аЯрЁБс>ўџ 24ўџџџ1џџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџџьЅСq`№ПЩbjbjqPqP .::Ц џџџџџџЄLLLLœœœАддддш АК Ж[ [ [ ! # # # # # # $phиnG -œс 9 "[ с с G LL9t m m m с HL8œ! m с ! m m „œm є а‡Ь| Фд) 4m ! Š 0К m F] Fm Fœm Д[ o m } ‰ X[ [ [ G G ] [ [ [ К с с с с ААА$дАААдАААLLLLLLџџџџ  FLOW CYTOMETRY MONITORING OF LEUKOCYTE-PLATELET AGGREGATES IN PTSD Anelko Vidovi1, Maja Vilibi2, Katja Gotovac1, Ante Sabioncello1, Vera Folnegovi-`malc2, Dragan Dekaris1 1Institute of Immunology, Rockefellerova 10, Zagreb; 2Vrap e Psychiatry Hospital, Bolni ka 32, Zagreb There is a growing evidence for influence of psychological factors on development of coronary artery disease (CAD). It is proposed that platelet-mediated mechanisms might form one of the pathways through which this association is formed. cit_af ref_bf(Steptoe, 2003 ref_num153)ref_af Whole blood flow cytometry is a valuable method for assessing platelet function and measurement of circulating leukocyte-platelet aggregates (LPA) provides a sensitive marker for in vivo platelet activation. We have recently introduced this method in our laboratory and our preliminary study obtained on 14 PTSD patients and 8 aged matched healthy controls showed reproducible results comparable with recent literature. We assume that increased leukocyte-platelet aggregation may frequently be induced in chronic stress and may accumulate to confer increased cardiovascular disease risk on susceptible individuals. A clean blood draw and gentle handling of specimens are required to avoid spontaneous platelet activation. Samples were drawn using 20 gauge needles in Citrate theophylline adenosine dipyridimole (CTAD) Vacutainers which chelates Ca2+ and increases intracellular cAMP, keeping platelets "quiet". 10 (L of blood was immediately added to 90 (L solution containing CD45 FITC, CD14 PE (Leucogate) and CD42a PerCP (platelet specific antibody) monoclonal antibodies. After 20 minutes of incubation at room temperature in the dark, samples were fixed using 900 (L of 0,5% paraformaldehyde and stored at 4(C for at least 2 hours. We analyzed samples with Becton Dickinson LSR II flow cytometer using CD45 and FSC threshold gating to get rid of unwanted events in unlysed and unwashed whole blood. Monocytes, neutrophils and lymphocytes could be distinguished based on linear forward versus side light-scatter properties, but identification of gated populations was improved by using a leukocyte-specific antibodies. Proportions of CD42a positive events in monocyte, neutrophil and lymphocyte gates were measured. Our results did not reveal any significant difference between PTSD patients and healthy controls although percentages of monocytes platelets aggregates tend to be higher in PTSD patients. This may be due to small sample sizes so further investigation is needed. Furthermore, bŠЈЊЦШцш  < > ^ b d Ь Ю .  №ьсЦЉtttttW*CJH*KHOJQJ\^JaJmHsH5h Оh О5>*CJKHOJQJ\^JaJmHsHh Оh ОCJaJhOjhЅUmHnHsHuŠb . 0 ХЦЧШЩ§јёёхн觧§gd О$a$gd О $„`„a$gd ОЄgd Оgd О Ш§§ 3 ƒ…ЦЧюяХЦ№ёХЦЧШЩѓшмшЮшЮшЮшРшОшГЏЋЇhOhWYЭhЅh ОhЅCJaJU jА№h Оh ОCJaJ jm№h Оh ОCJaJh Оh ОCJH*aJh Оh ОCJaJh Оh О<CJaJetter choice of monoclonal antibodies and multiparameter capabilities of LSR II flow cytometer might improve our detection of LPA and reveal possible risk for CAD in people under prolonged stress. ,1hА‚. 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