Effect Of Tirofiban On The Expression Of Platelet Membrane-Bound CD40 Ligand, SCD40L And MMP-2 In Plasma In Patients With Acute Myocardial Infarction After PCI

Nowadays the prevalence of the coronary heart disease(CHD) is becoming higher year by year.It is one of the important factors which affect people's health.Acute myocardial infarction(AMI) with high mortality and bad prognosis is the most serious type of CHD. The rupture of atherosclerotic plaque and thrombosis result in AMI . Platelets play an important role in the process of thrombosis .The acute and subacute thrombotic applications after percutaneous coronary intervention(PCI) therapy due much to the prethrombotic state caused by platelet activation and adhesion .So antiplatelet therapy is crucial to prevent the thrombotic events after PCI. Recently investigations indicate that overexpression of matrix metalloproteinases (MMPS) after PCI is associate with the thrombotic events after PCI. Glycoprotein (GP) IIb/IIIa antagonists are potent inhibitors of platelet aggregation that provide marked protection from ischemic events in patients undergoing PCI.This drug is the inhibitors of the so-called final common pathway of platelet aggregation. Tirofiban is one kind of GPIIb/IIIa antagonists. CD40 ligand (CD40L, also known as CD154), a transmembrane protein, was originally identified on CD4+T cells, but it was also recently found on activated platelets. CD40L is cryptic in unstimulated platelets but is rapidly presented to the platelet surface after platelet stimulation. The surface-expressed CD40L is subsequently cleaved over a period of minutes to hours, generating a soluble fragment termed sCD40L that remains trimeric. Studies on the cellular distribution of CD40L indicate that >95% of the circulating sCD40L exists in platelets. This suggests that platelet stimulatory events must be considered in the biological and pathological context of CD40Lfunction. It is now generally accepted that the CD40L-CD40 interaction is an initial event in atherothrombosis,leading in turn to the activation of several proinflammatory mediators.In fact,CD40L may promote the expression of MMPS and induce procoagulant activity in vascular cells,and the absence of CD40L affects the stability of arterial thrombi and delays arterial occlusion in vivo.In recent years,MMPS and sCD40L has been recognized biological markers of the atherosclerotic plaque's unstable statement. MMP-2 is a member of the MMPS'family.Studies shows that the serum level of MMP-2 and its activity is enhanced in AMI patients or after PCI.Platelet CD40L,sCD40L and MMP-2 are all associated with AMI and PCI .In this article ,we intend to observe the how tirofiban affects the expression of platelet CD40L,sCD40L and MMP-2 after the AMI patients received PCI therapy.To investigate what roles the platelet CD40L,sCD40L and MMP-2 play in the process of AMI and the formation of thrombotic applications after PCI.At the same time,to find out the association among the three factors.Maybe the results will supply more information about the function of GPIIb/IIIa antagonists and help us make good use of these drugs.Objectives:In this experiment, We want to investigate the problems followed:1,Observe the changes of platelet CD40L expression ,serum sCD40L and MMP-2 level before and after PCI in AMI patients. 2.Explore the associations among platelet CD40L ,sCD40L and MMP-2.3.Assemble the effects of Tirofiban on the three factors after PCI in AMI patients.Material and method: 1.Groups: Divide the patients who were diagnosed as AMI and accepted the PCI and transplanted the Rapamycin eluted stents in six hours after sickening between April, 2005 and August, 2006 in CardiacMedical Department of our hospital, the second hospital of Jilin university, into two groups : control group and tirofiban treatment group. 2.Medication: Both contol group and tirofiban treatment group were routinely treated with aspirin and clopidogrel before PCI . During the process of the PCI both the two groups were treated with heparin 8000u-10000u.In treatment group ,apply Tirofiban before transplanting stents, loading dose is 10ug/kg, intravenous injection within three minutes, and last for 24 hours intravenous drip by 0.15ug /kg /min maintenance dose. 3,Sample collect: all the patients were drawn peripheral venous blood for six milliliter before PCI and at 0 hour ,36 hours after PCI.(1)Two milliliter was put into test tubes containing 106mmol/L sodium citrate ,immediately transferred to the laboratory,centrifuged at 800r/min for 5minutes at room temperature, and thereafter used for obtaining platelet-rich plasma(PRP).(2)A separate aliquot of blood without any additives was immediately immersed in melting ice and allowed to clot for one hour before centrifugation (3500r/min at 4℃for 10minutes).The serum supernatant was stored at -80℃to be tested sCD40L.(3)The last 2 ml blood was put into test tubes without any additives and centrifuge at 3000r/min for 5 minutes, then took upper stratum blood serum and put into clean test tube to be tested MMP-2. Collected samples were preserved in icebox at 4℃. 4.platelet CD40L detection: The platelets were immunostained with fluorescein isothiocyanate conjugated anti-CD40L then the platelets were measured by flow cytometry within six hours of sampling and analysed by CellQuest Software.5,sCD40L detection: Concentrations of sCD40L were determined using a commercially available enzyme immunoassay according to the manufacturer's instructions .6.MMP-2 detection: Using the method of SDS-Page- Zymograph, detect the level ofMMP-2. 7.Statistical analysis: For clinical data,variables were compared by use of theχ2 test.An ANOVA for repeated measures followed by pairwise comparisons (Bonferroni t test).Differences in biochemical variables between groups at each collection time were analyzed by Student's unpaired t test.Correlations were accessed by deflective correlation analysis.Statistical significance was indicated by a value of P<0.05.All calculations were performed using the computer program SPSS 13.0.0.Results: 1.⑴The expression of platelet CD40L and serum sCD40L level in control group and tirofiban treatment group were different at 0.5hour before PCI and 0 hour ,36 hours after PCI. Both the expression of platelet CD40L and serum sCD40L level in two groups at 0 hour after PCI were the highest,and at 0.5 hour before PCI were the lowest .⑵Compared with the control group,both the expression of platelet CD40L and serum sCD40L levels were significantly decreased in treatment group(P<0.001).2.The serum MMP-2 level in control group and tirofiban treatment group was different at 0.5hour before PCI and 0 hour ,36 hours after PCI. The serum MMP-2 level at 36 hours after PCI was the highest,and at 0.5 hour before PCI were the lowest in control groups,and that at 0 hour after PCI was the highest,and at 0.5 hour before PCI was the lowest in treatment group.⑵Compared with the control group, serum sCD40L levels was significantly decreased in treatment group(P<0.001).3.There is positive correlation between platelet CD40L and serum sCD40L and no correlations between MMP-2 and platelet CD40L or serum sCD40L.Conclusions: 1.There were a lot of platelets being activated during the process of PCI.Platelet CD40L,serum sCD40L and MMP-2 may play important roles in the process of AMI and the formation of thrombotic applications afterPCI.2.The platelet may be the main resourse of the soluable CD40L in serum.3.A possible mechanism for tirofiban's prevention from the thrombolic applications after PCI in AMI patients is to induce the expression of platelet CD40L,serum sCD40L and MMP-2 level.