Novel Redox-Associated Mechanisms Preventing Alcoholic Fatty Liver

项目总结 在美国，与肝硬变相关的死亡是主要死亡原因之一，酒精性脂肪肝(AFLD)是 这些死亡中的48%。尽管它对健康和经济产生了深远的影响，但AFLD的管理仍然是一个具有挑战性的前景 因为：(1)没有有效的诊断工具或生物标记物来评估个体对AFLD的易感性或 进展到更严重的临床条件，以及(Ii)在过去40年中没有开发出新的治疗实体。这个 这项研究项目的长期目标是阐明保护肝脏免受AFLD影响的氧化还原相关机制。 从而确定针对这种疾病的潜在的新的预防和/或治疗靶点。氧化应激在 AFLD的发病机制涉及多条途径。导致氧化应激发展的一个主要因素是 谷胱甘肽(GSH)是肝脏中含量最丰富的非蛋白硫醇。GCLM基因敲除(KO)的初步研究 小鼠表明，肝脏GSH缺乏~85%可保护小鼠免受慢性乙醇引起的脂肪变性 行政管理。这些小鼠还表现出：(I)代谢乙醇和乙醛的能力增强，(Ii)持续的氧化应激 和诱导核因子-红系2相关因子2(NRF2)靶基因，以及(Iii)重要的是持续激活AMP-2。 活化蛋白激酶(AMPK)途径及相关脂类代谢基因的变化。这项研究项目将利用 GCLM KO模型对我们的初步研究进行了扩展，并调查了我们的工作假设，即慢性GSH耗竭导致 氧化还原激活AMPK通路，作为中心环节，触发保护机制，防止AFLD。我们建议 目的：阐明氧化还原相关机制(S)在KO肝细胞中维持AMPK活性，鉴定AMPK催化亚基异构体。 KO肝细胞中参与乙醇相关代谢和应激反应的依赖通路及其作用 GSH缺乏对KO小鼠AMPK途径和保护性表型的肝脏和肝外影响的比较。这个 这些研究的结果将提供有关关键信号和代谢途径的重要机制信息 对酒精性肝损伤的保护作用。预计这些新的知识将揭示新的治疗靶点。 酒精性肝损伤的治疗，如酒精性肝病。此K01奖项将允许申请者获得先进的知识和 通过整合跨学科资源在酒精研究方面的研究能力。申请人已组装了一份 由一批杰出的导师和顾问组成的咨询委员会。Vasilis Vasiliou博士(主要导师)是一位 在酒精代谢和毒性领域公认的领导者。Wajahat Zafar Mehal博士(共同导师)是一位著名的肝病专家， 肝脏免疫学家，在酒精性和非酒精性脂肪性肝病方面有丰富的专业知识。Michael Harris Nathanson博士(联合- Mentor)是耶鲁大学肝脏中心主任，是研究肝病信号通路的世界领先者之一。这个 该计划将征集赵宏宇博士(顾问)的专业知识，他是统计学应用方面的世界领先者之一 方法采用分子生物学方法。咨询委员会的每个成员都有在学术上取得成功的令人敬畏的培训记录 独立科学家和他们将指导申请者的职业和学术发展以及实施 拟议的实验。

PROJECT SUMMARY Cirrhosis-associated death is one of the leading causes of death in the United States and alcoholic fatty liver disease (AFLD) accounts for 48% of these deaths1. Despite its profound health and economic impact, the management of AFLD remains a challenging prospect because: (i) there are no effective diagnostic tools or biomarkers to assess individual susceptibility to AFLD development or progression to more severe clinical conditions, and (ii) no new therapeutic entities have been developed in the past four decades. The long term goal of this research project is to elucidate the redox-associated mechanisms involved in protecting the liver against AFLD and thereby identify potential novel preventive and/or therapeutic targets against this disease. Oxidative stress plays a central role in many pathways involved in the pathogenesis of AFLD. A major factor contributing to the development of oxidative stress is the depletion of glutathione (GSH), the most abundant non-protein thiol in the liver. Our preliminary studies using GCLM knockout (KO) mice demonstrate that ~85% deficiency in hepatic GSH renders mice protected from steatosis induced by chronic ethanol administration. These mice also exhibit: (i) enhanced capacity to metabolize ethanol and acetaldehyde, (ii) persistent oxidative stress and induction of nuclear factor-erythroid 2–related factor 2 (NRF2) target genes, and (iii) importantly, sustained activation of the AMP- activated protein kinase (AMPK pathway and associated changes in lipid metabolizing genes. This research project will utilize the GCLM KO model to expand upon our preliminary studies and investigate our working hypothesis that chronic GSH depletion induces redox activation of the AMPK pathway that serves as the central link triggering protective mechanisms that prevent AFLD. We propose to: elucidate redox-associated mechanism(s) sustaining AMPK activation in KO hepatocytes, identify AMPK catalytic subunit isoform- dependent pathways involved in ethanol-associated metabolic and stress response in KO hepatocytes, and determine the contribution of hepatic versus extrahepatic effects of GSH deficiency in modulating AMPK pathway and the protective phenotype of KO mice. The findings from these studies will provide important mechanistic information regarding key signaling and metabolic pathways involved in protection against alcohol-induced liver damage. It is anticipated that such new knowledge will reveal novel therapeutic targets for the treatment of alcohol-induced liver injury, such as AFLD. This K01 Award will allow the applicant to acquire advanced knowledge and research competency in alcohol research through an integration of interdisciplinary resources. The applicant has assembled an advisory committee composed of an outstanding group of mentors and consultants. Dr. Vasilis Vasiliou (primary mentor) is a recognized leader in the field of ethanol metabolism and toxicity. Dr. Wajahat Zafar Mehal (co-mentor) is a renowned hepatologist and liver immunologist who has extensive expertise in alcoholic and non-alcoholic fatty liver disease. Dr. Michael Harris Nathanson (co- mentor) is the Director of the Yale Liver Center and is among the world leaders in studying signaling pathways in liver disease. The program will enlist the expertise of Dr. Hongyu Zhao (consultant), who is among the world leaders in the application of statistical methods in molecular biology. Each member of the advisory committee has a formidable record of training academically successful independent scientists and they will mentor the applicant's career and academic development as well as the implementation of proposed experiments.