Protease Activated Receptor and Thrombomodulin Signaling by Coagulation Proteases

凝血蛋白酶激活的蛋白酶受体和血栓调节蛋白信号传导

基本信息

批准号：
10418775
负责人：
ALIREZA R. REZAIE
金额：
$ 43.7万
依托单位：
OKLAHOMA MEDICAL RESEARCH FOUNDATION
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-04-01 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10418775
关键词：
Acetylation Adherens Junction Anti-Inflammatory Agents Anticoagulants Binding Biological Assay Blood Vessels Cell Adhesion Molecules Cell Proliferation Chlorhexidine Coagulation Process Complex Cytoplasm Cytoplasmic Tail Data Deposition Disease Down-Regulation Endothelial Cells Epidermal Growth Factor Epigenetic Process Evaluation Extracellular Matrix Proteins Fibrosis Funding Gluconates Glycoproteins Goals Guanosine Triphosphate Phosphohydrolases HMGB1 Protein Human Immunofluorescence Immunologic Impairment Inflammation Inflammatory Injections Injury to Peritoneum Intraperitoneal Injections Lead Lectin Ligands Maintenance Mediating Membrane Mesothelial Cell Methods Modeling Monitor Mutagenesis Myocardial Infarction N-terminal Neoplasm Metastasis PAR-1 Receptor Pathway interactions Pattern recognition receptor Peptide Hydrolases Peritoneal Peritoneal Mesothelial Cell Permeability Pharmacology Phenotype Phosphorylation Play Production Prostaglandins I Protein C Proteinase-Activated Receptors Proteins Public Health Recombinants Regulation Reporting Research Role Sepsis Series Signal Pathway Signal Transduction Site Solid Specificity Stimulus Stroke System TLR2 gene TLR4 gene TNF gene Thrombin Thrombin Receptor Thrombomodulin Thrombosis Tight Junctions Tissue Sample Transforming Growth Factor beta Transmembrane Domain Vascular Endothelial Cell Western Blotting activated Protein C activated protein C receptor alpha-Thrombin base chondroitin sulfate glycosaminoglycan cofactor cytokine extracellular inhibitor novel therapeutics pathogen prevent protein activation receptor response thrombotic

项目摘要

Project Summary Thrombomodulin (TM) is a type-1 integral membrane receptor which was identified and cloned as a receptor for thrombin on endothelial cells. It is a multi-domain glycoprotein comprised of five distinct domains that include an N-terminal lectin-like domain, followed by six epidermal growth factor (EGF)-like domains, a membrane proximal Ser/Thr rich domain harboring chondroitin sulfate glycosaminoglycans, a single spanning transmembrane domain and a cytoplasmic tail. TM switches the specificity of thrombin from a procoagulant to an anticoagulant protease by binding to exosite-1 of thrombin via its EGF-like domains 4, 5, and 6 (TM456), thereby enabling thrombin to activate protein C to activated protein C (APC). The exosite-1-dependent interaction of thrombin with TM on endothelial cells can inhibit the thrombin recognition of protease-activated receptor 1 (PAR1) and procoagulant substrates. TM also exerts antiinflammatory functions through its lectin-like domain by unknown mechanisms. Recent results have indicated an association between loss of TM expression and uncontrolled cell proliferation and metastasis. Based on our preliminary data in this application, we hypothesize that thrombin can function as a true ligand for TM to elicit direct exosite-1-dependent antiinflammatory signaling responses in vascular endothelial cells. We hypothesize that this function of TM not only plays a cytoprotective role in response to inflammatory stimuli, but it may also contribute to regulation of the vascular tone, maintenance and stabilization of the barrier permeability function of the vasculature under steady-state conditions. We further hypothesize that TM prevents thrombin from interacting with PAR1 on microvascular endothelial cells, thereby TM directly transmitting the protective signaling function of thrombin via its cytoplasmic domain. We have prepared a series of TM receptor constructs, recombinant APC and thrombin derivatives and TM-null endothelial cells to investigate the following three Specific Aims: Aim 1 will investigate the hypothesis that thrombin can function as a true ligand for TM to elicit direct cytoprotective signaling responses in endothelial cells. Aim 2 will investigate the mechanism by which components of the thrombin-TM-APC signaling axis inhibit the proinflammatory signaling function of high mobility group box 1 (HMGB1) in endothelial cells. Aim 3 will develop a chlorhexidine gluconate (CG)-induced peritoneal injury model and evaluate the mechanism of the cytoprotective function of the thrombin- TM-protein C signaling axis in this model.

项目概要血栓调节蛋白 (TM) 是一种 1 型整合膜受体，被鉴定并克隆为内皮细胞上的凝血酶受体。它是一种多结构域糖蛋白，由五个不同的包含 N 端凝集素样结构域，随后是六个表皮生长因子的结构域 (EGF) 样结构域，一种含有硫酸软骨素的近膜 Ser/Thr 丰富结构域糖胺聚糖、单个跨越跨膜结构域和细胞质尾部。 TM开关通过与 exosite-1 结合，凝血酶从促凝剂到抗凝剂蛋白酶的特异性通过其 EGF 样结构域 4、5 和 6 (TM456) 激活凝血酶，从而使凝血酶能够激活蛋白 C 转变为活化蛋白 C (APC)。凝血酶与 TM 的外位点 1 依赖性相互作用内皮细胞可以抑制蛋白酶激活受体 1 (PAR1) 的凝血酶识别促凝血底物。 TM 还通过其凝集素样结构域发挥抗炎功能未知的机制。最近的结果表明 TM 表达缺失与以及不受控制的细胞增殖和转移。根据我们在此应用程序中的初步数据，我们假设凝血酶可以作为 TM 的真正配体发挥作用，从而引发直接的 exosite-1 依赖性血管内皮细胞的抗炎信号反应。我们假设这个函数 TM 不仅在响应炎症刺激时发挥细胞保护作用，而且还可能有助于调节血管张力、维持和稳定屏障稳态条件下脉管系统的渗透性函数。我们进一步假设 TM阻止凝血酶与微血管内皮细胞上的PAR1相互作用，从而TM 通过其胞质结构域直接传递凝血酶的保护性信号传导功能。我们制备了一系列TM受体构建体、重组APC和凝血酶衍生物， TM 无效内皮细胞研究以下三个具体目标：目标 1 将研究假设凝血酶可以作为 TM 的真正配体，引发直接的细胞保护信号传导内皮细胞的反应。目标 2 将研究组成部分的机制凝血酶-TM-APC信号轴抑制高迁移率基团的促炎信号功能内皮细胞中的框 1 (HMGB1)。目标 3 将开发一种葡萄糖酸氯己定 (CG) 诱导的腹膜损伤模型并评价凝血酶的细胞保护功能机制该模型中的 TM-蛋白 C 信号轴。