Proteomics of Oxidant-Induced Mitochondrial Damage in an Animal Model of NASH

描述(由申请人提供)： 与肥胖和II型糖尿病相关的非酒精性脂肪性肝炎(NASH)正在成为慢性肝病最常见的病因之一。最近的证据支持NASH的“两次打击”假说，其中“第一次打击”涉及胰岛素抵抗导致肝细胞中脂肪的积累，随后是“第二次打击”，后者导致线粒体功能障碍以及氧化和亚硝化应激。因此，线粒体被认为在NASH的发展中起核心作用，能量代谢紊乱和过量的氧化剂产生有助于病理。氧化应激导致线粒体损伤的机制可能涉及线粒体蛋白质的翻译后修饰。线粒体蛋白质的氧化修饰将损害线粒体的生物能量学，并导致氧化剂的进一步增加，而氧化剂被认为在单纯性脂肪肝向NASH的发展过程中发挥关键作用。NASH中线粒体的这些变化背后的分子途径和靶点是未知的，也是本应用的重点。在阐明NASH的分子机制和开发靶向疗法方面，一个阻碍进展的因素是缺乏合适的实验动物模型。初步数据表明，喂饲高脂饮食的小鼠复制了伴有线粒体功能障碍的NASH的关键组织病理学特征。基于这些观察，我们推测，不能适应肥胖相关的代谢变化会导致线粒体氧化剂的过度产生，从而导致蛋白质的翻译后修饰，线粒体功能受损，以及NASH。这个探索性项目的第一个目标是描述和量化NASH的发展和线粒体损伤对高脂饮食的反应。这一模型的使用将有助于未来利用转基因和/或基因敲除小鼠模型来研究肝脏线粒体免受代谢应激的保护机制。第二个目的是研究NASH对肝脏线粒体蛋白质组的翻译后修饰。利用新的蛋白质组学方法完成这些研究，将能够识别NASH中代谢途径和/或蛋白质中的分子缺陷，这些缺陷可以被线粒体特异性治疗策略用于治疗和预防。与公共健康相关(外行描述)-研究报告称，肥胖在世界上的流行率不断上升。肥胖与严重的健康问题有关，包括心脏病和糖尿病，并被认为是发展为慢性非酒精相关肝病的最常见风险因素。这项应用中的研究很重要，因为它们将确定导致肥胖相关肝病的潜在因素，这将导致发现新的治疗方法。

DESCRIPTION (provided by applicant): Nonalcoholic steatohepatitis (NASH) associated with obesity and type II diabetes is emerging as one of the most common etiologies of chronic liver disease. Recent evidence supports a "two-hit" hypothesis for NASH with the "first-hit" involving the accumulation of fat in hepatocytes as a consequence of insulin resistance followed by the "second-hit", which entails mitochondrial dysfunction and oxidative and nitrosative stress. Thus, the mitochondrion is postulated to play a central role in the development of NASH with disrupted energy metabolism and excess oxidant production contributing to pathology. The mechanism by which oxidative stress leads to mitochondrial damage is likely to involve the post-translational modification of mitochondrial proteins. Oxidative modifications to mitochondrial proteins will impair mitochondrial bioenergetics and lead to further increases in oxidants, which are proposed to play a key role in the progression from simple fatty liver to NASH. The molecular pathways and targets that underlie these changes to mitochondria in NASH are unknown and the focus of this application. One factor that has hindered progress in elucidating the molecular mechanisms of NASH and development of targeted therapeutics is the lack of appropriate experimental animal models. Preliminary data indicate that feeding a high-fat diet to mice reproduces the key histopathologic features of NASH with mitochondrial dysfunction. Based on these observations it is hypothesized that the inability to adapt to the metabolic alterations associated with obesity results in the overproduction of mitochondrial oxidants, which leads to post-translational modification of proteins, impaired mitochondrial function, and NASH. The first aim of this exploratory project is to characterize and quantify the development of NASH and mitochondrial damage in response to a high-fat diet in mice. The use of this model will facilitate future studies with transgenic and/or knockout mouse models to investigate mechanisms of protection from metabolic stress in liver mitochondria. The second aim is to characterize post-translational modifications to the hepatic mitochondria proteome in response to NASH. Completion of these studies using novel proteomic approaches will enable the identification of the molecular defects in metabolic pathways and/or proteins in NASH that can be targeted by mitochondria-specific therapeutic strategies for treatment and prevention. Public health relevance (lay description) - Studies report an increasing prevalence of obesity in the world. Obesity is associated with serious health problems including heart disease and diabetes and is recognized as the most common risk factor for development of chronic non alcohol-related liver diseases. The studies in this application are important as they will identify the underlying factors responsible for obesity-related liver diseases, which will lead to the discovery of new treatments.