Mechanisms of In Vivo Ethanol-Induced Mitochondrial Depolarization

该项目研究酒精如何导致线粒体功能障碍和肝脏疾病。它将填补了解酒精性肝病的关键空白。 理由：该项目解决了生物医学研究和公共卫生的一个重要问题。酒精性肝病（ALD）影响美国超过250万人，每年治疗和生产力损失超过15亿美元。乙醇损害肝脏的机制还远未明确，在开发有效的治疗方法之前，我们必须填补知识上的巨大空白。该研究的重点是了解乙醇导致线粒体功能障碍的机制，在活体动物中使用最先进的活体多光子/共聚焦显微镜技术，并有望填补理解ALD机制的关键空白。活体共聚焦/多光子显微镜是一种新技术，将在拟议的研究中开发和优化，作为一个强大的新工具，在活体动物的细胞器和分子水平上研究乙醇毒性。由于对酒精的生物反应的一些重要特征不能在体外系统中重现，因此这种技术是必不可少的。拟议的项目将立即设立3个新的全职职位，包括2名博士后研究员和1名技术员，此外还将部分支持另外两个职位。总共3.3个全职员工将通过ARRA资金创建或保留。这些招聘预计将迅速，没有困难，从候选人名单，PI已经有。已提交详细预算，并将其列入这一请求。在两年的ARRA支持结束时，调查人员将完成前两个具体目标。首先，他们将表征与SIAM和肝脂肪变性相关的乙醇处理后体内线粒体去极化的剂量依赖性、时间过程和恢复。其次，他们将评估乙醇脱氢酶（ADH），微粒体乙醇氧化系统（MEOS）和乙醛脱氢酶（ALDH）在乙醇诱导的线粒体去极化中的作用。在ARRA的支持下，2年的工作将开发和应用活体多光子显微镜的新技术，并为进一步研究阐明线粒体功能障碍导致ALD病理生理学的机制奠定坚实的基础。在为期2年的ARRA资助期间，将向国家和国际会议提交4-6篇摘要，并向同行评审期刊提交2-4篇论文。这是响应RFA-AA-08-002（线粒体在酒精诱导的组织损伤中的作用）的应用程序。该申请的优先级得分（179）是非资助RFA申请中最好的。R 01的RFA支付线为171，R21的RFA支付线为183。PI可以解决审查人员的所有主要问题。但是，由于这是一份RFA申请，PI不允许回应批评、修改申请和重新提交。

Summary: The project studies how alcohol causes mitochondrial dysfunction and liver disease. It will fill a critical gap in understanding alcoholic liver disease. Justification: The project addresses a problem important for biomedical research and public health. Alcoholic liver disease (ALD) affects more than 2.5 million people in the U.S. and costs over $1.5 billion/year for treatment and lost productivity. Mechanisms by which ethanol damages the liver are far from clear, and striking gaps in our knowledge must be filled before effective therapies can be developed. The study focuses on understanding the mechanisms by which ethanol causes mitochondrial dysfunction in living animals using state-of-the-art intravital multiphoton/confocal microscopy techniques and is expected to fill a critical gap in understanding the mechanisms of ALD. Intravital confocal/multiphoton microscopy is a novel technology that will be developed and optimized in the proposed study as a powerful new tool to investigate ethanol toxicity at organelle and molecular levels in living animals. Since ,some important features of the biological response to alcohol cannot be reproduced in in vitro systems, such technology is essential. The proposed project will create 3 new full-time positions immediately, including 2 postdoctoral fellows and one technician, in addition to partial support of two other positions. Total 3.3 FTEs will be created or retained through this ARRA funding. These hires are expected to be made quickly and without difficulty from a candidate list that PI already has. A detailed budget has been submitted and included with this request. At the end of two years of ARRA support, the investigators will complete the first 2 specific aims. First, they will characterize the dose-dependency, time course and recovery of mitochondrial depolarization in vivo after ethanol treatment in relation to SIAM and hepatic steatosis. Second, they will evaluate the role of alcohol dehydrogense (ADH), the microsomal ethanoloxidizing system (MEOS) and acetaldehyde dehydrogenase (ALDH) in ethanol-induced mitochondrial depolarization. The work during 2 years of ARRA support will develop and apply new technologies of intravital multiphoton microscopy and lay a solid foundation for further research to elucidate the mechanisms by which mitochondrial dysfunction contributes to the pathophysiology of ALD. During the 2-Year ARRA funding period, 4-6 abstracts will be submitted to national and international meetings and 2-4 papers will be submitted to peer-reviewed journals. This is an application in response to RFA-AA-08-002 (The Role Of Mitochondria In Alcohol-Induced Tissue Injury). The priority score of this application (179) is the best among those non-funded RFA applications. The RFA payline for R01 was 171 The RFA payline for R21 was 183. The PI can address all major concerns of the reviewers. However, because this is a RFA application, the PI is not allowed to respond to the critiques, revise the application, and resubmit.