins was concentration dependent and it was caused by hydrophobic bonding. 

From these measurements we have calculated the binding constant of these dyes with 

BSA protein, which was in order 4 > 3 > 2 > 1. The application of these biologically active 

dyes (1, 2, 3, 4) on breast cancer cell lines showed that these fluorescent dyes were 

able to stain all cellular components (cytoplasm, nucleus, lipid vesicules). The result 

fluorescence images of breast cancer cells staining were aqua tyrkys fluorescence studied 

by fluorescence microscope. Nucleus we have costained by DAPI dye, specific for DNA 

staining. Dye (3) as well as dye (4) destroyed nuclei of breast cancer cells after 3 hours 

and all studied dyes have cytotoxic effect on studied breast cancer cells after 24 hours. 


Lectures 

MOLECULar MODELING INSIGHT INTO CATALYTIC MECHANISMS OF 

GLYCOSYLTraNSFEraSES 

Igor Tvaroška 

Institute of Chemistry, Slovak Academy of Sciences, 845 38 Bratislava, Slovakia 

N- and O-linked oligosaccharide chains of glycoproteins play a crucial role in a number 

of biological processes. These compounds are found throughout biological systems and 

have been implicated in molecular recognition events such as bacterial, viral, and cellcell 

adhesion, inflammation, and tumor invasion. Numerous glycosyltransferases, that 

catalyze the addition of a specific glycosyl residue from sugar nucleotide to an acceptor 

substrate, were validated as prime targets for therapeutic intervention in human diseases. 

Effective inhibitors of these enzymes are not yet available and development of inhibitors 

for a specific glycosyltransferase is, therefore, of great interest. Though transition state 

analogs are valued tools for drug discovery as potent and specific inhibitors of enzymes, 

up to date, the ability to generate transition state analogs of glycosyltransferases has 

lagged behind. A transition state analog is a stable compound that structurally resembles 

the three-dimensional structure and charge distribution of a substrate(s) portion of 

the unstable transition state of an enzymatic reaction. Knowledge of the geometry and 

charge distribution of transition state provides blueprint for the design of the transition 

state analog inhibitors. It is obvious, that design of transition state analog inhibitors of 

an enzyme requires knowledge of the mechanism of the enzymatic reaction and the 

structure of transition state. Therefore, we have investigated the catalytic mechanism for 

inverting and retaining glycosyltransferases employing high level ab initio, DFT, and hybrid 

QM/MM calculations [1-6]. These results provided detailed insight into the mechanism 

of the monosaccharide transfer catalyzed by glycosyltransferases and revealed the main 

structural features of the transition states. The purpose of this paper is to summarize 

the structural insights into the catalytic mechanism of glycosyltransferases inferred from 

these calculations. 

Acknowledgements: This work was supported by the grants from the Slovak Research 

and Development Agency No. APVV-0607-07 and the Slovak Grant Agency VEGA No. 

2/0128/08. 


105

Lectures 

LivING COLOUrs: ILLUMINaTE YOUr aSSaYS WITH CLONTECH 

Jiří Vašák 

KRD molecular technologies s.r.o., Saratovská 26, 841 02 Bratislava, Slovakia 

With the Living ColorsR Fluorescent Proteins, you have access to the largest selection 

of fluorescent proteins on the market. Choose from 20 fluorescent proteins in 

6 distinct colors: far red, red, orange, yellow, green, and cyan. The Living Colors fluorescent 

proteins provide a valuable, noninvasive approach for investigating biological 

events in living cells and tissues. They are some of the most widely used reporters 

in biological research. Ideal for monitoring gene expression and protein localization 

in vivo, in situ, and in realtime. Excellent reporters to monitor promoter activity. 

Effective markers to visualize specific tissues, whole cells, or subcellular organelles. 

Most of our fluorescent proteins mature rapidly in vivo, permitting detection within hours 

of transfection. Each emits a distinct wavelength which is easily detected by fluorescence 

microscopy or flow cytometry. This makes it easy to perform multiplex experiments - that 

is, combine multiple fluorescent proteins for the simultaneous detection of two or more 

events in the same cell or cell population. With the appropriate filters and excitation 

sources, you can resolve as many as three, and in some cases four, fluorescent proteins 

at the same time. 


Lectures 

DIffERENCIES BETWEEN TWO ENDOXYLANASES frOM GH5 

Mária Vršanská, Katarína Šuchová, Vladimír Puchart and Peter Biely 

Department of enzymology of Carbohydrates, Institute of Chemistry, 

Center for Glycomics, SAS, Bratislava 

Endoxylanase A(XynA) produced by plant pathogen Erwinia chrysanthemi and endoxylanase 

IV(XynIV) isolated from Trichoderma reesei RUT C-30 are classified into glycoside 

hydrolase family 5 (GH5). A detailed study of both endoxylanases showed on differencies 

in their mode of action on variety of plant glucuronoxylans and linear xylooligosaccharides. 

The hydrolytic action of XynA was found to be absolutely dependend on the presence of 

4-O-methyl-D-glucuronosyl (MeGlcA) side residues in both oligosaccharides and polysaccharides. 

Neutral linear xylooligosaccharides are resistant towards enzymatic action of 

XynA. As a rule, the enzyme attacked the second glycosidic linkage following the MeGlcA 

branch towards the reducing end. The productive complex of XynA with glucuronoxylan 

is formed in such a way that the MeGlcA residue is bound to the xylopyranosyl residue 

accommodated at the hypothetical subsite -2. Depending on the distribution of MeGlcA 

residues on glucuronoxylan main chain, XynA generated series of shorter and longer 

aldouronic acids in which the MeGlcA is linked exclusively to the second xylopyranosyl 

residues from the reducing end. Endoxylanase Trichoderma reesei XynIV also from GH5 

do not recognize uronic acid chains as substrate determinants. XynIV showed lover afinity 

toward deacetylated glucuronoxylan than to acetylated beechwood glucuronoxylan 

or to rhodymenan, a sea alga β-1,3-β-1,4-xylan. The behaviour on polymeric substrates 

suggested that XynIV prefers xylan with irregularities in the main chain such as the replacement 

of β-1,4-linkages by β-1,3-linkages as it is in rhodymenan, or substitution of 

xylopyranosyl residues as it is in acetylglucuronoxylan. [1- 3 H]-Linear xylooligosaccharides 

are exclusively cleaved by XynIV at the first glycosidic linkage from the reducing end, to 

give [1- 3 H]-xylose as the only radioactive product. All experimental evidence suggests 

that the substrate-binding site of XynIV is composed of three subsites -II, -I and +I with 

a serious steric barrier at the position of the hypothetical subsite +II. Our data point to 

a great diversity among endoxylanases belonging to the GH5 family. 


107

Lectures 

ATOrvaSTATIN CHANGES MEMBraNE LIPID FLUIDITY IN 

MITOCHONDRIA ISOLATED frOM varIOUS TISSUES OF raTS 

Iveta Waczulíková 1 , Oľga Uličná 2 , Oľga Vančová 2 , Jarmila Kucharská 2 , 

Veronika Ilovská 1 and Libuša Šikurová 1 

1 

Faculty of Mathematics, Physics and Informatics, 2 Pharmacobiochemical Lab., 

3 rd Med. Clinic, Faculty of Medicine, Comenius University, Bratislava, Slovakia 

Atorvastatin is a potent cholesterol-lowering agent with well-described benefits and 

adverse effects manifested in clinical practice. Hypercholesterolemia leads to alterations 

in lipid composition and fluidity of blood cell plasma membranes and atorvastatin was 

reported to reverse these alterations. Little is known how the hypercholesterolemic 

condition and its therapy may influence mitochondrial membrane properties. We have 

examined the effect of two selected doses of atorvastatin on membrane lipid fluidity 

and function of mitochondria isolated from liver, heart and skeletal muscles of control 

and hypercholesterolemic rats. Wistar rats were on a high cholesterol diet for 8 weeks. 

Atorvastatin was administered per os either at a low or high dose (10/80 mg/kg/day, resp.) 

for 4 weeks. Fluidity was assessed spectrofluorometrically and functional parameters of 

mitochondria with a Clark oxygen electrode. 

Hypercholesterolemic condition affected the investigated properties of mitochondria in 

a tissue-dependent manner. This is likely the reason of an ambiguous action of atorvastatin 

on membrane fluidity. Atorvastatin seems to eliminate certain types of membrane 

damage induced by hypercholesterolemia, if it is controlled by an appropriate dosage. 

However, on average, it tended to fluidize the membranes and compromise capacity of 

mitochondrial respiratory chain as well as the rate of ATP formation in mitochondria in 

both, healthy and hypercholesterolemic rats. 

Acknowledgement: Supported by the grants: VEGA 1/0328/10 and 1/0293/08. 


Lectures 

HOW CAN OXIDATIVE STRESS AND DNA STABILITY BE INFLUENCED 

BY DIET AND PHYSICAL ACTIVITY? 

Karl-Heinz Wagner and Oliver Neubauer 

Department of Nutritional Sciences, Emerging Field Oxidative Stress and DNA Stability, 

University of Vienna, Austria 

Oxidative stress-induced DNA damage and insufficient DNA repair may play an important 

role in the etiology of cancer, diabetes and arteriosclerosis. Participation in regular 

physical activity as well as plant based diet reduces the risk of developing such and 

other lifestyle-dependent diseases. Clear evidence for protective effects was observed 

for several foods although no correlations between oxidative/antioxidative parameter 

and DNA relevant biomarker can be observed. 

Interestingly, acute and strenuous exercise may induce oxidative stress via enhanced 

formation of reactive oxygen (ROS) and nitrogen species (RNS) leading to oxidatively 

modified lipids, proteins and nucleic acids and possibly to lifestyle dependent diseases. 

Exercise-induced DNA damage might either be a consequence of oxidative stress or 

inflammatory processes or it is causally involved in inflammation and immunological 

alterations after strenuous prolonged exercise (e.g. by inducing lymphocyte apoptosis and 

lymphocytopenia). Currently, only a few data have investigated the influence of exercise 

on DNA stability and damage with conflicting results, small study groups and the use of 

different sample matrices or methods and result units. The presentation will address 

results of food based studies as well as the effects of exercise of various intensities and 

durations on DNA stability, focusing on human population studies. It will consider the 

type of exercise conducted (field or laboratory based) and the intensity performed (i.e. 

competitive ultra/endurance exercise or maximal tests until exhaustion). The findings 

suggest that competitive ultra-endurance exercise (>4 h) does not induce persistent DNA 

damage. However, when considering the effects of endurance exercise (

Lectures 

LECTINS frOM PATHOGENS: MYSTERY OF LIFE 

Michaela Wimmerová 

National Centre for Biomolecular Research & Department of Biochemistry, Faculty 

of Science, Masaryk University, Brno, Czech Republic 

Protein/carbohydrate interactions play a crucial role in host cell recognition by pathogens. 

Many of them have at disposal a wide arsenal of proteins that selectively recognize host 

cabohydrate epitops. In addition, while a general protein/saccharide binding is usually 

very weak, pathogens have evolved proteins that can bind even monosaccharides with 

unusually high affinities. 

Contribution will be focused on some examples of lectins from pathogens, which can 

mediate recognition and adhesion to host cells so that they could be important virulence 

factors. The combination of binding experiments (isothermal titration microcalorimetry, 

surface plasmon resonance,...) and X-ray crystallography approaches is used to decipher 

the thermodynamical and structural basis for high affinity binding of these lectins to host 

carbohydrates. Site-directed mutagenesis in combination with structural and functional 

studies is used for understanding particular amino acids for sugar specificity and preference. 


Lectures 

FUNCTION AND PROPERTIES OF HEarT AND KIDNEY MITOCHONDRIA 

(MIT) IN SPONTANEOUSLY HYPERTENSIVE (HYP) raTS: INFLUENCE OF 

CAPTOPRIL AND NIFEDIPINE 

Attila Ziegelhöffer 1 , Jana Mujkošová 1 , Oľga Uličná 2 , Iveta Waczulíková 3 , Miroslav ferko 1 , 

Norbert Vrbjar 1 , Štefan Polák 4 , Tanya Ravingerová 1 and Adriana Adameová 5 

1 

Inst Heart Res SAS, 2 Pharmacobioch Lab III rd Med Clin MF UK, 3 Dept Biomed Physics 

FMFI UK, 4 Inst Histol Embryol MF UK, 5Inst Pharmacol Toxicol, PaF UK, Bratislava 

There are only scarce data available about HYP-induced changes in function and properties 

of heart MIT and multi-organ studies also involving the effect of anti-HYP treatment 

with captopril (CAP) or captopril + nifedipine (CAPNI) are missing completely. In present 

study we complete this lack by estimating O 2 

consumption and oxidative phosphorylation 

(Oxygraph Gilson), DNP stimulated Mg-ATPase activity and membrane fluidity 

(fluorescence probe DPH) in MIT isolated from heart and kidney of 16 weeks old HYP 

rats treated for the next 4 weeks with CAP (80 mg.kg -1 daily) or CAPNI (80 CAP+10 NI 

mg.kg -1 daily). HYP induced a moderate elevation of MIT ATP production in response to 

HYP-induced increase in energy demands (ED) of the heart. By removing the HYP, both 

treatments also decreased the ED and normalized the MIT ATP production. In kidney MIT 

HYP decreased the ATP production by depressing the values of OPR. Both treatments, 

particularly that with CAPNI even deepened this damage. 

Acknowledgements: Grants: VEGA 1/0620/10; 2/0173/08.. 


111

Lectures 

THE ROLE OF ANGIOTENSIN II AND OXYTOCIN IN REGULATION OF 

ADIPOCYTE CELL SIZE 

Katarína Kršková, Daniela Ježová, Lucia Gajdošechová, 

Miroslava Eckertová and Štefan Zorad 

Institute of Experimental Endocrinology, Slovak Academy of Sciences, Bratislava 

It is well recognized that adipocytes produce a number of proteins collectively named 

adipokines and thus adipose tissue plays a central role in development of cardiovascular 

and metabolic disorders. It has been shown that hypertrofic adipocytes produce more 

proinflammatory and insulin sensitivity decreasing adipokines (Zorad et al. 2006). This 

study presents treatment with angiotensin II receptor type I blocker (ARB) or with oxytocin 

in order to manipulate adipose tissue morphology with stimulated production of 

insulin sensitizing adipokines. 

Chronic ARB treatment reduced adipose tissue mass, adipocyte size, leptin and TNFα 

gene expression and leads to an elevation of serum adiponectin concetration, adiponectin 

and PPARγ gene expression in adipose tissue without a change in food intake. Oxytocin 

treatment did not affect the adipose tissue mass and food intake in rats. Despite that, the 

adipocyte diameter was significantly decreased and it was accompanied by a significant 

elevation of gene expression of aP2, PPARγ, GLUT4, leptin, ACE and CD31. 

Our results demonstrate positive effects of renin-angiotensin system inhibition on fat 

tissue remodeling and insulin sensitivity. Our oxytocin data suggests a new approach to 

modulate adipose tissue morphology in order to stimulate expression of insulin sensitivity 

markers. We assume that oxytocin increases both adipogenesis and angiogenesis 

in adipose tissue. 

This work was supported by grant VEGA 2/0162/08. 


POSTERS 


113

Posters 

I. BIOCHEMISTry aND MOLECULar BIOLOGY of 

NErvOUS SYSTEM 


Posters 

1. 

LABOraTORY BIOMarKERS IN ISCHEMIC STROKE AND 

DEPRESSION IN HUMAN PATIENTS 

Daniel Čierny 1 , Stanislav Celec 1 , Mária Kovalská 1 , Peter Kaplán 1 , Igor Ondrejka 2 , 

Egon Kurča 3 and Ján Lehotský 1 

1 

Department of Medical Biochemistry, 2 Psychiatric clinic, 2 Neurological clinic, 

Jessenius Fac Med, Comenius University, Martin, 03601, Slovakia 

Due to the extreme complex pathogenesis of ischemic stroke, prognostic value of 

laboratory parameters is still matter of debate. A number of brain biomarkers for the 

diagnosis of ischemic stroke have been evaluated for clinical use in the past several 

years. Neurobiological basis of depression is not yet clarified and it is not yet clear the 

etipathogenesis of poststroke occurred depression. We present here laboratory examinations 

of plasma derievd analytes in the group of ischemic stroke patients, in the group 

of patients with depression compared with the group of apparently healthy subjects. 

Laboratory results indicate for a time dependent association between ischemic damage 

and biochemical parameters in postischemic plasma. It also suggests for their use as an 

additional predictors of tissue damage progression. 

Supported by: VEGA 0049/09, COST B30, VVCE 0064/07, MZ 2007/55UK-16. 


115

Posters 

2. 

IS IT POSSIBLE TO IMPROVE DEMYELINATION DISEASES MONITORING BY 

DETERMINATION OF SOME ENZYME ACTIVITIES CHaraCTERISTIC FOR 

THE CENTraL NErvOUS SYSTEM? 

Monika Ďurfinová 1 , Marta Brechtlová 1 , Ľubica Procházková 2 , Peter Kukumberg 2 , 

Ľubomír Kuračka 1 and Branislav Líška 1 

1 

Department of Chemistry, Biochemistry and Clinical Biochemistry, LF UK Bratislava, 

2 

2 nd Department of Neurology, LF UK Bratislava 

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system 

(CNS). It is the most common neurological disease characterized by recurrent relapses 

and/or progression that are attributable to multifocal inflammation, demyelination and 

axonal pathology within the CNS. MS lesions are typically located in the periventricular 

white matter of the brain as well as in superficial areas of the spinal cord. Since MS 

lesions are not routinely biopsied, the cerebrospinal fluid (CSF) is used for measurements 

of various soluble markers. We have started determination of enzyme activities 

in the CSF of MS patients, which have some relationship to the function of the CNS 

tissue – phosphodiesterase 3 , ,5 , -cAMP and K + -paranitrophenyl-phosphatase. These 

enzyme activities were at first monitored in model experiments in vitro. We suppose 

that K + -paranitrophenylphosphatase might have a connection with the nerve impulse 

transmission. Interestingly, the part of phosphodiesterase 3 , ,5 , -cAMP activity, which is 

calcium-calmodulin dependent, was inhibited by the conditions of activated oxidative 

stress. However, several questions still remain to be elucidated, e.g.: if this CNS inflamatory 

damage could infuence releasing of these enzymes into CSF and if these enzymes 

can reflect the intensity of the patological process. 


Posters 

3. 

SELECTED GENE POLYMORPHISMS IN ISCHEMIC STrOKE aND 

DEPRESSED HUMan paTIENTS frOM CENTraL SLOvaKIA 

1 

Andrea Evinová, 1 Eva Babušíková, 2 Pavol Adamík, 2 Igor Ondrejka, 3 Egon Kurča, 

3 

Milan Grófik and 1 Ján Lehotský 

Comenius University, Jessenius Faculty of Medicine, Department of Medical 

Biochemistry 1 , Psychiatry Clinic 2 , Neurological Clinic 3 , Martin, Slovakia 

The neurobiological basis of ischemic stroke and postischemic depression is not yet clarified. 

We have focused on several genes which could be connected with the pathogenesis 

of ischemic stroke and depression. Angiotensin converting enzyme (ACE), as a part of 

the renin-angiotensin system, is important factor in blood pressure regulation. Incidence 

of D allele is associated with higher protein level and its activity. Serotonin-transporterlinked 

promoter (HTTLPR) is a d egenerate polymorphic region in the gene that codes the 

serotonin transporter. Its polymorphism is connected with neuropsychiatric disorders. 

Likewise, S allele is associated with the functional reduction as compared to L allele. 

Glutathione-S-tranferases (GST) detoxify a broad range of xenobiotics and carcinogens. 

The most studied polymorphisms are in families of GSTM1, GSTT1 and GSTP1. In our 

study we tested the gene polymorphisms in three groups of patients, with: i) ischemic 

stroke ii) depression, and iii) healthy controls. In the DD polymorphism of ACE gene, the 

depressed pacients exhibit higher frequency of DD genotypes in comparison to controls. 

Stroke patients exhibit only non-significantly lower frequency in comparison to controls. 

So far, we did not observe any significant differences in allelic frequency for HTTLPR 

gene neither for depressed nor ischemic groups in comparison to controls. Our results 

suggest that ACE DD genotype is increased in depressed pacients. Analysis of selected 

polymorphisms of GSTM1 and GSTT1 genes show higher frequency of GSTM1 null and 

GSTT1 wild genotypes. 

This work was supported by the VEGA 49/09 and MZ-2007/55-UK-16 


117

Posters 

4. 

An anaLYSIS of THE IMPaCT of CNS ISCHEMIa ON MITOCHONDrial 

rESPIraTOry COMPLEXES 

Mária Chomová and Peter Račay 

Institute of Medical Biochemistry, Jessenius Faculty of Medicine in Martin, 

Comenius University in Bratislava 

Mitochondria are responsible for aerobic respiration and ATP synthesis by oxidative 

phosphorylation. In addition to being crucial for energy production and metabolic 

pathways, they also play key roles in the signal cell network. Ischemia/reperfusion is 

a multifactorial process that leads to disturbance of key mitochondrial functions and can 

initiate a cascade of events, which result in tissue damage and finally in the cell death. 

So the existence of mitochondrial signal network evokes numerous responses and effects, 

which depend on the signal nature, cell environment and the individual ability of 

the cell to deal with the signals. In this regard, the goal of the work was to contribute to 

understanding the different processes in mitochondria after ischemic attack, particularly 

in connection with mitochondrial respiratory complexes I and IV. The impact of 15 min 

global brain ischemia followed by 1, 3, 24 and 72 hours reperfusion on complexes I and 

IV in cortex and hippocampus was studied by spectrophotometric determination of their 

enzymatic activities by using two electron acceptors, decylubichinone and ferricyanide 

and two ways of membrane permeabilisation (sonification vs detergent). The noticed 

variations were analyzed by electrophoretic methods SDS PAGE and two- dimensional 

native BN PAGE/SDS PAGE. The posttranslation modifications of selected structural 

subunits of respiratory complexes were monitored by Western blotting and immunodetection 

of 3-nitrotyrosines as markers of oxidative damage to proteins. Add to this, 

the interest was focused on processes of the p53-mitochondrial pathway of apoptosis 

where the levels of p53 protein as well as pro- and antiapoptotic members of Bcl 2 family 

were monitored. A possible involvement of the complex I structural subunit GRIM-19 in 

apoptotic processes was also studied. 


Posters 

5. 

THE ROLE OF MAP-KINASE PATHWAY IN GLOBAL ISCHEMIA/ 

REPErfUSION INJURY OF raT BraIN afTER INDUCED 

HYPERHOMOCYSTEINEMIA 

Mária Kovalská 1 , Martina Pavlíková 1 , Zuzana Tatarková 1 , Peter Kaplán 1 , 

Dušan Dobrota 1 , Marian Adamkov 2 and Ján Lehotský 1 

1 

Department of Medical Biochemistry, 2 Department of Histology and Embryology 

Comenius University, Jessenius Fac Med Martin 

An altered cross-talk in intracellular signaling pathways is presumed in the mechanisms 

in ischemic injury. The MAPK/ERK is a signal transduction route that couples intracellular 

responses to cell surface receptors. ERK protein is part of the cascade leading to survival 

of neurons after injury. Tolerance induced by preconditioning (IPC) is tissue adaptation 

to sub-lethal ischemia. Hyperhomocysteinemia (hHcy) is one of the risk factor, which 

could have negative impact on the onset/progression of ischemia/reperfusion injury 

(IRI). In these experiments, we have studied changes in MAPK pathways and related 

enzymes after global IRI in forebrain homogenate. In addition, the effects of: i) ischemic 

preconditioning (IPC) and ii) hyperhomocysteinemia (hHcy) on IRI-associated alternations 

of protein levels of MAPK pathway was determined. Global forebrain ischemia was 

induced by 4-vessels occlusion. Rats were preconditioned by 5 min of sub-lethal ischemia 

and 2 days later, 15 min of lethal ischemia with reperfusion period of 1h, 3h, 24h and 

72h was induced. hHCy was induced by twice a day subcutaneous injection of homocysteine 

(0.45 µmol/g). Immunohistochemical as well as Western blot analysis identified 

MAPK/ERK protein in injured areas. The highest level of the protein was detected in the 

reperfusion time after IPC. Converse effect was observed in the reperfusion time after 

induced hHcy. This suggests that adaptive mechanisms in the MAPK signal transduction 

machinery might have a potential role in tissues response subjected to IRI and in the 

phenomenon of tolerance. 

Acknowledgments: Supported by VEGA 0049/09, VVCE 0064/07, UK-16/2007 and 

UK/10/2010. 


119

Posters 

6. 

ANATOMICAL DISTRIBUTION OF DYING CELLS WITHIN ADULT raTS 

ROSTraL MIGraTORY STREAM 

Marcela Martončíková, Kamila Lievajová, Juraj Blaško, 

Judita Orendáčová and Enikő Račeková 

Institute of Neurobiology, Slovak Academy of Sciences, Košice, Slovak Republic 

The rostral migratory stream (RMS) is a pathway along which newborn cells originated 

from the subventricular zone migrate toward the olfactory bulb where they differentiate 

into interneurons. This migratory pathway consists of three parts in caudo-rostral 

direction: the vertical arm, the elbow and the horizontal arm. The precursor cells are 

able to proliferate during migration in the RMS. It has been reported that these cells may 

also undergo apoptotic cell death. Dying cells has been observed along entire extent 

of the RMS. The number and distribution of dying cells in the RMS varies according to 

the methodology used. Commonly applied methodology for identification of apoptotic 

cells is a terminal dUTP nick-end-labelling (TUNEL). Another marker for detection of dying 

cells is a fluorochrome, Flouro Jade-C (FJ-C), which stains all degenerating neurons, 

regardless of mechanism of cell death. Studies dealing with the cell death in the RMS 

assess the total number of positive cells mainly in whole extent of the RMS. In this 

study we focused on the distribution of FJ-C positive cells in individual anatomical parts 

of the RMS. We found that the number of dying cell is the highest in the caudal part 

of the RMS – in the vertical arm. The lowest number of dying cells was observed in the 

middle part of the RMS –in the elbow. In the rostral part of the RMS- in the horizontal 

arm the number of these cells was higher than in the elbow. Differences in the number 

of dying cells in the individual parts of the RMS are probably associated with different 

developmental origin of these parts. 

Acknowledgments: Supported by VEGA grants: 2/0147/09; 2/058/08. 


Posters 

7. 

SECRETORY PATHWAYS SPCA1- CA2+ PUMP EXPRESSION AS ParT OF 

ISCHEMIC PRECONDITIONING IN raT FOREBraIN 

Martina Pavlíková, Mária Kovalská, Monika Kmeťová Sivoňová, 

Zuzana Tatarková and Ján Lehotský 

Department of Medical Biochemistry Jessenius Faculty of Medicine, 

Comenius University, Martin, Slovakia 

Neural cells of the brain have important secretory functions. They secrete many neurotransmitters 

and secretory proteins. The Golgi apparatus, as part of secretory pathways 

(SP), is recognized Ca2+ store, which regulates secretion by SPCA- Ca2+ATPase. In addition, 

SP are involved in stress sensing and transduction of apoptotic signals. Ischemic 

preconditioning (IPC) is an important phenomenon of adaptation of tissue to sub-lethal 

short-term ischemia, which results in increased tolerance of tissue to the lethal ischemia. 

In this study we have investigated the changes in SPCA1 expression after global 

ischemic injury (IRI) in hippocampal and cortical areas. In addition, the effects of IPC on 

IRI associated alternations of mRNA and protein levels of SPCA1 were determined. Rats 

were preconditioned by 5 min of sub-lethal ischemia and 2 days later, 15 min of lethal 

ischemia followed by reperfusion for 1h, 3h and 24h. RT-PCR and Western blot analysis 

clearly detected expression of SPCA gene in injured area after IRI insult. In addition, 

tissue response on the level of mRNA was maximal in the reperfusion period in both 

paradigms. IPC did not change significantly the expression profile, however magnitude 

of cell response was elevated. Protein level of SPCA was highest in the reperfusion time. 

Our results showed that IPC affects gene expression and SPCA translation in forebrain 

induced by ischemia. This suggests for a potential role of SPCA regulated secretory processes 

in adaptation of neuronal circuits as response to preischemic challenge. 

Acknowledgement: This work was supported by grants VEGA 0049/09, UK 10/2010, 

VVCE 0064/07. 


121

Posters 

II. BIOTECHNOLOGY 


Posters 

8. 

CHaraCTERIZATION OF BACTERIOPHAGES 

INFECTING CronoBACTer STraINS 

Hind Al Alami, Ľubomíra Tóthová, Michal Kajsík, Jana Gajdošová, 

Hana Drahovská and Ján Turňa 

Department of Molecular Biology, Faculty of Natural Sciences, Comenius University, 

Mlynská dolina 1, 842 15 Bratislava, Slovensko 

Cronobacter spp. (previously known as Enterobacter sakazakii) is an opportunistic 

pathogen, implicated in food-borne diseases especially in neonates and infants. Although 

the microorganism is widely distributed in the environment, dried infant milk formula 

has been implicated as the vehicle of transmission in many clinical manifestations. The 

current threat of antibiotic-resistant bacteria has renewed the interest in exploring 

bacteriophages as biocontrol agents. 

The aim of the our study was to isolate set of bacteriophages infecting Cronobacter 

strains and to assess their suitability to be used as antimicrobial agents in food industry. 

Six bacteriophages infecting Cronobacter were isolated from wastewater samples of 

three Bratislava wastewater treatment plants (Petržalka, Vrakuňa and Devínska Nová 

Ves) by propagating on indicator strains. We have determined the host specificity of 

bacteriophages on the collection of 16 strains. We have observed, that bacteriophages 

were able to infect 3-13 strains, three phages has specificity restricted to C. sakazakii 

species, the other three bacteriophages posses the broader host range specificity; they 

were able to inhibit growth of 11-13 strains of all Cronobacter species. By restriction 

mapping we observed different profiles for each phage. 


123

Posters 

9. 

EffECT of METHYL jaSMONaTE ON THE prODUCTION of 

saNGUINarINE aND ITS prECUrSOrs IN OPIUM POPPY SUSPENSION 

CULTUrES 

Andrea Balažová 1 , Víťazoslava Blanáriková 1 , Jindra Valentová 2 , 

František Bilka 1 and Ivana Holková 1 

1 

Department of cellular and molecular biology of drugs, FaF UK Bratislava 

2 

Department of chemical theory of drugs, FaF UK Bratislava 

Suspension cultures of opium poppy are not able to produce morphinan alkaloids but 

synthesize and store sanguinarine within a single cells. The morphinans and sanguinarine 

share common biosynthetical pathway, leading to (S)-reticuline as a crutial intermediate. 

The biosynthesis of these alkaloids begins with the conversion of tyrosine to both 

dopamine and 4-hydroxyphenylacetaldehyde. The suspension cultures of opium poppy 

enhenced their sanguinarine production after treating with three different concetrationes 

of methyl jasmonate (10, 100, 1000 μmol/l). The presence of elicitor in nutrient medium 

of suspension cultures also led to the changes in levels of sanguinarine precursors (DOPA, 

dopamine, tyrosine and tyramine). The suspension cultures were exposed to elicitror 

treatment for 2, 4, 6, 8, 10, 24 and 48 h. The maximum content of sanguinarine was 

detected after 8 h exposure to elicitor at concetration 100 μmol/l (5.8-fold increase over 

the control). The suspension cultures produced the highest amount of DOPA between 

2-6 h after elicitation with all three concetrationes of methyl jasmonate. The level of 

dopamine increased gradually in elicited cultures. The maximal content of dopamine was 

detected after 8 h exposure to elicitor at concentration 10 μmol/l (3.24-fold increase). 

The level of tyrosine started to increase later than the levels of DOPA and dopamine. The 

accumulation of tyrosine was positively affected mainly by methyl jamonate at concetrationes 

10 and 100 μmol/l. The levels of tyramine have not been changed significantly 

in elicited suspension cultures. 


Posters 

10. 

COMParISON OF SANGUINarINE PRODUCTION OF PAPavEraCEAE 

faMILY PLANTS IN VITro CULTURES 

František Bilka, Andrea Balažová, Andrea Bilková, Víťazoslava Blanáriková, 

Ivana Holková and Marián Vanko 

Department of cellular and molecular biology of drugs, Faculty of Pharmacy, 

Comenius University in Bratislava 

Intact plants of Papaveraceae family are producers of the whole range of benzylisoquinoline 

alkaloids, which are used in pharmaceutical industry. In vitro cultures derived from 

plants of Papaveraceae do not have the ability to produce so broad spectrum of alkaloids 

– active is only biosynthetic pathway leading to sanguinarine. We derived in vitro cultures 

of Papaver somniferum, Eschscholtzia californica, Chelidonium majus and Macleaya 

cordata. Their sanguinarine production abilities were tested and compared. Also the 

effect of elicitation on the sanguinarine production was studied. The lowest amounts of 

sanguinarine from all cultures tested were accumulated in suspension cultures of opium 

poppy (0.45 – 0.55 mg in 1 g of fresh weight). Eschscholtzia calicornica, Chelidonium majus 

and Macleaya cordata cultures produced similar amounts of sanguinarine (18.0 – 22.7 mg; 

20.5 – 26.3 mg; 15.4 – 20.3 mg in 1 g of fresh weight, resp.). In elicitation studies we used 

biotic (Botrytis cinerea hydrolysate) and abiotic (CuSO 4 

, methyljasmonate) stressors. In 

all cultures treated the increase in sanguinarine accumulation was observed. Among the 

elicitors used the most effective was B. cinerea hydrolysate. From all cultures tested the 

most intensive response was observed in opium poppy cultures (increse to almost 9 mg 

of sanguinarine in 1 g of fresh weight), although the amount of sanguinarine in elicited 

poppy cultures was lower than in non-elicited cultures of the other cultures. 


