2008 Barcelona - European Society of Human Genetics

Molecular and biochemical basis of disease
predict the TF binding the selected regions . This prediction was experimentally
analyzed by EMSA with nuclear extract of Jurkat T-cell
line. The sequences of Cy5-labeled ds oligonucleotides flanked the
polymorphic loci and were 39bp (C-33T) and 35bp(C-590T) long . Results:
It was predicted that TFs Oct-1, CREB, C/EBP and GATA could
selectively bind C-33T polymorphism locus while MZF-1, Sox-5, PU .1,
AP-2, GR, VDR, E2F-1 and T3R-alpha could bind C-590T locus . We
observed that C-33T EMSA probes associated with two specific complexes
. The TFs slightly discriminated between -33C and -33T probes .
The unlabeled consensus ds oligonucleotides for the Oct-1 completely
inhibited the lower shift band . Both bands were not inhibited by the oligonucleotides
corresponding to the C/EBP, CREB and GATA binding
sites. As for C-590T polymorphism, two specific bands were observed
in the sample with -590C probe rather than -590T probe . They were
not inhibited by the unlabeled concurrent probes for the predicted TF
binding . conclusions: Oct-1 transcription factor could be responsible
for the phenotypic effect of C-33T IL-4 promoter polymorphism . The
factor responsible for the C-590T polymorphism does exist but should
yet be identified. The work was supported by the RFBR grant 08-04-
01535 .
P05.105
Effect of semax treatment on expression of the neurotrophins
and their receptors in ischemic rat hippocampus
V. V. Stavchansky1 , L. V. Dergunova1,2 , A. B. Botsina2 , T. V. Tvorogova2 , V. I.
Skvortsova2 , S. A. Limborska1,2 ;
1 2 Institute of Molecular Genetics RAS, Moscow, Russian Federation, Institute of
Stroke RSMU, Moscow, Russian Federation.
Neurotrophins are known as natural neuroprotectors involved in proliferation,
differentiation and survival of neuronal and glial cells . We
analyzed the effect of synthetic polypeptide Semax (Met-Glu-His-Phe-
Pro-Gly-Pro) upon expression of neurotrophins Bdnf, Ngf, Nt3 and
their receptors TrkB, TrkA, TrkC, p75 in rat hippocampus after global
cerebral ischemia . The study was carried out on 2-3-month-old male
Wistar rats (n=85) . After 15 minutes of irreversible bilateral common
carotid artery occlusion the animals were exposed to intraperitoneal
injection of either Semax or saline 1, 4 and 8 hours after the occlusion .
Ischemic rats with saline injection were used as control groups . The
mRNA expression of neurotrophins and its receptors was assessed
by relative quantification using real-time RT-PCR . Gapdh was used
as the reference gene . The neurotrophins’ transcription was increased
compare with the level of these transcripts in control animals: for Bdnf
at 1h, 2h and 12h; for Ngf at 1h, 12h and 24h and for Nt3 at 1h and
12h after operation . Under Semax treatment the mRNA expression of
neurotophin’s receptors was increased compare with control as well:
for TrkB at 30 min, 1h and 24h, for TrkC at 8h and 12h, and for p75
at 2h, 4h and 12h after occlusion . The level of TrkA mRNA was decreased
at 1h, 2h, 4h, and 24h after surgery; at 8h it was increased .
It could be suggested that neuroprotective effect of Semax is possibly
mediated by neurotrophins and its receptors .
P05.106
semax and its c-terminal Pro-Gly-Pro tripeptide change the
transcription profile of growth factors and their receptors genes
after focal cerebral ischemia in rats
V. G. Dmitrieva1 , L. V. Dergunova1 , O. V. Povarova2 , V. I. Skvortsova2 , S. A.
Limborska1 ;
1 2 Institute of Molecular Genetics RAS, Moscow, Russian Federation, Institute of
Stroke RSMU, Moscow, Russian Federation.
Neuroprotective polypeptide Semax is used for acute therapy of stroke .
The effect of Semax and its C-terminal PGP tripeptide on mRNA expression
of 84 genes representing growth factors and their receptors in
the lesioned cortex of rat brain after 3 h of permanent middle cerebral
artery occlusion (pMCAO) was analyzed using the Rat Neurotrophin
and Receptors gene array (“RT² Profiler PCR Array, Superarray).
Under Semax treatment in the lesioned cortex mRNA expression of
19 genes was altered, the change in transcription of 7 genes of these
19 also was observed under ischemic conditions . PGP promoted the
transcriptional alteration of 23 genes . The list of genes which mRNA
expression was changed under Semax treatment only partly coincided
with the list of genes that was changed under PGP treatment; the direction
of such transcription alteration was the same . The most marked
(two times and more) alteration of mRNA expression under Semax and
PGP treatment was observed for Lif, Galr1, TrkA and p75 . Some of the
genes which mRNA expression were changed by Semax treatment
only: Fos, IL10ra, Il1b and Tgfb1; and by PGP treatment only: Fgf2,
Hctr1, Tacr1, Ntf5 and Ptger2 . Hence it was suggested that Semax
effects observed earlier do not reflect only PGP activity. Obviously the
mechanisms of action of these peptides are not the same and they
each have their own specific effects. From the list of genes altered
by Semax or PGP two types of growth factor targets were suggested:
neural tissue (neurons and glial cells) and vascular cells .
