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Mechanism of Procoagulant Activity Appearing on the Surface of Activated Platelet and New Development of Anttithrombotic Therapy by Its Inhibition.

活化血小板表面促凝活性的机制及其抑制抗血栓治疗的新进展。

基本信息

批准号：
15590771
负责人：
GOTO Shinya
金额：
$ 2.11万
依托单位：
TOKAI UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

项目摘要

Fibrin formation subsequent to the accumulation of platelet at site of disrupted atheroma play an-important role in the onset of arterial thrombosis such as acute myocardial infarction. We have developed a new ex vivo assay system, demonstrating procoagulant activity appearing on the surface of platelet formatting thrombi on the surface of immobilized collagen under arterial blood flow condition, which was composed from ex vivo flow chamber system as well as an epi-fluorescent vidio-microscopy. Human blood samples containing various concentrations of specific antithrombin agent of Argatroban were perfused on the immobilized type I collagen at a wall shear rate of 1,500 s^<-1>. Platelet, which was rendered fluorescent by addition of mepacrine, was fluorescinated as green, while fibrin monomer formed on the surface of activated platelets, which was fluorescent as red with the use of AlexFluor595 conjugated anti-fibrin monomer monoclonal antibody. We have shown that specific thrombin inhibitor, which has been considered as anticoagulant, could prevent the three-dimensional growth of platelet thrombi by blocking the function of thrombin generated on the surface of activated platelets. We have also shown that another thrombin inhibitor hirulog, which block the function of thrombin irreversibly, more efficiently prevented the three-dimensional growth of platelet thrombin. We have demonstrated the close relationship between platelet thrombus formation and the activation of coagulant. cascade, suggesting the platelet-derived procoagulant activity as a new target of antithrombotic treatment.

血小板在动脉粥样硬化破裂部位聚集后形成的血栓在动脉血栓形成如急性心肌梗死的发病中起重要作用。我们开发了一种新的体外测定系统，该系统由体外流动室系统和落射荧光显微镜组成，用于在动脉血流条件下，在固定化胶原表面形成的血小板血栓表面显示促凝血活性。将含有不同浓度的阿加曲班特异性抗凝血酶剂的人血液样品以1，500 s-1的壁剪切速率灌注在固定的I型胶原上<-1>。通过加入米帕林而呈现荧光的血小板被荧光化为绿色，而在活化的血小板表面上形成纤维蛋白单体，其通过使用AlexFluor 595缀合的抗纤维蛋白单体单克隆抗体而被荧光化为红色。我们已经证明，特异性凝血酶抑制剂，这已被认为是抗凝剂，可以阻止三维生长的血小板血栓通过阻断功能的凝血酶产生的活化血小板表面。我们还发现，另一种凝血酶抑制剂hirulog可不可逆地阻断凝血酶的功能，更有效地阻止血小板凝血酶的三维生长。我们已经证明了血小板血栓形成与凝血剂的激活之间的密切关系。级联，提示血小板衍生的促凝血活性作为抗血栓治疗的新靶点。