Platelet

New GenerationofBlood ComponentsProfessor A pourazarMedical Science University of IsfahanIsfahan IRAN

James BlundellBlundell’s transfusion devices included the impellor (A),which consisted of a cup, tube , and syringe; and thegravitator (B), consisting of a receptacle held high abovethe patient with an attached tube through which the bloodwas injected into the patient.

Plastic Blood Bags &Component Separation

Transition from Blood Banking toTransfusion Medicine.Recent technological advancements andautomation has redefined componentpreparationBetter equipments used for preparation ofcomponents and their storageQuality assurance and GMPIncreased demand of componentsChemotherapy for variety of malignant diseasesOrgan transplantSurgical interventionsImmunodeficiency diseases

Blood Cells Have Different Sizesand DensitiesPlateletRed Blood CellWhite Blood Cell

Blood components preparation

Whole BloodLight SpinHeavy SpinPlatelet rich plasmaHeavy SpinPacked Red CellsPlatelet poor plasmaPlatelet concentrateFresh frozen plasma CryopptCryo poor plasmaFractionationLight spin 2000 x g for 3 min, Heavy spin 5000 x g for 5 min, g = 28.38 x R(rpm/1000)

“Soft” Spin}Platelet Rich Plasma}Packed RedCells

“Hard” Spin} Plasma} BuffyCoat}Packed RedCells

Blood Components¢ Cellular components• Red cell concentrate• Platelet concentrate• Granulocyte concentrate¢ Plasma components• Fresh frozen plasma• Plasma• Cryoprecipitate¢ Plasma fractions

Whole blood derived componentsProduct Quantity CollectedRBCs 280+-60 mlPlatelets 5.5 x 10 9 in 50-60mlFFPlasma 200-250 mlCryoprecipitate 10-20mlGranulocytes 1.25 x 10 9 Gr in 20ml

Advantages of ComponentsS Storage of each component underoptimal conditions to maintain functionfor clinical use of bloodS Improvement in therapeutic conditionsS Avoid volume overloadS Leucoreduction of cellular productsS Optimization of resource management

Clinical Efficacy of ProductsAdverse Effectsshort term/long termAchieving intended outcome(save,prolong, improve life)

MODIFIED PRODUCTS¢ Packed Red Cells with additives¢ Frozen red Cells¢ Saline washed red cells¢ Leucodepleted red cells

Why leucodepletion ?¢ Leucocytes in blood components are responsible for manycomplications associated with blood transfusion as¢ FNHTRs 6.8% after red cell transfusion• 37.5% after platelet transfusion¢ Formation of antibodies that make future transfusions lesslikely to be effective and more likely to cause a reaction.”¢ Transmission of Viruses like CMV, HTLV 1 and 2 and someprions carried inside leucocytes¢ Increased perioperative infections and increased cancerrecurrence rate.¢ Transfusion associated acute lung injury.

Approximate Number ofLeucocytes in Cellular Blood ComponentsWhole blood 10 9Red cell concentrate 10 8 -10 9Platelet concentrate 10 7 -10 8Apheresis platelets 10 6Fresh frozen plasma

Leucocyte Reductionrequired to obtain benefitsS To reduce febrile non-hemolytic

Methods¢ Centrifugation¢ Freeze / Thaw¢ Filtration -Most common used technique• Filters• 1 st generation filters. Removes large clots and particulate debris• 2 nd generation filters. Designed to remove micro-aggregates of fibrin,platelets and leucocytes from red cell concentrates• 3 rd generation filters. Designed specifically to remove free leucocytes.• Filtration• Bedside during the transfusion• Component processing laboratory

Pre storage Leuco reduction• Early WBC removal reduces accumulationof cytokine & other breakdown products• WBCs containing bacteria and virusesremoved before fragmentation• Reduction in micro aggregate formation &haemolysis• Consistent WBC removal (

Leucodepleted ComponentExtraction¢ An automated blood componentprocessing system in whichcentrifuged whole blood is separatedinto 3 components in a single step¢ A process which provides optimumblood component separation withsimultaneous leukodepletion of the same

Optipress II¢ A multifunctional, fully automatedblood component extractor belongingto the second generation¢ Designed to supply the blood bankswith high quality product and GMP s,ensuring automation, efficiency andcost and time management¢ Wide range of protocols withIntegrated Tube sealers

