A Novel Form of Tissue Factor and Cardiovascular Disease

描述(申请人提供)：组织因子(TF)是一种跨膜糖蛋白，调节止血，被认为在介导动脉血栓形成中发挥关键作用。在动物模型中，转铁蛋白还介导动脉损伤的内膜反应和缺血再灌注损伤所致的损伤程度。Tf被认为是以单一形式存在的，其活性依赖于它在细胞膜上的插入。最近在组织和循环中存在的微粒中发现了转铁蛋白。此外，还发现了一种选择性剪接形式的转铁蛋白(AsTF)，它不包括外显子5，并且含有一个框架移位，它产生一个唯一的没有跨膜结构域的C-末端。AsTF的活性需要暴露在脂质中，而不是插入到脂质双层中。AsTF已经从血液中分离出来，是体内和体外血栓的主要成分。研究人员推测，asTF在调节动脉血栓形成、调节正常心脏止血、调节血管内膜增生和对缺血-再灌注的反应中发挥关键作用。这项建议将研究asTF和全长TF(Fltf)在细胞培养和基因工程小鼠中的调节和生物学作用(S)。目的1建立asTF在平滑肌细胞、巨噬细胞、内皮细胞和心肌细胞中表达的时间进程，并确定选择性剪接是否表现出激动剂、组织和发育特异性。它还将确定，作为一种可溶性分子，asTF是否与血管细胞特异结合，并表现出与其启动凝血能力不同的激动剂特性。目的2确定asTF在止血、血栓形成和TF介导的损伤反应中的作用。将产生一种或两种形式的转铁蛋白受内源性转铁蛋白启动子调控的小鼠。研究人员将检验如下假设：1)fltf对于胚胎存活至关重要；2)astf对于动脉损伤部位的显著血栓形成至关重要；3)astf在调节动脉损伤的内膜反应方面发挥关键作用；4)fltf在介导正常止血方面发挥主导作用，但astf在心肌毛细血管水平上发挥重要作用；5)astf和fltf在介导缺血再灌注损伤方面发挥互补作用；以及6)astf介导正常的细胞功能，包括生长、迁移和黏附。这些研究将为一种可能在心血管疾病中发挥关键作用的新型循环转铁蛋白提供见解。

DESCRIPTION (provided by applicant): Tissue factor (TF) is a transmembrane glycoprotein that regulates hemostasis and is thought to play a critical role in mediating arterial thrombosis. In animal models, TF also mediates the intimal response to arterial injury and the extent of damage resulting from ischemia-reperfusion injury. TF was thought to exist as a single form whose activity was dependent upon its insertion in the cell membrane. TF has recently been found in microparticles that are present in tissues and in the circulation. In addition, an alternatively spliced form of TF (asTF) has been identified that excludes exon 5 and contains a frame shift that generates a unique C-terminus without a transmembrane domain. asTF activity requires exposure to lipids, but not insertion into a lipid bilayer. asTF has been isolated from blood and is a major component of ex vivo and in vivo thrombi. The investigators hypothesize that asTF plays critical roles in regulating arterial thrombosis, in mediating normal cardiac hemostasis, and in mediating intimal hyperplasia and the response to ischemia-reperfusion. This proposal will examine the regulation and biologic role(s) of asTF and full-length TF (flTF) in cell culture and in genetically engineered mice. Aim 1 will establish the time course of asTF expression in smooth muscle cells, macrophages, endothelial cells, and cardiocytes and determine whether alternative splicing exhibits agonist, tissue and developmental specificity. It will also determine whether, as a soluble molecule, asTF binds specifically to vascular cells and exhibits agonist properties distinct from its ability to initiate coagulation. Aim 2 will establish the role of asTF in hemostasis, thrombosis, and in TF-mediated responses to injury. Mice will be generated in which one or both forms of TF are regulated by the endogenous TF promoter. The investigators will test the hypotheses that: 1) flTF is critical for embryonic survival; 2) asTF is critical for generating significant thrombosis at sites of arterial injury; 3) asTF plays a pivotal role in regulating the intimal response to arterial injury; 4) flTF plays the dominant role in mediating normal hemostasis, but asTF is important in mediating hemostasis at the level of the myocardial capillaries; 5) asTF and flTF play complementary roles in mediating ischemia-reperfusion injury; and 6) asTF mediates normal cell functions, including growth, migration, and adhesion. These studies will provide insights into a novel form of circulating TF that may play a key role in cardiovascular disease.