125

Posters 

11. 

DESIGN of an EXPrESSION SYSTEM for THE PRODUCTION OF 

RECOMBINANT HUMAN THROMBIN IN ESCherICHIA COLI 

Michaela Kandričáková 1 , Stanislav Stuchlík1 and Ján Turňa 1,2 

1 

Comenius University, Faculty of Natural Sciences, Department of Molecular Biology, 

2 

Department of Molecular Biology SAV Bratislava 

Thrombin is a plasmatic protease, which plays a key role in blood coagulation. This form of 

active prothrombin is characterised by many biological functions including procoagulation 

and anti coagulation effects. The polyfunctionality of thrombin has received increasing 

interest in biofarmaceutical and biomedical research. One of the possible ways to gain 

recombinant human thrombin is to use expression systems based on E.coli cells. E.coli 

offers many adventages over other host organisms and therefore remains the most 

commonly used host for the production of heterologous proteins. Prethrombin-2, an 

activation intermediate of thrombin, is often expressed as a precursor of prothrombin. 

Presumably, the use of this gene assures higher transcription and translation efficiency 

compared to prothrombin gene mainly because of the smaller size of the mRNA transcript. 

During the preparation of prethrombin-2 syntetic gene the known sequence will be used 

with optimized E.coli codon usage ensuring higher expression yields. 

The aim of this work is the design of proper expression system for the production of 

recombinant human thrombin in the E.coli host cells. The study is focused on selection of 

proper gene sequence with codon usage optimized for E.coli, selection of suitable expression 

system and in the next step we would like to optimise physiological conditions for the 

foreign protein expression and subsequent verify expression of recombinant thrombin. 


Posters 

12. 

HETEROLOGOUS EXPRESSION OF ROYAL JELLY APALBUMINS IN E. COLI 

Tatiana Kraková, Jozef Šimúth and Katarína Bíliková 

Department of Molecular Apidology, Institute of Molecular Biology, SAS Bratislava 

The honeybee royal jelly (RJ) contains many valuable components and biologically active 

proteins and peptides. Major proteins of RJ, designated as apalbumins belong to a protein 

family consisting of nine members with M r 

of 49-87 kDa. Apalbumin1, apalbumin2 and 

apalbumin3 account for 90% of the RJ protein content and are 88% identical in amino 

acid sequence. Minor RJ proteins are mainly homologues of apalbumins, antimicrobial 

peptides and enzymes. RJ proteins display multifunctional features. For example, apalbumin1, 

apalbumin2 stimulated tumor necrosis factor alpha release. We have focused our 

attention to the molecular characterization of apalbumin2, the second most abundant 

protein of RJ. Apalbumin2 is a basic protein of 52.7 kDa with eight isoelectric variants in 

the range of pI 7.5-8.5. For first time we purified royal jelly and characterized minority 

homologue of apalbumin2 named as apalbumin2a (48.6 kDa) with antibiotics activity 

against P. larvae, the inducer of American foulbrood of honeybee colony. 

The aim of our study was a biotechnological preparation of apalbumin1 and apalbumin2 

and their homologues by heterologous expression in E. coli with the goal of study their 

antibiotic and immunostimulatory properties. For these purposes we cloned appropriate 

cDNA or their of PCR fragments into pQE30 and pET28b(+) expression vectors and 

transformed into E. coli DH5α and/or BL21-CodonPlus(DE3)-RIL. We present the molecular 

properties of recombined apalbumins and their homologues as well. 


127

Posters 

13. 

EXPRESSION, PURIFICATION AND FUNCTIONAL CHaraCTERIZATION 

OF TWO FORMS OF agroBACTerIUM SP. STraIN CP4 EPSPS GENE IN 

ESCherICHIA COLI FOR HORIZONTAL GENE TraNSFER STUDIES 

Mahesh Madyagol, Stanislav Stuchlík and Jan Turňa 

Comenius University, Department of Molecular Biology, Faculty of Natural Sciences, 

Bratislava, SLOVAKIA 

The Agrobacterium sp. strain CP4 aroA gene encodes an enzyme called 5-enol-pyruvylshikimate-3-phosphate 

(EPSP) synthase. The enzyme is widely involved in glyphosate 

tolerant transgenic plants because it is the primary target of the nonselective herbicide 

glyphosate. We have cloned this gene and constructed a system for the high level expression 

of a recombinant form of this enzyme by amplifying the aroA gene from the 

genetically modified maize genomic DNA and the coding sequence of EPSPS gene was 

successfully subcloned in a plasmid-Escherichia coli system. Furthermore, the truncated 

form of CP4 EPSPS synthase has been created in order to compare in vitro glyphosate 

sensitivity between the two forms of enzymes. The cells containing the plasmid carrying 

both forms of EPSPS gene exhibited enhanced tolerance to herbicide glyphosate, 

compared to the control. The resulting plasmids produced the EPSP synthase in large 

quantities which has been purified to homogeneity and enzyme activity assay has been 

carried out. The study of horizontal gene transfer (HGT) is planned in the future work. 


Posters 

14. 

CONFORMATION TraNSITIONS OF CYTOCHROME C IN DEEP EUTECTIC 

SOLVENTS 

Jozef Parnica 1 , Lukáš Kandráč 1 and Marián Antalík 1, 2 

1 

Department of Biochemistry, Faculty of Science, P.J. Safarik University, Kosice, 

Slovak Republic, 2 Institute of Experimental Physics, SAS, Kosice, Slovak Republic 

Original deep eutectic mixtures (DES) were formed by mixing two components together in 

different molar ratios that had a melting point much lower than any of the constituents, 

until homogenous, colorless liquid was formed. Deep eutectic solvents were also included 

in the advanced ionic liquids. They have recently attracted much attention as “green” 

alternatives to conventional organic solvents in various fields including biochemistry and 

separation processes due to their unique properties such as high thermal stability, negligible 

vapor pressure and non-flammability. In our work, we applied DES’s as alternative 

solvents to determinate the stability and conformation transitions of a model protein, 

ferricytochrome c. We used urea, malonic acid, and sorbitol as hydrogen bond donors 

by UV-Vis spectroscopy and circular dichroism spectroscopy in mixtures of substituted 

quaternary ammonium salts e.g. hydroxyethyltrimethyl-ammonium (choline) chloride. 

We have found DES’s as suitable solvents for characterization new types of conformation 

states of ferricytochrome c that demonstrate their advantages and unique properties, 

and suggest opportunities for new applications. 

Acknowledgement: This work was supported within the projects Nos., 26220120001, 

26220220005, 2622022033 in frame of SF EU, Centre of Excellence of SAS Nanofluid and 

VEGA 0056, 0038 and 0079. 


129

Posters 

15. 

PrODUCTION of rECOMBINaNT prOTEINS USING HIGH CELL DENSITY 

CULTUrES of ESCHErICHIa COLI IN BIOreaCTOrs 

Lucia Pánčiová, Zdenko Levarski, Pavol Utekal, Stanislav Stuchlík and Ján Turňa 

Comenius University Faculty of Natural science, Department of Molecular Biology 

Mathematical modeling of processes, information techniques and database generation 

have received increasing interest of biotech industry in recent years. For the generation 

of predictable mathematical models and databases of fermentation processes a vast 

amount of data is required to precisely characterize the growth dynamics and microorganism 

metabolism. These data are generated using systematic approaches. Systems 

of metabolic engineering are an ideal way for the development of bacterial strains 

producing bioproducts or aminoacids. Production of recombinant proteins using high 

cell density cultures of Escherichia coli became one of the most often used methods in 

biotech industry, mainly because of the high volumetric productivity associated with this 

method, but also because of the vast amount of data available regarding to host genetics 

and physiology. Application of different cultivation techniques results in achievement 

of cell densities higher than 100 g of dry cell weight per liter of cultivation media. The 

most common problems during cultivation of cells to high densities are limited oxygen 

solubility in cultivation medium, formation of growth inhibitory by-products, accumulation 

of acetate and CO 2 

and generation of heat by the cells. The basic parameter of the 

growth kinetics observation is the specific growth rate, which can be different for every 

microorganism, type of the cultivation media or cultivation technique used. In this work, 

we have focused on the construction of recombinant strains of E.coli, suitable cultivation 

media selection, definition of crucial cultivation parameters, optimization of gene 

expression, monitoring of growth kinetics and finally determination of cell viability and 

biomass concentration. 


Posters 

16. 

SYNTHESIS AND SUrfaCE MODIFICATION OF ZINC OXIDE 

naNOParTICLES 

Michaela Šimšíková 1 and Marián Antalík 1,2 

1 

Department of Biochemistry, Faculty of Science, P.J. Safarik University, Kosice, 

Slovak Republic; 2 Institute of Experimental Physics, SAS, Kosice, Slovak Republic 

Zinc oxide nanostructures are very attractive for their unique properties and biocompatibility 

and in consequence of this have great potencial for applications in biosystems, 

for example biolabeling, biosensoring, delivery systems and others, which can be used 

in genetics, pathology, criminology, safety of food and many other industries. For these 

bioapplications are necessary surface modifications, for example with water soluble 

thiols like stabilizating agents, which can made to the nanostructures to better suit their 

integration with biological systems, leading to such interesting properties as enhanced 

aqueous solubility, bio-recognition or applicability for biological systems.. 

For synthesis of ZnO nanoparticles in aqueous solution we used three different concentrations 

of 11-mercapto-undecanoic acid MUA (0.1; 0.01 and 0.005 M) as stabilizing 

agent. The coating of nanoparticles with MUA could allow their solubility in a variety of 

organic solvent and the covalent binding through hydroxyl groups present in its structure. 

The PL spectra indicated that the optical properties of ZnO nanoparticles were changed 

with the insertion of MUA, and showed significant influence of this surface modification 

on intensity. 

Acknowledgements: This work was financially supported by the research grants VEGA 

No. 0038, 7055, 0056; VVGS PF 12/2010/CH: Slovak Academy of Science in frame of CEX 

NANOFLUID, projects ESF 26220120021 and NFP 26220220005. 


131

Posters 

17. 

CLONING, EXPrESSION aND aNTIMICrOBIal aCTIvITY of THE HUMan 

caTHELICIDIN LL-37 

Csaba Tóth 1 , Roland Pálffy 1 , Juraj Gašperík 2 , Stanislav Stuchlík 1 and Ján Turňa 1 

1 

Department of molecular biology, Faculty of Natural Sciences, Comenius University, 

2 

Institute of Molecular Biology, SAS Bratislava 

Antimicrobial peptides are short polypeptides involved in the innate immunity system 

and they can be found among all classes of life. They demonstrate antimicrobial activity 

against Gram-negative and Gram-positive bacteria, viruses, fungal pathogens and cancerous 

cells. LL-37 is a multifunctional peptide and the only antimicrobial peptide from 

the cathelicidin family found in human. It is mainly expressed in myeloid cells, where it 

is located in specific granules, but it was also described in inflamed skin, testis, wound 

fluid, lung epithelia, sweat and saliva. Apart from its’ wide spectrum of bactericidal activity, 

LL-37 also plays an important role in the regulation of the inflammatory response, 

neutralization of LPS and the promotion of wound healing. 

As the therapeutical and commercial importance of these peptides is rising, simple 

expression and purification systems will be needed for their large-scale production. 

We have designed and prepared a cost-effective and efficient expression system for 

the production of LL-37 in fusion with ketosteroid isomerase in E. coli cells. After the 

purification and activation of the peptide, we have tested its’ biological activity against 

various bacterial pathogens. 


Posters 

18. 

CHaraCTERIZATION OF BACTERIOPHAGES INFECTING 

SALMoneLLA enTerICA 

Ľubomíra Tóthová, Hind Al Alami, Jana Lintnerová, Hana Drahovská and Ján Turňa 


Bratislava, Slovakia 

Food-borne infections caused by Salmonella enterica represent a major healthcare concern 

worldwide. Bacteriophages are viruses infecting specifically bacteria. With regard 

to their characteristics, they can be considered as a potential antibacterial agents. In 

these applications bacteriophages are suitable to prevent or reduce colonization and 

diseases in livestock, to decontaminate carcasses and other raw products, to disinfect 

equipment and contact surfaces, etc. 

The aim of our study was to isolate bacteriophages infecting Salmonella, which can be 

used for elimination of these pathogenic bacteria in food technology. Bacteriophages 

were isolated from wastewater samples of three Bratislava wastewater treatment 

plants (Petržalka, Vrakuňa and DevínskaNováVes) by propagating on indicator strains. 

Six bacteriophages infecting Salmonella were isolated and used for further investigation. 

We have determined the host specificity of bacteriophages on the collection of 16 

strains belonging to different salmonella serotypes. Two different testing methods, the 

cross test and the colorimetric microplate test, were used rendering similar results. 

We have observed, that bacteriophages were able to infect 4-14 indicator strains and 

that all strains were sensitive to at least one bacteriophage. The chromosomal DNA of 

bacteriophages was isolated and characterized by restriction cleavadge. We observed 

different restriction profiles for each phage. 


133

Posters 

19. 

PrODUCTION of TWO rECOMBINaNT aLCOHOLDEHYDrOGENaSES 

SUITaBLE for BIOTraNSformaTION of C-6 aLDEHYDES INTO 

COrESPONDING aLCOHOLS 

Pavol Utekal, Lucia Pánčiová, Stanislav Stuchlík and Ján Turňa 

Department of Molecular biology Prif UK Bratislava 

C-6 aldehydes and alcohols contribute to the fresh green odor in plants and are widely 

used in perfumes and in food technology. Important member of this family is trans-2- 

hexenol. Carbonyl compounds such as aldehydes and alcohols are reduced by chemical 

methods in industry but it is not appropriate for production of compounds used in 

food industry. Therefore, in recent decades biocatalysis is used for these purposes. The 

enzymes suitable for reduction of aldehydes are oxidoreductases, which catalyze the 

reduction of carbonyl groups of aldehydes and alcohols. The most suitable enzyme from 

this class is alcoholdehydrogenase (ADH), which is the last enzyme involved in lipoxygenase 

pathway in several species of higher plants, which converts natural trans-2-hexenal 

to trans-2-hexenol. In the case of redox reactions catalyzed by oxidoreductases, which 

often require stoichiometric oxidation or reduction of costly coenzymes such as NAD(P) 

and FAD, efficient coenzyme recycling must be accomplished if a given application is to 

be practical from an economic standpoint. Formate dehydrogenase (FDH) is one of the 

frequently used biocatalysts for NADH regeneration. This work is focused on the cloning, 

expression, purification and measurements of enzyme activity of two enzymes - ADH 

from Saccharomyces cerevisiae and FDH from Candida boidinii. 


Posters 

20. 

EUKarYOTIC EXPrESSION as an INDISPENSaBLE TOOL for 

prEParaTION of naTIve DIMErIC forMS of NK CELL C-TYPE LECTIN- 

LIKE rECEPTOrs 

Ondřej Vaněk 1 , Petra Celadová 1 , Jan Bláha 1 , Daniel Kavan 2 , 

Petr Pompach 2 and Karel Bezouška 1,2 

1 

Department of biochemistry PřF UK Praha, 2 Department of immunology 

and gnotobiology MBÚ AVČR Praha 

Natural killer cells are able to recognize and kill a variety of tumor and infected cells. The 

recognition is mediated by wide repertoire of cell surface receptors, 

both activation and inhibitory, belonging to two main structural families: immunoglobulinlike 

and C-type lectin-like. While ligands for the Ig-like receptors were shown to be MHC 

gp I proteins, ligands only for some of the lectin-like receptors are known up to date. 

Both families share relatively weak binding characteristics and in the case of lectin-like 

receptors, in vitro oligomerization clearly improves binding through increase in avidity. 

However, these lectin-like receptors were described mostly as dimeric in vivo and this 

minimal oligomer would be also the best for structural studies. Moreover, there is no 

crystal structure known of extracellular domain of any of these receptors in its full-length 

natural dimeric form; probably because prokaryotic expression of native dimers is rather 

difficult. Here we present successful design, cloning and production of dimeric forms 

of some mouse, rat and human NK cell lectin-like receptors belonging to NKRP1 and Clr 

families. Full-length extracellular receptor parts were expressed via transient transfection 

of HEK293 cell lines in suspension culture either alone or as a cleavable fusion with Fc 

fragment of human IgG1 to promote native dimerization. This fusion strategy resulted in 

cleaved pure covalent dimers of e.g. rat Clrb and purely dimeric Fc fusion of rat NKRP1B 

proteins and their structural characterization is currently under way. 


135

Posters 

III. BIOINFORMATICS 


Posters 

21. 

phiGENOME – a WEB-baSED GENOME brOWSEr INTENDED FOR 

DISPLAY OF PHaGE GENOMES 

Matej Stano and Ľuboš Kľučár 


Graphical representations of genomes convey quick insight into organization of coding 

sequences and other functional regions within genomes. Applications intended for generation 

of graphical representations of genomes are genome browsers. 

We present phiGENOME, a web-based genome browser providing highly intuitive and 

interactive maps of phage genomes stored in phiSITE – database of gene regulation in 

bacteriopages. It provides dynamic genome maps built on the Adobe Flash platform as 

well as an accessory semi-graphics based on precisely formatted HTML tags. phiGENOME 

is a integral part of phiSITE web interface, therefore it was optimized for visualization of 

phage genomes with emphasized display of cis-regulatory elements. Graphical content 

is generated dynamically; required annotation data are retrieved promptly from the 

MySQL back-end. The data transfer from the database to the genome browser is supplied 

via XML files. 

Genome maps acquired from phiGENOME allow graphically subtle and lucid inspection 

of phage genomes. Individual genome features are displayed as coloured boxes; genes: 

blue, promoters: red, operators: green, terminators: yellow. All actions in this browser 

can be comfortably performed with mouse. Mouse drag enables continuous sliding along 

genome; mouse scroll zooms in (up to the level of nucleotide sequence) and out; mouse 

over a feature displays tool-tip with feature details; mouse click highlights selected feature 

and opens options menu for further actions. phiGENOME is an innovatory application 

according to the mode and purpose how the Flash technology was employed. It is freely 

accessible at phiSITE web interface: http://www.phisite.org. 


137

Posters 

IV. GENOMICS 


Posters 

22. 

SITE SPECIFIC INTEGraTION OF BACTERIOPHAGE µ1/6 INTO THE 

STrePTOMYCES aureoFACIenS CHROMOSOME 

Jarmila Farkašovská and Andrej Godány 

Institute of Molecular Biology, Slovak Academy of Sciences, Bratislava 

Phage integrases are enzymes that mediate unidirectional site-specific recombination 

between two recognition sequences, the phage attachment site, attP, and the bacterial 

attachment site, attB. Site-specific recombinases can be classified into two major families 

based on amino acid sequence homology and catalytic residues, either tyrosine or 

serine. The properties of site-specificity and efficiency make phage integrases excellent 

tools for the manipulation of DNA. 

The phage µ1/6 integrates site-specifically into the chromosome of tetracycline producing 

Streptomyces aureofaciens strains. The region of µ1/6 genome involved in this process 

have been identified. The deduced 416 amino acid protein encoded by orf5 displays 

significant similarity with site-specific recombinases of the tyrosine family (sometimes 

referred to as integrase family). The phage attachment site (attP) was localized downstream 

of the putative integrase gene. The junctions (attL and attR sites) of the prophage 

with the host genome have been determined, allowing identification of the host attachment 

site (attB), which has a common 45-nucleotide core region with attP. In attB 

this core sequence consists of the 3 ’ end of the putative tRNA (Thr) (ACA) gene. To prove 

the functionality of the putative integrase gene (orf5), an integrative vector pCTint was 

constructed. This integration plasmid, consisting of the µ1/6 putative integrase gene 

and the phage µ1/6 attP, and a selectable marker (thiostrepton resistance gene for 

streptomycete) inserted stably into the chromosome of S. aureofaciens and S. lividans. 

The µ1/6 integrase with a deduced molecular mass of 47,5 kDa was overproduced in 

Escherichia coli and further analyzed. 


139

Posters 

23. 

STUDY OF THERMOTOLEraNCE ISLAND IN CronoBACTer SPP 

Jana Gajdošová 1 , Natália Kamodyová 1 , Kristína Benedikovičová 1 , Hana Drahovská 1 , 

Eva Kaclíková 2 and Ján Turňa 1 

1 


Mlynská dolina 1, 842 15 Bratislava, Slovensko 

2 

Food Research Institute, Priemyselná 4, Bratislava 

Cronobacter spp. is an opportunistic pathogen associated with sporadic cases of infections 

in infants. Dried infant milk formula has been identified as a potential source of the 

microorganism and the mode of transmission. The spread of Cronobacter in dried foods 

is promoted by its higher resistance than other Enterobacteriaceae to environmental 

stresses, including heating, drying or osmotic stress. 

The aim of our work was to characterize DNA region, which is present in some Cronobacter 

strains and which contributes to their elevated survival at 58°C. The thermotolerance 

island was sequenced in C. sakazakii LMG 5740, the length of the region was 19 kbp 

and 22 orfs with the size 141 – 2850 bp were detected. The greatest part of the region 

contained a cluster of conservative genes, most of them have significant homologies with 

bacterial proteins involved in some type of stress response, including heat, oxidation 

and acid stress. Similar organization of the region was observed in all thermotolerant 

strains. By rt-PCR approach we detected high expression throughout all thermotolerance 

gene cluster in both stationary and exponentially grown bacteria. Part of the thermotolerance 

genomic island covering orfD-G was cloned to pUC21 under lac promoter and 

transformed to E. coli host strain. Recombinant strain showed 80% increased D 58 

value 

comparing with control strain containing the vector only. 


Posters 

24. 

DNA POLYMOrPHISMS of SELECTED rEPair GENES aND rISK of LUNG 

caNCEr 

Lucia Letková 1 , Tatiana Matáková 1 , Erika Halašová 2 , 

Anna Drgová 1 and Dušan Dobrota 1 

1 

Department of Medical Biochemistry; 2 Department of Medical Biology, 

Comenius University, Jessenius Faculty of Medicine, Martin, Slovakia 

The presence of polymorphisms of repair genes contributes to individual susceptibility 

to the development of lung cancer. We investigated the association between polymorphisms 

in DNA repair genes hOGG1 (Ser326Cys) and XRCC1 (Arg399Gln), which play an 

important role in DNA base excision repair. In this case-control study, we recruited 309 

lung cancer cases and 339 healthy controls. Genomic DNA was extracted from peripheral 

blood by phenol/chloroform extraction. The genotypes of hOGG1 and XRCC1genes were 

determined by PCR-RFLP method. The differences in the distribution of genotypes and 

alleles between cases and controls were analysed using x 2 -test. The odds ratio (OR) and 

95% confidence intervals (95% Cl) were adjusted for the risk of developing lung cancer, 

alone polymorphisms or their combinations. The presence of polymorphisms of genes 

hOGG1 and XRCC1 in women represents elevated risk of lung cancer compared to men. 

The frequency of the genotype XRCC1 Gln/Gln (OR=2,32; P=0,19) represented 2-fold 

higher risk of lung cancer compared with genotype Arg/Arg in women. Genotypes Arg/ 

Arg+Ser/Cys (OR= 2,45; P=0,19), Arg/Gln+Ser/Ser (OR=2,3; P=0,08), Arg/Gln+Cys/Cys 

(OR=2,45; P=0,76) had a 2-fold increase risk of lung cancer compared with Arg/Arg+Ser/ 

Ser genotype. The frequency of the Gln/Gln+Ser/Ser (OR=7,36; P=0,012) genotypes was 

statistically significantly higher compared with Arg/Arg+Ser/Ser genotype. In summary, 

variants alleles Cys326 and Gln399 are associated with reduce enzyme activity of genes 

and DNA repair capacity. The presence of these alleles leads to increase susceptibility 

at risk of developing lung cancer. 


141

Posters 

25. 

EXPrESSION of traNSCrIPTION faCTOrs aND MICrOrNaS 

IN TGF-SS1–INDUCED EMT of BENIGN prOSTaTE EPITHELIal CELLS 

E. Lincová/Slabáková 1,2 , Zuzana Pernicová 1,2 , Eva Slavíčková 1,2 , 

Alois Kozubík 1,2 and Karel Souček 1 

1 

Dept. of Cytokinetics, Institute of Biophysics AS CR, Brno, Czech Republic. 

2 

Dept of Exp. Biology, Masaryk University, Faculty of Science, Brno, CZ. 

Epithelial-mesenchymal transition (EMT), in which epithelial cells become motile mesenchymal 

cells, is viewed as an essential early step facilitating dissemination of tumour 

cells. MicroRNAs (miRNAs) are small non-protein-coding regulators of gene expression 

that play an important role in tumorigenesis and, depending on their targets, can 

function as tumour suppressors or oncogenes. Members of the miR-200 family, which 

regulate expression of ZEB transcription factors, have been found downregulated during 

EMT, suggesting an important role in inhibition of EMT. The aim of this study is to 

analyze the kinetics of expression of selected transcription factors and miRNAs in the 

EMT of prostate cells in order to identify key molecules responsible for tumorigenicity 

and invasivity of prostate cancer. 

In our experimental system, we induced EMT by TGF-ß1 treatment in BPH-1 cells derived 

from non-tumorigenic prostate epithelial cell lines. Among the genes analyzed, SNAI2/ 

Slug was rapidly and strongly upregulated. Changes in expression levels of ZEB1 and 

ZEB2 mRNA and miRNA of miR-200 family are observed after extended periods of TGF-ß1 

treatment and correlate with E-cadherin expression and progression of EMT but may not 

be responsible for rapid upregulation of EMT-driving transcription factors. 

Acknowledgements: This work was supported by grants No. 310/07/0961 and 303/09/H048 

of the Czech Science Foundation, IGA MZD 9956-4/2008, IGA MZD 9600-4/2008, by the AS 

CR, grants no. AV0Z50040507 and AV0Z50040702 and by grant no. CZ.1.07/2.3.00/09.0020. 


Posters 

26. 

THE ASSOCIATION BETWEEN EGF 61 G/A POLYMORPHISM AND 

COLORECTAL CANCER DEVELOPMENT 

Silvia Mahmood, Tatiana Matáková, Lucia Letková, Monika Kmeťová Sivoňová, 

Jozef Hatok and Dušan Dobrota 

Department of Medical Biochemistry, Jessenius Faculty of Medicine, 

Comenius University, Martin, Slovakia 

The epidermal growth factor (EGF) gene has been demonstrated to participate in the 

pathogenesis or maintenance of several human cancers of epithelial origin. The purpose 

of the present case-control study was to investigate the association of the single-nucleotide 

polymorphism of EGF + 61 G/A with the susceptibility to colorectal cancer (CRC) 

in a population in the north of Slovakia. We have analyzed the genotype distribution of 

this polymorphism in 120 patients (65 [54,2%] men and 55 [45,8%] women, mean age 

64,2 years) and 110 healthy subjects, using the PCR-RFLP technique. Differences in allele 

and genotype frequencies were evaluated by the Chi-square (c 2 ) and Fisher exact tests. 

Our data suggests that EGF +61 G/A polymorphism may be used as a genetic susceptibility 

marker for CRC. With the aim to inhibit cancer growth and to reduce the risk of 

metastasis we adapt an avascular tumour growth model to simulate the effects of drug 

application on multicell spheroid (MCS) [1, 2]. 

Acknowledgement: This work was supported by project “Creating a new diagnostic algorithm 

for selected cancer diseases” co-financed from EC sources and European Regional 

Development Fund. 

References 

(1) Mahmood, M., Mahmood, S. Dobrota, D. (2010): A numerical algorithm for avascular 

tumor growth model. Math. Comp. Sim., Vol. 80 (6): 1269-1277. 

(2) Ward JP, King JR (2003): Mathematical modelling of drug transport in tumour MCS 

and monolayer cultures. Math. Biosci., 181(2):177-207. 


143

Posters 

27. 

SPErm DNA INTEGrITY aSSESMENT USING DIfferENT COMET aSSay 

prOTOCOLS 

Milena Matejovičová 1 , Michaela Králíková 1 , Jitka Melounová 1 , Martina Vodová 1 , 

Jana Žáková 2 and Igor Crha 2 

1 

Department of Biochemistry, Medical Faculty MU, Brno, 2 Center for Reproductive 

Medicine, Dept. of Obstetrics and Gynecology, Brno 

DNA fragmentation in the individual cells can be measured using comet assay. The 

method is based on the electrophoresis of nuclear DNA immobilized in agarose gel that 

follows after the cell lysis and DNA denaturation. Cells containing damaged DNA have 

appearance of the comet: round „head“ representig unfragmented DNA and elongated 

„tail“, which intensity and lenght depends on both the extent of DNA fragmentation and 

fragment size. Specificity of the comet assay application on sperm consists in particullar 

organization of sperm chromatine, which is much more compact than in somatic cells 

and it contains a lot of disulfide bonds between DNA and protamines. For assesing DNA 

dammage in sperm cells, effective lysis and DNA denaturation without background 

damaging effect to DNA are crucial. 

A method of modified alkaline/neutral comet assay was used. We studied basal DNA 

damage in sperms from subjects undergoing semen analysis in the Center of Assisted 

Reproduction (CAR). Semen was liquified for 1 hour at room temperature and 1 ml aliquotes 

were frozen in liquid nitrogen. DNA damage level of frozen sperm samples was 

compared with the damage in a fresh samples, analyzed on a day of collection. No differences 

between fresh and frozen material were found. Next, we compared different 

methods of cell lysis. Two different protocol types were applied: 1) proteinase K digestion 

during lysis and 2) lithium diodosalicylate (LIS) as a detergent for cell membrane and 

chromatine protein core destruction. In both protocols, dithiothreitol (DTT) was used 

for disulfide bonds reduction. In the protocol with proteinase K, we have found higher 

level of DNA damage when using DTT than without this agent. The least damaged DNA 

was observed in assays with LIS and DTT. 


Posters 

28. 

COMParaTIve STUDY of HYPOMETHYLaTING 

aCTIvITIES of 5-azaCYTIDINE CONGENErs 

Marika Matoušová, Ivan Votruba, Miroslav Otmar and Helena Mertlíková-Kaiserová 



Cancer cells often display aberrant hypermethylation which leads to transcriptional 

inactivity of various gene groups (such as tumor suppressor genes) and is therefore 

responsible for genomic instability. The use of hypomethylating agents as anticancer 

drugs is intended for reactivate methylation-silenced genes by decreasing the overall 

methylation level. Since epigenetic therapy is expected to be more specific, less toxic 

and more effective than standard chemotherapy, the search for new hypomethylating 

agents is a top-priority task. 

The goal of this study was to compare hypomethylating activity of a series of 5-azacytidine 

congeners vs. decitabine and zebularine, the well-established DNA methyltransferase 

inhibitors. Quantitative methylation-specific PCR was employed to detect the efficiency 

of individual agents on thrombospondin-1 and cyclin-dependent kinase inhibitor 2B 

hypermethylated gene loci. Overall changes in DNA methylation level were quantified 

by direct detection of 5-methylcytosines by HPLC using digested genomic DNA. 

2´-Deoxy-5,6-dihydro-5-azacytidine was identified as a promising drug candidate for 

epigenetic therapy. It has similar hypomethylating activity as decitabine, the most effective 

hypomethylating drug tested, and is less cytotoxic. 

Acknowledgments: Research project of the Institute #OZ40550506; Project #1M0508 by 

the Ministry of Education, Youth and Sports of the Czech Republic. 


145

Posters 

V. CELL REGULATIONS aND SIGNAL 

TraNSDUCTION 


Posters 

29. 

THE ROLE OF 14-3-3 PROTEIN IN REGULATION OF GLUCOSE 

TraNSPORTER GLUT4 TraNSLOCATION TO ADIPOCYTE PLASMA 

MEMBraNE 

Lucia Gajdošechová, Miroslava Eckertová, Katarína Kršková and Štefan Zorad 

Institute of Experimental Endocrinology, SAS, Bratislava, Slovakia 

Basic mechanism of insulin action is translocation of GLUT4 transport vesicles (GTV) from 

inner cell compartments to plasma membrane. Defect of this mechanism leads to development 

of insulin resistance and diabetes. Beta isoform of 14-3-3 protein binds to Thrphosphorylated 

AS160, the substrate of insulin-activated Akt protein kinase, thereby reducing 

of AS160 GTP-ase activity and subsequently activating GTV translocation in adipose tissue. 

The main purpose of the present study was to evaluate GLUT4 and GLUT1 transporters 

protein content in epididymal adipose tissue plasma membranes of young obese Zucker 

rats and correlate it with the amount of 14-3-3 protein in fat tisssue total homogenate. 

Zucker rats represent a model of obesity and insulin resistance with mild hyperglycemia. 

Based on unchanged GLUT1 and elevated GLUT4 content in plasma membrane fraction 

of obese animals we assume normal insulin sensitivity with regard to glucose transporter 

translocation in adipose tissue of 3-month-old obese Zucker rats. Augmented GLUT4 

translocation seems to be due to an enormous increase in 14-3-3 protein which might play 

a crucial role in glucose transporter activation. Surprisingly, the serum concentration of 

adiponectin was significantly elevated in obese animals despite of decreased mRNA level 

in white adipose tissue. The elucidation of 14-3-3 protein regulation, e.g. by adiponectin, 

await for further investigation. 