Post Storage leucoreductionBedside filtrationUse of leukocyte filters

Leucodepleted blood components1. To prevent NHFTR2. To prevent HLA Alloimmunization*3. To prevent transmission ofleucotropic viruses (vCJD, CMV,HTLV-1, EBV)

Leucodepleted blood componentsPossible Conditions Under Review¢ To prevent Transfusion related immunosuppession¢ TRALI¢ Tumour Occurrences¢ Leucotropic Latent Viruses Reactivation CMV & HIV¢ Reducing postoperative wound infections andmortality

Clinical conditions which benefitfrom leucoreductionThalassaemia majorAplastic anaemiaSickle cell anaemiaLeukaemiaPatients for organ transplantationPatients on DialysisPatients with multiple/chronic infections

Irradiated BloodBlood irradiated with 25Gy destroys ability ofdonor lymphocytes inblood product toproliferate in therecipient and causeGVHD¢ Blood productsrequiring irradiation:WB, PRC, RDP, SDP¢ FFP and CRYO need not beirradiated

csCSCell Separator

Cell separators typesIFC – intermittent flow¢ Mobile¢ Single venepuncture¢ Disadvantage - Greater ECV causing adverse effectsbecause of replacement fluids¢ Open/closed depending on sets in useCFC-continuous flow¢ Double / Single venepuncture¢ Less ECV (extracorporeal volume)¢ Open/closed depending on sets in use

Cell separator derivedApheresis components¢ Apheresis constitutes a procedure in whichdonor/patient blood is processed to take away orremove a specific portion of blood.¢ The remaining blood is returned back to thedonor/patient¢ Automated cell separation and collection techniquesmore consistent and reproducible than manualmethods¢ Prescribed platelet doses are easily attained throughapheresis, as automation provides the control to targethigh yield products

Principles of ApheresisAnticoagulantAddedRemaining bloodComponents recombinedand returnedPlasmaPlateletsWhole Blood Lymphocytes Whole Bloodvein Granulocytes veinErythrocytesOptical Sensors Components separated Centrifugal forceby centrifugation and blood flow rateselectively removedDesiredoptimuminterfaceefficiency

Apheresis bloodcomponentsPlatelet – PlateletpheresisRed Cells-erythrocytapheresis/Red cell pheresisLeucocytes -leucopheresis or granulocytapheresisPlasma -PlasmapheresisPeripheral blood stem cells -PBSC

Apheresis DerivedBlood ComponentsProductQuantity CollectedPlatelets 3.0-8.0 x 10 11Plasma400-600 mlRBCs180-360 mlFFP/Cryoprecipitate 500 –700 mgGranulocytes 1.0 –3.0 x 10 10PBSCVariable

Plateletpheresis¢ It is the most common andsimplest form ofApheresis¢ Apheresis Platelets knownas Single Donor Platelets¢ Depending on therequirement, machineused and donor pre-count,one or multipletherapeutic doses can becollected

Plateletpheresis- Donor¢ Donor should meet all criteriaof W.B. donation¢ Donor should not have takenaspirin containing medicinewithin 36 hours.¢ Platelet count should be150.000.µl or more¢ Interval between theprocedures should be 72hours.¢ Procedure should not bedone more than 2 times in aweek or 24 times in a year.¢ Interval should be 3 mths ifred cells removed

Pooled Random Donor PlateletsAppear to be Similar toApheresis PlateletsHowever, There are Important differences

Lower Incidence ofTransfusion ReactionsPlatelet Component % Reactions Typical WBC LevelPooled Platelet 37.5 10 9Pooled Platelet28.0 10 5 - 10 6White Cell ReducedApheresis 20.0 10 6 - 10 7Apheresis White CellReduced0.0 10 4 - 10 5

Apheresis Platelets ReducePatient Exposure to MultipleDonorsApheresis Platelets:One DonorPooled Platelets:6-8 Donors

Patient Exposure Can Be DrasticallyReduced by Choosing ApheresisPlateletsNo. of Donor Exposures100806040200Apheresis Platelets Pooled Platelets2 4 6 8 10 12 14 16 18 20 22 24 26 28 30DaysExample data assuming a patient required platelettherapy every other day for one month