P05.107
Novel mutations in myoclonin1/EFHC1 in sporadic and familial
juvenile myoclonic epilepsy
M. Alonso 1 , M. Medina 2 , T. Suzuki 3 , R. Durón 2,4 , I. Martínez 1,4 , J. N. Bailey 4 , D.
Bai 4 , Y. Inoue 5 , I. Yoshimura 6 , S. Kaneko 6 , M. Montoya 2 , A. Ochoa 1 , A. Jara 1 , M.
Tanaka 2 , J. Machado 2 , S. Fujimoto 7 , M. Ito 8 , S. Hamano 9 , K. Sugita 10 , Y. Ueda 11 ,
M. Osawa 12 , H. Oguni 12 , F. Rubio 1 , K. Yamakawa 4 , A. Delgado-Escueta 4 ;
1 National Institute of Neurology and Neurosurgery Manuel Velasco Suárez,
Mexico City, Mexico, 2 National Autonomous University of Honduras, Tegucigalpa,
Honduras, 3 RIKEN Brain Science Institute, Saitama, Japan, 4 UCLA,
Los Angeles, CA, United States, 5 National Epilepsy Center, Shizuoka, Japan,
6 Hirosaki University, Hirosaki, Japan, 7 Nagoya City University, Nagoya, Japan,
8 Shiga Medical Center for Children, Moriyama, Japan, 9 Children´s Medical Center,
Saitama, Japan, 10 Ichikawa General Hospital, Shiga, Japan, 11 University of
Miyazaki, Miyazaki, Japan, 12 Tokyo Woman´s Medical University, Tokyo, Japan.
Background: Juvenile myoclonic epilepsy (JME) accounts for 3 to 12%
of all epilepsies . In 2004, the GENESS Consortium demonstrated 4
missense mutations in Myoclonin1/EFHC1 of chromosome 6p12 .1
segregating in 20% of Hispanic families with JME . Objective: To examine
what percentage of consecutive JME clinic cases have mutations
in Myoclonin1/EFHC1 . Methods: We screened 44 consecutive
patients from Mexico and Honduras, and 67 patients from Japan using
heteroduplex analysis and direct sequencing. Results: We found five
novel mutations in transcripts A and B of Myoclonin1/EFHC1 . Two novel
heterozygous missense mutations (c .755C>A and c .1523C>G) in
transcript A occurred in a singleton from Mexico and another singleton
from Japan . A deletion/frameshift (C .789del .AV264fsx280) in transcript
B was present in a mother and daughter from Mexico . A non-sense
mutation (c .829C>T) in transcript B segregated in 4 clinically and 7 epileptiform-EEG
affected members of a large Honduran family . The same
non-sense mutation (c .829C>T) occurred as a de novo mutation in a
sporadic case . Finally, we found a three-base deletion (-364^-362del .
GAT) in the promoter region in a family from Japan . Conclusion: Nine
percent of consecutive juvenile myoclonic epilepsy cases from Mexico
and Honduras clinics and three percent of clinic patients from Japan
carry mutations in Myoclonin1/EFCH1 . These results represent, the
highest number and percentage of mutations found for a juvenile myoclonic
epilepsy causing gene of any population group .
P05.108
Long Qt syndrome: two Novel Exonic KCNH mutations
K. Banihashemi 1 , S. Saber 2 , E. Zaklyazminskaya 3 , M. Houshmand 4 , M. Eftekharzadeh
5 , F. Fazelifar 6 , M. Rostami 2 , M. Khosroheidari 2 ;
1 GPEF, Ministry of Science, Research and technology, Tehran, Islamic Republic
of Iran, 2 Special Medical Center, Tehran, Islamic Republic of Iran, 3 Russian
Academy of Science, Moscow, Russian Federation, 4 NRCGEB, Tehran, Islamic
Republic of Iran, 5 Tehran Arrhythmia center, Tehran, Islamic Republic of Iran,
6 Rajaii Heart Hospital, Tehran, Islamic Republic of Iran.
Introduction . K Channels made with the KCNH2 protein are active in
the heart muscle, where they transport potassium ions out of cells . The
gene contains 15 exons spanning approximately 19 kb on chromosome
7q35 . A feature of K channelopathy is pronounced prolongation
of the QT interval and arrhythmia that can lead to sudden death .
Materials and methods . The subjects studied included a 6 members
family with 3 out of them involved with LQTS .Genomic DNA was extracted
according to a standard method, and then KCNH were PCRamplified
and sequenced for identifying LQTS-causing mutations.
Results . Two exonic novel mutation of KCNH in the exon 6 which was
present also both in his mother and his elder sister and also One more
novel intronic mutation .
Discussion . The present data, combined with those from previous
studies, give more information on KCNH2 mutations which affect loop
and pore proteins . The proband , a 16 years old boy with novel muta-