Ackowledgement: This work was supported by grant VEGA 2/0162/08. 


147

Posters 

30. 

ChaNGES IN COfILIN PHOSPHOrYLaTION DUrING THE aPOPTOSIS of 

LEUKEMIC JURL-MK1 CELLS 

Dana Grebeňová, Michaela Pluskalová, Zbyněk Hrkal and Kateřina Kuželová 

Institute of Hematology and Blood Transfusion, Prague, Czech Republic 

Cofilin is a key mediator of actin dynamics which is involved in processes requiring changes 

in the cytoskeleton structure, e. g. cell migration, cell division and adhesion to the extracellular 

matrix. It promotes actin filament severing and depolymerization, facilitating the 

breakdown of existing filaments and the enhancement of filament growth from newly 

created barbed ends. Cofilin phosphorylation at Ser3 is known to prevent its activity. 

We explored the effect of two antileukemic drugs (suberoylanilide hydroxamic acid and 

imatinib mesylate) on JURL-MK1 cell adhesivity to fibronectin and on cofilin activity. 

Under certain conditions, both compounds were able to enhance the cellular adhesivity 

and cofilin phosphorylation (inactivation) increased as expected. To the contrary, higher 

drug doses induced the apoptosis which was accompagnied by a decrease in cellular 

adhesivity to fibronectin and by F-actin disassembly. Surprisingly, cofilin phosphorylation 

at Ser3 markedly increased even in these conditions. This increase could be at least 

partly inhibited by the apoptosis inhibitor Q-VD-OPh, but not by the inhibitor of ROCK, 

one of the main regulators of cofilin activity. We speculate that some prominent actin 

structures have to be protected from cofilin-mediated destruction in order to assure 

the cell disintegration into apoptotic bodies. The phosphorylated (inactive) cofilin was 

concentrated in small distinct spots distributed throughout the cytoplasm. 

Ackowledgement: This work was supported by the grant No 301/09/1026 from the Grant 

Agency of the Czech Republic. 


Posters 

31. 

CHaraCTERIZATION OF a GENE ENCODING a SMALL REGULATORY RPOE 

– DEPENDENT RNA IN SALMoneLLA enTerICA SEROvar TYPHIMURIUM 

Dagmar Homerová, Bronislava Řežuchová, Henrieta Škovierová and Ján Kormanec 

Institute of Molecular Biology, Slovak Academy of Sciences, Bratislava 

Gene regulation of bacteria is a complex process in which proteins have been believed 

as the only relevant regulators for a long time. In recent years, the family of small noncoding 

RNA (sRNA) with the important role especially in post-transcription control 

was described. Enterobacterial pathogen Salmonella enterica serovar Typhimurium 

(S. Typhimurium) is an attractive model for studying sRNAs, since the research could 

clarify many processes affecting the regulation of bacterial pathogenicity. So far, more 

than 70 sRNAs with various roles in control of gene expression have been identified in S. 

Typhimurium. Among them, small antisense RNA MicA, activated by sigma factor RpoE, 

strongly represses the mRNAs of two porins, OmpA and LamB. To understand the role 

of MicA we have investigated its expression and role in virulence of S. Typhimurium. In 

vitro transcriptional analysis revealed presence of a single promoter, micAp, with the 

high similarity with the consensus RpoE promoter sequence showing clear dependence 

upon this sigma factor. Activity of micAp increased towards stationary phase and was 

induced by several stresses including heat shock, cold shock, acid stress, oxidative stress, 

ethanol, polymixin B, and by degS-specific stress elicited by unfolded C-terminal outer 

membrane protein OmpA. Lack of micA elicited an RpoE-dependent envelope stress 

response. Although in vitro phenotypic analysis revealed no significant differences between 

wild type and micA mutant strains, in vivo studies showed that the micA mutant 

is more virulent in the mouse model. 

Ackowledgement: This work was supported by the VEGA grant 2/0104/09 from Slovak 

Academy of Sciences. 


149

Posters 

32. 

MOLECULar MECHaNISMS INvOLvED IN POTENT PLaTINUM (IV) 

COMPLEX-MEDIaTED COLON caNCEr CELL SENSITIZaTION TO TraIL- 

INDUCED aPOPTOSIS 

Iva Jelínková 1,2 , Olga Vondálová Blanářová 1,2 , Jiřina Hofmanová 1,2 , Petr Sova 3 , 

Alois Kozubík 1,2 and Vaculová Alena 1,2 

1 

Department of Cytokinetics, Institute of Biophysics, AS CR, v.v.i., Brno; 2 Faculty of 

Science, Masaryk University Brno; 3 R&D, Pliva-Lachema, a.s., Brno, Czech Republic 

Platinum (II) complexes such as cisplatin are used in the therapy of many solid cancers, 

but their application is limited due to serious side effects and/or cancer cell resistance. 

Recently, platinum (IV) adamantylamine ligand-containing complex LA-12 has been introduced 

and shown as highly effective in many cancer cells including colon. TRAIL (TNFrelated 

apoptosis inducing ligand), a cytokine that belongs to the TNF family, is a potent 

and promising anticancer agent that triggers apoptosis in many cancer but not in most 

normal cells. However, some cancer cells are resistant to its effects. We demonstrated 

that LA-12 was very effective in potentiation of TRAIL-induced apoptosis in colon cancer 

cells. Molecular mechanisms involved in cell sensitization were investigated, with a particular 

focus on the death receptors, caspases, and mitochondria. We demonstrated that 

LA-12 was responsible for significant up-regulation of surface and total TRAIL receptor 

2 (DR5) protein level. Combined treatment of colon cancer with both agents resulted in 

enhanced caspase activation, and mitochondrial apoptotic pathway engagement. Our 

results showed that LA-12 is a very promising agent to be used in combined cancer therapy 

with TRAIL, and suggested the intracellular targets for the therapeutic interventions. 

Acknowledgements: This work was supported by grants No. 1QS500040507 IGA AS CR 

and 301/07/1557 GACR. 


Posters 

33. 

TraIL-INDUCED aPOPTOSIS caUSES aCTIvaTION of pro-SUrvival 

paTHWaYS IN NON-aDHErENTLY grOWING COLON caNCEr CELLS 

Lenka Kočí, Martina Hýžďalová, Alena Vaculová, 

Jiřina Hofmanová and Alois Kozubík 

Department of Cytokinetics, Institute of Biophysics, Academy of Sciences of the Czech 

Republic, v.v.i., Kralovopolska 135, Brno 61265, Czech republic 

Resistance of transformed epithelial cells to the deachment-induced apoptosis (anoikis) 

promotes cancer cell invasion and metastasis. We studied the effects of TNF-related 

apoptosis inducing ligand (TRAIL) on cytokinetic parameters and adhesive properties of 

the cell lines derived from human foetal (FHC cells) and human adenocarcinoma (HT-29 

cells) colon tissues in association with anoikis induction. 

We detected the significant decrease of TRAIL-induced apoptosis in the non-adherently 

growing HT-29 cells in comparison with the adherent cultivation. Based on this finding we 

focused our attention to detail mechanisms of the cell survival under TRAIL treatment. 

We confirmed our hypothesis of activation of pro-survival pathways, actually PI3K/Akt 

and MAPK/ERK, which are connected with focal adhesion kinase (FAK) phosphorylation. 

Increased phosphorylation of Akt and ERK kinases and also enhanced expression of FLIP 

and Mcl-1 proteins as downstream molecules of PI3K/Akt pathway were observed during 

non-adherent cultivation. 

Taken together, our data suggested that the decrease of the TRAIL-mediated apoptosis 

of colon epithelial cells induced by non-adherent type of cultivation is connected with 

activation of pro-survival pathways. 

Acknowledgements: This work was supported by grants 305/09/1526 GACR, 303/09/ 

H048 GACR, and 524/07/1178 GACR. 


151

Posters 

34. 

PrEParaTION aND fUNCTIONal anaLYSIS of PHOSPHOrYLaTION 

MUTaNT forMS of THE traNSCrIPTION faCTOr NFI 

Gabriel Kollárovič, Miroslava Kretová, Peter Baráth and Katarína Luciaková 

Laboratory of Molecular Biology, Cancer Research Institute, SAS, Bratislava 

Nuclear factors I (NFI) are transcription factors and has been implicated in many cellular 

processes such as embryonic development, cell differentiation and transformation 

processes. In vertebrates, NFI proteins are encoded by four isogenes and exist 

in multiple splice variant and can act both as activators and repressors. Recent data 

from the functional proteomic studies identify NFIs as a substrate for ATM/ATR kinase 

in response to DNA damage. Such genomic instability results in activation of a protein 

cascade(s) that leads to the cell cycle arrest. Regarding our previous data on the role of 

NFI in growth-regulated gene expression, a logical question arises whether NFI proteins 

indeed play a role in the ATM/ATR signaling pathway. For this purpose specific mutations 

in the ATM/ATR phosphorylation sites in the NFI was prepared to study its effects on 

the expression of target genes. The serine to alanine point mutations were prepared by 

PCR. Simultaneously restriction site was introduced for restriction enzyme SacI for rapid 

screening of clones bearing the mutated forms of NFI. 

Acknowledgements: This work was supported by VEGA grant No. 2/0074/08. 


Posters 

35. 

GENEraTION aND CHaraCTErIZaTION MONOCLONal aNTIBODIES 

agaINST ENDOSIaLIN, THE POTENTIal marKEr of TUMOr 

aNGIOGENESIS 

Soňa Kontseková, Anna Repič, Monika Baráthová, 

Katarína Polčicová and Jaromír Pastorek 

Institute of Virology, SAS, Dúbravská cesta 9, 84505 Bratislava 

Endosialin, also known as TEM1, or CD248, was first discovered with the monoclonal 

antibody (mAb) Fb5 as an experimental approach to identify new targets for anti-cancer 

strategies. It was described as a transmembrane glycoprotein selectively expressed on 

tumor endothelium, but its expression is barely detectable in normal human tissue. 

Recent experiments have challenged the endothelial expression of endosialin and suggested 

an expression by activated fibroblasts and pericytes. Subsequent studies have 

also confirmed that endosialin is upregulated in blood vessels in wide range of tumors 

and its expression is restricted to capillaries, which means that endosialin is probably 

involved in vascular reorganisation. It has been functionally implicated in angiogenesis, 

a process characterized by vascular branching and sprouting, which is necessary for tumor 

expansion and progression. In this work, we generated monoclonal antibodies against 

endosialin, characterized their isotypes and binding epitopes. We also analysed the 

activity of antibodies in a set of imunological assays and characterized the most stabile 

antibody VIII-16 with potential usage in next studies of endosialin function. 

Acknowledgements: This work was supported by VEGA 2/0210/09 and TRANSMED. 


153

Posters 

36. 

THE ROLE OF NFI IN P21 GENE EXPRESSION 

Miroslava Kretová, Ľudmila Šabová and Katarína Luciaková 

Laboratory of Molecular Biology, Cancer Research Institute SAS Bratislava 

p21 protein was originally identified as an inhibitor of cell cycle dependent kinases (CDK). 

p21 plays an important role in cell cycle arrest at the G1/ S checkpoint in response to 

DNA damage. p21 also modulates various processes such as cell growth, differentiation 

and apoptosis. p21 gene expression is mainly regulated at transcription level. The key 

regulator of p21 gene expression is tumor suppressor protein p53. However, expression 

of p21 may be either p53-dependent or p53-independent, which results in a network of 

control mechanisms influencing the cell cycle. Transcription factor NFI (nuclear factor-I) is 

a repressor of p21 transription. NFI and Sp1-binding sites, which play an important role 

in p21 gene expression, were identified in the p21 proximal promoter. The transforming 

growth factor β (TGF-β) activates p21 gene in G1 phase of the cell cycle. Molecular 

mechanism of the stress-induced expression of p21 is not well understood. TGF-β 

may activate signaling pathways affecting the target gene expression either directly 

by phosphorylation of Smad proteins or indirectly by activation of the MAPK signaling 

pathway. The aim of our work is to identify the signaling pathway(s) playing a role in the 

regulation of p21 gene in serum starved cells and in TGF-β-treated cells. Transfection of 

recombinant constructs bearing mutations and deletion of NFI binding sites in the p21 

promoter was used to measure the level of p21 expression in HaCaT, HCT-116 and JEG-3 

cells during cellular stress. 

Acknowledgements: This work was supported by VEGA grant No. 2/0074/08. 


Posters 

37. 

StrICT CONTrOL of aurICIN prODUCTION IN StrePTomyces 

aureofaCIens CCM 3239 INvOLvES a fEEDBaCK MECHaNISM 

Peter Kutaš, Ľubomíra Fecková, Alena Reháková, 

Renáta Nováková and Ján Kormanec 



In Streptomyces aureofaciens CCM 3239, we have previously identified a type II polyketide 

synthase gene cluster, aur1, responsible for production of the angucycline-like antibiotic 

auricin. We found out, that auricin was produced at very specific stage and afterwards 

it was degraded to a non-active metabolite(s). Two regulatory genes, aur1P and aur1R, 

whose deduced products share significant similarity with two different types of bacterial 

regulatory proteins, were investigated in order to reveal tight regulation in S. aureofaciens 

CCM 3239. Expression of the auricin biosynthetic genes is under control of the 

pathway-specific positive regulator Aur1P that belongs to the family of response regulators 

of bacterial two-component signal transduction systems. Transcriptional analysis 

revealed that the activity of the identified aur1Ap promoter is dramatically decreased 

in later stages of stationary phase and the promoter is directly activated by the auricin 

pathway specific activator Aur1P at the entry to stationary phase. Aur1P was shown to 

bind specifically the aur1Ap promoter and this binding was abolished by the presence 

of auricin and/or its intermediates. In addition, the aur1Pp promoter was negatively 

regulated by the TetR family Aur1R repressor, and its binding to the promoter was also 

obolished by the presence of an auricin intermediate(s). The results indicate specific 

feed-back mechanism of auricin production in S. aureofaciens CCM 3239. 

Acknowledgements: This work was supported by the Slovak Research and Development 



155

Posters 

38. 

ALTERED CALCIUM SIGNALING IN PERIPHEraL BLOOD MONONUCLEar 

CELLS OF CHRONIC KIDNEY DISEASE PATIENTS 

Ingrid Lajdová 1 , Viera Spustová 1 , Adrián Okša 1 and Dušan Chorvát Jr. 2 

1 

Slovak Medical University, Department of Clinical and Experimental Pharmacotherapy, 

2 

International Laser Centre, Department of Biophotonics, Bratislava 

A rise in the intracellular free calcium concentration ([Ca 2+ ] i 

) is a key signal in the initialization 

of wide range of cellular events. The elevation of [Ca 2+ ] i 

have been found in association 

with chronic kidney disease (CKD). The aim of this study was to investigate the [Ca 2+ ] i 

, 

intracellular calcium reserves, capacitative calcium entry and function of purinergic P2X 7 

receptors in peripheral blood mononuclear cells (PBMCs) of early-stage CKD patients. 

The study involved 22 healthy volunteers and 22 patients in CKD stage 2-3. The peripheral 

blood mononuclear cells (PBMCs) were separated by the Ficoll gradient centrifugation, 

and free intracellular calcium was measured using the Fluo-3 AM fluorimetry. The P2X 7 

pore function was evaluated by using a fluorescent dye ethidium bromide. 

Our results demonstrate that 1) [Ca 2+ ] i 

, intracellular calcium stores and the capacitative 

calcium entry were significantly increased already in early stages of CKD; 2) purinergic 

P2X 7 

receptors participate in intracellular calcium homeostasis regulation in CKD. 

Acknowledgements: Supported by grants No. APVT-21-033002 and VG SZU 19-90-07 


Posters 

39. 

EXPRESSION OF MICROPHTHALMIA-ASSOCIATED TraNSCRIPTION 

faCTOR CRITICALLY REQUIRES ACTIVE SWI/SNF CHROMATIN 

REMODELING COMPLEX 

Ľubica Ondrušová and Jiří Vachtenheim 

Laboratories of Molecular and Cell Biology, University Hospital Bulovka, 

Prague 8, Czech Republic 

Transcription factor MITF (microphthalmia-associated transcription factor) plays a central 

role in the expression of melanocyte-specific genes, lineage determination, and survival 

of embryonic, adult and malignant melanocytes. Here we show that the expression of 

MITF requires the presence of active SWI/SNF chromatin remodeling complex, which 

contains either Brg1 or Brm as a catalytic subunit. The SWI/SNF components were expressed 

in melanoma cell lines and only one cell line with Brg1 loss was found (SK-MEL-5 

cells). In several Brm/Brg1-expressing melanoma cell lines, knockdown of Brg1 severely 

compromised MITF expression with a concomitant downregulation of MITF targets and 

decreased cell proliferation. Although Brm was able to substitute for Brg1, sequential 

knockdown of both Brm and Brg1 in 501mel resulted in loss of proliferation and viability. 

Binding of Brg1 or Brm to the promoter of MITF was confirmed by chromatin immunoprecipitation. 

Furthermore, microarray analysis revealed that osteopontin, IGF1 and 

survivin, the proteins known to be associated with tumor progression, were reduced in 

Brg1-depleted 501mel cells, suggesting that loss of SWI/SNF function negatively affects 

survival pathways besides the MITF cascade. Our results demonstrate an essential role 

of SWI/SNF for MITF expression in melanoma cells and suggest that a tissue-aimed inactivation 

of the SWI/SNF complex might become an effective approach in the therapy 

of melanoma. 


157

Posters 

40. 

MCL-1 as a rEGULaTOr of aPOPTOSIS IN CML CELL LINE aND 

PErIPHEral BLOOD MONONUCLEar CELLS 

Barbora Brodská, Petra Otevřelová and Aleš Holoubek 

Institute of Hematology and Blood Transfusion, U Nemocnice 1, 

128 20 Prague 2, Czech Republic 

Mcl-1 is a Bcl-2 family protein which can act as an apical molecule in apoptosis control, 

promoting cell survival by interfering at an early stage in a cascade of events leading to 

release of cytochrome c from mitochondria. Mcl-1 has a short half life and is a highly 

regulated protein. We investigated changes in the expression of this protein during 

combined treatment with demethylating agent decitabine (DAC) and histone deacetylase 

inhibitor SAHA (Vorinostat). These drugs represent epigenetic forces regulating 

gene and protein expression in cells, affecting cell cycle and cell death. In our experiments, 

DAC alone causes substantial increase of p21WAF1 expression, higher levels of 

proteins p53 and Puma were also detected, but the viability of the cells, as measured 

by MTT test, decreased only minimally, and we did not detect any remarkable apoptotic 

effect. While in CML-T1 cells we observed a slight decrease in Mcl-1 expression and 

PARP fragmentation, none of these effects have been found in lymphocytes. Addition 

of SAHA simultaneously with DAC supressed DAC-induced p21WAF1 up-regulation and 

substantial down-regulation of Mcl-1 protein expression together with mitochondrial 

membrane (MM) depolarization and PARP fragmentation was also detected. The extent 

of MM depolarization and the cell viability decrease observed in both CML-T1 cells and 

PBMC after DAC+SAHA treatment implies that this combination has a synergistic effect 

on apoptosis and Mcl-1 seems to play a crucial role in this process. 

Acknowledgement: This work was supported by the grants IGA NS 9637-4 and IGA 

MZOUHKT2005 from the MHCR. 


Posters 

41. 

NovEL GUaNIDINE DErivaTIvES aND evaLUaTION of THEIr DNA 

BINDING affINITIES aND POSSIBLE aNTICaNCEr effECT 

Jana Plšíková 1 , Ján Kovaľ 2 , Jaromír Mikeš 2 , Mária Kožurková 1 , Peter Fedoročko 2 , 

Ladislav Janovec 3 , Ján Ungvarský 3 and Danica Sabolová 1 

1 

Department of Biochemistry UPJŠ Košice, 

2 

Department of Biology and Ecology UPJŠ Košice, 

3 

Department of Organic chemistry UPJŠ Košice 

Acridine derivatives represent a well known class of multi-faceted anticancer agents that 

generally interfere with DNA and inhibit important regulatory enzymes such as topoisomerases. 

In order to identify novel anticancer drugs, we evaluated the mechanism 

of action of a novel series of 1`,1``-(acridin–3,6–diyl)–3`,3``-dialkyldiguanidines (etyl 

to hexyl). The affinity of studied compounds with DNA was investigated by a variety of 

techniques including UV-VIS, fluorescence, CD spectroscopy and electrophoresis. Binding 

constants for the DNA-drug complexes were determined from spectrofluorimetric titrations, 

(K = 1,25 – 5,26 ×10 5 M -1 ). Moreover, these compounds were capable of inhibiting 

topoisomerase I. Biological activities of studied derivatives were determined using flow 

cytometric methods after 24, 48 and 72 h co-incubation with leukemic cancer cell line 

HL-60. The most profound effect on different cell parameters was observed after 72 h 

incubation with the pentyl and hexyl derivate. We detected a significant increase in caspase-3 

activity, increase of percentage of cells with dissipated mitochondrial membrane 

potential, cell accumulation in G1 phase of cell cycle and enhanced DNA fragmentation. 

In summary, the studied derivatives are capable of interacting with ctDNA thought 

intercalation and inhibiting topoisomerase I. They also display a significant anticancer 

effect in HL 60 cells. 

Acknowledgement: This work was supported by VEGA 1/0053/08, 1/0097/10 and by the 

Slovak Research and Development Agency under contract no. VVCE-0001-07. 


159

Posters 

42. 

HISTONE aCETYLaTION of LEUKEMIC Jurl-MK1 CELLS WITHIN SAHA 

treaTMENT 

Michaela Pluskalová, Dana Grebeňová, Zbyněk Hrkal and Kateřina Kuželová 

Institute of Hematology and Blood Transfusion, Prague, Czech Republic 

The basic unit of chromatin, the nucleosome, is formed by about 146 bp of DNA wrapped 

around the histone octamer. The histones play an important role in the arrangement of 

nucleosomes. N-terminal tails of histones undergo multiple reversible posttranslational 

modifications, which have been proposed to form the so-called ´histone code´. This code 

(epigenetic information) regulates the binding of transcription complexes and determines 

gene transcription rate. The best understood histone modification is the lysine acetylation 

which occurs as a result of mutually opposed activities of histone acetyltransferases 

(HAT) and histone deacetylases (HDAC). 

Suberoylanilide hydroxamic acid (SAHA) is the first member of the group of HDAC inhibitors 

which is used for treating patients with cutaneous T-cell lymphoma. While it is 

also evaluated in clinical trials for the treatment of other oncological and hematological 

diseases, the mechanism of its action is largely unexplained. We found that the effect 

of SAHA on the leukemic cell line JURL-MK1 are strongly dose-dependent: an increase 

in the cellular adhesivity to fibronectin and cell cycle arrest was observed for subtoxic 

SAHA concentration while the apoptosis was triggered at higher doses. Using specific 

antibodies against the individual acetylation sites, we identified dose-dependent changes 

in the acetylation of histones H2A, H2B and H4 induced by SAHA treatment. We also 

detected changes in the cellular localization of these histones (transfer from nuclear to 

cytoplasmic fraction) due to SAHA. 

Acknowledgement: This work was supported by grant 301/09/1026 from the Grant 

Agency of the Czech Republic. 


Posters 

43. 

RegulaTION of EXPrESSION of PLaKOGLOBIN, a KEY DESMOSOMal 

CONSTITUENT, BY arYL HYDrOCarBON rECEPTOr aND CaMP 

SIGNaLING 

Jiřina Procházková 1,2 , Lenka Umannová 1,2 , Alois Kozubík 1 , 

Miroslav Machala 2 and Jan Vondráček 1,2 

1 

Department of Cytokinetics, Insitute of Biophysics AS CR, Brno, 2 Department of 

toxicology, pharmacology and immunology, Veterinary Research Insitute, Brno 

Aryl hydrocarbon receptor (AhR) has been originally described as a transcription factor 

mediating the toxic effects of dioxin-like compounds, but its signaling can be triggered 

also by endogenous agents, such as second messenger cyclic adenosine monophosphate 

(cAMP). Using an in vitro model of liver progenitor cells, participating both in liver 

regeneration and in hepatocarcinogenesis, we first investigated the potency of cAMP 

and another protein kinase A activator, forskolin, to activate AhR signaling pathway, 

as analyzed at the level of: i) AhR nuclear uptake; ii) AhR protein degradation; and iii) 

induction of model target gene Cyp1a1. We further analyzed the potency of cAMP and 

forskolin, alone or in combination with dioxin to modulate expression of Jup gene encoding 

plakoglobin protein, which plays a key role in establishment of intercellular junctions. 

Using WB-F344 cells, we found that cAMP and forskolin may transiently activate AhR 

signaling. As both compounds were able to modulate TCDD-induced changes in Cyp1a1 

and Jup expression, our results support the existence of a possible link between cAMPactivated 

signaling and AhR in regulation of both structural and signaling functions of 

desmosomal and adherent junctions. 

Acknowledgement: This work was funded by the Czech Science Foundation, grants No. 

524/09/1337 and 305/09/1526]. 


161

Posters 

44. 

CharaCTErIZaTION of SarP rEGULaTOry GENE INvOLvED IN 

POSITIve rEGULaTION of an aNGUCYCLINE-LIKE POLYKETIDE 

aNTIBIOTICS aurICIN GENE CLUSTEr IN StrePTomyces aureofaCIens 

CCM 3239 

Alena Reháková, Renáta Nováková , Ľubomíra Fecková, 

Peter Kutaš and Ján Kormanec 



Streptomyces aureofaciens CCM 3239 is gram-positive soil bacterium undergoing a complex 

developmental life cycle. During aerial mycelium growth S. aureofaciens CCM 3239 

produces their secondary metabolites including antibiotic auricin. The auricin gene cluster 

was named aur1. Synthesis of auricin is strictly regulated due to toxicity of auricin and 

its regulation is under the complex control of several regulatory genes. The product 

of one of these regulatory genes (sa9) belongs to the family of Streptomyces antibiotic 

regulatory proteins (SARPs). We characterized this gene and its impact on auricin 

production. By using the S1-nuclease mapping we identified a single promoter, sa9p, 

which expressed in a narrow stage during growth in rich liquid Bennet medium. The sa9 

gene was disrupted by a homologous recombination in S. aureofaciens CCM3239. The 

resulting mutant strain had dramatically decreased auricin production. In addition we 

confirmed a direct effect of the aur1P regulatory gene upon sa9 expression. The auricin 

pathway-specific activator Aur1P directly interacted with the sa9p promoter, indicating 

a cascade regulation of auricin production. 

Acknowledgement: This work was supported by the Slovak Research and Development 



Posters 

45. 

THE COMLEX NETWORK REGULATORY CIRCUITS IN THE REGULATION 

OF SIGMA faCTORS INVOLVED IN DIffERENTIATION AND STRESS 

RESPONSE IN STrePTOMYCES CoeLICOLor A3(2) 

Bronislava Řežuchová, Beatrica Ševčíková, Dagmar Homerová and Ján Kormanec 



Genome of the gram-positive soil bacterium Streptomyces coelicolor is a twice as large as 

the E. coli genome and contains 7 825 genes including 66 genes encoding sigma factors 

of RNA polymerase that is the largest number among the so far examined bacteria. In 

B. subtilis a general stress response is under the control of a single sigma factor, SigB. Its 

gene is located in the operon together with rsbW and rsbV genes. The product of rsbW is 

an anti sigma factor of SigB, which serves as serine kinase that binds and phosphorylates 

its negative regulator so-called anti-anti sigma factor RsbV. However, genome sequence 

analysis in S. coelicolor has shown nine sigB homologues, as well as putative 48 anti-sigma 

factors (RsbW homologues) and 18 antagonists of anti-sigma factors (RsbV homologues). 

Bacterial two hybrid system was used to investigate and characterize interaction between 

nine known homologues SigB and 15 suggested anti-sigma factors, and between these 

anti-sigma factors and 9 selected anti-anti sigma factors. This method allows detect even 

relatively weak protein-protein interaction. The efficiency of interactions was quantified 

by measuring β-galactosidase activities in liquid cultures. The interactions were 

confirmed by native PAGE. The results revealed very complex way of regulation, where 

several sigma factors interact with different anti-sigma factors that additionally interact 

with several subclasses of anti-sigma factor antagonists. 

Acknowledgement: This work was supported by the VEGA grant 2/0104/09 from Slovak 



163

Posters 

vI. GLYCOMICS 


Posters 

46. 

DifferENCES IN INTEraCTION of LECTINS SPECIfICaLLY rECOGNIZING 

SIaLIC aCID rESIDUES WITH SUrfaCE of P-GP NEGaTIve or POSITIve 

L1210 CELLS 

Tatiana Kurucová, Helena Kavcová, Kristína Rogozanová, Lucia Messingerova, 

Danica Mislovičová 1 , Albert Breier and Zdena Sulová 

Institute of Molecular Physiology and Genetics SAS, Bratislava, Slovakia 

1 

Institute of Chemistry SAS, Bratislava, Slovakia 

Multidrug resistance (MDR) of mouse leukemic cell line L1210/VCR (R) obtained by 

adaptation of L1210 cells (S) to vincristine (VCR) is accompanied by overexpression of 

P-glycoprotein (P-gp). Selection with VCR that confers overexpression of P-gp is also inducing 

various kinds of metabolic alteration. We described recently several differences 

in cell surface saccharides between R and S cells. 

Aim of this study was to resolve in exist differences in interaction of Triticum vulgaris 

lectin (WGA), Maackia amurensis lectin (MAA), Sambucus nigra lectin (SNA) with cell 

surface of S, R cells and L1210 cells that express P-gp due to transfection of cells with 

plasmid containing human gene encoding this protein (T cells). While MAA was found to 

be nontoxic for S, R and T cells and SNA is exerting small cell damage effect, WGA induced 

concentration depended cytotoxic effect. Agglutination of cells by this lectin is more massive 

to cells expressed P-gp. WGA lectin interact more potently with P-gp positive cells 

R and T as with P-gp negative S cells. Spectrum of cell membrane glycoprotein targets 

of lectins were assessed by lectin blot methods. Our data indicated that application of 

lectins enable to study the differences cell surface saccharides. 

Acknowledgements: This work was supported by: APVV-0084-07, VVCE-0064-07, VEGA- 

2/0123/10, VEGA-2/0155/09. 


165

Posters 

47. 

THE prESENCE of P-GLYCOPrOTEIN IN L1210 CELLS DIrECTLY INDUCES 

DOWN-rEGULaTION of CELL SUrfaCE saCCHarIDE-tarGETS of 

CONCanavaLIN A 

Zdena Sulová 1 , Peter Ditte 2 , Tatiana Kurucová 1 , Eva Poláková 1 , Kristína Rogozanová 1 

Lucia Škvarková 1 , Ján Sedlák 3 , Jaromír Pastorek 2 and Albert Breier 1 

1 

Institute of Molecular Physiology and Genetics, 2 Institute of Virology, 3 Istitute of 

Experimental Oncology, SAS, Bratislava, Slovak Republic 

Overexpression of P-glycoprotein (P-gp), a plasma membrane drug transporter (an 

ABCB1 member of the ABC transporter family), is the most prevalent cause of multidrug 

resistance in cancer tissues. Lectin Concanavalin A (ConA) induces massive cell death of 

L1210 leukemia cells (S). Cell sublines of L1210 in which P-gp overexpression was induced 

by selection with vincristine (R) or by stable transfection with a plasmid encoding fulllength 

human Pglycoprotein (T) were less sensitive to ConA. Both P-glycoprotein-positive 

cell lines exhibited typical P-glycoprotein-mediated multidrug resistance. Resistance of 

R and T cells to ConA was associated with lower binding of ConA as compared to S cells 

when analyzed by the following methods: i) SDS PAGE and electrobloting of proteins in 

the crude membrane fraction followed by detection with biotinylated ConA and avidinperoxidase; 

ii) fluorescent cytometry or confocal microscopy of the intact cells with 

surfaces labeled by FITC-ConA. These data indicated that the presence of P-glycoprotein 

in L1210 cells independently on mode of its expression induced down-regulation of cell 

surface saccharide-targets of ConA. Therefore, this feature may be considered as a secondary 

cellular response on P-glycoprotein expression. 

Acknowledgements: This work was supported by: APVV-0084-07, VVCE-0064-07, VEGA- 

2/0123/10, VEGA-2/0155/09. 


Posters 

viI. mEMBraNE BIOCHEMISTRY AND 

BIOENERGETICS 


167

Posters 

48. 

EPITOP of Iva-520 MONOCLONal aNTIBODY ON THE 

BovINE SPErm CD46 MOLECULE 

Jana Antalíková, Jana Jankovičová, Katarína Michalková, 

Michal Simon and Ľubica Horovská 

Institute of Animal Biochemistry and Genetics, Slovak Academy of Sciences, 

Ivanka pri Dunaji, Slovak Republic, Centre of Excellence of Slovak Research and 

Development Agency „Biomembranes 2008“. 