Reduce Risk of Transfusion-Transmitted DiseaseApheresisPlateletsPooledPlatelets*Hepatitis B 1:63,000 1:7,875Hepatitis C 1:103,000 1:12,875HIV 1:493,000 1:61,625HTLV-11:641,0001: 80,125Aggregate Risk1:34,0001:4,250*BASED ON POOLING 8 PLATELET CONCENTRATES

Improved Patient Responsewith apheresis platelets¢ Became refractory less frequently¢ Became refractory after a higher numberof transfusions¢ Became refractory after a longer timeperiod¢ Maintained corrected count incrementsover the course of treatment

Speciality Platelets¢ HLA Matched Platelets• Are only available with Apheresis platelets. HLAmatched platelets are often necessary whenpatients become refractory to other platelettransfusion options¢ Directed Donor Platelets• Can only be achieved with apheresis platelets.

Concurrent Red Cells:¢ cRBC Collection System can effectivelyand safely collect a standardized unit ofRBCs concurrently with platelet and plasmaproducts.¢ cRBC prestorage leukoreduced productsare suitable for transfusion following 42 daystorage.

Concurrent PlasmaCollection¢ 2-3 units of Plasma can becollected along withplatelets¢ Max 500 ml depending onthe donor weightBenefits¢ Good quality testedplasma¢ Less donor exposure¢ Useful as FFP¢ Contains Ig, coagulationfactors¢ Protein replacement¢ High volumes forfractionation

Granulocyte collection¢ Infrequent –better antibiotics-use of growth factors¢ Neonatal septicemia¢ Uncontrolled adult septicemia¢ G-CSF/GM-CSF prior --- better yield¢ Adverse effect of HES / steroids avoided

Adult stem cells¢ An adult stem cell, or somatic stem cell, is• an undifferentiated cell• found among differentiated cell in a tissue or organ.¢ The cell can• renew itself• differentiate to yield the major specialized cell types of thetissue or organ.¢ Primary roles:• maintain and• repair the tissue in which they are found.¢ Reside in specific areas of each tissue where• they remain non-dividing (quiescent) until• activated by disease or tissue injury.¢ Haematopoietic stem cells are the most studied.

PBSC’s COLLECTIONFor allogenic PBSC’s collection anticubital veins& for autologous PBSC’s collection C.V accesscatheters used.¢ Cytokines increase release of CD 34 cells inPeripheral BloodPBSC’s collection should be timed when thenumber of CD34+ is maximal and collection to becontinued until an adequate number of cells havebeen obtained.Most commonly used targets are >2 x 10 6 CD34Cells per Kg.

Diseases already cured with stem cells¢ Acute leukemia¢ Chronic leukemia¢ Myelodysplastic syndromes¢ Stem cell disorders¢ Myeloproliferative disorders¢ Lymphoproliferativedisorders¢ Phagocyte disorders¢ Liposomal storage disorders¢ Inherited erythrocyteabnormalities¢ congenital inheritedimmune systemdisorders¢ Platelet abnormalities¢ Plasma cell disorders¢ Other malignanciesbreast,renal cellcarcinoma

Most recent applicationsDiabetesHeart DiseaseLiver DiseaseMuscular DystrophyParkinson's DiseaseSpinal Cord InjuryStrokeAlzheimer's Disease

Therapeutic Apheresis¢ A pathogenic substance existing in bloodcontributing to a disease or its symptomscan be more effectively removed byapheresis than by the body’s ownhomeostatic mechanisms¢ Plasma exchange if the substancecirculates in plasma¢ Cytapheresis if component is cellular

Therapeutic ApheresisVarious Applications¢ Cytapheresis¢ Adoptive Immunotherapy¢ Photopheresis¢ Plasmapheresis

Therapeutic Apheresis¢ Direct medical care of patient¢ Clearly defined policies- responsibilities¢ Trained technical staff¢ Facility of handling emergency situation¢ Attention on patients medication schedule¢ Documentation¢ Written informed consent

Therapeutic CytapheresisErythroctapheresis¢ Sickle cell disease¢ Malaria with hyper parasitaemiaLeukapheresis¢ Leukemia with hyper leukocytosis¢ Lymphoproliferative disorders¢ Pt may become unresponsive in long termtherapy

