PAR4 mediates platelet activation in venous thromboembolism

项目摘要/摘要 静脉血栓形成(VT)及其主要并发症肺栓塞(PE)通常被归类为 静脉血栓栓塞(VTE)。这种疾病每年影响近60万人，是全球最大的 终末期心血管疾病和死亡的原因。VT由三个主要因素推动，称为 Virchow三联症：高凝、血管内皮细胞功能障碍和血流停滞。它现在被认为是 血小板发挥着关键的启动作用，其机制刚刚被揭开。高凝状态 和凝血酶生成是VT的主要危险因素，凝血酶激活的蛋白酶激活的受体4 (PAR4)促进促凝血血小板、磷脂酰丝氨酸(PS)暴露和随后的凝血酶 一代。我认为血小板PAR4是VTE的重要贡献者。这一提议的科学前提是 是基于我的初步数据显示低反应性PAR4单核苷酸多态(SNP)(PAR4- P310L)位于PAR4的细胞外环3，在GWASMeta分析中与VTE的风险较低相关。 此外，具有同源PAR4-P322L多态的小鼠血小板在 对凝血酶激活的反应，最终表现为PAR4受体反应性降低。这样做的目的是 建议1)确定PAR4如何使用互补小鼠促进VT的启动和繁殖 模型，2)验证PAR4作为治疗VT的靶点3)确定PAR4在VT中如何调节血小板功能。我 假设血小板上的PAR4信号是VT的驱动因素，并且减少了PAR4信号或药物作用 抑制将提供对VTE的保护。我的目标是确定PAR4在PAR4启动和繁殖中的作用。 静脉血栓形成。我将使用3个互补的VT小鼠模型来建立PAR4作为主要贡献者 不同血栓形成条件下的血栓形成。为了确定PAR4反应性水平 血栓繁殖所必需的，我将使用3个不同水平的PAR4活性的小鼠模型：野生型， PAR4-P322L模拟人类SNP，PAR4-/-完全去除PAR4。此外，这项提案 目的明确PAR4激活在静脉血栓栓塞症中如何调节血小板反应。我将在微流控系统中使用人血 以及上述具有不同水平的PAR4反应性的小鼠模型来确定 PAR4影响室性心动过速的发生。完成后，该提案将确定一个未确定的 PAR4介导的血小板促凝血活性在VTE中的作用机制，可提供新的治疗方法 静脉血栓栓塞症患者的目标。

PROJECT SUMMARY/ABSTRACT Venous thrombosis (VT) and its major complication, pulmonary embolism (PE), are often grouped together as venous thromboembolism (VTE). This disease affects nearly 600,000 individuals per year and is the leading cause of end-stage cardiovascular disease and death. VT is promoted by three major factors, known as Virchow’s triad: hypercoagulability, endothelial cell dysfunction, and stasis of blood flow. It is now recognized that platelets play a critical initiating role, and the mechanism is just now being uncovered. Hypercoagulation and thrombin generation are major risk factors for VT, and thrombin-activated protease activated receptor 4 (PAR4) promotes procoagulant platelets, phosphatidyl serine (PS) exposure, and subsequent thrombin generation. I propose that platelet PAR4 is an important contributor to VTE. The scientific premise of this proposal is based my preliminary data showing that a hypo-reactive PAR4 single nucleotide polymorphism (SNP) (PAR4- P310L) located in extracellular loop 3 of PAR4 is associated with a lower risk for VTE in a GWAS meta-analysis. Further, mouse platelets with a homologous PAR4-P322L polymorphism exhibit decreased aggregation in response to thrombin activation and ultimately show decreased PAR4 receptor reactivity. The goals of this proposal are 1) to determine how PAR4 contributes to VT initiation and propagation using complementary mouse models, 2) validate PAR4 as a target to treat VT 3) define how PAR4 modulates platelet function in VT. I hypothesize that PAR4 signaling on platelets is a driver of VT, and reduced PAR4 signaling or pharmacological inhibition will offer protection from VTE. I aim to determine the role of PAR4 in the initiation and propagation of venous thrombosis. I will use 3 complementary mouse models of VT to establish PAR4 as a major contributor of clot development under different thrombotic conditions. In order to determine the level of PAR4 reactivity necessary for thrombus propagation, I will use 3 mouse models with differing levels of PAR4 activity: wild-type, PAR4-P322L mimicking the human SNP, and PAR4 -/- to completely remove PAR4. Additionally, this proposal aims to define how PAR4 activation modulates platelet response in VTE. I will use human blood in microfluidic chambers as well as mouse models with varying levels of PAR4 reactivity described above to determine how PAR4 influences the development of VT. Upon completion, this proposal will identify an undetermined mechanism behind PAR4-mediated platelet procoagulant activity in VTE, which can provide a novel therapeutic target for patients with VTE.