TGF BETA INDUCED DECREASES OF ENDOTHELIAL INTEGRITY

由于内皮细胞的变化导致细胞周转运增加 形状和细胞间隙的形成被认为是 许多疾病状态中溶质通量增加的主要途径 包括在肺动脉高压期间发现的血管重塑。 内皮细胞的形状和间隙的形成被认为是由 改变竞争性粘合剂平衡的信号转导途径 和收缩力。这项资助中概述的实验将 研究细胞间粘附是否减少， 细胞之间的相互作用，有助于细胞间隙的形成 加入细胞因子转化生长因子β后， （TGF-β）。TGF-β是一种细胞因子， 形状的融合单层，并伴随着增加 磁导率我假设以下机制来解释TGF-β 1。 β诱导的形状变化和细胞间隙的形成。转化生长因子 β将通过减少Ca++依赖性细胞间促进细胞分离 粘连这种Ca++依赖性细胞-细胞粘附的减少， 认为是由钙粘蛋白介导的，将是减少的结果， 钙粘蛋白亲嗜性结合的亲和力，而不是减少的结果 钙粘蛋白表面表达。然后，在发生细胞分离之后， 粘附连接解体。此外，虽然TGF-β诱导 细胞形状的改变依赖于基础向心张力，TGF-β 不会增加内皮细胞的收缩活性， 证明了介质，如凝血酶。这一假设将是 通过完成以下具体目标进行测试：1）确定Ca++是否 依赖性或Ca++非依赖性细胞-细胞粘附减少， 内皮细胞的TGF-β。钙粘蛋白表达的变化， 其被认为介导Ca++依赖性细胞粘附， 在用TGF-β治疗后测定。2)为了确定动力学 内皮细胞暴露后粘附连接解体 单层转化为TGF-β。3)为了确定蛋白质的磷酸化， 酪氨酸激酶的粘附连接是信号的一部分 TGF-β诱导内皮细胞增生的转导途径 单层渗透性4)为了确定这两种变化的贡献， 在内皮细胞收缩状态以及重组的 肌动蛋白细胞骨架转化为TGF-β介导的细胞形状变化。

Increases in pericellular transport due to changes in endothelial cell shape and the formation of intercellular gaps has been implicated as the major pathway for increased solute flux in a number of disease states including vascular remodeling found during pulmonary hypertension. Endothelial cell shape and gap formation is believed to be controlled by signal transduction pathways that alter the balance of competing adhesive and contractile forces. The experiments outlined in this grant will investigate if decreases in cell-cell adhesion, which tethers endothelial cells to one another, contributes to the formation of intercellular gaps following the addition of the Cytokine transforming growth factor beta, (TGF-beta). TGF-beta is a cytokine which is known alter endothelial cell shape in confluent monolayers and concomitantly increases the permeability. I hypothesize the following mechanism to explain the TGF- beta induced shape change and the formation of intercellular gaps. TGF- beta will promote cell separation by decreasing Ca++ dependent cell-cell adhesion. This decrease in Ca++ dependent cell-cell adhesion, which is believed to mediated by cadherins, will be the result of a decrease in the affinity of cadherin homophilic binding and not the result of a decrease in cadherin surface expression. Then, after cell separation has occurred, the adherens junction disassembles. Moreover, while the TGF-beta induced change in cell shape is dependent on basal centripetal tension, TGF-beta will not increase endothelial cell contractile activity as has been demonstrated for mediators such as thrombin. This hypothesis will be tested by completing the following specific aims: 1) To determine if Ca++ dependent or Ca++ independent cell-cell adhesion is decreased in endothelial cells by TGF-beta. Changes in the expression of cadherins, which are believed to mediate Ca++ dependent cell adhesion will also be determined following treatment with TGF-beta. 2) To determine the kinetics of adherens junction disassembly following exposure of endothelial cell monolayers to TGF-beta. 3) To determine if phosphorylation of proteins in the adherens junction by tyrosine kinases are part of the signal transduction pathway in TGF-beta induced increases in endothelial monolayer permeability. 4) To determine the contribution of both changes in endothelial cell contractile state as well as reorganization of the actin cytoskeleton to TGF-beta mediated changes in cell shape.