Ethanol toxicity and NO-dependent mitochondrial damage

描述（由申请人提供）：涉及活性氧（ROS） 高浓度乙醇的肝毒性。若干机制 例如线粒体GSH浓度降低、抑制 呼吸和对缺氧的敏感性增强。重点 这项建议的目的是研究乙醇对肝线粒体的影响， 通过一氧化氮（NO）、活性氮（RNS） 和ROS。先前的研究和初步数据表明， NO的作用可以在线粒体中证明， 呼吸功能和导致细胞色素c释放的过程。 虽然许多由乙醇引起的线粒体缺陷也可以被认为是 由RNS引起的机械联系尚未被研究。概念发展 NO在乙醇上转化为线粒体毒素 通过与ROS反应消耗。支持这一假设的主要发现 1）iNOS在体内乙醇消耗时被诱导，其产物NO， 线粒体呼吸的有效调节剂2）这与 相反，增加酪氨酸硝化-RNS的标志物3）NO抑制 细胞色素c从线粒体释放，而RNS如过氧亚硝酸盐， 促进这种促凋亡因子的释放4）慢性乙醇消耗 导致肝细胞对缺氧应激更敏感。这些数据 导致了一种假设，即导致酒精的关键机制 肝毒性是通过线粒体功能NO依赖性修饰。 这一概念将通过在野外追求以下具体目标来检验 型和INOS敲除消耗乙醇的小鼠：1.确定影响 慢性饮酒增加NO对线粒体呼吸的影响， 抗氧化能力和ROS/RNS形成。2.确定慢性的影响 饮酒对细胞色素c释放和线粒体蛋白的影响 在分离的细胞器和肝细胞中的修饰。3.确定 NO依赖性对线粒体蛋白质合成的影响及 慢性消耗乙醇后肝细胞缺氧应激。

DESCRIPTION (provided by applicant): Reactive oxygen species (ROS) are involved in the hepatotoxicity of high concentrations of ethanol. A number of mechanisms such as decreased concentrations of mitochondrial GSH, inhibition of respiration and enhanced susceptibility to hypoxia have been invoked. The focus of this proposal is to examine the effects of ethanol on hepatic mitochondrial function mediated through nitric oxide (NO), reactive nitrogen species (RNS) and ROS. Previous studies and preliminary data have shown that a biphasic effect of NO can be demonstrated in mitochondria in terms of control of both respiratory function and the processes leading to cytochrome c release. Although many of the mitochondrial defects elicited by ethanol can also be caused by RNS a mechanistic link has not been examined. The concept developed in this proposal is that NO is converted to a mitochondrial toxin on ethanol consumption by reaction with ROS. Key findings in support of this hypothesis are 1) iNOS is induced on ethanol consumption in vivo and its product, NO, is a potent regulator of mitochondrial respiration 2) this is associated with increased tyrosine nitration-a marker of RNS 3) NO, in contrast, inhibits cytochrome c release from mitochondria, whereas RNS such as peroxynitrite, promotes release of this pro-apoptotic factor 4) chronic ethanol consumption leads to greater sensitivity of the hepatocyte to hypoxic stress. These data have led to the hypothesis that a critical mechanism contributing to alcohol hepatotoxicity is through NO-dependent modification of mitochondrial function. This concept will be tested by pursuit of the following Specific Aims in wild type and INOS knock out mice consuming ethanol: 1. Determine the effects of increased NO by chronic alcohol consumption on mitochondrial respiration, antioxidant capacity and ROS/RNS formation. 2. Determine the effect of chronic alcohol consumption on cytochrome c release and mitochondrial protein modification in isolated organelle and hepatocytes. 3. Determine the NO-dependent effects on mitochondrial protein synthesis and the response of hepatocytes to hypoxic stress after chronic consumption of ethanol.