Therapeutic Cytapheresis- IILymphocytapheresis¢ Produces immunosuppression¢ RA, SLE, renal transplant rejectionPlateletpheresis¢ Myeloproliferative disorders¢ Polycythemia Vera¢ To avoid thrombotic or hemorrhagiccomplication

Adoptive ImmunotherapyImmunocompetent mononuclear cells¢ Stimulated by cytokines¢ Reinfused ---- tumour regressionLymphocytes harvested¢ Cultured in IL-2¢ Produce LAK cells¢ Renal cell Ca, melanoma, colorectal Ca

Adoptive ImmunotherapyAutologous monocytes¢ Stimulated by interferon gamma¢ Anti-tumour activityGene Therapy¢ Lymphocytes harvested¢ Transduction of missing gene¢ Reinfusion

Photopheresis¢ Patient ingests psoralen ----binds DNA ofall nucleated cells¢ Leucocytes harvested¢ Exposed to UVA radiation¢ UVA activates psoralen¢ Prevents replication¢ T-cell lymphoma¢ Other immune mediated diseases –scleroderma, RA, chr heart transplantrejection, HIV/AIDS

Factors Removed byTherapeutic Plasmapheresis¢ Immune complexes¢ Auto antibodies or alloantibodies¢ Auto antibodies causinghyper viscosity¢ Immune mediators¢ Protein bound toxins¢ Lipoproteins¢ Platelet aggregating factors¢ SLE¢ F VIII inhibitors¢ Waldenstrom’smacroglobulinemia¢ Goodpasteur’s ,Myasthenia gravis, /GB¢ Barbiturate poisoning¢ Hypercholesterolemia¢ TTP

The ALYX Component Collection System forcollection of two units of Leucodepleted RedCells¢ One continuous process tocollect, separate, andleukoreduce¢ Closed kit includesattached solutions andfilter for precise, efficientloading; no extra steps forthe operator¢ Leukoreduction is fullyautomated and can occurconcurrently withpostdonation care

The ALYX Component Collection SystemFrom donor to leukoreducedcomponents an average of just35 minutes onlyContinuous processing withintermittent draw and returnresults in an average donor timeof 22 minutesSystem-integrated leukoreductiontakes an average of 5 minutesfor 2 units of red cells

The ALYX Red Cell ProcedureQuality, speed and simplicity in double red cell collection

The ALYX Red Cell ProcedureQuality, speed and simplicity in double red cell collection

The ALYX Red Cell ProcedureQuality, speed and simplicity in double red cell collection

Autopheresis C¢ Auto C is an automated donorPlasmapheresis system¢ Designed to collect fresh frozenPlasma (sensors to monitor each phaseof the process)¢ Combined principle of• Membrane Filtration• Centrifugation¢ Process consists of 2 cycles• Collection Cycle• Reinfusion Cycle

Key Features¢ Collects 600-700 ml cell free plasma in 35minutes¢ Extra corporeal volume - 200 ml¢ Automatic flow control system to protectvascular integrity.¢ Continuous monitoring of venous pressureto control pump speed.¢ Hb detector to monitor red cellcontamination

Advantages of Auto - C¢Donor safety• Single arm process• Short procedure time• Low extracorporeal volume ( preventshypovolemia)• Continuous pressure monitoring ( optimizesthe blood flow within flow capacity limits)• Low anticoagulant usage for enhanceddonor safety• On-line processing reduces reinfusion errors

Pathogen inactivatedcomponents¢ Eliminates the concern about window periodinfectivity¢ Effective against any new,unknown andexisting pathogen therby reducing residualrisk of viral inf¢ Destroy at least one element of the virus sothat remaining virus fragments lack thestructure and components to infect therecipient

Pathogen inactivation¢ Heating---denatures viral proteins andN.acids¢ Ethanol fractionation¢ Irradiation¢ Photoinactivation –alters DNA/member lipoproteins¢ Treating with chemicals as psoralens¢ UV inactivation

Intercept blood system forplateletsInactivates viruses, bacteria, parasites andleukocytes by¢ Targeting of pathogen by amotasalen¢ Docking between the bases of both DNAor RNA¢ Illumination with UV light forms inter standcross links so that DNA/RNA can not unzipand replicate( Does not harm mature platelets)

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