Regulation of Podocyte Function By Angiotensin II

描述（由申请人提供）：血管紧张素II （ANGII）对足细胞功能的调节：肾小球上皮细胞（足细胞）是肾小球滤过屏障的关键组成部分。足细胞损伤与蛋白尿和进行性肾功能丧失有关。大量证据表明肾小球损伤至少部分是由ANGII通过1型（AT1）和2型（AT2）受体亚型介导的。在肾小球内，ANGII受体在足细胞、内皮细胞和系膜细胞上表达。足细胞同时表达AT1和AT2受体。该受体系统的激活可能调节足细胞功能，并且在疾病状态下可能促进足细胞损伤。在许多细胞类型和动物模型中，ANGII的损伤促进作用是由AT1受体介导的，并可能受到AT2受体的负调节。在这方面，我们的初步数据表明，atl诱导的Gq α亚基（Galphaq）的激活在体外和体内介导ANGII的破坏作用中起着关键作用。基于这一初步工作，我们假设：AT1受体在肾小球足细胞中具有促进损伤的作用，这种作用主要通过galphaq依赖途径介导，并由AT2受体的相反作用调节。在拟议的研究中，我们将围绕三个具体目标来研究这一假设。首先，我们将利用药理学和分子生物学技术确定AT1和at2依赖性信号通路，这些信号通路调节培养足细胞的增殖、蛋白质合成和凋亡。基于我们的初步结果，在第二个具体目标#2中，我们将使用小鼠肾素启动子在足细胞中表达一种组成型活性形式的Galphaq (GalphaqQ>L)，从而确定Galphaq偶联途径在促进足细胞损伤中的作用。最后，我们将确定AT1受体在促进肾小球损伤特别是足细胞中的作用，以及足细胞AT2受体在调节AT1和galphaq依赖性肾损伤中的作用。在这些研究中，我们将使用转基因技术特异性地在肾小球足细胞上过表达AT1或AT2受体。在确定转基因在未处理小鼠中的效果后，我们将使用我们的转基因动物来研究以下因素引起的肾损害的严重程度：2. ANGII输注；2 .足细胞中AT2受体和GalphaqQ>L的共表达；足细胞损伤使用我们实验室开发的模型。这些研究应该为足细胞ANGII受体在疾病状态下促进肾小球损伤中的作用提供新的、重要的见解。了解调节肾小球疾病过程的生化机制可能为制定治疗决策提供理论依据，并可能导致肾小球疾病治疗新策略的发展。

DESCRIPTION (provided by applicant): Regulation of podocyte function by angiotensin II (ANGII): Glomerular epithelial cells (podocytes) are a critical component of the glomerular filtration barrier. Podocyte damage is associated with proteinuria and progressive loss of kidney function. A large body of evidence indicates that glomerular damage is mediated, at least in part, by ANGII through type 1 (AT1) and type 2 (AT2) receptor subtypes. Within the glomerulus, ANGII receptors are expressed on podocytes,endothelial cells and mesangial cells. Podocytes express both AT1 and AT2 receptors. Activation of this receptor system presumably regulates podocyte function and, in disease states, may promote podocyte injury. In many cell types as well as animal models, the injury promoting effects of ANGII are mediated by AT1 receptors and may be negatively modulated by AT2 receptors. In this regard, our preliminary data suggest that ATl-induced activation of the Gq alpha-subunit (Galphaq) plays a key role in mediating the damaging effects of ANGII both in vitro and in vivo. Based on this preliminary work, we hypothesized that: AT1 receptors have injury promoting effects in glomerular podocytes that are largely mediated through Galphaq-dependent pathways and are modulated by the opposing actions of AT2 receptors. In the proposed studies, we will investigate this hypothesis focusing on three specific aims. First, we will identify AT1- and AT2-dependent signaling pathways that regulate proliferation, protein synthesis, and apoptosis in cultured podocytes using pharmacological and molecular biological techniques. Based on our preliminary results, in second specific aim #2, we will define the role of Galphaq-coupled pathways in promoting podocyte injury in vivo by expressing a constitutively active form of Galphaq (GalphaqQ>L)in podocytes using the mouse nephrin promoter. Lastly, we will determine the role AT1 receptors in promoting glomerular injury specifically in podocytes as well as the role of podocyte AT2 receptors in modulating both AT1- and Galphaq-dependentrenal damage. In these studies, we will use transgenic techniques to overexpress either AT1 or AT2 receptors specifically on glomerular podocytes. After determining the effect of the transgenes in unmanipulated mice, we will use our transgenic animals to investigate the severity of renal damage induced by: 1. infusion of ANGII, 2. co-expression of AT2 receptors and GalphaqQ>L in podocytes, and 3. podocyte injury using a model developed in our laboratory. These studies should provide new, important insights into the role of podocyte ANGII receptors in promoting glomerular injury in disease states. Understanding the biochemical mechanisms that regulate glomerular disease processes may provide a rationale strategy for making therapeutic decisions and could lead to the development of novel strategies for treating glomerular diseases.