THROMBIN SIGNALING IN HEMOSTASIS AND THROMBOSIS

描述(改编自摘要) 这是针对RFA HL-99-015提交的三份合作报告之一 动脉血栓形成。申请者提案的总体目标是定义 激活血小板的信号机制，从而提供一种 治疗发展和风险因素识别的框架。目标是 这个单独的项目的目的是定义凝血酶信号在 止血和血栓形成。凝血酶通过蛋白水解酶激活血小板 G蛋白偶联受体(PAR)。申请人应当使用存在以下缺陷的小鼠 这些受体问：1)血小板中的凝血酶信号是必要的吗 正常止血吗？PAR3和PAR4是已知的凝血酶受体 参与小鼠血小板活化。MPAR3似乎没有 介导跨膜信号转导，但作为mPAR4的辅助因子 低浓度凝血酶的切割和激活。因此， 申请者期望PAR4是小鼠凝血酶信号传递所必需的 血小板。申请者将使用PAR4缺陷小鼠来测试其重要性 凝血酶信号在止血中的作用。2)衰减或消融 血小板中的凝血酶信号抑制血栓形成？人类血小板使用PAR1 和PAR4用于凝血酶信号转导，目前尚不清楚是否抑制PAR1 和/或PAR4可用于预防或治疗血栓形成。第3-段 缺陷小鼠血小板类似于PAL抑制的人血小板 (两者都依赖于凝血酶信号的PAR4)。PAR4缺陷小鼠血小板 很可能被证明是类似于人类血小板，其中所有凝血酶信号 已经被屏蔽了。申请者将使用PAR3和PAR4缺陷小鼠来 确定是否部分或完全抑制凝血酶信号转导 血小板可预防动脉和微血管血栓形成。3)是 凝血酶或其他重要的蛋白水解酶激活内皮细胞 止血和血栓形成？申请人假设激活了 内皮细胞PAR1和PAR2通过促进血栓形成和炎症 血小板与白细胞滚动粘连。申请者将决定 PAR1和/或PAR2缺陷小鼠在 炎症、微血管和动脉血栓形成。4)做功能增益 血小板G蛋白偶联受体(GPCRs)突变促进血栓形成？ 在调节细胞的基因中发现血栓前突变 在编码血浆蛋白的基因中，行为已经落后于此。 一些人类疾病是由GPCRs的功能获得突变所介导的。 申请者将使用携带PAR4功能获得突变的小鼠来 确定这样的GPCR突变，单独或与 其他基因，可能是血栓形成的基础。

DESCRIPTION (Adapted from abstract) This is one of three collaborative ROls submitted in response to RFA HL-99-015 on Arterial Thrombosis. The applicants proposal's overall goal is to define the signaling mechanisms by which platelets are activated, thereby providing a framework for therapeutic development and risk factor identification. The goal of this individual project is to define the role of thrombin signaling in hemostasis and thrombosis. Thrombin activates platelets via proteaseactivated G protein-coupled receptors (PARs). The applicants shall use mice deficient in these receptors to ask: 1) Is thrombin signaling in platelets necessary for normal hemostasis? PAR3 and PAR4 are the thrombin receptors known to participate in mouse platelet activation. It appears that mPAR3 does not mediate transmembrane signaling but instead acts as a cofactor for mPAR4 cleavage and activation by low concentrations o f thrombin. Thus the applicants expect PAR4 to be required for thrombin signaling in mouse platelets. The applicants will use PAR4-deficient mice to test the importance of thrombin signaling in hemostasis. 2) Does attenuation or ablation of thrombin signaling in platelets inhibit thrombosis? Human platelets use PAR1 and PAR4 for thrombin signaling, and it is unknown whether inhibition of PAR1 and/or PAR4 would be useful for preventing or treating thrombosis. PAR3- deficient mouse platelets are analogous to PARl-inhibited human platelets (both rely on PAR4 for thrombin signaling). PAR4-deficient mouse platelets will likely prove analogous to human platelets in which all thrombin signaling has been blocked. The applicants will use PAR3- and PAR4-deficient mice to determine whether partial or complete inhibition of thrombin signaling in platelets protects against arterial and microvascular thrombosis. 3) Is endothelial cell activation by thrombin or other proteases important in hemostasis and thrombosis? The applicants hypothesize that activation of endothelial PAR1 and PAR2 promotes thrombosis and inflammation by promoting platelet and leukocyte rolling and adhesion. The applicants will determine whether mice deficient in PAR1 and/or PAR2 are protected in models of inflammation and microvascular and arterial thrombosis. 4) Do gain-of-function mutations in platelet G protein-coupled receptors (GPCRs) promote thrombosis? The discovery of prothrombotic mutations in genes that regulate cellular behaviors has lagged that in genes encoding the plasma proteins. Several human diseases are mediated by gain-of-function mutations in GPCRs. The applicants will use mice bearing gain-of-function mutations in PAR4 to determine if such GPCR mutations, alone or in combination with mutations in other genes, might be a basis for thrombophilia.