PDl: Function in thrombus formation and antithrombotic action of inhibitors in m

PDl：m 中抑制剂的血栓形成功能和抗血栓作用

基本信息

批准号：
8401639
负责人：
Bruce Furie
金额：
$ 40.98万
依托单位：
BETH ISRAEL DEACONESS MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-09-01 至 2017-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8401639
关键词：
Active Sites Amino Acids Antibodies Antigens Antiphospholipid Syndrome Bacitracin Binding Blood Circulation Blood Coagulation Disorders Blood Platelets Blood Vessels Cells Cessation of life Cloning Complex Cysteine Deep Vein Thrombosis Disulfides Endoplasmic Reticulum Enzymes Extracellular Protein Family Fibrin Fibrinolytic Agents Generations Glycoproteins Human Imagery In Vitro Infusion procedures Injury Instruction Integrin beta3 Integrins Isomerase Lasers Link Location Mentors Methods Modeling Monitor Monoclonal Antibodies Mus Myocardial Infarction Nature Nobel Prize Oxidoreductase Paralysed Perinatal Pharmacology Platelet Activation Platelet aggregation Play Property Protein Biosynthesis Protein C Deficiency Protein Disulfide Isomerase Proteins Pulmonary Embolism Quercetin Rest Roentgen Rays Role Rutin Shapes Stroke Structure Sulfhydryl Compounds Surface TFPI Testing Thioredoxin Thromboembolism Thromboplastin Thrombosis Thrombus Time United States Venous cremaster muscle design disulfide bond factor V Leiden fetal in vivo inhibitor/antagonist intravital fluorescence microscopy intravital microscopy mortality mouse model mutant neonatal death novel prevent prototype yeast protein

项目摘要

PROJECT SUMMARY (See instructions): To determine whether extracellular PDl plays a role in thrombus formation, PDl expression, platelet accumulation, and fibrin generation were monitored in the blood vessels of mice following laser-induced arteriolar injury. A time-dependent increase in PDl antigen was observed in the thrombus following injury. Infusion of bacitracin or a blocking monoclonal antibody to PDl into the circulation completely inhibited platelet thrombus formation and fibrin generation. These results indicate that PDl is required in vivo for both fibrin generation and platelet thrombus formation. The objective in this project is to determine whether inhibitors of PDl represent a new class of antithrombotic agents. In Aim #1, the current project proposes to Identify the substrates of PDl that participate in the initiation of thrombus formation by the expression of mutant forms of thiol isomerases into experimental thrombi ex vivo and the trapping of disulfide-linked complexes of thiol isomerases and their substrates. In Aim #2, the xray crystal structure of human PDl and PDl in complex will be determined. These structures will include PDl and PDl bound to an anfi-PDI Fab, PDl complexed to quercetin 3-rutinoside and to quercetin, and PDl complexed to beta3 integrin. In Aim #3, antithrombotic properties of quercetin will be tested in mice. The pharmacology of quercetin as an antithrombotic agent will be initially studied by direct visualization of thrombus inhibifion using the laserinduced thrombus formation model in the cremaster muscle of the mouse using intravital microscopy. The inhibition of thrombosis-initiated fetal/neonatal demise by quercetin will be tested in two mouse models: perinatal thrombosis with homozygous Factor V Leiden and heterozygous TFPI deficiency, and consumptive coagulopathy with homozygous protein C deficiency. If quercetin is an active antithrombotic, we will determine whether quercetin, which inhibits both platelet aggregation and fibrin generation, is superior to standard antithrombotic agents. PDl inhibitors will be compared to conventional antithrombotic agents in mouse models of thrombosis. These will include death or paralysis induced by pulmonary embolism, and by intravital study of thrombus formation associated with acquired anti-phospholipid syndrome.

项目摘要（参见说明）：为了确定细胞外PD1是否在血栓形成中发挥作用，在激光诱导的小动脉损伤后的小鼠血管中监测PD1表达、血小板积累和纤维蛋白生成。在损伤后的血栓中观察到PD1抗原的时间依赖性增加。将杆菌肽或针对PD1的阻断性单克隆抗体输注至循环中完全抑制了血小板血栓形成和纤维蛋白产生。这些结果表明体内纤维蛋白生成和血小板血栓形成都需要PD1。该项目的目的是确定PD1抑制剂是否代表一类新的抗血栓剂。在目标#1中，当前项目提出通过将硫醇异构酶的突变形式表达到离体实验血栓中以及捕获硫醇异构酶及其二硫键连接的复合物来鉴定参与血栓形成起始的PD1底物。基材。在目标#2中，将确定人PD1和PD1复合物的X射线晶体结构。这些结构将包括PD1和与抗PDI Fab结合的PD1，PD1 与槲皮素3-芸香苷和槲皮素复合，并且PD1与β3整联蛋白复合。在目标#3中，将在小鼠中测试槲皮素的抗血栓特性。槲皮素作为抗血栓剂的药理学将首先通过使用活体显微镜在小鼠提睾肌中使用激光诱导血栓形成模型直接观察血栓抑制作用来研究。槲皮素对血栓引发的胎儿/新生儿死亡的抑制作用将在两种小鼠模型中进行测试：纯合因子 V Leiden 和杂合 TFPI 缺乏的围产期血栓形成，以及纯合蛋白 C 缺乏的消耗性凝血病。如果槲皮素是一种活性抗血栓药物，我们将确定抑制血小板聚集和纤维蛋白生成的槲皮素是否优于标准抗血栓药物。 PD1抑制剂将与常规抗血栓药物进行比较小鼠血栓形成模型。这些包括由肺栓塞以及与获得性抗磷脂综合征相关的血栓形成的活体研究引起的死亡或瘫痪。