ENaC Transport in Insulin Resistance: Role of Insulin & IGF-1 Receptors

描述(申请人提供)：高血压在美国影响6000万成年人，胰岛素抵抗综合症在这些人中大约有一半会导致一种盐敏感型高血压。在胰岛素抵抗综合征中，外周阻力引起的代偿性高胰岛素血症通过直接作用于肾小管而增加钠滞留，从而增加血压。在集合管的主细胞中表达的上皮性钠通道(ENaC)是这一过程的基础。然而，在胰岛素抵抗综合征中，传播胰岛素诱导的ENaC刺激的主细胞上的受体尚未建立。关于胰岛素受体在调节肾脏钠重吸收中的作用，实验研究得出了相互矛盾的结果。另一方面，一些研究表明，激活胰岛素样生长因子-1(IGF-1)受体可以通过ENaC增强钠的重吸收。这项建议的目的是确定在胰岛素抵抗模型中特异性调节ENaC活性的机制。R01补助金将为首席研究人员(PI)提供必要的资源，以检验这一假说，即外周胰岛素抵抗引起的高胰岛素血症通过激活胰岛素和/或IGF-1受体，启动下游信号级联，以及增强ENaC介导的钠转运而导致盐敏性高血压。为了验证这一假设，本文提出了两个具体目标。目的1确定胰岛素和/或IGF-1受体在胰岛素抵抗小鼠模型中对血压、钠重吸收和ENaC介导的钠转运的作用。使用野生型和主要细胞特异性胰岛素或IGF-1受体基因敲除小鼠，PI将在这些小鼠中诱导胰岛素抵抗，然后测量全身血压、有或没有ENaC特异性抑制剂时的尿钠排泄，以及钠在体外微灌流集合管中的转运。目的2确定胰岛素介导的胰岛素和/或IGF-1受体激活刺激ENaC介导的主细胞钠转运的细胞机制。PI将测量对照和胰岛素抵抗野生型小鼠的受体激活；测量胰岛素抵抗野生型和基因敲除小鼠的ENaC表达；并使用一种新技术从野生型和基因敲除小鼠分离主细胞，比较ENaC介导的电流和受体信号通路。拟议目标的预期结果是确定刺激ENaC所需的受体和受体后信号通路，从而确定胰岛素抵抗综合征中的盐敏感型高血压。我们预计，这些结果将通过确定适当的目标来预防和治疗与胰岛素抵抗相关的疾病的高血压，如肥胖症和2型糖尿病，从而显著推动这一领域的发展。这项研究的创新之处在于，我们将直接比较胰岛素和IGF-1受体，并使用新技术挑战单边的高胰岛素血症范例，简单地激活集合管中表达ENaC的主细胞中的胰岛素受体。

DESCRIPTION (provided by applicant): Hypertension affects 60 million adults in the United States, and the insulin resistance syndrome induces a salt- sensitive form of hypertension in approximately half of these individuals. In the insulin resistance syndrome compensatory hyperinsulinemia provoked by peripheral resistance increases sodium retention and hence, blood pressure via direct effects on the renal tubule. The epithelial sodium channel (ENaC) expressed in principal cells of the collecting duct is fundamental to this process. However, the receptors in principal cells which propagate insulin-induced ENaC stimulation in the insulin resistance syndrome are not established. Experimental studies have yielded conflicting results on the role of the insulin receptor in mediating renal sodium reabsorption. On the other hand, several studies have suggested that activation of the insulin-like growth factor-1 (IGF-1) receptor can enhance sodium reabsorption via ENaC. The objective in this proposal is to determine the mechanisms that specifically regulate ENaC activity in models of insulin resistance. The R01 Grant will provide the necessary resources for the principal investigator (PI) to test the hypothesis that hyperinsulinemia provoked by peripheral insulin resistance induces salt-sensitive hypertension via activation of the insulin and/or the IGF-1 receptor, initiation of downstream signaling cascades, and enhancement of ENaC- mediated sodium transport. To test this hypothesis, two specific aims are proposed. Aim 1 is to define the contribution of the insulin and/or IGF-1 receptor to blood pressure, sodium reabsorption, and ENaC-mediated sodium transport in mouse models of insulin resistance. Using wild-type and principal cell-specific insulin or IGF-1 receptor knockout mice, the PI will induce insulin resistance in these mice and then measure systemic blood pressure, urine sodium excretion in the presence and absence of an ENaC-specific inhibitor, and sodium transport across in vitro microperfused collecting duct. Aim 2 is to determine the cellular mechanisms by which insulin-mediated activation of the insulin and/or IGF-1 receptor stimulates ENaC-mediated sodium transport in principal cells. The PI will measure receptor activation in control vs. insulin-resistant wild-type mice; measure ENaC expression in insulin-resistant wild-type vs. knockout mice; and compare ENaC-mediated currents and receptor signaling pathways using a novel technique to isolate principal cells from wild-type and knockout mice. The expected outcomes of the proposed aims are to identify the receptors and post-receptor signaling pathways required for stimulation of ENaC and hence, salt-sensitive hypertension in the insulin resistance syndrome. We anticipate that these results will significantly advance the field by identifying appropriate targets for the prevention and treatment of hypertension in diseases associated with insulin resistance, such as obesity and Type 2 diabetes mellitus. The proposed research is innovative in that we will directly compare the insulin and IGF-1 receptor and employ novel techniques to challenge the unilateral paradigm of hyperinsulinemia simply activating the insulin receptor in ENaC-expressing principal cells of the collecting duct.