Microvascular Endothelial Cells in the Pathogenesis of Chronic Kidney Injury

描述（由申请人提供）： 我的长期目标是了解肾小管周围毛细血管如何促进肾纤维化，这是所有慢性肾脏疾病的最终共同途径。拟议的研究将调查机制，控制管周毛细血管通透性，炎症过程的主要组成部分，导致纤维化，重点是粘附连接蛋白血管内皮钙粘蛋白（VE-钙粘蛋白）和血管生成信号分子胎盘生长因子（PIGF）的调节。初步数据显示，在慢性肾损伤的单侧输尿管梗阻（UUO）模型中，VE-钙粘蛋白和PIGF的水平增加。VE-钙粘蛋白的上调可以控制纤维化，而PIGF似乎是促纤维化的。本研究旨在探讨慢性肾损伤过程中VE-钙粘蛋白的表达和功能变化，验证VE-钙粘蛋白表达改变可调节肾微血管通透性和纤维化的假说，并确定PIGF是否通过改变VE-钙粘蛋白的功能发挥作用：（1）阐明慢性肾损伤过程中肾脏VE-钙粘蛋白的表达、定位和分子相互作用的变化。UUO研究将通过免疫染色和免疫金电子显微镜研究VE-钙粘蛋白表达和细胞及亚细胞定位的变化，还将测量VE-钙粘蛋白磷酸化和与相关蛋白（包括连环蛋白）的结合。(2)验证VE-钙粘蛋白上调通过限制肾微血管通透性和间质炎症降低肾纤维化严重程度的假设。UUO研究将使用VE-钙粘蛋白杂合子（）小鼠和小干扰RNA（siRNA）来研究VE-钙粘蛋白表达水平如何影响慢性肾损伤的进展。(3)研究PlGF诱导VE-钙粘蛋白功能变化的假设，该功能增加微血管通透性并促进纤维化。使用PIGF -/-小鼠的UUO研究将确定VE-钙粘蛋白定位或磷酸化的变化是否与PIGF对肾纤维化的调节相关。细胞培养研究将测试PlGF是否直接影响VE-钙粘蛋白的表达和功能。 这项研究对公共卫生有很大的潜在影响，因为九分之一的美国人目前患有慢性肾脏疾病，50万人患有终末期疾病，并通过透析或肾移植进行终身治疗。因此，迫切需要慢性肾病的替代治疗。

DESCRIPTION (provided by applicant): My long-term objective is to understand how renal peritubular capillaries contribute to renal fibrosis, the final common pathway of all chronic kidney diseases. The proposed studies will investigate mechanisms that control peritubular capillary permeability, a major component of the inflammatory process leading to fibrosis, focusing on the adherens junction protein vascular endothelial cadherin (VE-cadherin) and its regulation by the angiogenic signaling molecule placental growth factor (PIGF). Preliminary data show that levels of VE-cadherin and PIGF are increased in the unilateral ureteral obstruction (UUO) model of chronic renal injury. Up-regulation of VE-cadherin may control fibrosis, whereas PIGF appears to be pro-fibrotic. The following aims will investigate how the expression and function of VE-cadherin change during chronic renal injury, test the hypothesis that altering VE-cadherin expression can modulate renal microvascular permeability and fibrosis, and determine whether PIGF acts by altering the function of VE-cadherin: (1) Delineate the changes in renal VE-cadherin expression, localization, and molecular interactions during chronic renal injury. UUO studies will investigate the changes in expression and cellular and subcellular localization of VE-cadherin by immunostaining and immuno-gold electron microscopy, and will also measure VE-cadherin phosphorylation and binding to associated proteins including catenins. (2) Test the hypothesis that up-regulation of VE- cadherin reduces the severity of renal fibrosis by limiting renal microvascular permeability and interstitial inflammation. UUO studies will use VE-cadherin heterozygote () mice and small interfering RNA (siRNA) to investigate how the level of VE-cadherin expression affects the progression of chronic renal injury. (3) Investigate the hypothesis that PIGF induces changes in VE-cadherin function that increase microvascular permeability and promote fibrosis. UUO studies using PIGF -/- mice will determine whether changes in VE- cadherin localization or phosphorylation are associated with modulation of renal fibrosis by PIGF. Cell culture studies will test whether PIGF directly affects VE-cadherin expression and function. This research has a large potential impact on public health, because one in nine Americans currently have chronic kidney disease, and 500,000 have end-stage disease and are on life-long therapy by dialysis or renal transplantation. Thus, alternative treatments for chronic kidney disease are urgently needed.