Membrane cofactor protein (MCP/CD46) serves as a cofactor for the factor I-mediated 

cleavage of C3b and C4b complement components deposited on self tissues and probably 

plays also a role in reproduction. In Human and also New World monkeys the unique 

pattern of sperm CD46, different from other cells and tissues were detected. The bovine 

sperm isoform remains unclear. Using the monoclonal antibody (mAb) IVA-520 against 

bovine CD46 (obtained after intrasplenic immunisation of BALB/c mice with intact bull 

spermatozoa) the expression of CD46 on bovine spermatozoa has been detected. By SDS- 

PAGE of detergent solubilised sperm proteins and immunoblotting with mAb IVA - 520 

we found out two bands with molecular weight of 53 and 43 kDa - different from common 

blood cells and tissue isoforms (46-52 lower band, 60-68 upper band), resistant to 

digestion with N-glycosidase F and disappearing after neuraminidase treatment (not in 

other cells and tissues). The treatment of suspension of whole spermatozoa with neuraminidase 

decreased the binding ability of mAb IVA-520 on sperm CD46 molecule. We 

suppose the posttranslational modification of the molecule are sperm-specific, the part 

of bovine CD46 epitope recognised by mAb IVA - 520 is formed probably by the serine, 

threonine and-proline rich region (STP) of MCP molecule and the sialic acid probably 

takes part in an epitope conformation of mAb IVA-520. 

Acknowledgement: This work was supported by grant VEGA-2/0001/09 and in part by 

the grant VVCE-0064-07. 


Posters 

49. 

DIMETHYL-OXALYLGYCINE MODULATES GENE EXPRESION AND PROTEIN 

LEVELS OF THE SODIUM CALCIUM EXCHANGER IN HEK 293 CELL LINE 

Cagala M. 1 , Lencesova L. 1,2 , Hudecova S. 1 , Csaderova L. 2 , Sirova M. 1 , 

Cholujova D. 3 , Kopacek J. 4 , Pastorekova S. 2,4 and Krizanova O. 1,3 

1 

IMPG, Center of Excellence for Cardiovascular Research, SAS, Vlarska 5, 

833 34 Bratislava, Slovak Republic 2 MMC, SAS, Vlarska 3-7, 831 01 Bratislava, 

Slovak Republic 3 IEO, SAS, Vlarska 7, 833 91, Bratislava, Slovak Republic 4 IV, 

Center of Excellence for Cardiovascular Research, SAS, Dubravska cesta 9, 

845 05 Bratislava, Slovak Republic 

Up to now a little is known about the effect of hypoxia on the NCX1 expression and 

function. Therefore we studied, how dimethyl-oxalylglycine (DMOG), an activator and 

stabilizator of the HIF-1α could affect expression of the NCX1 in HEK 293 cell line. We also 

tried to determine, whether this activation can result in the development of apoptosis in 

HEK 293 cells. We have found that DMOG treatment for 3 hours significantly increased 

gene expression and also protein levels of the NCX1. This increase was accompanied 

with the decrease in intracellular pH. Wash-out of DMOG did not result in decrease of 

NCX1 mRNA and protein to original, control levels, although pH returned to control 

values. In the promoter region of the NCX1 we did not find consensus sequence for 

HRE, but we have found consensus NF-kB sequence in this region of the NCX1. Using 

luciferase reporter assay we observed increase in NCX1 promoter activity after DMOG 

treatment, which suggests that NF-kB is involved in DMOG induced upregulation of 

the NCX1. Moreover, we also showed that this upregulation might be involved, at least 

partially, in the induction of the process of apoptosis. Nevertheless, further experiments 

are needed to clarify this issue. 

Ackowledgement: This work was supported by grants APVV 51-0397-07, VEGA 2/0082/10 

and TRANSMED. 


169

Posters 

50. 

DEHYDROERGOSTEROL ELUCIDATES STEROL UPTAKE PROCESS IN YEAST 

S. CereVISIae 

Peter Kohút 1 , Martin Valachovič 1,2 , Lucia Hronská 1 and Ivan Hapala 1 

1 

Institute of Animal Biochemistry and Genetics, Slovak Academy of Sciences, 

Moyzesova 61, 90028 Ivanka pri Dunaji, Slovakia 

2 

Medical University Vienna, Christian Doppler Laboratory for Infection Biology, 

Max F. Perutz Laboratories, Vienna, Austria 

Yeast Saccharomyces cerevisiae is a facultative anaerobic organism - it can grow either 

in the presence or absence of oxygen. However, during anaerobic growth it is unable to 

synthesize unsaturated fatty acids and ergosterol, which have to be supplied externally. 

Uptake of external sterol in yeast consists of three steps: 1) passage through the cell 

wall, 2) entry into plasma membrane and 3) integration into metabolism, but molecular 

mechanisms of these individual steps are largely unknown. Sterol uptake can be efficiently 

studied with fluorescent sterol probes. Dehydroergosterol (DHE) is a naturally 

fluorescent sterol and structural similarity between DHE and ergosterol – native yeast 

sterol – ensures that the data acquired using DHE as a probe in the study of sterol uptake 

in yeast reflect metabolic processes taking place under physiological conditions. In our 

experiments we used DHE to analyze molecular details of sterol uptake in the yeast S. 

cerevisiae, particularly to reveal the role of Aus1p and Pdr11p proteins in this process. 

These proteins are members of the ABC transporter family and they were previously 

identified in a wide-scale screen as proteins involved in sterol uptake in S. cerevisiae. We 

used mutants in Aus1p and Pdr11p putative sterol transporters to separate the first two 

steps in sterol uptake process. Fluorescent properties of DHE enabled us to distinguish 

between membrane-incorporated and cell wall-associated sterol. Our results indicate 

that Aus1p and Pdr11p are required for entry of sterols into the plasma membrane and 

not for internalization of sterols as suggested in some studies. 

This work was supported by grants APVT-51-029504 and VVCE-0064-07 


Posters 

51. 

MYCOBaCTErial EPOXIDEHYDrOLaSE EPHD IS INvOLvED IN 

faTTY aCID METaBOLISM 

Jan Madacki 1 , Katarína Mikušová 1 , Mary Jackson 2 and Jana Korduláková 1 

1 

Department of Biochemistry, Faculty of Natural Sciences, Comenius University, 

Bratislava, Slovakia; 2 Department of Microbiology, Immunology, and Pathology, 

Colorado State University, Fort Collins, CO, USA 

The cell envelope of mycobacteria is a crucial determinant of virulence and drug resistance 

in mycobacteria. The major components of the mycobacterial envelope are mycolic 

acids – very long branched α-alkyl-, β-hydroxy- fatty acids, which play an important role 

in the integrity of the mycobacterial cell, as well as in pathogenicity of mycobacteria. 

Experimental observations, accumulated during several decades allowed to draw a general 

scheme for the mycolic acid biogenesis. However, many key questions regarding the 

molecular basis of the biosynthetic routes leading to the mature mycolates still have to 

be answered. 

We investigated the function of the protein EphD in the non pathogenic strain of 

Mycobacterium smegmatis. The recombinant protein was produced in E. coli and using 

the generic substrate we showed that EphD displays the epoxide hydrolase activity in 

vitro. Phenotypic analysis of M. smegmatis carrying the disrupted copy of ephD orthologue 

revealed that this protein is involved in the fatty acid metabolism, particularly in 

the biogenesis of mycolic acids. 

Acknowledgement: This work was supported by Slovak Grant Agency VEGA (grant No. 

1/0223/08) 


171

Posters 

52. 

ISOFORMS OF AMP-ACTIvaTED PROTEIN KINASE SUBUNITS IN 

LYMPHOCYTES AND OBESITY 

Boris Lakatoš, Lucia Bialešová and Eva Harnišová 

Institute of Biochemistry, Nutrition and Health Protection, Department of Biochemistry 

and Microbiology, Faculty of Chemical and Food Technology, Slovak University of 

Technology, Radlinského 9, 812 37 - Bratislava, Slovakia 

Obesity becomes to be a real problem in developed countries and it seems that its 

prevalence increases. One of consequences of obesity is the increasing incidence of 

metabolic syndrome - a state of metabolic dysregulation characterized besides obesity 

also with resistance to insulin, changes in blood lipids level, early atherosclerosis, changes 

in blood tension, which can result to the appearance of civilization diseases. Although 

the obesity could be caused by different factors, studies in last decades pointed out on 

fundamental mechanisms leading to the defects in lipid metabolism, which are regulating 

energy fluxes in animal metabolism. It is known that 5´-AMP-activated protein kinase 

(AMPK) a sensor of metabolic status plays the key regulatory role. Structurally is AMPK 

heterotrimeric complex consisting of catalytic α subunit and regulatory β and γ subunits 

existing in several isoforms (α 1 

, α 2 

, β 1 

, β 2 

, γ 1 

, γ 2 

, γ 3 

). AMPK is widely distributed in all body 

tissues. In this work we followed expression of AMPK subunits isoforms in human lymphocytes 

isolated from blood of undergraduate students of FCHFT SUT. Simultaneously 

we measured basic biochemical parameters of lipid, glucose and energy metabolism. 

The obtained values were used to create correlation dependence with body mass index 

(BMI) and physical activity. 

We found expression of genes for α 2 

, β 2 

and γ 3 

subunits in all blood samples regardless of 

gender, BMI or physical activity of probands. Sequential analysis of obtained PCR products 

for α 2 

subunit of AMPK did not show any polymorphisms. Biochemical parameters 

showed rather weak, if any, correlation with BMI. 

Acknowledgements: This work was supported by the grant VEGA, nr. 1/0589/08. 


Posters 

53. 

DISTRIBUTION AND BIOCHEMICAL CHaraCTERIZATION OF CD52-LIKE 

MOLECULE IN BULL EPIDIDYMIS 

Katarína Michalková, Michal Simon, Jana Antalíková and Ľubica Horovská 

Institute of Animal Biochemistry and Genetics SAS Ivanka pri Dunaji 

The aim of this study was to analyze the distribution and biochemical properties of the 

antigen in bull genital tract using monoclonal antibody IVA-543 produced in IABG SAS. 

CD52 is a GPI linked protein with a molecular weight of about 25 to 29 kDa, it is expressed 

on all lymphocytes, and in male genital tract. Except humans, CD52 has been described 

in mice, rats, chimps and dogs. Characteristic common to all species is very similar nature 

of expression in cells of the epididymal epithelium, the protein in the genital tract 

is produced post-testicular and constitutes the main surface glycopeptide of the sperm 

from the corpus and cauda epididymis. Our results revealed that CD52-like molecule in 

the reproduction system of bull is produced by epididymal epithelial cells and secreted 

into the lumen. Immunohistochemical analysis confirmed that the antigen is synthesized 

in the epididymis downstream, in the largest quantities it can be detected in the cauda 

epididymal tissue. The similar results we obtained by western blot analysis of the cauda 

epididymal fluid. FACS analysis and the indirect immunofluorescence of sperm from of 

epididymis confirmed these results, in the proximal part of epididymis CD52-like molecule 

is present on 4.73 % of sperm and in the distal part it is present on up to 99.45 % of the 

sperm. Bovine antigen expression in the distal part of the epididymis points out that 

the bull sperm acquire CD52-like molecule on their surface during their transit through 

the epididymis, especially in the distal part of this organ. Western blot analysis showed 

that the molecular weight of this bull epididymal antigen derived from tissue, fluid or 

from sperm is ranging from 22 to 25 kDa. 

Acknowledgments: This work was supported by grants VEGA-2/6023/27 and 

APVV-VVCE-0064-07. 


173

Posters 

54. 

IDEBENONE ACTIvaTION OF GLYCEROL-3-PHOSPHATE OXIDATION IN 

LIVER MITOCHONDRIA frOM CONTROL AND HYPERTHYROID raTS 

Hana Rauchová 1,2 , Martina Vokurková 1,2 and Tomáš Soukup 1 

1 

Institute of Physiology, Academy of Sciences of the Czech Republic, 

2 

Centre for Cardiovascular Research, Prague, Czech Republic 

A synthetically prepared analog of coenzyme Q (CoQ) with lower hydrophobicity, idebenone 

(IDE; hydroxydecyl-ubiquinone), was found to be very effective in animal experiments and 

human replacement therapy when the synthesis of CoQ was decreased. In our previous 

studies, we found that mitochondrial FAD-linked glycerol-3-phosphate dehydrogenase 

(GPDH; EC 1.1.99.5) from brown adipose tissue is activated by IDE. However, in most 

mammalian tissues expression of GPDH is highly depressed and enzyme activity is very 

low. Thyroid hormones are known to have a marked influence on GPDH activity; they 

especially induce GPDH activity in liver. The aim of our study was to test to what extent 

IDE can activate glycerol phosphate (GP) oxidation measured as oxygen uptake and GP 

cytochrome c oxidoreductase activity in mitochondria from control and hyperthyroid rat 

liver. We found the significant increase of GP-dependent oxygen uptake as well as the 

activation of enzyme activity. Our results indicate that IDE may be used for the activation 

of the GP shuttle catalyzing the interconversion between dihydroxyaceton phosphate and 

GP (formed by GPDH together with its cytosolic NADH-dependent counterpart) through 

which NADH from cytosol can be oxidized by the mitochondrial respiratory chain to contribute 

to the maintenance of the high rate of glycolysis. It might be important in the case 

when mitochondrial Complex I is impaired and energy production must be maintained. 

Acknowledgement: This work was supported by the GACR (303/09/0570) and Ministry 

of Education, Youth and Sport (1M6798582302 and AV0Z 50110509). 


Posters 

55. 

EffECT OF ATOrvaSTATIN ON BIOENERGETICS OF THE LIVER 

MITOCHONDRIA ON a HIGH-LIPID DIET 

Oľga Uličná 1 , Oľga Vančová 1 , Jarmila Kucharská 1 , Peter Božek 2 , 

Iveta Waczulíková 3 and Libuša Šikurová 3 

1 

Pharmacobiochemical laboratory 3rd Dept of Int Medicine Med Fac UK Bratislava, 

2 

Dept of Clin Biochem and Hematol St Michal Hospital Bratislava, 3 Dept of Nuclear 

Physic and Biophysic Math Phys Inf Fac UK Bratislava 

Statins impair hepatocellular cholesterol production by inhibiting the synthesis of mevalonate. 

Mevalonate is a precursor not only of cholesterol but also of ubiquinone, which 

is an important carrier of the mitochondrial respiratory chain. 

The aim of the study was to evaluate the oxidative phosphorylation in liver mitochondria 

in rats on a high-lipid diet. We investigated the toxicity of atorvastatin on the liver 

mitochondria. Male Wistar rats (b.w. 210-270g) were divided into four groups: 1. control 

group on the Larsen´s diet, 2. hypercholesterolemic (HCh) on a high-lipid diet (Larsen´s 

diet containing 4% of cholesterol and 10% of a saturated fat) 3. and 4. HCh treated with 

atorvastatin at the dose of 10 and 80 mg.kg -1 , respectively. After 8 weeks, total cholesterol 

(tChol) and triacylglycerols (TAG) were determined in the plasma and liver tissues. 

Liver mitochondria were isolated by differential centrifugation. Parameters of oxidative 

phosphorylation were measured on an oxygraph Gilson 5/6H. We found an increased 

content of TAG and tChol in the plasma and liver tissues of HCh group. Atorvastatin at the 

high dose lowered tChol and TAG in the plasma and liver tissues. Parameters of oxidative 

phosphorylation were significantly impaired in the liver mitochondria of HCh rats. The 

high dose of atorvastatin worsened bioenergetics of the liver mitochondria of HCh rats, 

but the low dose had no effect. Our results suggest that administration of low doses of 

atorvastatin does not exert a negative effect on liver under the high-fat dietary conditions. 

Acknowledgements: Supported by the grants VEGA 1/0328/10 and 1/0293/08. 


175

Posters 

56. 

EffECT OF PAMAM G4 DENDRIMER ON LIVER MITOCHONDRIA 

OXIDATIVE PHOSPHORYLATION AND METABOLIC CONTROL IN 

EXPERIMENTAL DIABETES 

Oľga Vančová 1 , Oľga Uličná 1 , Katarína Šebeková 2 , 

Magdalena Labieniec 3 and Cezary Watala 3 

1 

Pharmacobiochemical laboratory 3rd Dept of Int Medicine LFUK Bratislava, 

2 

Department of Preventive and Clinical Medicine SZU Bratislava, 3 Department of 

Haemostasis and Haemostatic Dis Med Univ of Lodz, Poland 

Prolonged exposure to hyperglycaemia causes non-enzymatic glycation of proteins and 

can lead to production of reactive oxygen species. 

Polyamidoamine dendrimer PAMAM G4, a strong nucleophilic molecule with 64 free 

primary amino groups at its surface appears as an effective scavenger of excessive 

glucose in diabetes. 

The aim was to study the ability of PAMAM G4 to lower the concentration of plasma 

glucose, to suppress long-term parameters of hyperglycaemia and to improve oxidative 

phosphorylation in the liver mitochondria. 

Experimental diabetes mellitus was evoked by an injection of streptozotocin in male Wistar 

rats. After 7 days half the rats were administered PAMAM 64 i.p. daily. After 8 weeks the 

concentration of glucose, the end-products of advanced glycation and the end-products 

of advanced protein oxidation in plasma and glycated haemoglobin in blood were determined. 

Liver tissue was used for mitochondria isolation and parameters of oxidative 

phosphorylation were measured using volt-amper method with a Clark oxygen electrode. 

Our results, for the first time in vivo experiment, show that PAMAM G4, regardless of 

its known cytotoxicity, significantly reduced hyperglycaemia and all the measured longterm 

markers of hyperglycaemia in diabetic rats. This positive effect is not sufficient to 

restore the impaired mitochondrial function in experimental diabetes. 

Acknowledgement: Supported by grant VEGA 1/0328/10. 


Posters 

57. 

BIOCHEMICAL AND MOLECULar ANALYSIS OF NITraTE-RESISTANT 

MUTANT OF MethanothermobaCTer thermautotrophICus 

Monika Vidová, Zuzana Nováková and Peter Šmigáň 

Institute of Animal Biochemistry and Genetics SAV Bratislava 

In spite of many studies over the past decade, the processes of energy conservation 

in methanoarchaea have not yet been satisfactorily elucidated. To contribute to the 

solution of this complex problem, we started with a systematic genetic approach to the 

problem of energy conservation in methanoarchaea. This work presents a microbiological, 

biochemical and molecular analysis of a spontaneous mutant of Methanothermobacter 

thermautotrophicus resistant to nitrate. Nitrate inhibits the A 1 

cytoplasmatic domain of 

A 1 

A 0 

-ATP synthase. 

Nitrate inhibits methanogenesis in the wild-type cells in the presence of 30 mM nitrate; 

however, the nitrate-resistant mutant exhibited two times higher methanogenesis, even 

in the presence of 70 mM nitrate. While nitrate profoundly inhibited ATP synthesis driven 

by methanogenic electron transport in the wild type, only a slight inhibition was observed 

in the mutant strain. These results suggested a modification in the ATP-synthesizing 

system of the mutant strain. The sequence of the complete A 1 

A 0 

-ATP synthase operon 

(MTH952 – MTH961) in the wild-type and mutant strains was determined and compared. 

Two mutations leading to amino acid substitutions in two A 1 

A 0 

-ATP synthase subunits 

were identified – Ala 337 

Val in subunit A and Ala 292 

Ser in subunit B. Moreover, this study 

revealed the differential expression of several proteins that may contribute to nitrate 

resistance. The results imply that changes of nitrate sensitivities of nitrate-resistant 

mutant is due to mutational substitutions in the A 1 

A 0 

-ATP synthase operon. 


177

Posters 

vIII. NEW METHODOLOGIC PROCEDURES 


Posters 

58. 

DEVELOPMENT OF a DETECTION TOOL TO FOLLOW THE SPECIFIC 

ACTIVITY OF BUTYRYLCHOLINESTEraSE IN HUMAN PATIENTS 

Katarína Mrvová and Anna Hrabovská 


Odbojárov 10, 832 32 Bratislava 

Many hypotheses have been proposed in afford to explain a butyrylcholinesterase (BChE) 

inter-individual variability in humans, e.g., different expression levels; different catalytic 

properties. Yet, it has been impossible to study this topic due to the lack of an efficient 

detection tool. However, we have recently generated a selective and specific antibody 

against human BChE which allows us to address this problem. The aim of the project 

was to develop an ELISA assay for detection of BChE activity and use this tool to study 

the inter-individual BChE activity variation in humans. 

Human plasma was prepared from the capillary blood of 86 healthy human volunteers 

(age 20 – 23; BMI = 17,6-28,4). The Ellman’s assay was used at the conditions determined 

in our laboratory (see the abstract of D. Neuschlova). ELISA has been used to determine 

specific activity. All samples were tested in triplicates. 

In the first step we determined the conditions for the ELISA assay. The lowest saturating 

dilution of the primary antibody and the experimental serum dilution were determined 

from the saturation curves. In the second step, human sera were tested for the total 

BChE activity (Ellman’s assay) and for the specific BChE activity in excess plasma (ELISA) 

and compared. Our results suggest that an inter-individual BChE activity in humans is 

a caused by both, higher expression level and different catalytic properties. 

Acknowledgement: The project was supported by APVV grant SK-CZ-0028-09. 


179

Posters 

59. 

OPTIMALIZATION OF ELLMAN’S ASSAY TO STUDY 

THE KINETICS OF CHOLINESTEraSES 

Dominika Neuschlová and Anna Hrabovská 


Odbojárov 10, 832 32 Bratislava 

Ellman’s assay (EA) has been widely used in experimental research and clinical practice. 

The limitations of this method are however a high background in biological samples, 

instability of the dissolved substrate (thiocholine) and its sensitivity to the light exposure. 

The aim of this project was to determine the favorable conditions for EA in order 

to lower the background and the reagent instability and thus allow detecting even very 

low activities of cholinesterases. 

Human butyrylcholinesterase was chosen to study the conditions of EA. Butyrylthiocholine 

iodide was used as a substrate. Phosphate, HEPES and Ringer buffers were used at pH 

values 7,0; 7,5; 8,0; and 8,5. Stability of the substrate dissolved in each buffer was tested 

over the time. The full spectrum was followed in each buffer. Velocity of the color product 

production was followed as a function of time (v/t curve) and substrate concentration (v/s). 

The substrate was the most stable in the presence of HEPES buffer. The charts of full 

spectrums and the velocity dependences suggested the same kinetics of butyrylcholinesterase-catalyzed 

reaction of butyrylthiocholine in both phosphate and HEPES buffers. 

Based on our results we can conclude that Ellman’s assay performed in HEPES buffer, in 

contrary to phosphate buffer, is more suitable for measuring of cholinesterase activity. 

This is due to the lower background (raising from substrate instability) and unaffected 

kinetics. 

Acknowledgement: The project was supported by APVV grant SK-CZ-0028-09. 


IX. PATHOBIOCHEMISTRY AND TraNSLATION 

MEDICINE 

Posters 


181

Posters 

60. 

EffECT OF fraCTIONATED DOSES OF GAMMA raYS ON THE ROSTraL 

MIGraTORY STreaM Of aDULT raTS 

Soňa Bálentová 1 , Eva Hajtmanová 2 , Yvetta Mellová 3 , 

Ivana Kinclová 2 and Marián Adamkov 1 

1 

Institute of Histology and Embryology, Jessenius Faculty of Medicine in Martin, 

CU, Martin, Slovakia, 2 Department of Radiotherapy and Oncology, Martin Faculty 

Hospital, Martin, Slovakia, 3 Institute of Anatomy, Jessenius Faculty of Medicine 

in Martin, CU, Martin, Slovakia 

Ionizing radiation commonly used in the radiotherapy of brain tumours can cause adverse 

side effects to surrounding normal brain tissue. The adult mammalian subventricular 

zone (SVZ) of the brain lateral ventricles (LV) and their subsequent lateral ventricular 

extension, the rostral migratory stream (RMS), is one of the few areas, which retains the 

ability to generate new neurons and glial cells throughout life. Take into account the fact, 

that ionizing radiation is one of the strongest exogenous factors affecting cell proliferation, 

the aim of the present study was to investigate the occurence of radiation-induced 

alterations of apoptosis in the forebrain’s RMS using animal model of radiosurgery. Adult 

male Wistar rats were investigated 7, 14 or 21 days after whole-body irradiation with 

fractionated doses of gamma rays (the total dose of 3 Gy). Radiation-induced apoptotic 

cell death was determined by in situ labeling of DNA nick ends (TUNEL) and light microscopy 

evaluation of TUNEL-positive cells. However, the data from quantitative analysis of 

the numbers of apoptotic cells are still under evaluation, our preliminary results showed, 

that ionizing radiation clearly affect this neurogenic region. 

Acknowledgement: This work was supported by AV4/2026/08 Grant. 


Posters 

61. 

IS RESPIraTORY PATHWAY ACTING THROUGH NO-sGC? 

Ľudmila Capková, Alexandra Dávidová, Andrea Kucháriková and Nadežda Lukáčová 

Institute of Neurobiology, Slovak academy of Sciences, Košice 

Most effects of the nitric oxide (NO) are mediated by stimulation of soluble guanylyl 

cyclase (sGC) and subsequent increase in cyclic guanosine monophophate (cGMP) formation. 

NO/sGC/cGMP signalization was identified in neuronal pathways of the brain, 

but little is known about this signaling pathway in the spinal cord. The aim of our study 

was to find out, whether bulbospinal respiratory pathway is acting through NO-sGC 

signalization. This pathway begins in medulla and project to the phrenic motoneurons 

controlling diaphragm activity. The distribution of the neuronal nitric oxide synthase 

(nNOS), β1 subunit of soluble guanylyl cyclase (β1sGC) and synaptophysin (SYN) was 

explored in control animals and after C2-C3 spinal cord hemisection in upper part of the 

respiratory pathway. Retrograde tracer Fluorogold (FG) was used for identification of 

respiratory neurons in medulla. Two days after injection of FG into the phrenic nucleus 

(PN) at C4 level revealed amount of FG fluorescent neurons in the ventral respiratory 

group (VRG) mostly on contralateral side. We showed intense punctate nNOS and SYNpositive 

terminals of respiratory neurons in neuropile of PN in control animals and strong 

depletion of these terminals on contralateral side and almost entire depletion of nNOS 

and SYN-positive terminals on ipsilateral side of lesioned animals. Eight days after the 

hemisection we have revealed lightly ß1sGC fluorescent motoneurons of PN and around 

them a few nNOS fluorescent boutons on contralateral side. Almost no sign of ß1sGC 

immunopositivity could be seen ipsilaterally to the hemisection. These results together 

suggest that bulbospinal respiratory pathway is acting through NO-sGC. 

Ackowledgement: The experimental work was supported by VEGA Grant 2/0015/08. 


183

Posters 

62. 

THE effECT of naTUral POLYPHENOLS ON aDIPONECTINE LEvEL IN 

paTIENTS SUfferING frOM erECTILE DYSfUNCTION 

Monika Dvořáková 1 , Jana Muchová 1 , Branislav Trebatický 2 , 

Ján Breza 2 and Zdeňka Ďuračková 1 

1 

Department of Chemistry, Biochemistry and Clinical Biochemistry, Medical Faculty, 

Bratislava, Slovakia, 2 Department of Urology, Faculty of Medicine, 

Comenius University, Bratislava, Slovakia 

As patients suffering from erectile dysfunction (ED) are considered patients with temporary or 

permanent impotence. In the pathophysiology of ED psychological as well as organic factors 

(or both together) can take a part. The main role in the erection process plays a relaxation 

of smooth muscles inside the arterial system and cavernous tissue. It is controlled by spinal 

reflex and can be initialized by visual, osmatic or imaginary stimulation. The most important 

blood vessels relaxant is NO. By effect of free radicals, NO becomes more inactivated, what 

leads to lowered tissue relaxation caused by change in cGMP/cAMP ratio. Pycnogenol ® 

(Pyc) as a mixture of natural polyphenols and polyphenolic extract from gingko biloba leafs 

(EGb761) stimulate a constitutive NO synthase, induce vasodilatation and improve the 

microcirculation in the blood. Adiponectin is a protein hormone that modulates glucose 

regulation and fatty acid catabolism. Levels of the hormone are inversely correlated with body 

fat percentage. Levels of adiponectin are reduced in diabetics compared to non-diabetics. 

Weight reduction significantly increases circulating levels. Hypoadiponectinemia is an 

independent risk factor for developing of e.g. metabolic syndrome, cardiovascular disease 

and diabetes mellitus, which are often found as disorders associated to ED. To our double 

blind, randomized, placebo controlled study 52 patients suffering from ED were included. 

Patients were investigated before, one and three months after supplements administration 

and after termination of drugs supplementation. In addition, ED group of patients was split 

to diabetes and nondiabets groups to compare adiponectin levels. No change after Pyc or 

Egb 761 administration has been found in comparison to placebo group. Similarly, there has 

not been found a difference in ED patients with diabetes and ED patients without diabetes 

mellitus. ED patients have decreased levels of adiponectine compared to reference values. 

But what is the reason of this decrease is hard to say because of the complexity of the disease. 

Ackowledgement: This study was supported by VEGA grants of Ministry of Education of the 

Slovak Republic, and Mind and Health, civil association. 


Posters 

63. 

STUDY OF ANTIAPOPTOTIC PROTEINS RESPONSIBLE FOR DEVELOPMENT 

OF DRUG RESISTANCE IN ACUTE LEUKEMIA 

Jana Jurečeková, Jozef Hatok, Andrea Štefániková, Dušan Dobrota and Peter Račay 

Comenius University in Bratislava, Jessenius Faculty of Medicine in Martin, 

Department of Medical Biochemistry 

Deregulation of apoptosis disrupts the complex and delicate balance between cell proliferation, 

survival and cell death and plays a major role in the development of diseases 

such as cancer, and particularly acute leukemia. Malignant cells that have alterations in 

proteins involed in cell death signaling are very frequently resistant to chemotherapy 

and are difficult to treat with chemotherapeutic agents that primarily act by inducing 

apoptosis. Structural analysis of antiapoptotic proteins together with studies of their 

biochemical mechanisms have outlines strategies for generation of drugs, resulting in 

numerous novel chemical entities with mechanism-based activity. 

In presented study, the influence of ABT-737, the synthetic inhibitor of antiapoptotic 

proteins Bcl-2 and Bcl-xL, on induction of apoptosis and viability of leukemic cell lines 

HL-60 and K-562 was tested. Higher sensitivity of HL-60 (EC 50 

= 4,5 µM) probably correlates 

with mildly lower expression of Mcl-1, which overexpression may be responsible 

for resistance to ABT-737. Fragmentation of DNA was observed already after 3 hours of 

cultivation with ABT-737, so we confirmed that ABT-737 acts via induction of apoptosis. 

ABT-737 was also found to enhance the effects of chemotherapy in HL-60. 

Understanding of the core components of the apoptotic machinery at the molecular and 

structural levels may lead to creating a new era in cancer therapy where the intrinsic and 

acquired resistance of malignant cells to apoptosis can be pharmacologically reversed, 

reinstating natural pathways of cell suicide. 

Acknowledgements: This work was supported by the Grant UK/38/2010. 


185

Posters 

64. 

PROGNOSTIC SIGNIFICANCE OF MIr-21 AND MIr-143 EXPRESSION IN 

TISSUE SAMPLES OF COLORECTAL CarCINOMA AND COLORECTAL 

LIVER METASTASES 

Vlastimil Kulda 1 , Martin Pešta 2 , Ondřej Topolčan 2 , Lukáš Řehoř 1 , Martin Svatoň 1 , 

Václav Liška 3 , Václav Babuška 1 , Luboš Holubec 2 and Radim Černý 1 

1 

Department of biochemistry LF UK Plzeň, 2 Department of internal medicine 

II LF UK Plzeň, 3 Department of surgery LF UK Plzeň 

Some of the microRNAs, which are the endogenously expressed regulatory small 

noncoding RNA molecules, have an altered expression in colorectal cancer. The aim of 

our study was to assess the relationship between miR-21 and miR-143 expression and 

prognostic/clinicopathological features of colorectal carcinoma (CRC) and colorectal 

liver metastases (CLM) as well. The estimation was performed in 46 paired (tumor and 

control) tissue samples of CRC. Further we studied 30 tissue samples of CLM. MiR-21 

and miR-143 expressions were quantified using the qRT-PCR method. Relation of miR-21 

and miR-143 expression to DFI (disease free interval) (Wilcoxon; p=0.0026 and p=0.0191, 

respectively) was recorded. There was shorter DFI in patients with a higher expression 

of miR-21 and surprisingly also in patients with a higher expression of miR-143, which 

is a putative tumor suppressor. There was a higher expression of miR-21 and lower 

expression of miR-143 in CRC tissue in comparison with adjacent normal colon tissue 

(p

Posters 

65. 

THE FUNCTIONALITY of aPOPTOSOME APParaTUs aND THE 

EXPRESSION OF ITS rEGULaTOrS IN NON-SMaLL CELL LUNG 

caRCINOMA 

Erika Moravčíková 1 , Evžen Křepela 1 , Jan Procházka 1 , 

Jan Čermák 1 and Kamila Benková 2 

1 

Department of Pneumology and Thoracic Surgery, 2 Department of Pathology, 

University Hospital Bulovka, Prague, Czech Republic 

Deficient signaling in the apoptosome pathway may contribute to tumorigenesis and 

neoplastic progression of malignant tumors as well as to their chemo- and radioresistance. 

In the present study, we investigated the functionality of the apoptosome apparatus 

(AA), the expression of mRNAs encoding the activatable and inactivatable Apaf1 

protein variants, and the expression of AA regulators, including nucling/UACA, APIP, and 

procaspase-2 (PC-2) in human non-small cell lung carcinoma (NSCLC) cells and tissues. 

