Role of Insulin Receptors in the Kidney

胰岛素受体在肾脏中的作用

基本信息

批准号：
8293359
负责人：
Carolyn Mary Ecelbarger
金额：
$ 32.75万
依托单位：
GEORGETOWN UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-05-15 至 2014-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8293359
关键词：
Acute Blood Blood Pressure Breeding Cadherins Carrier Proteins Cells Centrifugation Defect Diabetes Mellitus Distal Distal convoluted renal tubule structure Dose Duct (organ) structure Epithelial Etiology Excretory function Fasting Financial compensation Generations Genotype Glucose Hour Hyperinsulinism Hypertension Immunohistochemistry Incidence Insulin Insulin Receptor Insulin Resistance Kidney Knock-out Lead Limb structure Maintenance Measures Metabolic syndrome Metabolism Molsidomine Mus Natriuresis Nitrates Nitric Oxide Nitric Oxide Donors Nitric Oxide Synthase Nitrites Obesity Organ Pancreas Pathology Pathway interactions Phosphatidylinositols Phosphorylation Phosphotransferases Plasma Play Production Proteins Receptor Signaling Regulation Renal tubule structure Role Saline Sgk protein Signal Pathway Signal Transduction Site Sodium Sodium Channel System Testing Thick Tissues Transgenic Mice Tubular formation Urine Vasodilator Agents Western Blotting Wild Type Mouse Zucker Rats benzamil blood pressure regulation cell type epithelial Na+ channel feeding inhibitor/antagonist insight mouse model public health relevance receptor receptor expression recombinase response saluretic transmission process uptake urinary

项目摘要

DESCRIPTION (provided by applicant): Obesity and diabetes lead to high circulating levels of insulin and high blood pressure (BP). However, mechanisms underlying these associations are in dire need of clarification. For example, in the kidney, insulin, through its own receptor, can lead to sodium retention, and activate the epithelial sodium channel (ENaC). Nonetheless, insulin can also increase the production of nitric oxide (NO), which has been shown to reduce the activity of ENaC. Thus, these two actions of insulin would be expected to have opposing effects on BP. To better elucidate the role of the insulin receptor in the kidney, we have developed an transgenic mouse with "knockout" (KO) or deletion of the insulin receptor (IR) in the distal tubule of the kidney (IRKO). These mice survive and grow normally, but have significantly higher basal BP. They are also impaired in their ability to rapidly excrete a NaCl load, as well as, have blunted rise in urinary nitrates plus nitrites (a urinary form of NO) excretion in the basal state and after insulin treatment. Thus our hypothesis is that IR in the kidney may have a previously unappreciated role in facilitating volume excretion and in the maintenance of normal BP. We further suggest these deficiencies arise directly as the result of impaired NO production with subsequent over-activity of ENaC. Three main aims are outlined below. Aim 1 is to determine whether reduced renal NOS activity in the distal tubule is a mechanistic determinant of the sodium-excretory defect and elevated BP in the IRKO mice. Renal nitric oxide synthase (NOS) activity will be measured in inner medulla, outer medulla, and cortex, as well as, microdissected proximal tubule, thick ascending limb, and collecting duct from fed and fasted IRKO and WT mice. We will also test the direct effects of insulin on NOS activity and NO levels in inner medullary collecting duct (IMCD) cultures. Finally, we will test whether molsidomine, an NO donor, restores NO levels, as well as normalizes BP and sodium excretion in IRKO mice. Aim 2 is to determine whether increased activity of the epithelial sodium channel (ENaC) is a mechanistic determinant of the sodium-excretory defect and elevated BP in the IRKO mice. For this aim, we will test whether benzamil, an antagonist of ENaC, abolishes differences in BP and natriuresis. ENaC regulation will be examined in native tissue and in primary IMCD cells. Finally, in Aim 3 we determine whether distinct IR signaling relating to ENaC activation and NO generation is altered in the IRKO mouse IMCD. Phosphorylation of critical proteins involved in IR signal transmission from the receptor to the activation of ENaC and nitric oxide synthase will be evaluated. These proteins include, but are not limited to, the insulin receptor substrate (IRS), phosphoinositide-3-kinase (PI-3K), and the serum and glucocorticoid-regulated kinase (SGK1). Overall, these studies will highly elucidate the role of insulin in the the distal tubule with regard to its role in blood control and sodium regulation. Moreover, they may provide insight into the pathology underlying hypertension associated with the metabolic syndrome. PUBLIC HEALTH RELEVANCE: Insulin resistance is associated with high circulating plasma levels of insulin, as well as, increased blood pressure; however, cellular mechanisms relating the two are not well understood. The kidney, a major regulator of blood pressure, expresses insulin receptors along the renal tubule; however, their role there is undefined. These studies are aimed at elucidating insulin's role in the distal tubule, which will hopefully allow for earlier and better targeted treatments in obesity and diabetes.

