Insulin, Renal Sodium Transport and Blood Pressure

描述(申请人提供)：在动物和人类中，高胰岛素血症都与高血压有关。此外，胰岛素已被证明会导致钠在肾脏中滞留。在细胞培养中，胰岛素直接增加阿米洛利敏感钠通道(ENaC)的钠转运能力，该通道通常位于肾脏连接小管和集合管。钠在肾小管远端不适当的滞留可能导致细胞外液体积增大和高血压。然而，目前尚不清楚高胰岛素血症对钠平衡的直接和间接影响。此外，高胰岛素血症对特定的肾脏钠转运体和/或通道的表达和调节的影响还没有得到充分的研究。针对其中许多蛋白质的抗体直到最近才出现。在这项建议中，我们计划测试胰岛素和“胰岛素增敏剂”，如PPAR-伽马激动剂，作为一个整体，将增加肾脏中表达的几种关键钠转运蛋白的蛋白质丰度的总体假设。我们将通过半定量免疫印迹和免疫组织化学检测这些蛋白的丰度和细胞位置的慢性和急性变化。具体目标1，我们计划评估循环胰岛素水平升高对肾小管致密斑后部分两种主要顶端钠转运蛋白的相对丰度的直接影响：1)阿米洛利敏感的上皮钠通道(ENaC)；2)硫氮敏感的Na-CI协同转运体(NCC)。在具体目标2中，我们将评估ENaC亚基和NCC蛋白丰度的变化与大鼠血压变化的相关性，以及对转运体或通道选择性利尿剂，即阿米洛利和多硫叠氮的敏感性。在特定的目标3中，我们将解决在胰岛素输注中观察到的ENaC亚单位或NCC蛋白丰度增加的候选细胞机制。在特定的目标4中，我们将评估NCC和ENaC亚单位在响应急性和慢性胰岛素暴露时细胞分布的相对变化。最后，在特定的目标5中，我们将研究饮食中PPAR-γ激动剂对正常大鼠和胰岛素抵抗、肥胖的Zucker大鼠所有主要肾脏钠转运蛋白调节的影响。这些研究有望使我们对胰岛素在钠平衡中的作用有一个开明的理解。

DESCRIPTION (provided by applicant): Hyperinsulinemia has been linked to hypertension in both animals and humans. Furthermore, insulin has been shown to result in sodium retention by the kidney. In cell culture, insulin directly increases sodium transport capacity of the amiloride-sensitive sodium channel (ENaC), normally located in the renal connecting tubule and collecting duct. Inappropriate retention of sodium in the distal portion of the tubule could result in expanded extracellular fluid volume and hypertension. However, it is not clear what are direct versus indirect effects of hyperinsulinemia with regard to sodium balance. Furthermore, the impact of hyperinsulinemia on the expression and regulation of specific renal sodium transporters and/or channels has not been aqequately studied. Antibodies against many of these proteins have only recently become available. In this proposal, we plan to test the overall hypothesis that insulin and "insulin sensitizing agents", such as PPAR-gamma agonists, will, as a whole, increase the protein abundances of several critical sodium transport proteins expressed in the kidney. We will examine both chronic and acute changes in the abundance and cellular location of these proteins by semi-quantitative immunoblotting and immunohistochemistry. For specific aim 1, we plan to assess the direct effect of increased circulating insulin levels on the relative abundances of the two primary apical sodium transport proteins of the postmacula densa portion of the renal tubule: 1) the amiloride-sensitive epithelial sodium channel (ENaC); and 2) the thiazide-sensitive Na-CI cotransporter (NCC). In specific aim 2, we will evaluate the correlation of changes in ENaC subunit and NCC protein abundances with changes in rat blood pressure, and sensitivity to transporter or channel selective diuretics, i.e., amiloride and polythiazide. In specific aim 3, we will address candidate cellular mechanisms for the increase in ENaC subunit or NCC protein abundances observed with insulin infusion. In specific aim 4 we will evaluate relative changes in cellular distribution of both NCC and ENaC subunits in response to acute and chronic insulin exposure. Finally, in specific aim 5, we will investigate the impact of dietary PPAR-gamma agonists on the regulation of all of the major renal sodium transporter proteins in normal rats and insulin resistant, obese Zucker rats. These studies will, hopefully, provide us with an enlightened understanding of the role of insulin in sodium balance.