First, the enzymatic analysis showed that in 2 of 6 examined NSCLC cell lines and in 18 

of 59 NSCLC tissues (from surgically treated patients) the cytochrome-c (cyt-c) plus dATP 

could trigger a significant increase in cytosolic caspase-3-like activity. Furthermore, the 

endogenous as well as the (cyt-c + dATP)-induced caspase-3-like activities were higher 

in NSCLC tissues as compared to matched lungs. Both in the tumors and the lungs, the 

expression of mRNAs encoding the activatable Apaf1-XL and -LC variants was higher 

than the expression of mRNAs encoding the inactivatable Apaf1-LN and -S variants. 

Interestingly, the expression of both nucling/UACA and APIP was downregulated in 

NSCLC tissues as compared to the lungs, but it did not correlate with the AA activation 

in NSCLC cell lines and tissues and lungs. The expression of PC-2 mRNA in the tumors 

was higher as compared to the lungs, but there was no correlation between PC-2 mRNA 

and the endogenous caspase-3-like activity in NSCLC tissues. The results of this study 

indicate that the expression of nucling/UACA, APIP and PC-2 did not importantly affect 

the functionality of AA in NSCLC. 

Acknowledgments: Supported by research projects (MZO 00064211 and NS/9715-3) from 

Ministry of Health, Czech Republic. 


187

Posters 

66. 

EffECT OF OMEGA-3 PUfa ON LIPID PROFILE AND OXIDATIVE STRESS IN 

HYPERCHOLESTEROLEMIC CHILDREN 

Iveta Ondrejovičová 1 , Jana Muchová 1 , Zuzana Paduchová 1 , 

Zuzana Nagyová 2 and Zdeňka Ďuračková 1 

1 

Institute of medical chemistry, biochemistry and clinical biochemistry, 

Medical Faculty, Comenius University, Bratislava, Slovak Republic 

2 

Juvenalia, s.r.o., Pediatric center, Dunajská Streda, Slovak Republic 

Hypercholesterolemia is defined as an impairment of lipid metabolism. It is characterized 

by an increased level of cholesterol above 6.2 mmol/L in adults and above 4.2 mmol/L in 

children. Several studies have proved that every fourth child in Slovakia older than 11 – 12 

years has increased cholesterol levels. The aim of our clinical study was to monitor changes 

in lipid profile in 35 children with a mild hypercholesterolemia (average age 16 ± 3.34 years), 

after daily supplementation with a nutritional product containing omega-3 polyunsaturated 

fatty acids (PUFA) (1000 mg EPA/DHA) and phytosterol esters (1300 mg) during 16 

weeks period. Oxidative stress also participates in pathogenesis of hypercholesterolemia 

and therefore we have monitored the effect of this product on markers of oxidative damage 

to lipids (lipid hydroperoxides, 8-isoprostanes) and the total antioxidative status. We 

collected samples before and after 8 and 16 weeks of administration of the supplement. 

Serum/plasma was prepared by a standard procedure and was stored at -20 ºC. The total 

cholesterol (TCH), LDL-cholesterol (LDL), HDL-cholesterol (HDL), triacylglycerols (TAG), 

C-reactive protein, lipoprotein-A, glucose, uric acid, as well as parameters of oxidative stress 

were analyzed in serum/plasma samples. After 16 weeks of supplement administration we 

have found out that levels of TCH (5.24–4.86 mmol/L, p

Posters 

67. 

ANALYSIS OF UraTE TraNSPORTERS SLC22A12 aND SLC2A9 IN PATIENTS 

WITH RENAL HYPOURICEMIA IN CZECH POPULATION 

Blanka Stibůrková 1 , Makoto Hosoyamada 3 , Kimiyoshi Ichida 4 and Ivan Šebesta 1,2 

1 

Charles University in Prague, First Faculty of Medicine, Institute of Inherited Metabolic 

Disorders and 2 Institute of Clinical Biochemistry and Laboratory Medicine; 3 Division of 

Pharmacotherapeutics, Faculty of Pharmacy, Keio University, Tokyo; 4 Department of 

Pathophysiology, Tokyo University of Pharmacy and Life Sciences, Japan 

Renal hypouricemia is a heterogeneous inherited disorder characterised by impaired 

tubular uric acid transport, reabsorption insufficiency and/or acceleration of secretion 

(OMIM #220150) with severe complications such as acute renal failure and nephrolithiasis. 

The most causative genes are SLC22A12 and SLC2A9. We have selected 14 patients from 

10 families for analysis of the SLC22A12 and SLC2A9 from the group of 569 hypouricemic 

cases. Sequence analysis of SLC22A12 revealed three transitions G366R, T467M, R477H and 

one deletion A416_L418del in seven heterozygotes, three compound hetero-zygotes and 

four homozygotes. Sequence analysis of SLC2A9 revealed three unpublished transitions 

G216R, D281H, N333S, one insertion with frame shifting change p.[I118HfsX27] and four 

published sequence variants G25R, V282I, R294H and P350L in two heterozygotes, five 

compound heterozygotes and two homozygotes. The function and immunohistochemistry 

analysis in Xenopus laevis oocytes including subcellular localization, colocalization and 

processing dynamics and transport of proteins are in process. 

Our finding supports the prediction that intact function of both SLC22A12 and SLC2A9 

transporters is necessary for normal urate reabsorption. Further detailed studies could 

clarify genotype/phenotype relations in hypo/hyper-uricemia and gout and in conditions 

related to hyperuricemia as well. 

Acknowledgement: Acknowledgement: Supported by grants MSM0021620806 and IGA 

MZ 11322 – 4 /2010, Czech Republic. 


189

Posters 

68. 

STUDY OF THE EffECT OF HISTONE DEACETYLASE INHIBITOR ON THE 

SENSITIVITY OF LEUKAEMIC CELLS TO THE CYTOSTATICS 

Andrea Štefániková 1 , Jozef Hatok 1 , Jana Jurečeková 1 , Ivana Plameňová 2 , 

Dušan Dobrota 1 and Peter Račay 1 

1 

Department of Medical Biochemistry, 2 Clinic of Haematology and Transfusiology, 

JLF UK Martin 

One of the possibilities how to overcome a chemoresistance of tumor cells is a combination 

of classic cytostatics with substances possessing a potential to block proliferation 

or induce apoptosis of these cells. Histone deacetylases are a group of enzymes that 

catalyze removing acetic acid residue from histones. This epigenetic process leads to 

condensation of chromatin and suppresion of transcription. The influence of histone 

deacetylases inhibitors is intensively studied nowadays, especially because of their connection 

with antiproliferative and antineoplastic effect. There has long been known an 

inhibitor of histone deacetylases class I - sodium butyrate. It is a short chain fatty acid 

that has effects at the molecular, cellular, and tissue level. By performing of in vitro MTT 

test, we have revealed polyresistance of leukaemic blasts isolated from blood of AML 

patient to almost all cytostatics tested. Addition of sodium butyrate in concentration of 

2mM led to significantly increased chemosensitivity of blasts. In search for molecular 

mechanism of butyrate action we have performed analysis of leukaemic cell line HL60 

treated with sodium butyrate. Our results open possibility that addition of butyrate 

increases chemosensitivity of blasts by alteration of expression of proteins involved in 

apoptosis initiation. 

Acknowledgement: This work was supported by grant UK/223/2010 to Andrea Štefaniková. 


Posters 

69. 

CONTENT OF faTTY ACIDS IN FOOD AND HEALTH STATUS 

Ladislav Vaško and Janka Vašková 

Department of Medical Chemistry, Biochemistry and Clinical Biochemistry, Faculty of 

Medicine, Pavol Jozef Šafárik University in Košice, 040 66 Košice 

Change in blood and tissue fatty acid composition join and trigger quite a few of pathological 

variations. Even incorrect lipid nutrition could be negatively affecting the health 

status. Know edge of fatty acid content and ratio is important to ensure suitable lipid 

nutrition depending on organism physiological needs to prevent morbidity. In case of 

congenital afflictions determination of fatty acid content elaborates diagnostics and 

predestinates diet treatment with defined fatty acids. Demand of cognizance the food 

fatty acid composition, either their influence on health state or prevention infers possible 

solution for right lipid nutrition including direct n-6 and n-3 polyunsaturated fatty acids 

ratio. In feeding trial on laying hens the addition of flax oil in first treatment group and 

fish oil in the second treatment group led to outstanding increase of polyunsaturated 

fatty acid content. It concerned, especially, n-3 acid content in blood, eggs and although 

in fat tissue of laying hens. Results attained n-6: n-3 ratio making possible for consumers 

to improve existing adverse n-6: n-3 ratio by eating such improved white meat and eggs. 

Acknowledgement: Study was supported by Slovak grant agency for Science VEGA 

1/0799/09. 


191

Posters 

70. 

EffECT of HUMIC aCIDS in VIVo 

Janka Vašková and Ladislav Vaško 

Department of Medical Chemistry, Biochemistry and Clinical Biochemistry, Faculty of 

Medicine, Pavol Jozef Šafárik University in Košice, 040 66 Košice 

Humic acids were described as prospective compounds utilized to ensure sufficient quantity 

of food for human population at a high economic profitability of agriculture production 

and moreover protection of the environment. In feeding trial on broiler chickens the 

effects of humic acids as feeding additives on production parameters and activities of 

antioxidant enzymes in blood were tested. 14 800 chickens from treatment group were 

compared to 142000 untouched control chickens. Treatment group showed significant 

decrease in mortality (0.95%) in comparison to control (3.5%), better of feed conversion 

(13%) and faster growth. Comparison of the results from blood sampling on days 14 and 

35 of treatment period showed enhanced cooperation of antioxidant enzymes as glutathione 

peroxidase, glutathione reductase, superoxiddismutase leading to overall lower 

levels of substrate peroxidation. Addition of humic acids to feed in treatment group in 

comparison to control showed better protection against radical generation. 

Acknowledgement: Study was supported by Slovak grant agency for Science VEGA 

1/0799/09. 


Posters 

X. PROTEOMICS AND ENZYMOLOGY 


193

Posters 

71. 

HEXaMEr formaTION trIGGErs a SWITCH frOM an INaCTIve TO an 

aCTIve CONformaTION IN HUMan MITOCHONDrial LON prOTEaSE 

Vladimír Pevala 1 , Jacob A. Bauer 1 , Javier García-Nafría 2 , Gabriela Ondrovičová 1 , 

Ľuboš Ambro 1 , Elena Blagova 2 , Vladimir M. Levdikov 2 , Anthony J. Wilkinson 2 , 

Keith S.Wilson 2 and Eva Kutejová 1 

1 

Institute of Molecular Biology, Department of Biochemistry, SAS, Dúbravská cesta 21, 

845 51 Bratislava, Slovakia, 

2 

Structural Biology Laboratory, Department of Chemistry, University of York, 

York YO10 5YW, UK 

Although Lon is one of the least complicated ATP-dependent proteases, a structure 

of the full-length protein still has not been determined. At present, structures have 

been solved of a fragment of the N-terminus, the small α-domain and the proteolytic 

domain. We determined the crystal structure of the proteolytic domain of the human 

mitochondrial Lon protease. Although the overall structure is very similar to the EcLon 

one, the conformation around the active site more closely resembled that seen in the 

Methanococcus jannaschii Lon structure. A detailed analysis of these three structures led 

us to propose a mechanism by which hexamer formation is coupled to a conformational 

transition at the active site, which converts the inactive conformation seen in the hLon 

structure to one resembling that seen in the EcLon one. To better understand the roles 

of the proteolytic domain in the overall functions of human Lon protease, we designed 

several point mutations in this domain based on the known Lon protease crystal structures. 

We then tested their influence on protease, peptidase, and ATPase activity as well 

as on oligomer formation and stability. 

Acknowledgement: This work was supported by VEGA 2/0141/08, APVV-0024-07 and 

by European Commission funding SPINE2–COMPLEXES project LSHG–CT–2006–031220. 

We thank the staff at the ESRF (beamline ID14-4) for provision of synchrotron facilities, 

and Johan Turkenberg and Sam Hart for assistance with data collection. 


Posters 

72. 

BIOCHEMICal CHaraCTErIZaTION of Rv1459c prOTEIN PUTaTIve 

GT-C GLYCOSYLTraNSferaSE frOM MYCOBaCTEria 

Milo Bystrický 1 , Martina Beláňová 1 , Mary Jackson 2 , 

Katarína Mikušová 1 and Jana Korduláková 1 

1 


Bratislava, Slovakia; 2 Department of Microbiology, Immunology, and Pathology, 

Colorado State University, Fort Collins, CO, USA 

Mycobacterium tuberculosis and related species of the order Actinomycetales carry 

a significant number of glycosyltransferases of the GT-C class. This group of glycosyltransferases 

comprises integral membrane proteins with dependency on polyprenylphosphate-linked 

sugar donors. No protein structure of the GT-C superfamily has yet 

been solved. In mycobacteria GT-C glycosyltransferases are believed to be involved in 

the later biosynthetic steps of key polysaccharides in the mycobacterial cell envelope. 

We focused on the functional characterization of the mycobacterial protein Rv1459c, 

putative GT-C glycosyltransferase encoded by the gene located in the cluster of the 

genes rv1459c-rv1456c. Homologues of the genes rv1459c - rv1456c are present in all 

mycobacterial species sequenced so far. Similar gene clusters were identified also in 

corynebacteria and nocardia - organisms with the cell wall structures very similar to that 

of mycobacteria. The glycosyltransferase Rv1459c was produced in E. coli in the form 

of soluble protein fused with the maltose binding protein. Heterologous production of 

the soluble Rv1459c allowed us to isolate this protein for crystallography purposes, as 

well as for the investigation of the interactions between Rv1459c and the subunits of 

the ABC transporter Rv1458c/Rv1457c/Rv1456c. 

Acknowledgement: This work was supported by Slovak Research and Development 

Agency (grant No. APVV-0499-07) 


195

Posters 

73. 

THE STUDY of rYaNODINE rECEPTOr 2 N-TErMINal rEGION 

rESPONSIBLE for HEart arrYTHMIas aND HEart faILUre 

Ľubomír Borko 1,2 , Vladena Bauerová-Hlinková 2 , Eva Hostinová 2 , 

Juraj Gašperík 2 and Jozef Ševčík 2 

1 

Department of Molecular Biology, PRIF UK, Mlynská dolina, 842 15 Bratislava 

2 

Institute of Molecular Biology, SAV, Dúbravská cesta 21, 845 51 Bratislava 

Ryanodine receptor (RyR) is a homotetramer composed of four subunits with a molecular 

weight of ~560 kDa. In humans there are three isoforms of RyR: RyR1, RyR2 and RyR3. 

RyR1 (expressed mostly in sceletal muscle) and RyR2 (in myocardium) are suggested to 

be of a vital importance. RyRs are localized in the membrane of sarcoplasmatic reticulum. 

The role of RyR is to transport Ca 2+ from sarcopasmatic reticulum to the myoplasm. 

The main RyR2 gating regulation mechanism is considered to be the domain-domain 

interaction between the N – terminal (aa ~ 1-600) and central region (aa ~ 2100-2500). 

A hypothesis was proposed that mutations in these regions cause regulation failure and 

thus lead to nonspecific Ca 2+ release which results in several heart diseases. Ryanodine 

receptor 2, has a domain structure, so we decided to prepare DNA fragments coding 

for residues 384-606, 391-606, 409-606, 1-606 and 1-655. These fragments were cloned 

into appropriate pET expression vectors for heterologous expression in E. coli strains. 

Recombinant polypeptides RyR2 409-606, NusA_409-606, Trx_409-606, Trx_391-606, 

Trx_384-606, 1-606 and 1-655 were obtained in high expression levels. Because of high 

rate of aggregation and instability of these recombinant proteins it was necessary to 

use appropriate detergents, pH values, buffers and salts. Isolation and purification of 

recombinant polypeptides was optimized to obtain soluble monomeric products. To 

study stability and folding we prepared a number of N-terminal mutations (1-606 fragment), 

which are connected to heart arrhythmias and failures. Our work is oriented to 

study domain-domain and domain-ligand interactions and structure determination of 

interacting regions by X-ray crystallography. 


Posters 

74. 

THE fUNCTIONal CHaraCTErIZaTION of THE PUTaTIve 

MYCOBaCTErial ABC traNSPOrTer MSMEG_6366 - MSMEG_6369 

Petronela Dianišková 1 , Jana Korduláková 1 , Henrieta Škovierová 1 , Devinder Kaur 2 , 

Mary Jackson 2 , Patrick J. Brennan 2 and Katarína Mikušová 1 

1 


Bratislava, Slovakia; 2 Mycobacterial Research Laboratories, Department of 

Microbiology, Immunology and Pathology, Colorado State University, 

Fort Collins, Colorado, USA 

The mycobacterial cell envelope differs substantially from the cell walls of other bacteria. 

This unique structure accounts for its unusually low permeability and resistance towards 

common antibiotics and thus enables mycobacteria to survive and multiply within the 

host. The main covalently linked structural element of mycobacterial cell wall consists of 

three entities – peptidoglycan, heteropolymeric arabinogalactan and highly hydrophobic 

mycolic acids. During the last years a number of enzymes involved in the biosynthesis 

of these components have been identified. In spite of this fact, a great challenge is to 

decipher the mechanism of the assembly of this complex structure including the transport 

of the cell wall intermediates across the plasma membrane. In our effort to identify the 

transport proteins that could be involved in this process we focused on a putative ABC 

transporter Rv3781-Rv3783 from M. tuberculosis H37Rv. Rv3781 and Rv3783 genes are 

located around glfT1, the gene encoding galactosyl transferase responsible for the initiation 

of the galactan biosynthesis. Here we demonstrate that orthologs of all three genes are 

co-transcribed in M. smegmatis mc 2 155 and we have also confirmed that recombinant 

Rv3781 ortholog (MSMEG_6366) and GlfT1 (MSMEG_6367) from M. smegmatis mc 2 155 

interact with each other suggesting that they form a complex that could be involved in 

the biosynthesis of the cell wall in mycobacteria. 

Ackowledgement: This work was supported by the Slovak Research and Development 

Agency under the contract No. RPEU-0012-06, by European Comission under contract 

LSHP-CT-2005-018923”NM4TB“, and by the grant AIDS-FIRCA TW 006487 from NIH, NIAID. 


197

Posters 

75. 

EffECT OF DROUGHT ON THE METABOLISM OF TOBACCO PLANTS 

(NICOTIana TABACUM L.) 

Veronika Doubnerová 1 , Lucia Miedzińska 1 , Jana Dobrá 2 , 

Radomíra Vaňková 2 and Helena Ryšlavá 1 

1 

Department of Biochemistry, Faculty of Natural Science, Charles University in Prague, 

2 

Institute of Experimental Botany AS CR 

Drought is one of the most significant types of abiotic stress worldwide, due to almost 

a third of the Earth surface is arid or semi-arid area. In this study the changes in enzyme 

activities of NADP-malic enzyme (EC 1.1.1.40; NADP-ME), phosphoenolpyruvate carboxylase 

(EC 4.1.1.31; PEPC) and pyruvate, phosphate dikinase (EC 2.7.9.1; PPDK) in tobacco 

plants (Nicotiana tabacum L., cv. W38) after drought were investigated. Enzyme activities 

in tobacco leaves were significantly increased during 11 days of stress, PEPC 2-fold, PPDK 

3,3-fold and NADP-ME 4-fold compared to control plants. The regulation of NADP-ME and 

PEPC activities were studied on transcriptional level by the real-time PCR method and 

on translational level-immunochemically. The amount of NADP-ME protein and mRNA 

for chloroplast NADP-ME isoform was increased, but mRNA for cytosolic isoform was 

not affected. The amount of PEPC protein was unchanged; amount of mRNA for PEPC 

was little decreased. Also regulation of PEPC activity was studied, because this enzyme is 

regulated at many levels, especially by phosphorylation. We suppose that the decreased 

activity after alkaline phosphatase treatment, activation by D-glucose-6-phosphate and 

increased ratio of activity at pH 7.1 and 8.1 means enhanced phosphorylation level of 

PEPC molecule in stressed plants. 

Acknowledgements: This work was supported by the grants of Ministry of Education of 

the Czech Republic (grants MSM0021620808 and 1M0505). 


Posters 

76. 

THERMAL STABILITY OF CYTOCHROME C AND α-LACTALBUMIN 

COMPLEXES 

Diana Fedunová 1 , Zuzana Flachbartová 2 , Jaroslava Bágeľová 1 , 

Zuzana Gažová 1 and Marián Antalík 1,2 

1 

Department of Biophysics, Institute of Experimental Physics SAS, Kosice, Slovakia 

2 

Institute of Chemical Sciences, Faculty of Science, P. J. Safarik University, 

Kosice, Slovakia 

Effective activation of apoptosis is one of the important tools for tumor cell treatment. 

Cytochrome c (cyt c) plays relevant role during early phase of apoptosis after release 

from mitochondria in vivo. A new approach is oriented to the study of the ability of cyt 

c to induce apoptosis by its transport from extracellular location. The active transport 

of cyt c to the cells requires complexation with compounds supporting this process. 

We have studied interactions of cyt c with α-lactalbumin (α-LA) in order to find optimal 

conditions for their complexation necessary for using these complexes in induction of 

programmed cell death. We have found that α-LA has only negligible effect on cyt c heme 

pocket within wide pH interval. The complex formation is accompanied by turbidity increase 

even at low protein concentrations. Thermal stability of the complex depends on 

protein concentration ratio and absolute concentration value. The properties of studied 

complexes depend strongly on ionic strength. 

Acknowledgements: This work was supported within the projects Nos. 26220120021, 

26220120001, 26220220005, 2622022033 in frame of SF EU, Centre of Excellence of SAS 

Nanofluid and VEGA 0056, 0038 and 0079. 


199

Posters 

77. 

Rep 34 

prOTEIN ENCODE BY PLaSMID pGP2 frOM aCetobaCTer 

Peter Grones, Zuzana Odnogová and Jozef Grones 

Comenium University, Department of Molecular Biology, Bratislava 

The Acetobacter estunensis Rep 34 

protein participates in the replication of bacterial small 

cryptic plasmid pGP2. The Rep 34 

protein (213 aa, 23.65 kDa) from Acetobacter plasmid 

pGP2, was cloned to the expression vector, that ensure fusion with a His-tag sequence 

(Rep 34 

His-tagged), over-expressed in Escherichia coli and purified by metal-affinity 

chromatography to yield a highly purified and active protein. On this purified protein 

number different activities and motifs were detected. DNA band-shift assays showed that 

the Rep 34 

His-tagged protein bound to the regulation region for replication on the linear 

double-stranded DNA. In the protein was determined phosphatase activity, ATPase activity 

and protein is possible to unwind double strand DNA. After fusion with GFP protein the 

accruement of protein expression with confocal microscopy was proven. 


Posters 

78. 

PRODUCTION OF 3HYDROXYPROPIONALDEHYD BY THE STraINS OF 

LACTOBACILLUS reuTerI 

Hana Kiňová Sepová, Andrea Bilková, František Bilka and Lýdia Bezáková 

Department of Cell and Molecular Biology of Drugs FaF UK Bratislava 

Eight bacterial strains were isolated from stomach mucus of breast fed lamb (C, D, and 

E) and kid (KO4b, KO4m, KO5, KG1, KG4). Four of these strains were identified by sequencing 

of 16S rDNA as Lactobacillus reuteri (E, KO4b, KO4m, and KO5) and tested for 

production of antimicrobial substance 3hydroxypropionaldehyd (3HPA). For this purpose 

PCR primers targeting gene of large subunit of glycerol dehydratase (GD), key enzyme 

involved in 3HPA synthesis, were designed. Presence of this gene-fragment was detected 

in the case of all four strains and the reference strain L. reuteri ATCC 55730. Sequencing 

of PCR product proved that it is fragment of gene of large subunit of GD. The ability to 

produce 3HPA was determined spectrophotometrically, positive reaction was noticed 

in the case of L. reuteri E and reference strain. Quantative analysis of 3HPA production 

by L. reuteri E and L. reuteri ATCC 55730 in aerobic and anaerobic conditions was determined 

spectrophotometrically by tryptophan method. The amount of 3HPA produced 

aerobically was in both strains higher than those produced anaerobically. In the case of 

L. reuteri E it was aerobically 1.60 mmol/L, anaerobically 0.58 mmol/L and in the case of 

L. reuteri ATCC 55730 it was aerobically 184.51 mmol/L and anaerobically 5.71 mmol/L. 

Strain L. reuteri E produced lower amounts of 3-HPA than probiotic strain L. reuteri 

ATCC 55730. However this does not exclude it from the group of potential probiotics; 

it has other important attributes that allow L. reuteri E to be a good probiotic strain for 

veterinary use. 


201

Posters 

79. 

THIOrEDOXIN SYSTEM IN StrEPTOMYCES 

Michaela Koháryová and Marta Kollárová 


Mlynská dolina CH-1, 842 15 Bratislava, Slovak Republic 

Thiol-disulphide bond balance is generally under control of Trx system and/or GSH/GSHR 

system in bacteria. Both systems are the main regulators of redox homeostasis, which 

can modulate biological activity of many proteins and also participate in protection 

against oxidative stress. S. coelicolor A3(2) (like other Streptomycetes) is a suitable model 

organism for studying Trx system, because lacks GSH/Grx system (Newton et al., 1996). 

In the most prokaryotic and eukaryotic organisms are multiple functions accomplished 

by single thioredoxin. The complete genome sequence of S. coelicolor A3(2) (Bentley et 

al., 2000) revealed several possible thioredoxin genes: three genes for thioredoxin, two 

for putative thioredoxins, one for thioredoxin reductase and two for putative thioredoxin. 

Their high sequence similarity can predicted similar physiological functions. All genes 

encode proteins with putative oxidoreductase activities, however their biological roles 

are not determined yet. It seems, that S. coelicolor has a very complex redox system, what 

can be responsible for multicellular development of Streptomyces. We prepared E. coli 

strains with overproduced thioredoxins (A, A2, A3) and also thioredoxin reductase TrxB. 

We purified the proteins and performed their functional analysis. After first screening 

we obtained TrxB protein crystals. 

Acknowledgements: This work was supported by project “BIOMAKRO1 ITMS:26240120003”, 

“BIOMAKRO2 ITMS:26240120027”, thanks to operating program Research and progression 

financed by Europe fund of regional development and also thanks to VEGA grant 1/03/09. 


Posters 

80. 

STUDIES OF NOVEL BIvaLENT TACRINE DERIvaTIVES TarGETING 

ACETYLCHOLINESTEraSES 

Mária Kožurková 1 , Danica Sabolová 1 , Slávka Hamuľaková 1 , Jana Janočková 1 , 

Jana Plšíková 1 , Pavol Kristian 1 , Ján Imrich 1 , Ladislav Janovec 1 , Ondrej Holas 2 , 

Miroslav Pohanka 2 and Kamil Kuča 2 

1 

Institute of Chemistry, Faculty of Sciences, P.J. Šafárik University, Košice, SK 

2 

Faculty of Military Health Sciences, University of Defence, Hradec Králové, CZ 

Derivatives of acridine/tacrine are effective drugs against Alzheimer disease (AD) which 

is associated with the progressive memory loss and other cognitive impairments. The 

primary approach to treating AD is delaying of symptoms of disease and increasing the 

acetylcholinesterases levels in the brain by using acetylcholinesterase inhibitors. The 

cholinesterase inhibitors have been identified according to their binding mode. 

Eleven compounds of acridine/tacrine derivatives were synthesized and their properties 

have been studied by UV-Vis, fluorescence spectrophotometry and circular dichroism. 

The most promising inhibitor of acetylcholine from the newly prepared compounds were 

derivative N-{2-[4-(acridine-9-yl) piperazino]etyl}-N-(1,2,3,4-tetrahydroacridine-9-yl) 

amine (IC 50 

= 0.003 ± 0.0006 µM) and N-(1,2,3,4-tetra-hydroacridine-9-yl)-N´-[2-(1,2,3,4- 

tetrahydroacridine-9-ylamino)butyl]thiourea (IC 50 

= 0.002 ± 0.0004 µM). These compounds 

were two-fold stronger inhibitors compared to standards (tacrine and 7-MEOTA). 

Acknowledgement: This work was supported by the Scientific Grant Agency VEGA 

1/0053/08 and 1/0097/10 (Slovak Republic) and Ministry of Defence– OVUOFVZ200805 

(Czech Republic). 


203

Posters 

81. 

ApplicaTION of CONCENTraTION fLUOrESCENCE matrICES TO THE 

DETECTION of fLUOrESCENCE METaBOLITES IN urINE 

Lucia Lichardusová, Jaroslav Kušnír and Mária Mareková 

Institute of Chemistry, Biochemistry, Medical Biochemistry and LABMED a.s., 

UPJŠ Košice 

Urine contains a variety of organic and inorganic chemicals, including a number of 

natural fluorophores, most of which are tryptophan metabolites. The alteration in the 

autofluorescence of urine could result from both physiological and pathological changes. 

The synchronous fluorescence spectrum (SFS) is considered to be the characteristic “fingerprint“, 

because it is unique for a given mixture and it characterizes it graphically as one 

unit. By introducing concentration parameter, the SFS form concentration fluorescence 

matrices which provide the information of the precise mixture ratios of the urine samples. 

In this work we present application of concentration fluorescence matrices to the detection 

of urine fluorophores, particularly tryptophan and tyrosine metabolites. The SFS 

include autofluorescence of individual fluorophores and their interaction. Every alternation 

of one fluorophor disturbs balance of fluorophores interaction and this is shown by 

alternation graphic record. The CSMF spectra were formed by the SFS at various Δλ. As 

the urine is a complex multi-component system, the shape and intensity of the spectra 

obtained under different Δλ changed distinctly and contained rich and varied information. 

Acknowledgement: This study was supported by VEGA 1/0402/10. 


Posters 

82. 

CRH TraNSGLYCOSYLASES CATALYZE INTER POLYMERIC LINKAGES IN 

FUNGAL CELL WALL 

Marián Mazáň 1 , Noelia Blanco 2 , Javier Arroyo 2 and Vladimír Farkaš 1 

1 

Institute of Chemistry - Center for Glycomics, Slovak Academy of Sciences, 

Dúbravská cesta 9, 84538 Bratislava, Slovakia 

2 

Departamento de Microbiología II, Facultad de Farmacia, 

Universidad Complutense de Madrid, 28040 Madrid, Spain 

The cell wall of Saccharomyces cerevisiae is an elastic structure that provides osmotic 

and physical protection and determines the shape of the cell. Crh1 and Crh2 are GPIlinked 

cell wall proteins required for the formation of linkages between chitin and β-1,3- 

glucose branches of β-1,6-glucan in the cell wall of budding yeast. According to the CAZY 

database, they belong to the glycoside hydrolase family 16. 

We developed a fluorescent in vitro assay system for determination of the transglycosylating 

activity of Crh proteins. His-tagged Crh1p and Crh2p of S. cerevisiae were heterologously 

expressed in Pichia pastoris, purified on Ni-column and their basic biochemical 

characteristics were investigated. 

The following properties of Crh proteins were determined: pH and temperature optima, 

donor and acceptor substrate specificity, effectors, thermal stability and mode of action. 

Acknowledgement: This work was supported by grant no. 2/0011/09 from the Slovak 

Grant Agency for Science VEGA. 


205

Posters 

83. 

DELETION of GLUTamaTE DECarBOXYLaSE GENE frOM trICHODErma 

virIDE F-534 STraIN 

Ľuboš Nižnanský, Svetlana Kryštofova and Ľudovít Varečka 

Department of biochemistry and microbiology, Faculty chemical and food technology 

Enzyme glutamate decarboxylase (GAD) is present in eukaryotic and prokaryotic organisms, 

where is playing role in different processes. GAD catalyses the conversion of L-glutamate 

to a non-coded amino acid 4-aminobutyrate (GABA). In yeasts, GAD is important for 

oxidative stress response to acid environment. In this study, we used electroporation 

to transfer the gad gene disruption cassette with hygromycin resistance into T. viride 

conidia and obtained 21 transformants. Deletion of gad gene was proved by GAD activity 

measurements and Southern analysis in two transformants (mutants 10 and 21). Mutants 

lacking GAD have delayed conidiation. Submerged mycelia formed pellet-shaped mycelia 

during submerged cultivation unlike of wild strain. Microscopic analysis of Δgad10 

and Δgad21 mutants revealed the increase in forming chlamydospores and thinning of 

hyphae. In submerged cultivation with sucrose as carbon source, growth rates of these 

mutants were lower than that of wild-type strain, while the growth on GABA as a carbon 

source showed the opposite pattern. Growth of Δgad10 was more rapid than that of the 

wild-type strain using GABA as carbon source but L-glutamate had an opposite effect. 

Respiratory quotients of these mutants were higher than that of wild-type strain. Thus, 

the gad gene seems to influence basic bioenergetic processes in this fungus with the 

impact on growth and conidiation. 

Acknowledgement: This work was supported by grant APVV nr. 0642-07 


Posters 

84. 

