PROTEIN S, C4BBP, AND THREE COAGULANT MODELS

PROTEIN S、C4BBP 和三种凝血剂模型

• 批准号: 2178912
• 负责人: FLETCHER B TAYLOR
• 金额: $ 37.36万
• 依托单位: OKLAHOMA MEDICAL RESEARCH FOUNDATION
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-12-01 至 1995-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2178912
• 关键词: Escherichia coli infections; baboons; binding proteins; blood coagulation; cell membrane; complement; complement pathway; cytokine; disease /disorder model; disseminated intravascular coagulation; epidermal growth factor; fibrinogen; flow cytometry; fluorescent dye /probe; inflammation; leukocytes; light microscopy; monoclonal antibody; platelets; protein C; protein S; protein structure function; scanning transmission electron microscopy; thrombin; thromboplastin; tissue /cell culture; tumor necrosis factor alpha; vascular endothelium; venous thrombosis

The main objective of these studies is to continue study of the influence of protein S and C4b-binding protein on the disseminated intravascular coagulant response in the lethal E.coli model of sepsis. Since these two factors also influence microvascular thrombotic and deep vein thrombotic responses to inflammatory stimuli, we plan to study the role of these two factors in these models as well. The central hypotheses is that protein S has two protective functions. The first is as an anticoagulant cofactor for activated protein C and the second is as an anticomplement cofactor for C4b-binding protein. Conversely, insufficiency of protein S relative to C4bBP can lead to an amplification of both the inflammatory and coagulant responses because they are linked and drive each other. Thus reduction of protein S by consumption or by neutralization by excess C4bBP can leave pro-inflammatory as well as procoagulant activity unchecked unless there is sufficient protein S to serve both its anticomplement and anticoagulant functions. Certain aspects of this hypothesis can be tested in vivo. Using the model of disseminated intravascular coagulant response to lethal E.coli we will intervene with C4bBP/protein S complex or protein S and monitor their effects on physiologic and laboratory parameters. These include C5b-9 and C4bc markers of complement system activation, thrombin-antithrombin complexes and fibrinogen markers of coagulant activation as well as cytokine and components of the protein C pathway (ie. free and bound protein S, protein C and C4bBP). We will determine if there is a pool of C4bBP/protein S complex bound to cell membranes with antibodies to the Gla domain of protein S by which these complexes might be anchored. We will monitor this by assaying for increases of the complex in plasma following infusion of antibodies. We also will study recruitment of radiolabeled complexes to these membranes before and after an inflammatory stimulus (ie. E.coli, TNF). We will study which domains of protein S participate with the appropriate antibodies. We will intervene with anti-C3b antibody and compare its anti -complement effects with those of the C4bBP/protein S complex. Using the models of the microvascular thrombotic response to sublethal E.coli and C4bBP and the deep vein thrombotic response to TNF and C4bBP, we will probe some of the same systems described above as well as features unique to each model. We will examine the role of tissue factor and platelets in the former and that of tissue factor and leukocytes in the latter. To give these studies more perspective, the pathologic responses of these two models will be divided into stages. This includes correlation of laboratory markers with light and electron microscopic studies of tissue. These studies should provide insights into the relationship between inflammation and coagulation in E.coli sepsis and offer new perspectives on diagnosis and treatment.

这些研究的主要目的是继续研究这些影响 S蛋白和C4b结合蛋白对弥漫性血管内皮细胞的影响 致死性大肠杆菌败血症模型中的凝血反应。因为这两个人 影响微血管血栓形成和深静脉血栓形成的因素 对炎症刺激的反应，我们计划研究这两个因素的作用 这些模型中的因素也是如此。 核心假设是，S蛋白具有两种保护功能。 第一种是作为活化蛋白C的抗凝剂辅因子，以及 二是作为C4b结合蛋白的抗补体辅因子。 相反，相对于C4bBP，S蛋白质不足会导致 炎症和凝血反应的放大，因为 它们是相互联系、相互推动的。从而降低了S的蛋白质含量 消耗或通过中和过量的C4bBP可以离开 未检查促炎和促凝血活性，除非 S是否有足够的蛋白质来提供其抗补体和 具有抗凝血功能。这一假设的某些方面可以是 在体内进行了测试。弥散性血管内凝血剂模型的应用 应对致死性大肠杆菌我们将干预C4bBP/蛋白S复合体 或蛋白质S，监测其对生理和实验室的影响 参数。这些包括补体系统的C5b-9和C4BC标志物 凝血酶-抗凝血酶复合体和纤维蛋白原标志物的活化 凝血剂激活与细胞因子和蛋白C组分 路径(即游离和结合蛋白S、蛋白C和C4bBP)。我们会 确定是否存在与细胞结合的C4bBP/蛋白S复合体 带有针对S蛋白GLA结构域的抗体的膜 复合体可能是锚定的。我们将通过化验来监测这一点 注射抗体后血浆中复合体的增加。我们 也将研究放射性标记的复合体在这些膜上的招募 炎症性刺激前后(即，大肠杆菌、肿瘤坏死因子)。我们会 研究S蛋白的哪些结构域与适当的 抗体。我们将用抗C3b抗体进行干预并比较其 抗补体作用与C4bBP/S蛋白复合体的作用相同。 利用微血管血栓对亚致死率的反应模型 大肠杆菌和C4bBP以及对肿瘤坏死因子和C4bBP的深静脉血栓反应， 我们将探讨上面描述的一些相同的系统以及 每个型号都有独特的功能。我们将研究组织因子的作用 前者为血小板，后者为组织因子和白细胞 后者。为了给这些研究更多的视角，病理学 这两个模型的响应将被划分为阶段。这包括 实验室标记与光镜和电子显微镜的相关性 对组织的研究。这些研究应该为我们提供对 大肠埃希菌败血症炎症与凝血的关系 为诊断和治疗提供新的视角。