Platelet Activation And Its Protein Kinase B Activity In Elderly Patients With Coronary Heart Disease

Protein kinase B (PKB) which was identified in 1991 is a serine/ threonine kinase. A key enzyme in PKB activation is phosphatidy linositide 3 -kinase (PI -3k) , which triggers the phosphorylation of PKB. Recently, PKB is more and more paid attention because it is involved in ischemic heart disease. It is proved that platelet plays an important role of the occurrence and development of coronary heart disease. Under some circumstances, platelet activation leads to throm-bocyte adhesion, aggregation and release. Many people have used platelet associated P - selectin as a sensitive, quantitative measure of platelet activation. P - selectin becomes a special molecule mark of platelet activation. Platelet antagonists could not effectively inhibit its activation. This study is to assess the effect of platelet activation and PKB signal pathway and to study the pathogenesis of coronary heart disease.MATERIALS AND METHODSObjective:30 cases of UAP and 32 cases of AMI are accorded with WHO diagnostic standard which worked out in 1979, except liver and kidneydisease, diabetes, hypertension and so on. They would not take platelet antagonists in two weeks. Normal control is 30 cases.Apparatus:Centrifuge, Flow Cytometer, Liquid Cintillation Counter, Fluorescence Sepectrophtometer, et al.Reagent:PAC - 1 FITC, ADP, TXB2 and 6 - keto - PGF1a ELISA kit, histone H2B, protein kinase A inhibitor peptide, [r - 32P) ATP (10mCixml-1).Methoed:Sample treat; UAP and AMI groups were given anticoagulants and platelet antagonist. Blood samples were drawn 10 min and 6 hour before and after the treatment. Centrifuge sample and obtain PRP.P - slectin assay: Take PRP to nurture 5 min in the tube containing ADP, then add 1 % PFA and be washed by PBS, and form suspension of platelet. 50ul platelet suspension is added to PAC - 1 FITC interact 30 min, then washed and centrifuged. To examine sample with Flow Cytometer in 30 min, to collect 10 000 platelet and calculate MFI. The software of Cell - Quest is used to analyze materials.TXB2 and 6 - keto - PGF1a assay: Examine sample with ELISA kit.PKB activity assay: in 0. 5 ml Eptupe, PKB activity is assayed for 10 min in 0. 5mol x L^Tris/Hcl pH 7.5,10mmol x L-1MgAC, Immol x L-12 - ME,1umol x L-1 protein kinase A inhibitor peptide, 400umol x L-1 32P - ATP and 0. 2mg x ml-1 histone H2B. Reaction products were spot on whatman P31 exchange piece. Test cpm with Liquid Cintillation Counter and measure protein by Lowrry.RESULT1. Compare with control, P - selectin and TXB2 in UAP patients and AMI patients were higher ( UAP :16.85 ±2.65Vs10.33 ± 2.47, P<0.05; 153.4±23.6 pgxml-1Vs129.4±18.4pgxml-1, P<0.05), (AMI: 17.86±2.08 Vs10.33± 2.47, P<0.01; 206.8±21.6pgxml-1Vs129.4± 18.4pgxml-1, P<0.01) , 6 - keto -PGF1a was evidently inhibited(UAP:25. 8 ±6 . 3pg x ml-1 Vs28.4± 7.3 pgxml-1, P<0.05;AMI:24.8±7.9 Vs28.4±7.3 pgxml-1, P<0. 05). But P - selectin was no differences occurred in the two group(P>0.05).2. After treatment, P - selectin was significantly inhibited (P< 0.001). TXB2/6 - keto - PGF1a was also inhibited (P < 0. 001) .3. The PKB of platelet membrane in UAP patients and AMI patients was higher than in control (UAP: 119.03 ± 20.98 Vs96. 17 ± 2 3. 31,P<0.001; AMI; 128. 98 ±26.65Vs96. 17 ±23.31,P?0. 05 ).DISCUSSIONRecently years, the study in the pathogenesis of coronary heart disease has made better progress. Besides focus of coronary atherosclerosis, platelet activation and coronary spasm being found was the other important pathologic feature. Related coronary heart diseases to platelet function were quiet close. Platelet activati...