Generating Mouse Mutants with Diabetic Nephropathy

描述（由申请人提供）： AMDCC的目标是开发糖尿病并发症的动物模型，忠实地再现在人类中观察到的糖尿病并发症。该建议将提供一种糖尿病肾病（DN）小鼠模型，重点是两个关键的保护内皮通路：eNOS和前列环素合酶（PGIS）。这些途径不仅在内皮细胞内共定位，而且它们的活性也通过过氧硝酸盐的细胞水平在生物化学上相互关联，并且两者都涉及人类糖尿病肾病、神经病、视网膜病和大血管疾病。在上一个资助周期中，范德比尔特AMDCC研究中心调查了小鼠糖尿病肾病（DN）的遗传基础。这些研究确定系统性eNOS缺失是一种关键的遗传修饰剂，可将C57 BL/6小鼠从耐药品系转化为对DN易感的品系。内皮型一氧化氮合酶（eNOS或NOSIII）的遗传破坏，而不是ApoE或LDLR与C57 BL/6中DN的显著加速相关，其特征不仅在于强烈的白蛋白尿，而且还在于显著的系膜血管溶解和扩张伴肾功能（GFR）降低。eNOS作为人糖尿病肾病风险的临床相关调节剂的参与得到了临床研究的支持，临床研究显示具有eNOS Glu 298 Asp多态性的糖尿病患者不仅表现出降低的eNOS活性，而且还表现出加速的肾衰竭风险{Noiri，2002 #6975; Shin Shin，2004 #8934}。越来越多的证据表明，内皮功能障碍与糖尿病并发症，特别是肾病和大血管疾病的发病机制有关{Schalkwijk，2005 #9302}。类似地，在前列环素合酶（PGIS）中也发现了多态性，尽管它们在糖尿病肾病进展中的具体作用尚未确定， 本研究的目的有两个：目的1：明确内皮型一氧化氮合酶在糖尿病肾病发生发展中的作用;目的2：明确内皮型前列环素合酶在糖尿病肾病发生发展中的作用。为了实现这一点，我们将产生条件性靶向（floxed）eNOS和PGIS等位基因，并将这些小鼠与Tie 2 mERCre小鼠杂交，允许特异性地从内皮暂时控制这些等位基因的缺失。这些研究应允许解剖这些生化途径在小鼠糖尿病肾病进展中的作用。

DESCRIPTION (provided by applicant): The goal of the AMDCC is to develop animal models of diabetic complications that faithfully reproduce diabetic complications observed in humans. This proposal will provide a model of mouse model of diabetic nephropathy (DN) focusing on two key protective endothelial pathways: eNOS and prostacyclin synthase (PGIS). These pathways are not only co-localized within the endothelial cells but their activity is also biochemically interrelated through cellular levels of peroxynitrate, and both that have been implicated in human diabetic nephropathy, neuropathy, retinopathy and macrovascular disease. In the previous funding cycle, the Vanderbilt AMDCC site investigated the genetic underpinnings of diabetic nephropathy (DN) of mice. Those studies identified systemic eNOS deletion as a critical genetic modifier that converts C57BL/6 mice from a resistant strain to one that is susceptible to DN. Genetic disruption of endothelial nitric oxide synthase (eNOS or NOSIII), but not ApoE or LDLR was associated with a marked acceleration of DN in C57BL/6, not only characterized by a robust albuminuria, but also by dramatic mesangiolysis and expansion with decrease renal function (GFR). The involvement of eNOS as a clinically relevant modifier for risk of human diabetic nephropathy is bolstered by clinical studies showing that diabetics with an eNOS Glu298Asp polymorphism not only exhibit decreased eNOS activity but also an accelerated risk of renal failure {Noiri, 2002 #6975; Shin Shin, 2004 #8934}. Accumulating evidence implicates endothelial dysfunction in the pathogenesis of diabetic complications, particularly nephropathy and macrovascular disease {Schalkwijk, 2005 #9302}. Similarly polymorphisms have been identified in prostacyclin synthase (PGIS), although their specific role in the progression of diabetic nephropathy has not been established, The present proposal has two specific aims: Aim 1 will determine the role of endothelial eNOS in the progression of diabetic nephropathy; while To determine the role of Endothelial prostacyclin synthase in the progression of diabetic nephropathy. To achieve this we will generate conditionally targeted (floxed) eNOS and PGIS alleles, and cross these mice with a Tie2mERCre mouse, allowing temporally controlled deletion of these alleles specifically from the endothelium. These studies should allow the dissection of the role of these biochemical pathways in the progression of diabetic nephropathy in mice.