Platelet ERO1alpha in thrombosis and hemostasis

项目摘要/摘要 据报道，仅在2009年，美国32%的死亡是由心血管疾病引起的，包括 急性冠脉综合征、中风/短暂性脑缺血发作和外周动脉疾病。在这些基础上 病理表现为血小板活性增强。血小板在止血和血栓形成中起着至关重要的作用。跟随 血管和组织损伤，血小板迅速附着在损伤部位并分泌颗粒状内容物， 重新招募和激活额外的血小板，最终通过相互作用形成血栓 整合素αIIb、β3和纤维蛋白原。由于整合素αIIbβ3在血小板血栓形成中起重要作用， 整合素的拮抗剂已经开发出来。然而，直接抑制与整合素αIIbβ3的配体结合 可能会损害止血并增加出血的风险。因此，已经提出了许多努力来发展 阻断整合素αIIbβ3功能或信号转导的抑制剂。 许多研究已经确定了硫醇-二硫键交换作为调节机制的作用。 对于整合素αIIbβ3。使用实时活体显微镜，我们和其他人证明了异构酶活性 细胞表面蛋白二硫键异构酶(Pdi)是一种典型的硫醇异构酶，在细胞表面蛋白的调节中起着关键作用。 血管损伤部位整合素β3的激活和血栓的形成，提示PDI是一种新的 血栓形成的治疗靶点。然而，阻断细胞外PDI会影响止血效果 通过尾巴出血时间评估小鼠的功能。这些结果证明有必要进一步研究如何 细胞外PDI活性在血管损伤后受到调节。 在内质网(ER)中，PDI催化硫醇-二硫化物的氧化、还原和异构化 蛋白质折叠和内质网氧化还原蛋白1α(ERO 1α)是负责接受电子的关键蛋白质 减少PDI以氧化PDI，恢复其活性。在这个全面的提案中，我们将检验这一假设 血小板表面ERO_1α通过调控α活性和β3调节整合素PDI的配体结合功能 促进血栓形成过程中的血小板聚集。拟议的研究将提供对 抑制整合素αIIbβ3配体结合功能的新途径--α-PDI 信号轴，并可能导致新的治疗策略的发展。

PROJECT SUMMARY/ABSTRACT It was reported that in 2009 alone, 32% of all deaths in the US resulted from cardiovascular disease, including acute coronary syndrome, stroke/transient ischemic attack, and peripheral artery disease. Underlying these pathologies is increased platelet activity. Platelets play an essential role in hemostasis and thrombosis. Following vascular and tissue damage, platelets rapidly adhere to the site of injury and secrete granular contents that recruit and activate additional platelets, culminating in the formation of thrombi through the interaction between integrin αIIbβ3 and fibrinogen. Because of the essential role of integrin αIIbβ3 in platelet thrombus formation, antagonists of the integrin have been developed. However, direct inhibition of ligand binding to integrin αIIbβ3 could impair hemostasis and increase the risk of bleeding. Thus, many efforts have been put forward to develop inhibitors blocking integrin αIIbβ3 function or signaling. Numerous studies have now characterized the role of thiol-disulfide bond exchange as a regulatory mechanism for integrin αIIbβ3. Using real-time intravital microscopy, we and others demonstrated that the isomerase activity of cell surface protein disulfide isomerase (PDI), a prototypical thiol isomerase, plays a critical role in regulating integrin β3 activation and thrombus formation at the site of vascular injury, thereby suggesting PDI to be a novel therapeutic target for the treatment of thrombosis. However, blocking extracellular PDI compromised hemostatic function in mice as assessed by tail bleeding times. These results warrant the need for further research into how extracellular PDI activity is regulated following vascular injury. In the endoplasmic reticulum (ER), PDI catalyzes thiol-disulfide oxidation, reduction, and isomerization during protein folding and ER oxidoreductin 1α (ERO1α) is the key protein responsible for accepting the electrons from reduced PDI to oxidize PDI, regenerating its activity. In this comprehensive proposal, we will test the hypothesis that platelet surface ERO1α modulates the ligand-binding function of integrin αIIbβ3 by controlling PDI activity and facilitates platelet aggregation during thrombosis. The proposed studies will provide mechanistic insight into an innovative approach to downregulate the ligand-binding function of integrin αIIbβ3 by inhibition of the ERO1α-PDI signaling axis and may lead to the development of novel therapeutic strategies.