ENDOTHELIAL CELL PLASMINOGEN ACTIVATOR INHIBITOR TYPE 2

异常的血栓形成和溶解与 几种心血管疾病，包括动脉粥样硬化， 血栓栓塞和痔疮病症。 纤溶 因此，该系统在维护 止血平衡 很明显，许多纤溶酶 血浆的成分或者来源于内皮细胞 (ECs)（例如，组织型纤溶酶原激活剂）或与EC结合 (e.g.,尿激酶样纤溶酶原激活剂），以及EC- 因此，必须精确调节介导的纤维蛋白溶解。 的 意想不到的发现是，培养的EC也产生两种 免疫学上不同的纤溶酶原激活物抑制剂，包括 类型1（派-1）和类型2（PAL-2）都引入了先前的 我们对这个问题的理解， 系统，并提出了改变纤溶系统的可能性， 各种治疗后的细胞活性，或血液中 某些人类疾病，反映了其中任何一种变化， 抑制剂的 虽然派-1最近已被广泛研究， 关于派-2生产的信息很少 的EC。 本申请的长期目标是开发 准确理解派-2在人类疾病中的作用。 这项建议的具体目的是确定这一机构的作用， 抑制剂调节EC介导的纤溶。 派-2将是 足够数量的纯化，以允许发展 派-2的特异性抗体和免疫测定 活性和抗原。 这些技术将被采用，在 结合免疫荧光研究，以确定其 蜂窝位置（例如，存在于胞质溶胶、膜部分中，或 细胞外基质）在汇合的脐静脉EC中，以及在 其他类型的EC。 几个结构特征（即，分子 重量、等电点、碳水化合物的存在） 将建立由融合的人EC产生的细胞， 与从组织细胞淋巴瘤细胞中纯化的派-2相比， U-937线 这些研究将提供一个框架， 采用免疫组化法测定派-2合成和分泌基础速率， 免疫沉淀实验补充的测定 使用生物合成标记的派-2。 几个影响 因素（例如，细胞生长、细胞传代、血清、内毒素， 单核因子等）派-2的生产， 研究了 这些研究对于阐明 PAI-2在多种需要EC介导的过程中的作用 蛋白水解，如凝块溶解，组织修复，血管生成， 等等，并准确地定义派-2对改变的 纤维蛋白溶解状态

Abnormal thrombus formation and dissolution are associated with several cardiovascular diseases including atherosclerosis and both thromboembolic and hemorrhogic conditions. The fibrinolytic system thus assumes pivotal importance in maintaining the hemostatic balance. It is clear that many of the fibrinolytic components of plasma either originate from endothelial cells (ECs) (e.g., tissue-type plasminogen activator) or bind to ECs (e.g., urokinase-like plasminogen activator), and that EC- mediated fibrinolysis must therefore be precisely regulated. The unexpected finding that cultured ECs also produce two immunological distinct plasminogen activator inhibitors, including both type 1 (PAI-1) and type 2 (PAL-2), introduces a previously unsuspected level of complexity to our understanding of this system, and raises the possibility that the altered fibrinolytic activity of cells following various treatments, or of blood in certain human diseases, reflect changes in either one of these inhibitors. Although PAI-1 has been studied extensively recently, little information is available concerning the production of PAI-2 by ECs. The long term objective of this application is to develop an accurate understanding of the role of PAI-2 in human disease. The specific aim of this proposal is to define the role of this inhibitor in regulating EC-mediated fibrinolysis. PAI-2 will be purified in sufficient quantities to permit the development of specific antibodies and immunologic assays for both PAI-2 activity and antigen. These techniques will by employed, in conjunction with immunofluorescence studies, to define its cellular location (e.g., presence in cytosol, membrane fraction, or extracellular matrix) in confluent umbilical vein ECs, as well as in other types of ECs. Several structural features (i.e., molecular weight, isoelectric point, presence of carbohydrate) of PAI-2 produced by confluent human ECs will be established and compared to PAI-2 purified from the histocytic lymphoma cell line, U-937. These studies will provide a framework to determine the basal rate of PAI-2 synthesis and secretion using immunologic assays complemented by immunoprecipitation experiments employing biosynthetically labeled PAI-2. The effect of several factors (e.g., cell growth, cell passage, serum, endotoxin, monokines, etc.) on the production of PAI-2 will then be investigated. These studies are essential to delineate the role of PAI-2 in a variety of processes that require EC-mediated proteolysis, such as clot dissolution, tissue repair, angiogenesis, etc., and to accurately define the contribution of PAI-2 to altered fibrinolytic states.