Renal Cortical Oxidative & Nitrosative Stress in IDDM

在大鼠1型糖尿病（DM）的早期阶段，超氧阴离子（O2 -）的加速产生通过过氧亚硝酸盐（ONOO）的形成促进了一氧化氮（NO）的降解，导致蛋白酪氨酸硝化和血管反应性的改变。糖尿病引起肾脏氧化应激的机制尚未确定。我们发现糖尿病大鼠的循环血管紧张素II （Angll）水平和肾皮质Angll 1型（AT 1）受体蛋白水平均显著升高。at1受体激活可引起O2。-通过NAD(P)H氧化酶活化生产。我们还发现了糖尿病大鼠肾皮质Hsp90水平降低的证据。新出现的证据表明，Hsp90是NO生成的重要步骤，至少在一定程度上起着限制O2的作用。-由NO合成酶（NOS）产生。在这些初步观察的基础上，我们假设NAD(P)H氧化酶激活和nos解偶联有助于DM患者肾皮质氧化和亚硝化应激，从而影响NO的生物利用度和作用，最终减少其对肾功能的影响。我们将利用stz诱导大鼠DM模型来评估：1)angll依赖性NAD(P)H氧化酶激活在肾皮质O2.-中的作用

During the early stage of type 1 diabetes mellitus (DM) in the rat, accelerated superoxide anion (O2 -) production fuels degradation of nitric oxide (NO) via peroxynitrite (ONOO.) formation, resulting in protein tyrosine nitration and alterations in vascular reactivity. The mechanisms engendering renal oxidative stress in DM have not been established. We have found that rats with DM exhibit significant increases in both circulating angiotensin II (Angll) levels and renal cortical levels of the Angll type 1 (AT 1) receptor protein. AT 1 receptor activation can provoke O2.- production via NAD(P)H oxidase activation. We have also found evidence of reduced Hsp90 levels in the renal cortex of rats with DM. Emerging evidence indicates that Hsp90 represents an essential step in NO production, functioning at least in part to limit O2.- production by NO synthase (NOS). On the basis of these preliminary observations, we hypothesize that NAD(P)H oxidase activation and NOS-uncoupling contribute to renal cortical oxidative and nitrosative stress in DM, with consequent effects on NO bioavailability and action that ultimately diminish its impact on renal function. The STZ-induced model of DM in the rat will be utilized to evaluate: 1) the role of Angll-dependent NAD(P)H oxidase activation in the renal cortical O2.- production during DM; 2) the contribution of Hsp90- or BH4-dependent NOS-uncouplin9 to renal cortical O2.- production during DM; and 3) the potential role of tyrosine nitration of guanylyl cyclase in altering in renal microvascular function during DM. Tin accord with RFA-DK-O2-O23, this application requests support for a collaborative partnership between two independent, established investigators - Pamela K. Carmines (a renal physiologist with expertise in the renal microvascular complications of DM)) and Jennifer S. Pollock (a protein chemist with particular expertise in NOS). Drs. Carmines and Pollock recently documented their success as collaborators in a joint publication providing much of the preliminary data for this project. Completion of the proposed work should position Dr. Pollock as an active diabetes researcher. Funding of this research partnership would also strengthen the ongoing efforts of Dr. Carmines' laboratory by facilitating access to expertise and technologies that are complementary to those currently available to her laboratory for studies of renal microvascular function and oxidative stress in DM. The proposed collaboration thus allows a biochemical, cellular, and functional approach that could unveil novel targets for therapeutic interventions that limit the development or consequences of oxidative and nitrosative stress during the early stage of DM, thereby delaying or preventing development of diabetic nephropathy.