CHaraCTERIZATION OF β-N-ACETYLHEXOSAMINIDASE IN LEavES OF 

TOBACCO PLANTS 

Helena Ryšlavá 1 , Veronika Doubnerová 1 , Robert Valenta 1 , Kateřina Kloudová 1 , 

Jana Trefancová 1 , Helena Synková 2 and Noemi Čeřovská 2 

1 

Department of Biochemistry, Faculty of Natural Science, Charles University in Prague, 

2 

Institute of Experimental Botany AS CR 

Plant glycosidases were characterized in seeds, but there is little information about these 

enzymes in leaves. In tobacco plants (Nicotiana tabacum L., cv. Petit Havana, SR1) the 

activity of α-glucosidase (EC 3.2.1.20), β-glucosidase (EC 3.2.1.21), α-galactosidase (EC 

3.2.1.22), β-galactosidase (EC 3.2.1.23), α-mannosidase (EC 3.2.1.24) and activity of β-Nacetyl-hexosaminidase 

(EC 3.2.1.52) were determined. The highest activity was found 

for β-N-acetyl-hexosaminidase, this enzyme was able to hydrolyze both chromogenic 

substrates p-NP-GlcNAc and p-NP-GalNAc. The products of the reaction were found to 

be non-competitive inhibitors of the reactions catalyzed by β-N-acetyl-hexosaminidase. 

The hydrolysis of another substrate -chitobiose was studied by capillary electrophoresis. 

The size of β-N-acetyl-hexosaminidase molecule was estimated at 245 000. Under biotic 

stress caused by viral infection enhanced activity of β-N-acetyl-hexosaminidase was found. 

Acknowledgements: This work was supported by the grants of Ministry of Education of 

the Czech Republic (grants MSM0021620808 and 1M0505). 


207

Posters 

85. 

DNA BINDING STUDY of 9-OXO-9,10-DIHYDroacrIDINcarBOXYHYDraZIDES 

as POTENT TOPOISOMEraSE I INHIBITOrs 

Danica Sabolová, Lucia Krajňáková, Jana Plšíková and Mária Kožurková 

Department of Biochemistry, Institute of Chemistry, Faculty of Science, 

P.J. Šafárik University, Moyzesova 11, SK-04167 Košice 

DNA, the basic genetic material, is one of the most enigmatic biomolecules and efforts 

to understand the subtleties of its behaviour from a structural and energetic perspective 

have not yet been fully rewarded. The pivotal role played by DNA in the synthesis 

of proteins as well as its own replication makes it an extremely important potential 

target for drugs, especially for anticancer, antibiotic and antiviral action. Regions of DNA 

involved in vital processes like origin of replication, promotion of transcription etc. are 

of particular interest as targets for such drugs. 

In this work, the interactions of acridone derivates 9-oxo-9,10-dihydroacridine-carbohydrazides 

with DNA were studied by a variety of spectroscopic techniques including 

UV-Vis spectrophotometry, fluorimetric titrations and circular dichroism. From spectrofluorimetric 

titrations the binding constants for DNA - drug complexes were determined 

(K= 1.2 × 10 5 M -1 – 5.1 × 10 5 M -1 ). The effect of investigated compounds on the thermal 

denaturation profiles of calf thymus DNA were also studied. By electrophoretic methods 

was determined, that the new drugs were able to inhibit the topoisomerase I. 

Acknowledgements: This work was supported by the Slovak Grant Agency VEGA, No. 

1/0053/08 and 1/0097/10. 


Posters 

86. 

IN VIVO CROSS-LINKING FOR IDENTIFICATION OF TELLURITE 

RESISTANCE-ASSOCIATED PROTEINS 

Jana Schubertová Aradská 1 , Dušan Blaškovič 1 and Ján Turňa 1,2 

1 

Department of molecular biology PriF UK Bratislava, 2 Department of molecular 

biology SAV Bratislava 

In spite of the fact that a number of tellurite-resistance determinants have been 

characterized, little is known about the mechanism responsible for this resistance. 

Determinant encoding resistance against pottasium tellurite was discovered in clinical 

isolate of Escherichia coli KL53 on a large conjugative plasmid pTE53. Analyses showed 

that genes terB, terC, terD and terE are essential for conservation of the resistance. In 

order to understand this mechanism we are looking for tellurite resistance associated 

proteins. For initial investigation of protein-protein interactions we decided to use in vivo 

chemical cross-linkings to effectively capture and identify interacting proteins. Chemical 

cross-linking stabilizes interactions through covalent bond formation, allowing the detection 

of protein-protein interactions in native cells that are weak and/or transient. The 

most commonly used cross-linking reagent is formaldehyde, a water-soluble and cell 

membrane-permeable molecule, which provides fast and reversible cross-linking reaction. 

With this in mind, we decided to investigate the effects of formaldehyde cross-linking. 

All four genes were cloned to pET expression system to produce His-tagged proteins 

and constucts were transformed to E. coli BL21(DE3). These strains were subequently 

co-transformed with plasmid pLK18 which contains the functional part of the tellurite 

resistance operon, encompasing terBCDEF. Strains were cultivated on medium with tellurite 

to induce oxidative stress and living cells were treated with formaldehyde. Crosslinked 

TerB, TerC, TerD and TerE proteins were purificated by Ni-NTA resin and analyzed by 

SDS-PAGE. In the next step we would like to identify associated proteins by MS analyses. 


209

Posters 

87. 

MULTIPLE prOTEaSES are SECrETED BY vEGETaTIve TrIChoderma 

VIride MYCELIUM CULTIvaTED WITH prOTEIN INDUCEr 

Martin Šimkovič, Anita Gdovinová, Zuzka Zemková and Ľudovít Varečka 

Department of biochemistry and microbiology, Faculty of chemical and food 

technology, Slovak Univerzity of Technology, Radlinského 9, 81237 Bratislava 

The cultivation of Trichoderma viride mycelium with purified protein substrate (bovine 

serum albumin, ovalbumin) resulted in the secretion of protease activity into the medium 

under submerged conditions. The secretion began within 30 h and the greatest secretion 

activity was observed after 72 h cultivation. The secretion continued upon the prolonged 

cultivation (up to 8 d) with lower secreted proteolytic activity. Gelatine zymography of 

the secreted protease revealed high-molecular weight protease(s) (~200 kDa) with high 

autolytic activity as the only secretory product. Low-molecular weight protease(s) involved 

in the mycoparasitic activity of Trichoderma spp. was seen after prolonged cultivation 

only, as a band with m.w. about 30 kDa. Expression of known Trichoderma spp. genes 

encoding secreted proteases Prb1, ProA showed that only Prb1 was expressed after 3-4 

days of cultivation, i.e., after fading out the early secretion phase. The secretory activity 

of the earlier phase was impaired by tunicamycin and brefeldin A and was significantly 

stimulated by uncoupler. The existence of biphasic fungal secretory response represents 

a yet not recognized process. The structure and the biological role of secreted proteases 

should be elucidated in the future. 

Acknowledgements: This work was supported by the VEGA Grant Agency project (No. 

1/0434/08) and by the Science and Technology Assistance Agency under the contracts 

Nos. APVV-0642-07, APVT-20-003904 and VVCE-0064-07. 


Posters 

88. 

PHYTOALEXINS REDUCE INSULIN AMYLOID AGGREGATION 

Katarína Šipošová 1 , Andrea Antošová 2 , Peter Kutschy 1 , 

Zuzana Daxnerová 3 and Zuzana Gažová 2 

1 

Institute of Chemical Sciences, 3 Institute of Biology and Ecology, Faculty of Science, 

P. J. Šafárik University, Košice, Slovakia, 2 Department of Biophysics, Institute of 

Experimental Physics SAS, Košice, Slovakia 

Amyloid aggregation, a generic behavior of proteins, is related to incurable human pathologies 

(amyloid-related diseases) associated with formation of amyloid deposits in 

the body. Insulin amyloid deposits have been observed in patients with diabetes after 

repeated subcutaneous insulin injection. The recent data confirm the toxic effect of aggregates 

on the cells, however, it was found that reduction of amyloid aggregates plays 

important role in prevention as well as therapy of amyloidosis. We investigated capability 

of phytoalexin derivatives to affect the insulin amyloid aggregation. Phytoalexins are 

low molecular weight secondary metabolites produced by plants after their exposure to 

biological or physical stress. By ThT and ANS fluorescence assays, the efficiency of derivatives 

to inhibit formation of amyloid aggregates and depolymerize pre-formed amyloid 

fibrils was studied. We identified very effective phytoalexin derivates, benzocamalexin 

and cyclobrassinin, with significant inhibiting and depolymerizing activities. For these 

derivatives, the determined IC50 and DC50 values are at low micromolar concentrations. 

Our data suggest the potential therapeutic use of the most effective phytoalexins in the 

reduction of insulin amyloid aggregation. 

Acknowledgements: This work was supported within the projects Nos. 26220220005, 

26220120033 and 26220120021 in frame of SF EU, Centre of Excellence of SAS Nanofluid, 

VEGA 0079, 0056 and VVGS PF 13/2010/Ch. 


211

Posters 

89. 

THE LysM DOMaIN IN SUrfaCE IMMUNOGENIC prOTEIN (Sip) aND ITS 

INfLUECE ON ELICITaTION of IMMUNITY agaINST StrePTocoCCus 

agalaCTIae 

Barbora Vidová, Michal Chotár and Andrej Godány 


The microbial pathogens are characterized by adapting their surface proteins in order to 

protect themselves against the host immune system; accordingly, there is a possibility 

of some variability also within the sip gene. This work was focused on determining of 

potential variability within the sip gene in the collection of bovine isolates of S. agalactiae 

by multiplex PCR method and characterizing the protein Sip on its nucleotide level, 

with the aim to study its probable variability and prepare the basis for future study 

of its epitope responsible for immunity elicitation. Concurrently, we also examined 

the presence of a peptidoglycan-binding anchor, the LysM domain motif. Because the 

identification of both variability and anchor motif is important for the identification of 

a potential epitope within the Sip, the immune responses to whole and partial Sip were 

also tested in the mice model. 

Acknowledgements: This work was supported by the VEGA grant no. 2/0121 of the Slovak 



Posters 

XI. REACTIVE SPECIES IN BIOMEDICINE 


213

Posters 

90. 

effECT of OMEGa-3 faTTY aCIDS ON PON 1 arYLESTEraSE 

aND laCTONaSE aCTIvITY IN CHILDrEN SUfferING frOM 

HYPErCHOLESTErOLEMIa 

Lucia Andrezálová, Zuzana Országhová, Jana Muchová and Zdeňka Ďuračková 


Faculty of Medicine, Comenius University Bratislava, Slovakia 

Hypercholesterolemia is characterized by an increased level of blood cholesterol and give 

rise to the development of atherosclerosis. The atherosclerosis process begins already in 

childhood and is the main risk factor that contributes to the pathology of cardiovascular 

diseases in adulthood. 

Paraoxonase 1 (Pon 1), HDL-associated enzyme with antioxidative properties, plays an 

important role in the antiatherogenic processes. The aim of this study was to determine 

PON 1 arylesterase and lactonase activity in 35 moderately hypercholesterolemic children 

(average age 16 ± 3.34) and study the changes in its activity during the daily administration 

of nutritional supplement (CW) comprising of omega-3 fatty acids (1000 mg EPA/ 

DHA) and phytosterol esters (1300 mg) over a 16 week period PON 1 arylesterase and 

lactonase activities were determined spectophotometrically towards phenylacetate and 

dihydrocoumarin in serum samples. CW supplementation did not significantly affect PON 1 

activities. While arylesterase activity was within the physiological range, lactonase activity 

was moderately reduced compared to healthy subjects. PON 1 activities were correlated 

with other biochemical parameters as well as with lipid peroxides and homocysteine. 

Acknowledgement: Authors wish to thank for partial financial support to Obsidian Research 

Limited, Pork Talbot (Great Britain), Mind and Health, civil association and MVTS and 

VEGA grants of Ministry of Education SR. 


Posters 

91. 

ANTIOXIDANT RESPONSE TO TREATMENT WITH NATUraL COMPOUNDS 

AND AN NO DONOR IN EXPERIMENTAL HYPERTENSION 

Ima Dovinová 1 , Zuzana Pakanová 2 , StanislavaVranková 1 , Oľga Pecháňová 1 , 

Soňa Čačányiová 1 , František Kristek 1 and Helena Paulíková 2 

1 

Institute of Normal and Pathological Physiology, SAS, Bratislava; 

2 

Department of Biochemistry and Microbiology, Faculty of Chemical and Food 

Technology STU, Bratislava 

SOD enzymes play an important role in cardiovascular tissue by protecting NO against 

oxidative inactivation by superoxide. Both NO and superoxide are free radicals with 

unpaired electrons and can react with one another. Studies with pharmacologically inhibited 

SOD1 and SOD3 showed that NO cannot be released from endothelium without 

oxidative degradation, that is, SOD enzymes play an important role in vasodilatation and 

in protection of NO in blood vessel walls. 

In this study, we observed the effect of flavonoids (Alibernet red wine extract), melatonin, 

and an NO-donor, PETN, on antioxidant response of blood vessels and the heart in 

animal hypertension models, SHR and SHR-cp. 

The experiments show that in young hypertensive rats the Alibernet extract increased 

the activity of SOD and NOS in the left ventricle of SHR and the total antioxidant status 

in plasma. 

In adult hypertensive rats, the NO donor, PETN, increased SOD and GPx activities and 

decreased left ventricle damage. 

The results indicate that the most remarkable effects of the Alibernet red wine extract 

in young hypertensive rats, and of the PETN NO donor in adult hypertensive rats are 

related to protective effects of SOD enzymes in the heart. 

Ackowledgement: The work was supported by VEGA Grant No 2/0066/08. 


215

Posters 

92. 

WILSON’s DISEaSE aND OXIDaTIve STrESS 

Marián Koláček 1 , Jana Muchová 1 , Eva Uhlíková 1 , 

Viera Kupčová 2 and Ladislav Turecký 1 

1 

Institute of medical chemistry, biochemistry a clinical biochemistry 

Faculty of medicine Comenius University, Bratislava 

2 

3 th internal clinic Faculty of medicine Comenius University 

Wilson’s disease (hepatolenticular degeneration) is autosomal hereditary recessive 

disorder of copper metabolism with disorder of copper excretion into the bile and its 

excessive storage in some organs. Disorder is caused by mutation of the gene encoding 

specific copper transport protein – ATP7B for ATPase type P. Mutated ATP7B inhibits 

copper excretion into the bile, stimulates its direct release into the blood and its storage 

in organs and this leads to increased free radicals production. Decreased production 

of functional ceruloplasmin increases iron toxicity and thus oxidative stress becomes 

further increased. 

Our goal was to find out how are selected parameters of oxidative stress affected by 

Wilson’s disease. The studied group consisted of 17 patients with diagnosed Wilson’s 

disease (9 women and 8 men). We determined total antioxidant capacity of blood plasma 

(TEAC), activity of a superoxide dismutase (SOD) in erythrocytes and nitrotyrosine – 

marker of damage of proteins by free radicals derived from nitrogen. 

TEAC, as well as SOD activity in erythrocytes of patients with Wilson’s disease were higher 

compared to control group (2,51 mmol/l vs. 1,22 mmol/l and 11,7 U/g Hb vs. 9,2 U/g 

Hb). We saw no significant difference in levels of nitrotyrosine of patients and controls 

(54,7 nmol/l vs. 53,2 nmol/l). 

The increase of TEAC of blood plasma in patients with Wilson’s disease may be a compensatory 

response to the decline of ferooxidase activity of ceruloplasmin in serum as 

well as a response to increased oxidative stress in liver. 


Posters 

93. 

AGE-RELATED CHANGES IN ACTIVITIES OF MITOCHONDRIAL ELECTRON 

TraNSPORT CHAIN COMPLEXES IN THE raT HEarT. 

Stanislav Kuka, Zuzana Tatarková, Peter Račay, Ján Lehotský, 

Dušan Dobrota and Peter Kaplán 

Department of Medical Biochemistry Jessenius Faculty of Medicine, 

Comenius University, Martin 

Oxidative damage of tissues by reactive oxygen species (ROS) has been implicated to be 

a major factor of aging process. Mitochondria are considered to be the main intracellular 

sources of ROS as well as the major target for their damaging effects. 

In our study we examined age-related changes in activities of electron transport chain 

complexes in cardiac mitochondria of adult (6 months), old (15 months) and senescent 

(26 months) male Wistar rats. Our results display non-uniform changes in activities of 

particular ETC complexes. While old rats showed no significant changes in activities of 

complexes I and II compared with adult control rats, in senescent rats the activities of 

both complexes were significantly decreased (P < 0.01). On the other hand, complex III 

activity was significantly lower already at 15 months (P < 0.05) and further decreased at 

age of 26 months (P < 0.01). The most pronounced age-related changes were observed 

in complex IV activity. Compared to adult, complex IV activity decreased by 21% (P < 

0.01) at old age and by 37% (P

Posters 

94. 

SELECTIve PHOSPHODIESTEraSE-3 INHIBITOr OLPrINONE aLLEviaTES 

OXIDaTIve LUNG INJUry INDUCED BY MECONIUM 

Daniela Mokrá 1 , Anna Drgová 2 , Rudolf Pullmann st. 3 and Andrea Čalkovská 1 

1 

Department of Physiology, 2 Department of Biochemistry, 3 Department of Clinical 

Biochemistry, Jessenius Faculty of Medicine, Comenius University, Martin 

In the pathophysiology of neonatal meconium aspiration syndrome, inflammation and 

oxidation play a key role. This study evaluated effects of phosphodiesterase (PDE)-3 

inhibitor olprinone on meconium-induced lung injury. 

Oxygen-ventilated adult rabbits received intratracheally meconium or saline (Sal, n=7). 

Thirty minutes after meconium instillation, animals were treated by olprinone (0.2 mg/kg 

i.v., Mec+Olp, n=7) or were left without treatment (Mec, n=8). All animals were oxygenventilated 

for additional 5 hours. Left lungs were saline-lavaged and differential WBC in 

the sediment was estimated. Right lungs were used to determine lung edema by wet/ 

dry weight ratio and oxidative damage by estimation of thiol content, conjugated dienes, 

thiobarbituric acid-reactive substances (TBARS), dityrosines and lysine-lipid peroxidation 

products concentrations, cytochrome c oxidase activity (COX) in mitochondria of lung, 

and total antioxidant status (TAS) in lung homogenate. In blood plasma, TBARS and TAS 

were determined. 

Meconium instillation increased lung neutrophils and edema formation and concentrations 

of oxidation markers and decreased concentration of TAS. Olprinone reduced the lung 

edema, lung neutrophils and several oxidation markers and increased TAS concentrations. 

Selective PDE-3 inhibitor olprinone effectively reduced oxidative lung injury in meconiuminstilled 

animals. 

Acknowledgments: Supported by Project “Center of Excellency in Perinatology” No. 

26220120016 co-financed by EC, and by Grant VEGA No. 1/0061/08. 


Posters 

95. 

METYLNICOTINAMIDE AND DNA OXIDATION DAMAGE IN raTS WITH 

STREPTOZOTOCINE INDUCED DIABETES MELLITUS 

Zuzana Országhová 1 , Zuzana Paduchová 1 , Ingrid Žitňanová 1 , Cezary Watala 2 , 

Jana Muchová 1 and Zdeňka Ďuračková 1 

1 



2 

Department of Haemostasis and Haemostatic Disorders, 

Medical University of Lodz, Poland 

Increased oxidative and glycooxidative stresses contribute to the development and progression 

of diabetes-associated complications. Under conditions of DM many important 

biomolecules including DNA, lipids or proteins are affected. N(1)methylnicotin-amide 

chloride (MNA), which is a derivative of nicotinamide, is an intensively studied agent 

possessing remarkable anti-inflammatory as well as antidiabetic properties. This effect is 

associated with scavenging of reactive forms of oxygen, in particular superoxide radical 

anion and hydroxyl radical. In this study we have studied the level of oxidative damage to 

DNA in lymphocytes and liver tissue as well as markers of glycation (AGEs) and oxidation 

(carbonyls) damage of proteins in streptozotocine induced diabetic rats. Administration 

of MNA in dose 200 mg/kg of weight during 7 weeks significantly decreased DNA damage 

in lymphocytes and level of protein carbonyls. All doses of MNA decreased DNA damage 

in liver tissue. Advanced glycation of proteins were not affected with MNA. Our results 

show possible beneficial protective role of MNA against oxidative stress and damage of 

important biomolecules under the conditions of DM. 

Acknowledgement: This study was partially supported by MVTS and VEGA grants. 


219

Posters 

96. 

AntioxidaNT capaCITY of SELECTED anaLOGS of NUCLEIC aCID 

COMPONENTS: COMParISON of CELL-frEE aND CELL-baSED aSSaYS 

Eliška Procházková, Petr Jansa, Lucie Čechová, Ivan Votruba and 

Helena Mertlíková-Kaiserová 



Overproduction of reactive oxygen species (ROS) is a phenomenon accompanying a number 

of human diseases as well as drug toxicity. Pharmacological potential of the analogs 

of nucleic acid components is of considerable interest and so is their ability to interfere 

with ROS production. In this study we used a series of poly-substituted pyrimidines 

and purines in order to explore their antioxidant capacity. Their ability of direct radical 

scavenging was first determined with use of the trolox equivalent antioxidant capacity 

(TEAC) assay. Consequently, the effects of the tested compounds on Fe 2+ /ascorbateinduced 

lipid peroxidation in rat liver microsomes were studied. Finally, ROS cellular 

levels were traced in compound-pretreated HepG2 cells exposed to oxidative stress by 

means of a redox-sensitive fluorescein-based probe CM-H 2 

DCFDA. Although remarkable 

antioxidant capacity of several compounds was detected in individual assays, we 

observed that the results largely depend on the method used and the data therefore 

have to be interpeted with caution. We suggest that these variations could be due to 

different physico-chemical properties of the compounds. 

Acknowledgement: This work was supported by the Research project of the IOCB 

OZ40550506 and the Project #1M0508 by the Ministry of Education, Youth and Sports 

of the Czech Republic. 


Posters 

97. 

EffECTIVENESS OF PHENOLS AS ANTIOXIDANTS AGAINST SUPEROXIDE 

raDICAL 

Beáta Veliká and Ivan Kron 

Department of Medical Chemistry, Biochemistry, Clinical Biochemistry and 

LABMED a.s, Faculty of Medicine, PJ Šafárik University in Košice, Slovakia 

Phenolic antioxidants are included in the category of free radical terminators. It is known, 

that phenols reduce rates of oxidation of organic mater by transferring a hydrogen atom 

from OH groups to the chain carrying ROO° radicals (Foti, 2007). It was proved, that the 

intramolecular hydrogen bonding has an important effect on the antioxidant properties 

of phenols. Various ortho-substituents may function as H-bond acceptors (Foti, 2007). 

For the study of antioxidant properties of selected compounds (resorcinol, hydroquinone, 

pyrocatechol and pyrogallol) we used a generator of superoxide radical (Beauchamp and 

Fridovich, 1971) with NBT as a detector after 5, 10, 15 and 20 min of UV illumination. These 

compounds are able to capture and acquit an electron, what depends on the structure 

of used compounds. The antioxidant activity of phenols is related to the number and 

position of hydroxyl groups on benzene skeleton. The best antioxidant activity showed 

dihydroxyderivatives in para- and ortho- position. On the other hand, prooxidant properties 

were recorded for resorcinol and pyrogallol in time and concentration dependance. 

Our study confirms the relationship between antioxidant properties and structure. 

Acknowledgement: VEGA 1/0624/08. 


221

Posters 

98. 

EffECT OF N-3 POLYUNSATUraTED faTTY ACIDS SUPPLEMENTATION ON 

raT LIVER IN DIffERENT THYROID STATUSES 

Martina Vokurková 1,2 , Hana Rauchová1,2 , Stanislav Pavelka 1 , 

Tomáš Soukup 1 and Narcis Tribulová3 

1 

Institute of Physiology, Academy of Science of the Czech Republic, 2 Centre for 

Cardiovascular Research, Prague, Czech Republic, 3 Institute for Heart Research, 

Slovak Academy of Sciences, Bratislava, Slovak Republic 

It is known that thyroid hormones influence lipid metabolism and oxidative stress. 

Epidemiological studies demonstrate that n-3 polyunsaturated fatty acids (PUFAs) consumption 

is associated with a reduced risk of atherosclerosis and hyperlipidemia. The aim 

of our study was to test how PUFAs supplementation in different thyroid statuses of rat 

can affect lipid metabolism and parameters of oxidative stress in the liver. The different 

thyroid statuses of the experimental groups (euthyroid, hyperthyroid and hypothyroid) 

were defined by the levels of the thyroid hormones in the plasma and an activity of liver 

mitochondrial glycerol-3-phosphate dehydrogenase. In this study, the hyperthyroid rats 

had significantly decreased levels of total plasma cholesterol and LDL-cholesterol than 

the hypothyroid rats. The levels of HDL-cholesterol were increased in the hyperthyroid 

rats in comparison with the euthyroid rats. The concentrations of liver thiol groups differed 

according to the thyroid statuses (they were the highest in the hypothyroid group 

and the lowest in the hyperthyroid group). However, no differences in the content of 

conjugated dienes were observed among the experimental groups. PUFAs supplementation 

thus, in our experimental design, modified neither lipid metabolism nor parameters 

of the oxidative stress in the liver. 

Acknowledgements: This work was supported by the GACR (303/09/0570) and Ministry 

of Education, Youth and Sport (1M6798582302 and AV0Z 50110509). 


Posters 

99. 

EffECT OF RENAL TraNSPLANTATION ON OXIDATIVE STRESS-RELATED 

BIOMarKERS 

Adéla Zdařilová 1 , Alena Rajnochová Svobodová 1 , Jana Zapletalová 2 , Pavel Štrebl 3 , 

Josef Zadražil 3 and Jitka Vostálová 1 

1 

Department of Medical Chemistry and Biochemistry, 2 Department of Medical 

Biophysics, Faculty of Medicine and Dentistry, Palacký University, Hněvotínská 3, 

775 15 Olomouc; 3 Department of Internal Medicine III University Hospital, 

I. P. Pavlova 6, 775 20 Olomouc, Czech Republic 

Patients after kidney transplantation (KT) have a high prevalence of oxidative stress (OS) 

that is associated with the excess of cardiovascular complications observed in this population. 

Determination of OS-related parameters together with specific kidney function 

markers is used to monitor graft function. 

Time-dependent changes in OS-related markers in patients (N = 39; male = 23; female 

= 16; mean age = 57 ± 10) before (day 0) and after KT (day 1, 7, 30, 90 and 180) were 

evaluated. Total antioxidant capacity (TAC), levels of advanced oxidation protein products 

(AOPP), lipid peroxidation as thiobarbituric acid-reactive substances (TBARS) and 

reduced glutathione (GSH), activities of glutathione peroxidase (GPX), catalase (CAT) and 

superoxide dismutase (SOD) and kidney function markers were measured. 

AOPP, TAC and TBARS were significantly decreased whereas GSH was significantly increased 

after KT. Antioxidant enzyme activities did not changed after KT during monitored period. 

Of specific kidney function markers glomerular filtration was significantly increased and 

creatinine level was significantly decreased after KT. 

Acknowledgements: This work was supported by grants MSM 6198959216 and MZ CR 

(NS/9964-4). 


223

Posters 

XIII. XENOBIOCHEMISTRY 


Posters 

100. 

BIOTraNSformaTION of SELECTED aNTHELMINTICS IN raT 

taPEWOrm HYMENOLEPIS DIMINUTa 

Hana Bártíková, Jana Firbasová, Ivan Vokřál, Lenka Skálová, Jiří Lamka, 

Vladimír Kubíček and Barbora Szotáková 

Faculty of Pharmacy, Charles University, Hradec Králové, Czech Republic 

In all organisms, drug metabolising enzymes (DME) serve as an efficient defence against 

the potential negative action of xenobiotics. While human and mammalian DME have 

been intensively studied for many years, DME of helminth parasites have been relatively 

little investigated so far in spite of the fact that helminth´ DME can preserve the parasite 

against the anthelmintic drug and could contribute to drug-resistance development. The 

present project was designed to evaluate DME activities in rat tapeworm (Hymenolepis 

diminuta), This tapeworm, infecting mammals and causing hymenolepiasis, is often used 

as model species for investigation into Cestodes as the life cycle of H. diminuta is relatively 

simple, involves rodents (rats primarily) as the definitive host and beetles (Tribolium or 

Tenebrio) as the intermediate host. The implementation of H. Diminuta breeding was 

the first task of our project. Rats were experimentally infected with H. diminuta cysticercoids 

obtained from T. molitor and then adult worms were collected from rat small 

intestine. Activities of oxidation enzymes, carbonyl-reducing enzymes and conjugation 

enzymes were tested in subcellular fractions of H. diminuta homogenate. The results 

showed the ability of H. diminuta enzymes to reduce effectively the carbonyl group of 

xenobiotics, e.g. anthelmintics flubendazole and mebendazole. Catalase, peroxidases, 

UDP-glucuronosyl transferase and glutathione-S-transferase activities were detected as 

well. These results document the ability of H. diminuta to detoxify different xenobiotics 

including certain anthelmintics. 

Acknowledgements: This project was supported by the Czech Science Foundation, grant 

No. P502/10/0217 and by SVV-2010-261-003. 


225

Posters 

101. 

INHIBITOry effECT of naTUral flavONOIDS ON EQUINE LIver 

GLUTatHIONE s-traNSferaSE 

Iva Boušová, Zuzana Průchová, Lucie Trnková and Jaroslav Dršata 

Department of Biochemical Sciences, Charles University in Prague, Faculty of 

Pharmacy, Hradec Králové 

Mammalian glutathione S-transferases (GST, EC 2.5.1.18), group of intracellular enzymes 

involved in detoxification of xenobiotics, belong to the most abundant cytosolic proteins. 

They catalyze conjugation of the thiol group of the glutathione (GSH) to electrophilic xenobiotics, 

and thereby defend cells against the mutagenic, carcinogenic, and toxic effects of 

these compounds. Several classes of GST isozymes differing in substrate specificity exist. 

Flavonoids are a group of polyphenolic compounds that are widely distributed in plant 

kingdom. They are known to possess antioxidant, anti-radical, antiviral, antibacterial, 

anti-inflammatory, and vasodilatory activity. Besides positive acting, flavonoids exert 

also pro-oxidative effects and are able to inhibit various enzymes (e.g. cyclooxygenase 

and glutathione peroxidase). We suppose that dietary flavonoids may inhibit GST and 

thus influence detoxification of xenobiotics. 

GST was incubated in the presence or absence of flavonoids (0-100 µM). Then its activity 

was measured using common spectrophotometric assay with 1chloro-2,4-dinitrobenzene 

adapted to 96-well plates. Inhibitory effect of studied flavonoids was characterized by IC 50 

values. The most pronounced inhibition was found in the case of gallocatechin gallate (IC50 

= 1.26 µM), which is the major polyphenolic constituent in the green tea. The obtained 

results indicated that GST can be a potential target for inhibitory effect of flavonoids. 

Acknowledgments: This study was supported by the Czech Science Foundation (GAČR, 

grant No. 524/09/P121). 


Posters 

102. 

MULTIDRUG RESISTANT P-GLYCOPROTEIN POSITIVE CELLS arE ALSO 

CROSS-RESISTANT TO CISPLATIN 

Lenka Gibalová 1 , Ján Sedlák 2 , Alena Reháková 1 , Martina Labudová 3 , 

Zdena Sulová 1 and Albert Breier 1 

1 

Institute of Molecular Physiology and Genetics SAS Bratislava, 2 Cancer Research 

Institute SAS Bratislava, 3 Institute of Virology SAS Bratislava 

P-glycoprotein (P-gp, a drug transporter of the plasma membrane) mediated multidrug 

resistance of neoplastic cells represents a real problem in the chemotherapeutic treatment. 

Mouse leukemia cells L1210 (S) and two drug resistant cell lines, in which the 

overexpression of P-gp was induced by i. selection with vincristine (R) or by ii. transfection 

of S cells with human gene for P-gp (T) are our experimental models. Cisplatin (cisPt) 

is a substance untransportable by P-gp. We found that R and T cells are also resistant 

to cisPt. However, cisPt resistance in R and T cells could not be reversed by verapamil 

(known P-gp inhibitor), that excluding responsibility of P-gp transport activity for this 

resistance. CisPt induced more pronounced entry to apoptosis in S cells than in R or T 

cells. While similar levels of Bax and Bcl-2 proteins were observed in P-gp negative and 

positive cells, cisPt induced a more significant decrease of the Bcl-2 levels in S cells than 

in R and T cells. Consistent with this, cisPt induced a larger decrease of the Bcl-2 content 

in the Bcl-2/Bax heterooligomer in S cells than in R cells. Moreover, cisPt induced 

apoptotic DNA fragmentation was observed in all three cell sublines. All observations 

indicated that expression of P-gp in L1210 cells is directly associated with reduction of 

cisPt induced apoptosis and consequently with resistance to this drug that is not connected 

with P-gp induced cisPt efflux. 

Ackowledgement: This work was supported by: VVCE-0064-07, APVV-0084-07 and VEGA 

2/0123/10, 2/0155/09. 


227

Posters 

103. 

THE effECTIvENESS of oraCIN IN ENHaNCING THE CYTOTOXICITY 

of DOXOrUBICIN THrOUGH THE INHIBITION of DOXOrUBICIN 

DEaCTIvaTION IN breaST caNCEr CELL LINE MCF-7 

Veronika Hanušová 1 , Lenka Vildová 1 , Věra Králová 2 , Ladislava Schröterová 2 , 

Lenka Trilecová 3 , Alena Pakostová 1 and Lenka Skálová 1 

1 

Department of Biochemical Sciences, Faculty of Pharmacy, Charles University, Hradec 

Králové, Czech Republic, 2 Department of Biology, Faculty of Medicine, 

Charles University, Hradec Králové, Czech Republic 3 Veterinary Research Institute, 

Brno, Czech Republic 

Doxorubicin (DOX) has played a key role in the treatment of breast cancer, but the clinical 

utility of this agent is strictly limited by his associated toxicities, especially cardiotoxicity. 

