PLATELET HIGH MOLECULAR WEIGHT KININOGEN

血小板高分子量激肽原

• 批准号: 3349536
• 负责人: ALVIN H SCHMAIER
• 金额: $ 6.28万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-07-01 至 1989-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3349536
• 关键词: alpha globulin; bradykinin; coagulation factor XI; coagulation factor XII; cofactor; exopeptidase; human subject; immunochemistry; kallikreins; molecular weight; monoclonal antibody; platelets; prostacyclins; protease inhibitor; radiotracer; vascular endothelium; zymogens

The objectives of this proposal are to characterize the mechanisms whereby platelet high molecular weight kininogen (HMWK) can be an extracellular regulator of contact phase zymogen activation, a possible intracellular inhibitor of platelet calpains, and an indirect regulator of endothelial function. The specific aims of this investigation are as follows: (1) Platelet HMWK present on the external membrane of activated platelets will be quantified and characterized by 125I-monoclonal anti HMWK Fab binding studies; (2) Platelet HMWK will be characterized in unstimulated and activated platelets as to its location and distribution by immunocytochemical staining using colloidal gold; (3) Purified HMWK subunit binding to platelets will be done to determine the portion of the HMWK molecule that binds to platelets and to determine if the affinity of 125I-HMWK-platelet binding is altered when platelets are activated; (4) Coordinate factor XI/XIa-HMWK platelet binding will be performed to determine if HMWK serves as their platelet receptor; (5) The molecular structure of platelet HMWK will be characterized by immunoblot to determine if it is different from plasma HMWK or if it is cleaved by platelet calpain; (6) The effect of purified platelet calpain on purified HMWK will be studied to determine if platelet calpain activates HMWK and produces an altered HMWK with a structure similar to the HMWK seen in platelets, and to determine if HMWk itself can be an inhibitor of platelet calpain; (7) The availability and function of bradykinin derived from platelet HMWK will be ascertained to determine whether it is directly secreted from platelets and to determine whether a sufficient amount can be made available from platelet HMWK to stimulate endothelial cell secretion of prostacyclin. These studies are intended to characterize the platelet surface as a site for contact phase zymogen activation through the expression of HMWK on the platelet external membrane; to demonstrate that HMWK is activated by and is an inhibitor of platelet calpain; and to determine that bradykinin from secreted platelet HMWK is sufficient to stimulate endothelial cell prostacyclin secretion. These objectives are consistent with the hypothesis that platelet HMWK is a mediator and regulator of the surface-activated defense reactions of the proteins of the Hageman factor pathways which participate in the initiation of coagulation, the inflammatory response, and local injury-site blood pressure control.

本提案的目的是确定各种机制的特点， 血小板高分子量激肽原（HMWK）可以是细胞外的 接触相酶原激活的调节剂，一种可能的细胞内 血小板钙蛋白酶抑制剂和内皮细胞的间接调节剂 功能 本研究的具体目的如下：（1） 存在于活化血小板外膜上的血小板HMWK将 通过125 I-单克隆抗HMWK Fab结合进行定量和表征 （2）血小板HMWK将在未受刺激和 活化血小板的位置和分布， 胶体金免疫细胞化学染色：（3）纯化的HMWK亚基 将进行与血小板的结合以确定HMWK的部分 与血小板结合的分子，并确定 当血小板被激活时，125 I-HMWK-血小板结合改变;（4） 将进行协调因子XI/XIa-HMWK血小板结合， 确定HMWK是否作为其血小板受体;（5）分子 血小板HMWK的结构将通过免疫印迹表征以确定 如果它不同于血浆HMWK或如果它被血小板裂解 （6）血小板钙蛋白酶对纯化HMWK的影响 进行研究，以确定血小板钙蛋白酶是否激活HMWK并产生 改变的HMWK具有与血小板中观察到的HMWK相似的结构， 确定HMWk本身是否可以是血小板钙蛋白酶的抑制剂;（7） 从血小板HMWK衍生的缓激肽的可用性和功能将是 确定其是否直接从血小板分泌， 以确定是否有足够的数量可以从 血小板HMWK刺激内皮细胞分泌前列环素。 这些研究旨在表征血小板表面作为一个位点， 对于通过HMWK在细胞膜上的表达进行的接触相酶原活化， 血小板外膜;证明HMWK被激活， 血小板钙蛋白酶抑制剂;并确定缓激肽从 分泌的血小板HMWK足以刺激内皮细胞 前列环素分泌 这些目标符合 假设血小板HMWK是一种介质和调节剂， Hageman因子蛋白的表面活化防御反应 参与启动凝血的途径， 炎症反应和局部损伤部位血压控制。