描述（由申请人提供）：肥胖和糖尿病会导致高循环胰岛素水平和高血压 (BP)。然而，这些关联背后的机制迫切需要澄清。例如，在肾脏中，胰岛素通过其自身的受体，可以导致钠潴留，并激活上皮钠通道（ENaC）。尽管如此，胰岛素也可以增加一氧化氮 (NO) 的产生，这已被证明可以降低 ENaC 的活性。因此，胰岛素的这两种作用预计会对血压产生相反的影响。为了更好地阐明胰岛素受体在肾脏中的作用，我们开发了一种转基因小鼠，其在肾脏远端小管中“敲除”（KO）或删除了胰岛素受体（IR）（IRKO）。这些小鼠存活并正常生长，但基础血压显着升高。它们快速排泄 NaCl 负荷的能力也受到损害，并且在基础状态和胰岛素治疗后尿硝酸盐和亚硝酸盐（一氧化氮的一种尿形式）排泄的上升减弱。因此，我们的假设是，肾脏中的 IR 在促进容量排泄和维持正常血压方面可能具有以前未被认识到的作用。我们进一步认为，这些缺陷是由于 NO 生成受损以及随后 ENaC 过度活跃而直接产生的。下面概述了三个主要目标。目的 1 是确定远端小管肾 NOS 活性降低是否是 IRKO 小鼠钠排泄缺陷和血压升高的机制决定因素。将测量喂食和禁食 IRKO 和 WT 小鼠的内髓质、外髓质和皮质，以及显微解剖的近端小管、粗升肢和集合管中的肾一氧化氮合酶 (NOS) 活性。我们还将测试胰岛素对内髓集合管 (IMCD) 培养物中 NOS 活性和 NO 水平的直接影响。最后，我们将测试一氧化氮供体吗多明是否可以恢复 IRKO 小鼠的一氧化氮水平，并使血压和钠排泄正常化。目标 2 是确定上皮钠通道 (ENaC) 活性的增加是否是 IRKO 小鼠钠排泄缺陷和血压升高的机制决定因素。为此，我们将测试 ENaC 拮抗剂苯扎米尔是否能消除血压和尿钠排泄的差异。将在天然组织和原代 IMCD 细胞中检查 ENaC 调节。最后，在目标 3 中，我们确定 IRKO 小鼠 IMCD 中与 ENaC 激活和 NO 生成相关的独特 IR 信号传导是否发生改变。将评估参与 IR 信号从受体传递到 ENaC 和一氧化氮合酶激活的关键蛋白质的磷酸化。这些蛋白质包括但不限于胰岛素受体底物 (IRS)、磷酸肌醇 3-激酶 (PI-3K) 以及血清和糖皮质激素调节激酶 (SGK1)。总的来说，这些研究将高度阐明胰岛素在远端小管中在血液控制和钠调节中的作用。此外，它们还可以深入了解与代谢综合征相关的高血压的病理学。公共卫生相关性：胰岛素抵抗与高循环血浆胰岛素水平以及血压升高有关；然而，与这两者相关的细胞机制尚不清楚。肾脏是血压的主要调节者，沿着肾小管表达胰岛素受体。然而，他们的角色尚未明确。这些研究旨在阐明胰岛素在远端小管中的作用，这有望为肥胖和糖尿病提供更早、更好的针对性治疗。