In present project was studied the possibilities of inhibition of the DOX reduction in breast 

cancer cell line MCF7, one possible strategy to improve DOX efficacy. DOX is deactivated 

mainly by the action of carbonyl reductases in MCF-7. Oracin (ORC, 6-[2-(2-hydroxyethyl) 

aminoethyl]-5,11-dioxo-5,6-dihydro-11H-indeno[1,2-c] isoquinoline) significantly inhibited 

DOX reduction in MCF-7 cytosol. The cytotoxicity of DOX, ORC and DOX+ORC combinations 

in MCF7 cells was assayed using three different end-point tests of cell viability. In 

all DOX+ORC combinations, only non-cytotoxic ORC concentrations were used to avoid 

the possible additive toxic effect of ORC. After a 48-hour exposition, DOX together with 

ORC was able to kill 55 % more cells than DOX alone. All DOX+ORC combinations were 

more toxic in rapidly proliferating cells than in slowly proliferating cells. ORC significantly 

increases DOX efficacy in MCF-7 cells, probably due to the inhibition of DOX reductases. 

Results suggest that concomitant use of ORC and DOX may improve the therapeutic index 

of DOX in the managements of breast cancer. 


Posters 

104. 

CYTOCHrOME b 5 

POTENTIaTES aCTIvITIES of CYTOCHrOMES 

P450 1A1 aND 1A2 TO OXIDIZE aNTICaNCEr drUG ELLIPTICINE TO 

PHarmaCOLOGICaLLY effICIENT METaBOLITES 

Věra Kotrbová 1 , Barbora Mrázová 1 , Eva Frei 2 and Marie Stiborová 1 

1 

Department of Biochemistry, Faculty of Science, Charles University, Albertov 2030, 

128 40 Prague 2, Czech Republic; 2 Department of Molecular Toxicology, 

German Cancer Research Center, 69 120 Heidelberg, Germany 

Ellipticine is an alkaloid exhibiting significant antineoplastic activities. Its mode of action 

is based mainly on cytochrome P450 (CYP)-mediated formation of covalent DNA 

adducts. Many CYP-dependent reactions have been shown to be stimulated by another 

microsomal protein, cytochrome b 5 

(b 5 

). Five ellipticine metabolites, 9-OH-, 12-OH-, 

13-OH-, 7-OH-ellipticine and the ellipticine N 2 -oxide, are generated by CYP1A1/2. The 

patterns and amounts of ellipticine metabolites vary significantly when b 5 

is present 

in the incubations. The formation of detoxication products of its oxidation (7-OH- and 

9-OH-ellipticine) is decreased, while generation of 13-OH- and 12-OH-ellipticine, the 

metabolites responsible for formation of DNA adducts, is increased considerably. The 

enhanced generation of ellipticine-DNA adducts in the presence of b 5 

in the system was 

confirmed by 32 P postlabeling. The treatment of rats with ellipticine resulted in elevated 

expression of b 5 

. Other proteins with or without heme in their molecules are without 

such effects on ellipticine oxidation. Cytochrome b 5 

affects also the kinetics of ellipticine 

oxidation by CYP1A1/2. In the case of the CYP1A1, the presence of b 5 

changes the kinetics 

of ellipticine oxidation to 12-OH- and 13-OH-ellipticine from hyperbolic to sigmoidal, 

whereas no cooperativity was observed with CYP1A2. 

Acknowledgements: Supported by GACR (203/09/0812, P301/10/0356), the Czech Ministry 

of Education (MSM0021620808, 1M0505) and GAUK (127208). 


229

Posters 

105. 

SELENITE-INDUCED CELL DEATH IN HUMAN COLON CANCER CELLS 

Věra Králová and Emil Rudolf 

Department of Medical Biology and Genetics, Faculty of Medicine in Hradec Králové 

Selenium compounds have been shown to play role in chemoprevention of tumours. 

Sodium selenite was reported to inhibit proliferation and induce different types of cell 

death in cancer cells in vitro. In our study we tested the effects of sodium selenite on 

cell proliferation and cell death in human colon cancer cell line HCT 116. Cell proliferation 

was measured by xCELLigence impedance method. Cell morfology was followed by 

time-lapse videomicroscopy. Caspase activity and mitochondrial membrane potential 

were determined by flow cytometry. Activation of DNA damage response was detected 

using western blotting and immunofluorescence. Sodium selenite at concentration of 10 

µM induced cell death in HCT 116 cells characterised by cell detachment and rounding, 

decrease in mitochondrial membrane potential and activation of caspases. Seleniteinduced 

cell death was prevented with antioxidant MnTMPyP and enhanced by depletion 

of reduced glutathion with BSO, which suggests role of oxidative stress. The process was 

accompanied by increase in ATM kinase and histone H2A.X phosphorylation. 

Acknowledgements: This work was supported by Charles University Grant Agency (GAUK) 

Research Project 129609. 


Posters 

106. 

EffECTS of flavONOIDS ON CYTOCHROMES P450 afTER Peroral 

SINGLE DOSE aDMINISTraTION TO maLE raTS 

Jitka Křížková, Kamila Burdová, Petr Hodek and Marie Stiborová 


Hlavova 2030, Praha 2, 128 43 

In recent years, the consumption and use of dietary supplements containing concentrated 

phytochemicals (e.g. flavonoids) increased dramatically. Flavonoids, as foreign compounds 

(xenobiotics), have great potential to modulate the activity of cytochrome P450s 

(CYPs), xenobiotic-metabolizing enzymes involved in the activation and detoxification of 

food and environmental carcinogens. Thus, the aim of this study was to investigate the 

effects of selected flavonoids on CYPs in rat liver and small intestine, as the two main 

organs responsible for xenobiotic metabolism, after p.o. single dose (60 mg/kg b.w.) 

administration to male rats. 

The expression and specific activity of CYP1A and CYP2B were determined using Western 

blotting technique and specific activity assays with alkyl-resorufin derivatives. The rats 

were sacrificed 24h, 48h and 72h after the single dose treatment. 

To evaluate the effects of flavonoids on CYPs along small intestine, the tissue was dissected 

into proximal (near pylorus), middle and distal parts. Of the two natural compounds, 

isoquercitrin was the most efficient CYP1A1 inducer in the proximal part of small intestine 

72h after the single dose treatment. Obtained data demonstrate the different effects of 

selected flavonoids on CYP expression in rat liver and small intestine in dependence on 

the time after the administration. Since these phytochemicals are xenobiotics, and thus 

they can increase the human risk of cancer development, their consumption in large 

quantities should be carefully considered. 

Acknowledgements: This work was supported by the Czech Science Foundation (Grant 

No. 305/09/H008) and the Grant Agency of Charles University (Grant No. 4909). 


231

Posters 

107. 

ActivITIES of drUG-METaBOLIZING aND aNTIOXIDaNT ENZYMES 

IN Haemonchus contortus STraINS rESISTaNT or SENSITIve TO 

aNTHELMINTICS 

Tamara Lasotová 1 , Hana Bártíková 1 , Ivan Vokřál 1 , Barbora Szotáková 1 , 

Vladimír Kubíček 1 , Jiří Lamka 1 , Marián Várady 2 and Lenka Skálová 1 

1 

Faculty of Pharmacy, Charles University, Hradec Králové, Czech Republic 

2 

Institute of Parasitology, Slovak Academy of Sciences, Košice, Slovakia 

Haemonchus contortus is one of the most pathogenic parasites of small ruminants (e.g., 

sheep and goat). The treatment of haemonchosis is complicated because of frequent 

resistance of H. contortus to common anthelmintics. The aim of this project was to 

compare the activities of antioxidant enzymes and biotransformation enzymes toward 

selected anthelmintics and model xenobiotics in three different strains of H. contortus: 

ISE strain (sensitive to common anthelmintics), BR strain (resistant to benzimidazole 

anthelmintics) and WR strain (resistant to all common anthelmintics). For this purpose, 

lambs were experimentally infected with H. contortus L3 larvae and then adults 

worms were collected from ovine abomasums. The biotransformation of anthelmintics 

albnedazole and flubendazole was tested in vitro (in subcellular fractions of H. contortus 

homogenate) as well as ex vivo (in living nematodes cultivated in flasks with medium). 

In vitro, the activities of several carbonyl-reducing enzymes, UDP-glucosyltransferase, 

glutathione-S-transferases, peroxidases, catalase, and superoxid dismutase toward model 

substrates were tested. The acquired data showed significant differences in tested activities 

between sensitive and resistant H. contortus strains. The altered activities of certain 

detoxifying enzymes could protect some parasites against the toxic effect of the drugs 

and contribute to drug-resistance of these parasites. 

Acknowledgement: This project was supported by the Czech Science Foundation, grant 

No. P502/10/0217. 


Posters 

108. 

cytochrOMES P450 1A1/2 OXIDIZE carCINOGENIC arISTOLOCHIC 

aCID I forMING ITS DETOXICaTION METaBOLITE aND DECreaSING 

LEvELS of aa-DNA aDDUCTS IN vivo 

Kateřina Levová 1 , Jana Šístková 1 , Eva Frei 2 , Volker M. Arlt 3 , David H. Phillips 3 , 

Heinz H. Schmeiser 2 and Marie Stiborová 1 

1 


128 40 Prague 2, Czech Republic; 2 German Cancer Research Center, 69 120 Heidelberg, 

Germany; 3 Institute of Cancer Research, Section of Molecular Carcinogenesis, Sutton, 

Surrey, SM2 5NG, United Kingdom 

Aristolochic acid (AA) causes the development of aristolochic acid nephropathy (AAN), 

the Balkan endemic nephropathy (BEN) and urothelial cancer. One of the common 

features of AAN and BEN is that not all individuals exposed to AA suffer from these diseases. 

We found that hepatic microsomes of wild-type (WT) mice oxidize AAI in vitro to 

detoxication metabolite, AAIa, while those of a HRN line were without this effect. Levels 

of AA-DNA adducts in organs of WT mice exposed to AAI were up to more than 10-fold 

lower than in those of HRN mice. To define the role of CYP enzymes in AAI oxidation, we 

used besides hepatic microsomes of these mouse models, also those of human and rat. 

Using correlations between the CYP catalytic activities provided with the set of human 

hepatic microsomes from 14 different human donors and the rates of formation of AAIa 

metabolite in the same set of human hepatic microsomes, we found a highly significant 

correlation coefficient between the rates of phenacetin O-deethylase, a marker for 

CYP1A2, and the levels of AAIa. The results demonstrate a major role of hepatic CYPs in 

AAI detoxication in vivo and that of CYP1A1/2 in this reaction in vitro. 

Acknowledgements: Supported by GACR (303/09/0472, 305/09/H008) and Ministry of 

Education (MSM 0021620808). 


233

Posters 

109. 

THE EffECT OF BENDIOCarB ON ANTIOXIDANT ParaMETERS IN MALE 

AND FEMALE ORGANS OF raBBIT 

Anna Sobeková 1 , Ľuboslava Lohajová 1 and Peter Javorský 2 

1 

Department of Chemistry, Biochemistry and Biophysics UVLF in Košice 

2 

Institute of Animal Physiology SAV Košice 

The toxicity of bendiocarb is believed to be connected with the acetylcholine esterase 

inhibition. However, the toxic effect of pesticides is cumulated from all their metabolites. 

Male and female rabbits were orally administered bendiocarb at a dose 5 mg.kg -1 body 

weight for 30 days. Administration of bendiocarb resulted in a significant alterations of 

antioxidant and detoxifying enzymes activities (SOD-superoxide dismutase, CAT-catalase, 

GPx-glutathione peroxidase) in liver and kidney of male and female rabbits. In the male 

liver, the activities of SOD and CAT were not affected by bendiocarb, but changes in SOD 

isoenzymes pattern were observed in the experimental groups. The activities of GPx 

were significantly decreased on the 3rd and the 10th day of an experiment. Significantly 

higher levels of TBARS indicate the oxidative stress of the organ. The activity of GPx does 

not change in female liver. In the kidney, the activities of all followed enzymes changed 

without significant increasing of TBARS. 

The inhibition of antioxidant defence system, increased TBARS values and changes in 

SOD isoenzyme pattern showed that the toxic effect of bendiocarb is not only in the 

acetylcholine esterase inhibition, but probably also in ROS production. ROS can be produced 

by cyclic biotransformation of bendiocarb and its metabolites. Our results also 

show the alterations in the level of some antioxidant parameters regarding variation in 

the response of male and female animals. 

Acknowledgement: This work was supported by grant project VEGA 2/0021/09. 


Posters 

110. 

INTEraCTION of PLaSMID DNA aND MITOCHONDria WITH CYCLIC 

CHaLCONE anaLOGUES 

Miroslava Štefanišinová 1 , Mária Kožurková 2 , Vladimíra Tomečková 1 and 

Mária Mareková 1 

1 

Department of Medical Chemistry, Biochemistry and Clinical Biochemistry, 

UPJŠ - Faculty of Medicine, Tr. SNP 1, 040 01 Košice, Slovak Republic, 

2 

Department of Biochemistry, Institute of Chemistry, UPJŠ - Faculty of Science, 

Moyzesova 11, 040 66 Košice, Slovak Republic 

The binding of small molecules to DNA has been of great interest due to the understanding 

of the drug-DNA interaction. Substituted chalcone (1) and its cyclic chalcone analogues: 

indanone (2), tetralone (3), benzosuberone (4) with dimethylamino substituent in 2, 4 

position are interesting as drugs with antitumor activity. The purpose of this study was 

to investigate the interaction of DNA with new ligands (1 – 4) using by spectroscopy 

methods. Ligand - DNA binding affinities and constants (K) of the DNA-drug complexes 

were determined by UV-Vis and fluorescence spectrophotometric titration and CD spectroscopy. 

Generally, these compounds bound to DNA and show significant decrease of 

fluorescence and bathochromic shift of excitation and emission maxima compared to the 

spectral characteristics of the free form of ligands in solution phase. The strong hypochromism, 

extensive broadening, red-shifting and increased stability of DNA double helix 

were observed when these derivatives where bound to DNA. The compounds have no 

circular dichroism spectrum when are free in the solution but they induced CD spectrum 

when they are in the complex with DNA. The helical band at 246 nm corresponding to 

the DNA unwound extent exhibited decrease for all the compounds in the same order 

as that of the binding affinity. This phenomenon could be due to the stabilization of the 

right-handed B-form of DNA by intercalation. 

Acknowledgments: This work was supported by grant project VEGA 1/0402/10 


235

Posters 

111. 

ASSOCIATION POLYMORPHISMS of GST, hOGG1 aND XRCC1 GENES 

WITH LUNG aDENOCarCINOMa 

Tatiana Matáková 1 , Erika Halašová 2 , Lucia Letková 2 

Anton Dzian 3 and Dušan Dobrota 1 

1 

Institute of Medical Biochemistry, JMF CU in Martin; 2 Institute of Medical Biology, 

JMF CU in Martin; 3 Clinic of Surgery, JMF CU and MFH in Martin; Slovak Republic 

A case-control study with 130 lung cancer cases and 290 controls was conducted. DNA 

was extracted from peripheral blood leukocytes, and the polymorphisms of GSTM1, 

GSTT1, GSTP1, XRCC1 and hOGG1 enzymes were determined by PCR-based methods. 

In this study we determined the genotype distribution of all these genes, and their combinations. 

Association between specific genotypes and the development of lung cancer 

were examined using logistic regression analysis to calculate odds ratios (OR) and 95% CI. 

The GSTT1 null genotype (OR=2.2; p=0.004) was associated with an elevated risk. The 

statistically significant correlation was found also for the combined genotypes of GSTT1 

null and GSTM1 positive (OR=2.2; p=0.04) and GSTT1 null and GSTP1 Ile/Val or Val/Val 

(OR = 2.9; p = 0.009). For hOGG1 and XRCC1 were no difference in the frequency of 

genotypes between matched cases and controls. The statistically significant correlation 

was found also for the combined genotypes of GSTT1 null and GSTM1 positive (OR=2.2; 

p=0.04) and GSTT1 null and GSTP1 Ile/Val or Val/Val (OR=2.9; p=0.009). 

The statistically significant correlation was found for the combined genotypes of GSTT1 

null and hOGG1 Ser/Ser (OR=2.2; p=0.02) and GSTT1 null and XRCC1 Arg/Arg (OR=2.7; 

p=0.03). 

The genotype of selected enzymes affects the risk for lung adenocarcinoma. 

Acknowledgments: This work was supported by Ministry of Health of the Slovak republic 

under the project No. 2007/48-UK-13 and project “Center of translational medicine” 

cofinanced from EC sources and European Regional Development Fund. 


Posters 

112. 

oxidaTION of 3-aMINOBENZaNTrONE, a HUMan METaBOLITE of 

carCINOGENIC 3-NITrOBENZaNTHrONE, BY HUMan aND 

RAT CYTOCHrOMES P450 

Jana Mizerovská 1 , Helena Dračínská 1 , Volker M. Arlt 2 , Heinz H. Schmeiser 3 , 

Eva Frei 3 and Marie Stiborová 1 

1 


128 40 Prague 2, Czech Republic, 2 Section of Molecular Carcinogenesis, Institute of 

Cancer Research, Brookes Lawley Building, Sutton, Surrey SM2 5NG, United Kingdom, 

3 

German Cancer Research Center, 69 120 Heidelberg 

3-Aminobenzanthrone (3-ABA) is the main reductive metabolite of the carcinogenic 

environmental pollutant 3-nitrobenzanthrone (3-NBA). 3-ABA was detected in the urine 

samples of salt mining workers exposed to diesel emissions, indicating that exposure to 

3-NBA can be detectable. In addition, both parental 3-NBA and its metabolite, 3-ABA, 

were found to induce biotransformation enzymes such as cytochrome P450 1A1 and 

NAD(P)H:quinone oxidoreductase (NQO1) in several tissues of rats i.p. treated with 

both compounds. 

Oxidation of 3-ABA was studied using human and rat recombinant CYP enzymes. The 

most potent enzymes oxidizing 3-ABA were identified to be CYP1A1 and CYP3A. Using 

these enzymes, the kinetic parameters such as K m 

and V max 

were determined. Here we 

also investigate the induction potential of 3-NBA to induce NQO1 and CYP1A1 in rats, 

after the 3-NBA intratracheal instillation. Expression of NQO1 and CYP1A1 protein and 

its enzymatic activity were induced by 3-NBA in liver, kidney and lung tissues of rats 

treated i.t. with a single dose of 0.2 and 2 mg/kg bw of 3-NBA. 

Acknowledgments: Supported by GACR (303/09/0472, 203/09/0812 and 305/09/H008) 

and the Czech Ministry of Education (MSM 0021620808). 


237

Posters 

113. 

METaBOLIC aCTIvaTION of carCINOGENIC BENZO[a]PYrENE BY 

CYTOCHrOME P450 1A1 IS DICTaTED BY COMPOSITION of THE MIXEDfUNCTION-MONOOXYGENaSE 

SYSTEM 

Michaela Moserová 1 , Miroslav Šulc 1 , Volker M. Arlt 3 , David H. Phillips 3 , 

Eva Frei 2 and Marie Stiborová 1 

1 

Department of Biochemistry, Charles University, Albertov 2030, 128 40 Prague 2; 2 

Department of Molecular Toxicology, German Cancer Research Center, 

69 120 Heidelberg; 3 Section of Molecular Carcinogenesis, Institute of Cancer Research, 

Brookes Lawley Building, Sutton, Surrey SM2 5NG, UK 

Carcinogenic benzo(a)pyrene (BaP) was investigated for its potential to generate DNA 

adducts and to induce cytochrome P450 (CYP)and NADPH:CYP reductase (POR) enzymes 

in livers. 

BaP induces expression of these enzymes in livers of experimental models, which leads to 

increase in their enzymatic activity. In addition, this compound is capable of generating 

DNA adducts, predominantly in livers of studied organisms. The stimulation effect was 

attributed to induction of CYP1A1/2 and/or enzymes, which are responsible for oxidative 

activation of BaP to the metabolites generating major DNA adducts in vitro. BaP is 

oxidized by hepatic microsomes from pretreated and control animals to six metabolites. 

BaP is also oxidized with purified CYP1A1 reconstituted with NADPH:CYP reductase 

generating only five metabolites, but forming two DNA adducts. Activation of BaP by 

CYP1A1 is dictated by composition of the mixed-function-monooxygenase system of the 

endoplasmic reticulum. 

Acknowledgments: Supported by Czech Science Foundation (303/09/0472, 305/09/ 

H008), Grant Agency of Charles University (127208) and Ministry of Education, Youth 

and Sports (MSM 0021620808). 


Posters 

114. 

THE STUDY ON CYTOCHROME B 5 

–MEDIATED STIMULATION 

OF ELLIPTICINE OXIDATION BY CYTOCHROME P450 3A4 TO ITS 

PHarMACOLOGICALLY MORE EffICIENT METABOLITES 

Barbora Mrázová 1 , Eva Martínková 1 , Radek Indra 1 , Eva Frei 2 and Marie Stiborová 1 

1 


128 43 Prague 2, Czech Republic 

2 

German Cancer Research Center, 69 120 Heidelberg, Germany 

Ellipticine alkaloid exhibiting significant antineoplastic activities. CYP3A4 was found to be 

the major enzyme responsible for oxidation of ellipticine to 13-hydroxy- and 12-hydroxyellipticine, 

two metabolites generating DNA adducts. Many CYP3A4-dependent reactions have 

been shown to be stimulated by another microsomal protein, cytochrome b 5 

(b 5 

). Beside 

12-hydroxy- and 13-hydroxyellipticine, other three ellipticine metabolites are generated 

by CYP3A4. Human b 5 

, purified rabbit hepatic b 5 

and apo-b 5 

reconstituted with heme were 

capable of stimulating oxidation of ellipticine by CYP3A4. The patterns and amounts of 

ellipticine metabolites vary significantly when b 5 

is present in the incubations. Whereas, 

the formation of detoxication product of ellipticine oxidation (9-hydroxyellipticine) is 

practically not influenced by b 5 

, generation of 13-hydroxy- and 12-hydroxyellipticine is 

increased significantly by these b 5 

proteins. The enhanced generation of ellipticine-DNA 

adducts in the presence of b 5 

in the system was confirmed by 32 P postlabeling. Other 

proteins with or without heme in their molecules are without such effects on ellipticine 

oxidation. The hyperbolic kinetics of ellipticine oxidation by CYP3A4 with or without b 5 

indicates no cooperativity. The results indicate that the presence of heme in b 5 

seems to 

be required for the stimulation of CYP3A4-mediated oxidation of ellipticine. 

Acknowledgments: Supported by GACR (P303/10/0356, 203/09/0812), the Czech Ministry 

of Education (MSM0021620808, 1M0505) and GAUK (1272080). 


239

Posters 

115. 

IMMUNODETECTION OF 11SS-HYDROXYSTEROID DEHYDROGENASE 

DURING PURIFICATION OF a NEW HUMAN MEMBraNE-BOUND 

CarBONYL REDUCING ENZYME 

Miloslava Netopilová, Libuše Černá, Lucie Škarydová and Vladimír Wsol 

Department of Biochemical Sciences, Faculty of Pharmacy, Charles University, 

Hradec Králové , Czech Republic 

The 11ß-hydroxysteroid dehydrogenase type 1 (11ßHSD1) is a membrane-bound microsomal 

protein that mainly catalyzes the conversion of inactive glucocorticoid cortisone 

to active cortisol. However, this enzyme has a broad substrate specificity and can also 

metabolize various xenobiotics with carbonyl groups, for example the cytostatic drug 

oracin. Oracin is reduced by microsomal enzymes to two enantiomers of 11-dihydrooracin 

(DHO). Stereospecificity of the conversion depends on type of tissues and enzymes. 

While human liver microsomes reduce oracin to (-)-DHO and (+)-DHO in a ratio of 60:40, 

purified human liver 11ßHSD1 change oracin more specifically to (-)-DHO (76%). Because 

11ßHSD1 is only known microsomal enzyme reducing oracin, it is supposed that the liver 

microsomes contain at least one other oracin carbonyl reducing enzyme with stereospecific 

production of (+)-DHO. The presence of 11ßHSD1 was determinated by immunodetection 

using Western blotting. The blot was incubated with a primary polyclonal rabbit antibody 

against human 11ßHSD1(1:1000, Abcam) and then with a secondary polyclonal swine 

antibody anti-rabbit IgG coupled to horseradish peroxidase (1:1000, Dako). About 30 

fractions were prepared using separation of human microsomes on Q-Sepharose. The 

highest reducing activity was found in the flow through fractions Q2-Q4 and in the fractions 

Q10-Q14; 11ßHSD1 was detected only in fraction Q11-Q14. The selected fraction 

Q12 was separated on the Phenyl-Sepharose column and the highest activity was in the 

fractions P10-P12. In these fractions, 11ßHSD1 was not found. 

Acknowledgement: This project was supported by the Grant Agency of Charles University, 

Grant No. 45508/C/2008. 


Posters 

116. 

PHYTOREMEDIATION – THE PROMISING METHOD FOR THE REMOvaL OF 

PHarMACEUTICAL RESIDUES frOM WASTEWATER 

Radka Podlipná a , Petra Šídlová b , Kotyza Jan c and Tomáš Vaněk a 

a 

Laboratory of Plant Biotechnologies, Joint Laboratory of the Institute of Experimental 

Botany, Academy of Science of the Czech Republic and 

Research Institute of Crop Production, Prague 

b 

Department of Biochemical Sciences, Faculty of Pharmacy UK, Hradec Králové 

c 

Faculty of Environmental Technology, Institute of Chemical Technology, Prague 

Recently there is an increasing concern about contamination of the environment with 

pharmaceutical residues and their metabolites. These contaminants are excreted into 

a hospital or municipal wastewater in considerable amount; in addition many of them 

are not completely degraded in sewage treatment plants and remains in water effluent. 

Although the environmental concentrations of these compounds are usually on low level 

(

Posters 

117. 

ELLIPTICINE CYTOTOXICITY TO HUMan THYrOID caNCEr CELL LINES 

Jitka Poljaková 1 , Tomáš Eckschlager 2 , Eva Frei 1 and Marie Stiborová 1 

1 


2 

Department of Pediatric Hematology and Oncology, 

2 nd Medical School, Charles University in Prague 

Ellipticines are plant alkaloids with an antineoplastic activity. The mode of action is based 

mainly on DNA intercalation, inhibition of topoisomerase II and formation of covalent 

DNA adducts mediated by cytochromes P450 (CYP) and peroxidases. Such ellipticine- 

DNA-adducts are formed in vitro, in human breast adenocarcinoma MCF-7, leukemia 

HL-60, CCRF-CEM, neuroblastoma IMR-32, UKF-NB-3, UKF-NB-4 cell lines and in vivo in 

rats exposed to ellipticine. Here, the cytotoxicity of ellipticine to human thyroid cancer 

cell lines BHT-101, B-CPAP and 8505-C was investigated. Furthermore, the effect of hypoxic 

conditions on the ellipticine toxicity to these cells was also examined. The toxicity 

of ellipticine to thyroid cancer cells cultivated under the standard conditions was higher 

than that to the cell lines grown under hypoxia (1% oxygen). Another target of this work 

was to evaluate the effect of ellipticine on expression of enzymes activating ellipticine, 

namely on CYP1A1, 1B1, 3A4, cytochrome b 5 

and peroxidases COX-1 and TPO. 

Acknowledgement: Supported by GACR (P301/10/0356) and Czech Ministry of Education 

(MSM0021620813). 


Posters 

118. 

QUALITY AND TIME-STABILITY OF HUMAN SKIN EXPLANTS 

Alena Rajnochová Svobodová 1 , Adéla Zdařilová 1 , Dana Kylarová 2 , 

Bohumil Zálešák 3 and Jitka Vostálová 1 

1 

Department of Medical Chemistry and Biochemistry, Palacký University Olomouc, 

2 

Department of Histology and Embryology, Palacký University Olomouc, 

3 

Department of Plastic and Aesthetic Surgery, Faculty Hospital, Olomouc 

Human skin obtained from plastic surgery has been used for isolation of skin cells such 

as keratinocytes, fibroblasts, melanocytes or endothelial cells for several decades. Single 

cell cultures, co-cultures or 3D cultures of these cells have been utilized for dermatological 

studies including phototoxicity and photoprotection investigation. However, these 

very simple systems are very different from human skin. Therefore cultivation of human 

skin organ culture (ex vivo skin) has been developed as a more suitable model system. 

The aim of this study was to determine explant usage period (quality and stability 

during cultivation). Morphological structure and antioxidant parameters (activity of 

superoxide dismutase (SOD), glutathione peroxidase (GPX), glutathione S-transferase 

(GST) and catalase (CAT) and level of thiobarbituric acid-reactive substances (TBARS) 

and glutathione (GSH) were evaluated. Epidermis did not show any significant changes 

in its morphology until 72 h cultivation. In dermis moderate acute inflammatory reaction 

was observed (48 - 72 h) but it disappeared after 120 h. Dermal papillae started to be 

reduced after 96 h and these changes became more significant after 120 h cultivation. 

TBARS level together with GSH amount and activity of SOD have markedly increased 

for the cultivation period (0 - 144 h). Activities of GPX and CAT have grown for 72 - 96 h 

and then decreased. Changes in GST activity did not have any trend and varied between 

70 - 160 % of control (0 h). 

In conclusion human skin explants can be utilized for various dermatological applications 

including photoprotection or metabolic and xenobiochemic studies. 

Acknowledgement: This work was financially supported by the grant MSM 6198959216. 


243

Posters 

119. 

OXIDATIVE STRESS IN TURKEYS CAUSED BY CHRONIC CADMIUM 

EXPOSURE 

Anna Sobeková 1 , Katarína Holovská 2 and Peter Javorský 3 

1 

Department of Chemistry, Biochemistry and Biophysics UVLF in Košice, 

2 

Department of Anatomy, Histology and Physiology UVLF in Košice, 

3 

Institute of Animal Physiology SAV Košice 

Heavy metals can cause oxidative stress by increasing the formation of reactive oxygen 

species (ROS), which render antioxidants incapable of defence against growing amounts 

of free radicals. The alterations of ROS scavenging enzyme activities are the consequences 

of this effect. The effect of single Cd and Cd+Zn on the antioxidant enzymes 

(SOD-superoxide dismutase, GPx-glutathione peroxidise, GR-glutathione reductase, 

GST-glutathione-S-transferase) and the content of thiobarbituric acid reactive species 

(TBARS) was evaluated in parenchymal tissues (liver, kidney) from turkeys. 

In the liver, the specific activity of SOD was significantly increased in Cd group. Mitochondria 

try to decrease intracellular ROS levels by decreasing to consume of oxygen via formation 

of „extremely swollen mitochondria“. Ultrastructural changes were confirmed by 

electron microscopy. TBARS values were not changed in Cd group. Antioxidant defence 

system were able to protect the hepatocytes against oxidative damage. 

Activity of SOD was significantly decreased in the kidney. Cadmium evoked the increase 

of activity of another determined antioxidant enzymes (GSHPx, GR, GST). Higher TBARS 

values indicate the oxidative stress in this organ. Zinc co-administration shows protective 

effect in both observed tissues. 

The alterations in the activities of antioxidant defence system and increased TBARS 

values showed that toxic effect of cadmium is not only in the high thiol group affinity, 

but also in ROS production. 


Posters 

120. 

SIMILarITY BETWEEN raT AND HUMAN ENZYMES INVOLVED IN 

OXIDATION 2-NITROPHENOL, a METABOLITE OF CarCINOGENIC 

2-NITROANISOLE 

Martina Svobodová, Markéta Martínková, Helena Dračínská and Marie Stiborová 

Department of Biochemistry, Faculty of Science, Charles University in Prague 

2-Nitrophenol (2-NP) is the main detoxication metabolite of 2-nitroanisole (2-NA) that 

is an important industrial pollutant and a potent carcinogen for rodents. The aim of this 

study was to investigate the efficiency of rat and human hepatic cytochromes P450 (CYPs) 

to metabolize 2-NP, to determine the metabolites formed during such a metabolism and 

to compare the efficiencies of rat CYPs with those of human. 2-NP is oxidized by both 

liver microsomes to one metabolite, 2,5-dihydroxynitrobenzene (2,5-DNB). To define 

the role of rat CYPs in 2-NP oxidation we investigated the modulation of this reaction 

by specific inducers and selective inhibitors of these enzymes. Using microsomes from 

Baculovirus transfected insect cells expressing recombinant rat and human CYP enzymes, 

we found that rat recombinant CYP2E1, 2C11, 2B1, 1A2, 1A1 and 3A4 were the most 

effective to oxidize 2-NP. Similarly, human CYP2E1, followed by CYP2A6, 2C6, 3A4 and 

2D6 were the most efficient to oxidize 2-NP. In addition, kinetics of oxidation of 2-NP by 

the most efficient enzyme catalyzing this reaction, CYP2E1, was investigated. The present 

study shows the similarity between human and rat hepatic enzymes metabolizing 

2-NP to 2,5-DNB and indicate that rat might serve as a suitable model to mimic the fate 

of 2-NP in human. 

Acknowledgements: Supported by GACR (303/09/0472) and the Czech Ministry of 

Education (MSM 0021620808). 


245

Posters 

121. 

OVEREXPRESSION OF P-GLYCOPROTEIN IN L1210/VCR CELLS IS 

ASSOCIATED WITH CHANGES IN SEVEraL ENDOPLASMIC RETICULUM 

PROTEINS 

Mário Šereš, Eva Poláková, Oľga Križanová, Zdena Sulová and Albert Breier 

Institute of Molecular Physiology and Genetics SAS Bratislava 

Multidrug resistant cells R, which express membrane drug efflux transporter – P-glycoprotein 

(P-gp, a member of ABC transporter family), was found to be also less sensitive to thapsigargin, 

inhibitor of sarco(endo)plasmic reticulum calcium pump (SERCA), in contrast 

to parental drug-sensitive cells L1210 (S). R cells was obtained from S cells by selection 

with vincristine (VCR). We have studied differences among S and R cells in expression 

of endoplasmic reticulum proteins involved in the regulation of calcium homeostasis 

and calcium-dependent processes. Amounts of mRNA encoding RyR and IP 3 

-receptor 

channels were found to be at similar in S and R cells. However, mRNAs encoding IP 3 

R1 

or 2 were decreased in resistant cells cultivated in the presence of VCR, while mRNA 

encoding RyR remained unchanged. The amount of mRNA encoding SERCA2 was lower 

in R cells as in S cells. This decrease was more pronounced when R cells were cultivated 

in the presence of VCR. Calnexin was believed to be active in P-gp maturation, but surprisingly, 

calnexin was found to be less expressed at the protein level in R as in S cells. 

We have also studied thapsigargin effect on expression of some endoplasmic reticulum 

proteins and P-glycoprotein. P-gp expression was decreased in presence of thapsigargin. 

Thus, S and R cells differ in the expression of endoplasmic reticulum proteins involved 

in the control of intracellular calcium homeostasis or calcium-dependent processes. 

Acknowledgement: This work was supported by: VVCE-0064-07, APVV-0084-07 and 

VEGA 2/0123/10, 2/0155/09. 


Posters 

122. 

EnvirONMENTal POLLUTaNTS as faCTOrs MODULaTING THE 

INflaMMaTOry rESPONSE aND fUNCTIONS of LUNG CELLS 

Lenka Umannová 1,2 , Miroslav Machala 2 , Alois Kozubík 1 and Jan Vondráček 1,2 

1 

Laboratory of Cytokinetics, Institute of Biophysics, ASCR v.v.i., 612 65 Brno, 


2 

Veterinary Research Institute v.v.i., 621 00 Brno, Czech Republic 

Chronic inflammation plays a central role in pathogenesis of lung cancer or chronic obstructive 

pulmonary disease, which belong among leading causes of death. Air pollution 

represents a significant risk factor for lung cancer incidence. The polluted atmosphere 

contains various matrices with carcinogenic potential including vehicle exhaust, side-stream 

tobacco smoke, coal combustion effluents and transient metals. In the present study, we 

analyzed mutual deregulation of enzymes involved both in xenobiotic metabolism and 

inflammatory responses, caused by environmental pollutant and important tobacco smoke 

constituent benzo[a]pyrene (BaP) and TNF-a in rat lung RLE-6TN alveolar type II cell line. 

Our results show that BaP modulates the expression/activity of enzymes induced by 

TNF-a that are involved in inflammatory responses such as prostaglandin-endoperoxide 

synthase 2 (COX-2) or inducible nitric oxide synthase (iNOS), leading to enhanced release 

of their products such as prostaglandin E2. Moreover, BaP increases expression of inflammatory 

cytokines induced by TNF-a in target cells. Important role in increased expression 

may play oxidative stress and activation of p38 mitogen activated protein kinases. These 

results demonstrate that environmental pollutants affect signaling pathways related to 

inflammatory response and may thus contribute to deregulated inflammation and lung 

epithelial damage. 

Acknowledgement: Supported by the Czech Ministry of Agriculture, grant No. MZe 

0002716202. 


247

Posters 

123. 

THE MECHANISM OF CYTOTOXIC EffECT OF NOVEL ACRIDINE 

INTERCALATORS 

Zuzana Vantová 1 , Helena Paulíková 1 , Mária Kožurková 1 , Danica Sabolová 1 , 

Ima Dovinová 1 , Pavol Kristián 2 , Ján Imrich 2 , Ján Ungvarský 2 and Ladislav Janovec 2 

1 

Department of Biochemistry and Microbiology, STU Bratislava, 

2 

Department of Biochemistry, UPJŠ Košice 

Acridines are well-known chemotherapeutic agents in cancer therapy. New class of acridine 

derivatives were synthesized – 2´,2´´-[(acridin-3,6-diyl)]-3´,3´´-dialkyl-1,3-diimidazolidin-4- 

on hydrochloride (AcrDIMs). These compounds bearing imidazolidinone rings, are capable 

to interact with DNA, mainly by intercalation and to inhibit activity of topoisomerase I. 

Their biological activity to inhibit cell proliferation of leukaemia cells (HL-60 a L1210 cells) 

and adherent ovarian cancer cell line A2780 was confirmed. Despite the lowest affinity 

to DNA (K = 1.9×10 5 M -1 ), 2´,2´´-[(acridin-3,6-diyl)]-3´,3´´-dihexyl-1,3-diimidazolidin-4-on 

was the most active derivative with an IC 50 

of 4.61±2.08 µM on a HL 60 cell line, with IC 50 

of 5.52±1.08 µM on a L1210 cell line and with IC 50 

of 30,83 ± 2,63 µM on a A-2780 cells 

after 48h of incubation.. It has been discovered that the cytotoxicity of AcrDIMs correlates 

with their hydrophobicity (uptake into cells was monitored). Although apoptosis 

of treated cells has not been confirmed it has been shown that dihexyl-AcrDIM arrested 

cell cycle, and HL-60 cells were accumulated in G 1 

phase after 48 h incubation. Together, 

our results support the hypothesis that AcrDIMs exert cytostatic properties by interaction 

with DNA inducing arrest of cell cycle and oxidative stress in cancer cells. 

Acknowledgement: This work has been supported by VEGA grant 1/0097/10 and 1/0053/08. 


Posters 

124. 

RETINOIC ACID-INDUCED DIffERENTIATION MODULATES THE 

APOPTOTIC EffECT OF SODIUM vaLPROATE IN HL-60 CELLS 

Jiří Vrba and Jitka Ulrichová 

Department of medical chemistry and biochemistry UP Olomouc 

Differentiation of myeloid leukemia cells results in resistance to various apoptotic stimuli. 

We examined whether human leukemia HL-60 cells differentiating by all-trans retinoic 

acid (ATRA) into neutrophil-like phenotype became resistant to the apoptogenic activity 

of valproate, an inhibitor of histone deacetylases. In undifferentiated HL-60 cells, valproate 

induced the G0/G1 cell cycle arrest and apoptotic changes including dissipation 

of the inner mitochondrial membrane potential, loss of plasma membrane asymmetry 

and/or integrity, activation of caspase-9 and caspase-3, and appearance of sub-G1 DNA. 

Cotreatment with ATRA enhanced the apoptotic response of undifferentiated cells to 

valproate. On the other hand, in HL-60 cells pretreated for 72 h with ATRA, valproate in 

combination with ATRA induced lower dissipation of mitochondrial membrane potential 

and weaker apoptotic changes in the plasma membrane while the DNA fragmentation 

was not attenuated in comparison with undifferentiated cells. We conclude that ATRAinduced 

differentiation modulates but does not abrogate the apoptotic response of 

HL-60 cells to sodium valproate, and nuclei are preferentially affected during apoptosis 

of differentiated cells. 

Acknowledgements: This research was supported by the Ministry of Education, Youth 

and Sports of the Czech Republic (MSM 6198959216). 


249


LIST of AUTHORS 

Names Pages E-mail address 

Adameová A. 111 

Adamkov M. 119, 182 

Adamík P. 45, 117 

Al Alami H. 123, 133 hendalalami@hotmail.com 

Ambro Ľ. 194 lubos.ambro@savba.sk 

Andrezálová L. 214 lucia.andrezalova@fmed.uniba.sk 

Antalík M. 55, 71, 129, 131, 199 

Antalíková J. 168, 173 Jana.Antalikova@savba.sk 

Antalová Ľ. 

Antošová A. 57, 211 

Arlt V.M. 233, 237, 238 

Arroyo J. 205 

Babušíková E. 41, 64, 101, 117 babusikova@jfmed.uniba.sk 

Babuška V. 186 

Bágeľová J. 199 bagelova@saske.sk 

Balažová A. 124, 125 balazova@fpharm.uniba.sk 

Balážová M. 36 simockova@gmail.com 

Bálentová S. 182 balentova@jfmed.uniba.sk 

Bánovčin P. 64 

Barák I. 42, 72 imrich.barak@savba.sk 

Barančík M. 43 Miroslav.Barancik@savba.sk 

Baráth M. 44 chemmbar@savba.sk 

Baráth P. 152 

Baráthová M. 153 

Barta A. 87 

Barták F. 97 

Bártíková H. 225, 232 hana.bartikova@faf.cuni.cz 

Bauer J.A. 194 jacob.bauer@savba.sk 

Bauerová-Hlinková V. 196 

Beláňová M. 195 belanova@fns.uniba.sk 

Benedikovičová K. 140 

Benej M. 46 martin.benej@gmail.com 

Beneš J. 97 

Benková K. 187 

Benkovičová V. 60 

Benson V. 47 

Bernard V. 61 

Bezáková L. 201 


251

List of Authors 

Bezouška K. 47, 83, 135 bezouska@biomed.cas.cz 

Bialešová L. 172 

Biely P. 37, 107 chempbsa@savba.sk 

Bíliková K. 48, 127 katarina.bilikova@savba.sk 

Bilka F. 124, 125, 201 bilka@fpharm.uniba.sk 

Bílková A. 125, 201 

Bílková Z. 93 

Bittšanský M. 45, 54 michal@bittsansky.net 

Blagová E. 194 

Bláha J. 135 

Blanáriková V. 124, 125 

Blanco N. 205 

Blaško J. 120 

Blaškovič D. 209 

Bobalová J. 96 

Borko Ľ. 196 borko.l.jr@gmail.com 

Borovanský J. 49 jborov@lf1.cuni.cz 

Bortlíček Z. 50 

Bouchal P. 50 bouchal@mou.cz 

Boušová I. 226 Iva.Bousova@faf.cuni.cz 

Božek P. 175 

Brechtlová M. 116 

Breier A. 51, 165, 166, 227, 246 breier@up.upsav.sk 

Brennan P.J. 197 

Breza J. 184 

Brodská B. 158 

Budinská E. 50 

Buchal R. 89 

Bullogh P. 72 

Burdová K. 231 

Bystrický M. 195 

Capková Ľ. 76, 183 capkova@saske.sk 

Cagala M. 169 martin.cagala@savba.sk 

Cehlar O. 95 

Celadová P. 135 

Celec S. 115 

Crha I. 144 

Csáderová L. 73, 169 

Čačányiová S. 215 

Čalkovská A. 218 

Čechová L. 220 

Čermák J. 187 



Černá L. 240 

Čeřovská N. 207 

Černý R. 52, 186 Radim.Cerny@lfp.cuni.cz 

Čierny D. 115 

Danchenko M. 58 

Dávidová A. 76, 183 

Daxnerová Z. 211 

Demnerová K. 63 

Dianišková P. 197 dianiskova@fns.uniba.sk 

Divoký V. 88 

Ditte P. 166 

Dobrá J. 198 

Dobrota D. 

41, 45, 54, 64, 91, 93, 101, 119, 141, 143, 

dobrota@jfmed.uniba.sk 

185, 190, 217, 236 

Doubnerová V. 198, 207 doubnerova@volny.cz 

Douděrová D. 38 dana.u@email.cz 

Dovinová I. 215, 248 ima.dovinova.@savba.sk 

Dračínská H. 237, 245 

Drahovská H. 123, 133, 140 drahovska@fns.uniba.sk 

Drgová A. 141, 218 

Dršata J. 226 

Ďuračková Z. 53, 84, 184, 188, 214, 219 

Ďurfinová M. 116 monika.durfinova@fmed.uniba.sk 

Dušenka R. 94 

Dvorsky R. 95 

Dvořáková M. 184 monika.dvorakova@gmail.com 

Dzian A. 59, 236 

Eckertová M. 112, 147 

Eckschlager T. 242 

Evinová A. 75, 117 peregrinova@jfmed.uniba.sk 

Fabianová E. 59 

Faltinová A. 55 andrea.faltinova@savba.sk 

Farkaš V. 56, 205 chemvfar@savba.sk 

Farkašovská J. 139 jarmila.farkasovska@savba.sk 

Fecková Ľ. 69, 155, 162 

Fedoročko P. 159 

Fedunova D. 199 fedunova@saske.sk 

Ferko M. 111 

Firbasová J. 225 

Fišerová A. 47 

Flachbartova Z. 95, 199 

Flores Ramirez G. 93 virugafl@savba.sk 


253


Forejt J. 80 

Frei E. 98, 229, 233, 237, 238, 239, 242 

Gaburjáková J. 55 

Gajdošechová L. 112, 147 lgajdosechova@azet.sk 

Gajdošová J. 123, 140 gajdosova7@gmail.com 

Gáll J. 89 

Garbis S.D. 50 

García-Nafría J. 194 

Gašperík J. 132, 196 

Gažová Z. 57, 199, 211 gazova@saske.sk 

Gdovinová A. 210 

Gibalová L. 51, 227 lenka.gibalova@savba.sk 

Gnipová A. 60 

Godány A. 139, 212 

Grebeňová D. 148, 160 grebenova@uhkt.cz 

Griač P. 36, 90 Peter.Griac@savba.sk 

Grobárová V. 47 

Grófik M. 117 

Grones J. 200 grones@fns.uniba.sk 

Grones P. 200 

Guerin M.E. 100 

Guzy J. 78 

Hajduk M. 55, 58 hajduch@savba.sk 

Hajtmanová E. 182 

Halašová E. 141, 236 

Hamuľáková S. 203 

Hanušová V. 228 Veronika.Hanusova@faf.cuni.cz 

Hapala I. 170 

Harnišová E. 172 

Hatok J. 59, 91, 143, 185, 190 hatok@jfmed.uniba.sk 

Hernychová L. 50 

Hirsch J. 44 

Hodek P. 231 

Hofmanová J. 150, 151 

Holas O. 203 

Holič R. 90 

Holková I. 124, 125 

Hollý P. 59, 

Holoubek A. 158 

Holovská K. 244 

Holubec L. 186 

Holý A. 79 



Homerová D. 149, 163 dagmar.homerova@savba.sk 

Horovská Ľ. 168, 173 

Horváth A. 60 horvath@fns.uniba.sk 

Hosoyamada M. 189 

Hostinová E. 196 

Hrabovská A. 61, 179, 180 hrabovska@fpharm.uniba.sk 

Hrkal Z. 148, 160 

Hronská L. 170 

Hrstka R. 50 

Hudecová S. 169 

Hudeček J. 62 hudecek@natur.cuni.cz 

Hulíková K. 47 

Huľo E. 59 

Husárová V. 45 

Hýžďalová M. 151 

Cholujová D. 169 

Chomová M. 75, 91, 118 chomova@jfmed.uniba.sk 

Chorvát Jr. D. 156 

Chotár M. 212 

Chudej J. 91 

Chudý P. 59, 91 

Chvojka J. 97 

Ichida K. 189 

Ilovská V. 108 

Imrich J. 203, 248 

Indra R. 239 

Indrák K. 88 

Ivanová M. 43 

Jackson M. 43, 100, 171, 195, 197 

Jamroškovič J. 42 

Jankovičová J. 168 

Janočková J. 203 

Janovec L. 159, 203, 248 

Jansa P. 220 

Janů P. 63 janup@vscht.cz 

Javorský P. 234, 244 

Jelínková I. 150 

Jeseňák M. 64 jesenak@gmail.com 

Ježová D. 112 

Jurečeková J. 59, 91, 185, 190 jurecekova@jfmed.uniba.sk 

Kaclíková E. 140 

Kačáni L. 65 laco.kacani@cemit.at 


255


Kádek A. 47 

Kajsík M. 123 

Kaliňák M. 66 michal.kalinak@stuba.sk 

Kamodyová N. 140 

Kandráč L. 129 

Kandričáková M. 126 kandricakovam@gmail.com 

Kaplán P. 75, 101, 115, 119, 217 

Karmažínová M. 67, 74 maria.drigelova@savba.sk 

Kaur D. 197 

Kavan D. 47, 83, 135 

Kavcová H. 165 

Kärner E. 52 

Kerná V. 45 

Kinclová I. 182 

Kiňová Sepová H. 201 kinovasepova@fpharm.uniba.sk 

Kliment J. 94 

Kloudová K. 207 

Klubicová K. 58 

Kľučár Ľ. 137 

Kočí L. 151 koci@ibp.cz 

Koháryová M. 202 koharyova@fns.uniba.sk 

Kohút P. 170 peter.kohut@savba.sk 

Koláček M. 216 marian.kolacek@fmed.uniba.sk 

Kollárik M. 99 

Kollárová M. 202 

Kollárovič G. 152 gabriel.kollarovic@savba.sk 

Kominková V. 103 

Koneracká M. 57 

Kontseková S. 153 virusona@savba.sk 

Kopáček J. 68, 169 virukopa@savba.sk 

Kopčanský P. 57 

Korduláková J. 100, 171, 195, 197 kordulakova@fns.uniba.sk 

Kormanec J. 69, 149, 155, 162, 163 jan.kormanec@savba.sk 

Kostecká Z. 70 kostecka1@azet.sk 

Kotrbová V. 229 verakotrbova@centrum.cz 

Kotyza J. 241 

Kovacech B. 95 

Kovaľ J. 159 

Kovalská M. 75, 94, 115, 119, 121 kovalskaM@post.sk 

Kovácsová M. 87 

Kozubík A. 142, 150, 151, 161, 247 

Kožurková M. 71, 159, 203, 208, 235, 248 maria.kozurkova@upjs.sk 



Krajčíková D. 72 daniela.krajcikova@savba.sk 

Krajňáková L. 208 

Kraková T. 127 tatiana.krakova@savba.sk 

Králíková M. 144 

Králová V. 228, 230 kralovav@lfhk.cuni.cz 

Krejci E. 61 

Kretová M. 152, 154 miroslava.kretova@savba.sk 

Kristek F. 215 

Kristián P. 203, 248 

Križanová O. 73, 169, 246 olga.krizanova@savba.sk 

Kron I. 221 

Kroužecký A. 97 

Kryštofová S. 206 

Kršková K. 112, 147 katarina.krskova@savba.sk 

Křen V. 47, 83 

Křenek K. 47 

Křepela E. 187 

Křížková J. 231 jitus.k@atlas.cz 

Kubíček V. 225, 232 

Kuča K. 203 

Kučerová J. 88 

Kucháriková A. 76, 183 

Kucharská J. 108, 175 

Kuka S. 101, 217 kuka@jfmed.uniba.sk 

Kukumberg P. 116 

Kulda V. 186 Vlastimil.Kulda@lfp.cuni.cz 

Kuncová J. 97 

Kupčová V. 216 

Kuračka Ľ. 116 

Kurča E. 81, 115, 117 

Kurucová T. 165, 166 tatiana.kurucova@savba.sk 

Kušnír J. 204 

Kutaš P. 69, 155, 162 peter.kutas@gmail.com 

Kutejová E. 194 eva.kutejova@savba.sk 

Kutschy P. 211 

Kuželová K. 148, 160 

Kvetňanský R. 73 

Kylarová D. 243 

Labienec M. 176 

Labudová M. 227 

Lacinová Ľ. 67, 74 lubica.lacinova@savba.sk 

Lajdová I. 156 ingrid.lajdova@szu.sk 


257


Lakatoš B. 172 boris.lakatos@stuba.sk 

Lamka J. 225, 232 

Lasotová T. 232 

Lauková M. 73 

Lehotský J. 75, 101, 115, 117, 119, 121, 217, lehotsky@jfmed.uniba.sk 

Lenčešová Ľ. 73, 169 

Letková L. 141, 143, 236 letkova@jfmed.uniba.sk 

Levarski Z. 130 

Levdikov V.M. 194 

Levová K. 233 katkalev@gmail.com 

Lievajová K. 120 

Lichá L. 152 

Lichardusová L. 204 lucialichardusova@gmail.com 

Lincová E. 142 lincova@ibp.cz 

Lintnerová J. 133 

Lipšová A. 49 

Liptaj T. 66 

Liška V. 186 

Líška B. 116 

Lohajová Ľ. 234 lohajova@uvlf.sk 

Lovecká P. 63 

Luciaková K. 152, 154 

Lukáčová N. 76, 183 lukacova@saske.sk 

Lukeš J. 60 

Macková M. 63 

Madacki J. 171 

Madyagol M. 128 ing.mahesh@gmail.com 

Mahmood S. 143 mahmood@jfmed.uniba.sk 

Machala M. 161, 247 

Májek P. 77 majek@adinis.sk 

Máleková Ľ. 103 

Manglová D. 83 

Mareková M. 78, 204, 235 maria.marekova@upjs.sk 

Martásek P. 38 

Martínková E. 239 

Martínková M. 245 

Martončíková M. 120 martoncikova@saske.sk 

Mašín J. 82 

Mašlanková J. 78 

Matáková T. 59, 94, 141, 143, 236 matakova@jfmed.uniba.sk 

Matějovič M. 97 

Matejovičová M. 144 matejovic@med.muni.cz 



Matoušová M. 145 matousova@uochb.cas.cz 

Mazáň M. 205 marian.mazan @savba.sk 

Mellová Y. 182 

Melounová J. 144 

Mertlíková-Kaiserová H. 79, 145, 220 kaiserova@uochb.cas.cz 

Messingerová L. 165 lucia.messingerova@savba.sk 

Miedzińska L. 198 

Mihola O. 80 mihola@img.cas.cz 

Michalík J. 81 michalik@mfn.sk 

Michalková K. 168, 173 Katarina.Fabryova@savba.sk 

Mikeš J. 159 

Mikušová K. 100, 171, 195, 197 mikusova@fns.uniba.sk 

Minárik G. 77 

Mislovicová D. 165 

Mizerovská J. 237 janna.mizerovska@seznam.cz 

Mokrá D. 218 mokrá@jfmed.uniba.sk 

Moravčíková E. 187 moravcikerika@yahoo.com 

Morová J. 82 jana.m@volny.cz 

Moserová M. 238 moserova@seznam.cz 

Mrázek H. 83 Hynek.mrazek@biomed.cas.cz 

Mrázová B. 229, 239 barunka.mrazova@seznam.cz 

Mrvová K. 179 mrvova.kaja@gmail.com 

Mudrochová M. 50 

Muchová J. 84, 184, 188, 214, 216, 219 jana. muchova@fmed.uniba.sk 

Muchová K. 42 

Mujkošová J. 111 

Müller P. 50 

Mullerová D. 72 

Nagyová Z. 84, 188 

Nalivaeva N.N. 41 

Nenutil R. 50 

Netopilová M. 240 netopilova@faf.cuni.cz 

Neubauer O. 109 

Neuschlová D. 180 dominika.neuschlova@gmail.com 

Nižnanský Ľ. 206 lubosniznansky@gmail.com 

Novák I. 97, 162 

Novák M. 95 

Novaková R. 69, 155 

Nováková Z. 96, 177 

Odnogová Z. 200 

Okša A. 156 

Ondrejka I. 45, 115, 117 


259


Ondrejovičová I. 84, 188 iveta.ondrejovicova@fmed.uniba.sk 

Ondriaš K. 103 

Ondrovičová G. 194 gabriela.ondrovicova@savba.sk 

Ondrušová Ľ. 157 lubica.ondrusova@gmail.com 

Orendáčová J. 120 

Oros R. 85 Roman.Oros@shimadzu.eu.com 

Országhová Z. 53, 214, 219 zuzana.orszaghova@fmed.uniba.sk 

Osička R. 82 

Otevřelová P. 158 Petra.Otevrelova@uhkt.cz 

Otmar M. 145 

Paduchová Z. 188, 219 

Pakanová Z. 215 

Pakostová A. 228 

Pálffy R. 132 

Pánčiová L. 130, 134 lucia.panciova@gmail.com 

Paris Z. 60 

Parnica J. 129 jozef.parnica@gmail.com 

Parohová J. 87 

Pastorek J. 68, 153, 166 

Pastoreková S. 68, 169 

Paulíková H. 86, 215, 248 helena.paulikova@stuba.sk 

Pavlíková M. 75, 94, 119, 121 Martina.Pavlikova@jfmed.uniba.sk 

Pavelka S. 222 

Pavlendová N. 42 

Pecháňová O. 87, 215 olga.pechanova@savba.sk 

Perez-Reyes E. 67 

Pernicová Z. 142 

Pešta M. 186 

Pevala V. 194 vladimir.pevala@savba.sk 

Phillips D.H. 233, 238 

Piterková L. 88 lucie.piterkova@upol.cz 

Plameňová I. 190 

Pláteník J. 89 jan.platenik@lf1.cuni.cz 

Plšíková J. 159, 203, 208 janaplsikova@gmail.com 

Pluskalová M. 148, 160 pluskalova@uhkt.cz 

Podhradský D. 71 

Podlipná R. 241 podlipna@ueb.cas.cz 

Pohanka M. 203 

Poláček H. 45 

Polák Š. 111 

Poláková E. 166, 246 

Polčicová K. 153 



Poleková L. 51 

Poljaková J. 242 Jitka.Poljakova@seznam.cz 

Poloncová K. 90 katarinapoloncova@hotmail.com 

Pompach P. 135 

Poturnajová M. 46 

Preťová A. 58 

Procházka J. 187 

Procházková E. 220 prochazkova@uochb.cas.cz 

Procházková J. 161 iris@ibp.cz 

Procházková Ľ. 116 

Průchová Z. 226 

Puchart V. 107 

Pullmann st. R. 218 

Qiang W. 72 

Racek J. 92 

Račay P. 59, 75, 91, 101, 118, 185, 190, 217 racay@jfmed.uniba.sk 

Račeková E. 120 

Rajdl D. 92 rajdl@fnplzen.cz 

Rajnochová Svobodová A. 223, 243 alf.svoboda@seznam.cz 

Rahydov N. 58 

Rauchová H. 174, 222 rauchova@biomed.cas.cz 

Ravingerová T. 111 

Reháková A. 69, 155, 162, 227 alenka.rehakova@gmail.com 

Repič A. 153 

Rogozanová K. 165, 166 

Roumeliotis T. 50 

Rozbeský D. 47 

Ru F. 99 

Rudolf E. 230 

Ryšlavá H. 198, 207 rysl@natur.cuni.cz 

Řehoř L. 186 

Řežuchová B. 149, 163 

Sabolová D. 159, 203, 208, 248 danica.sabolova@upjs.sk 

Sedláčková E. 89 

Sedlák J. 166, 227 

Sevcik J. 95 

Schmeiser H.H. 233, 237 

Schröterová L. 228 

Schubertová Aradská J. 209 jana.aradska@gmail.com 

Simon M. 168, 173 

Sivoňová M. 94, 121, 143 sivonova@jfmed.uniba.sk 

Skálová L. 225, 228, 232 skaloval@faf.cuni.cz 


261


Slavíčková E. 142 

Sobeková A. 234, 244 sobekova@uvlf.sk 

Souček K. 142 

Soukup T. 174, 222 

Sova P. 150 

Sprinzl M. 39 mathias.sprinzl@uni-bayreuth.de 

Spustová V. 156 

Stano M. 137 matej.stano@savba.sk 

Stiborová M. 98, 229, 231, 233, 237, 238, 239, 242, 245 stiborov@natur.cuni.cz 

Stibůrková B. 189 blanka.stiburkova@centrum.cz 

Struhárová I. 50 

Stuchlík S. 126, 128, 130, 132, 134 stuchlik@fns.uniba.sk 

Sulová Z. 51, 73, 165, 166, 227, 246 zdena.sulova@savba.sk 

Surdeníková L. 99 surdenikova@jfmed.uniba.sk 

Svatoň M. 186 

Svetlíková Z. 100 zsvetlik@gmail.com 

Svoboda J. 47 

Svobodová M. 245 svobodova.mar@seznam.cz 

Sýkora R. 97 

Synková H. 207 

Szemes T. 77 

Szotáková B. 225, 232 barbora.szotakova@faf.cuni.cz 

Šabová Ľ. 154 

Šebeková K. 176 

Šebesta I. 189 

Šebo P. 82 

Šereš M. 51, 246 mario.seres@savba.sk 

Ševčík J. 196 

Ševčíková B. 163 

Šídlová P. 241 

Šikurová L. 108, 175 

Šimkovič M. 210 martin.simkovic@stuba.sk 

Šimončíková P. 43 

Šimšíková M. 131 michaela.simsikova@gmail.com 

Šimúth J. 48, 127 

Šipošová K. 57, 211 katkasiposova@gmail.com 

Šírová M. 73, 169 

Šístková J. 233 

Škarydová L. 240 

Škodová I. 60 

Škovierová H. 149, 197 

Skrabana R. 95 Rostislav.Skrabana@savba.sk 



Škrha Jr. J. 89 

Škultéty Ľ. 58, 93 

Škvarková L. 166 

Šmigáň P. 96, 177 peter.smigan@savba.sk 

Šolcová M. 92 

Špitalský V. 77 

Šťavíková V. 

csbmb@seznam.cz 

Štefaniková A. 91, 185, 190 stefanikova@jfmed.uniba.sk 

Štefanišinová M. 235 

Štengl M. 97 milan.stengl@lfp.cuni.cz 

Štrebl P. 223 

Šuchová K. 107 

Šulc M. 238 

Švíglerová J. 97 

Tang J. 72 

Tatarková Z. 75, 94, 101, 119, 121, 217 tatarkova@jfmed.uniba.sk 

Trefancová J. 207 

Thimová M. 63 

Tomáška Ľ. 102 tomaska@fns.uniba.sk 

Tomášková N. 55 

Tomášková Z. 103 zuzana.tomaskova@savba.sk 

Tomečková V. 104, 235 vladimira.tomeckova@upjs.sk 

Topolčan O. 186 

Tóth C. 132 toth@fns.uniba.sk 

Tóthová Ľ. 123, 133 tothoval@seznam.cz 

Trachtulec Z. 80 

Trebatický B. 184 

Tribulová N. 222 

Trilecová L. 228 

Trnková L. 226 

Turecký L. 216 

Turňa J. 123, 126, 128, 130, 132, 133, 134, 140, 209 turna@cvtisr.sk 

Turner A.J. 41 

Tvaroška I. 44, 105 chemitsa2@savba.sk 

Uhlíková E. 216 

Uhrík B. 51 

Uličná O. 108, 111, 175, 176 olga.ulicna@fmed.uniba.sk 

Ulrichová J. 249 

Umannová L. 161, 247 umi@ibp.cz 

Unger Ch. 52 

Ungvarský J. 159, 248 

Urban P. 78 


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Urbániková Ľ. 55 

Utekal P. 130, 134 utekal@fns.uniba.sk 

Vaculová A. 150, 151 

Vadovič P. 93 

Vachtenheim J. 49, 157 

Valachovič M. 170 

Valenta R. 207 

Valentová J. 124 

Vančová O. 108, 175, 176 olgavancova@gmail.com 

Vaněk O. 135 

Vaněk T. 241 kenav3@seznam.cz 

Vanko M. 125 

Vaňková R. 198 

Vantová Z. 248 zuzana.vantova@gmail.com 

Várady M. 232 

Varečka Ľ. 206, 210 

Vašák J. 106 krd@krd.cz 

Vaško L. 191, 192 ladislav.vasko1@upjs.sk 

Vašková J. 191, 192 janka.vaskova@upjs.sk 

Veliká B. 221 bvelika@gmail.com 

Verébová K. 89 

Verner Z. 60 

Vidová B. 96, 212 barbora.vidova@savba.sk 

Vidová M. 177 monika.vidova@savba.sk 

Vildová L. 228 

Vodová M. 144 

Vojtěšek B. 50 

Vokřál I. 225, 232 

Vokurková M. 174, 222 martina@biomed.cas.cz 

Vondálová Blanářová O. 150 

Vondráček J. 161, 247 

Vostálová J. 223, 243 

Votruba I. 79, 145, 220 

Vranková S. 87, 215 

Vrba J. 249 vrbambv@seznam.cz 

Vrbjar N. 111 

Vršanská M. 107 Maria.Vrsanska@savba.sk 

Waczulíková I. 108, 111, 175 waczulikova@fmph.uniba.sk 

Wagner K.-H. 109 karl-heinz.wagner@univie.ac.at 

Watala C. 176, 219 

Weignerová L. 83 

Wendel M. 52 



Wilkinson A.J. 194 

Wilson K.S. 194 

Wimmerová M. 110 michaw@chemi.muni.cz 

Wsol V. 240 

Zadražil J. 223 

Zahradníková A. 55 

Zálešák B. 243 

Zapletalová J. 223 

Závišová V. 57 

Zdařilová A. 223, 243 alfa.baba@seznam.cz 

Zemková Z. 210 

Ziegelhöffer A. 111 usrdzigy@savba.sk 

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ISBN 978-80-88866-83-1 

9 788088 866831

XXII. BIOCHEMICKÝ ZJAZD - Jesseniova lekárska fakulta

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