Alcohol-Induced Mitochondrial Dysfunction and the Hepatocyte Clock

 描述（由申请人提供）：酒精性肝病（ALD）是慢性饮酒导致发病和死亡的主要原因。尽管对ALD的医学和科学知识有所增加，但对ALD的分子事件仍然知之甚少。我们实验室以前的研究表明，慢性饮酒后肝脏线粒体生物能量功能严重受损，是肝细胞坏死的重要致病因素。然而，负责线粒体损伤的机制仍然未知。我们认为肝细胞分子生物钟的破坏是慢性酒精诱导的线粒体功能障碍的基础。已发表的研究表明，生物钟影响肝脏氧化代谢（β-氧化），线粒体生物合成的关键调节因子（Pgc 1a）在外周组织中表现出昼夜节律。重要的是，我们发现Pgc 1a，Pgc 1b和Pdk 4的节律在酒精喂养的小鼠肝脏中显着减弱，这表明线粒体代谢灵活性和对能量供应和需求的时间依赖性波动的反应能力在慢性酒精暴露的肝脏中严重受损。与此一致，我们观察到，转录因子，核呼吸因子1（Nrf 1），这是必不可少的整合核和线粒体编码的基因转录表现出昼夜节律，这是废除了遗传破坏的昼夜节律在肝脏中。此外，我们发现，细胞色素c氧化酶的活性，线粒体呼吸链的末端和限速酶，显示了一个强大的昼夜节律，在对照小鼠的肝线粒体， 长期酗酒导致抑郁总之，这些新数据强烈表明，昼夜节律钟在一天中调节线粒体功能，而慢性酒精对这些节律的破坏会损害肝脏的生物能量功能。因此，我们的长期目标是检验这一假设，即慢性酒精介导的肝时钟中断是慢性酒精消费者肝线粒体功能障碍的关键机制。为了验证这一假设，我们提出了以下目标。在目标1中，我们将确定肝细胞时钟是否直接调节线粒体生物能量学并影响肝脏中慢性酒精诱导的线粒体功能障碍。在目标2中，我们将确定酒精介导的肝细胞时钟改变和肝线粒体生物能量学损伤是否可以用靶向时钟的药物（ROR配体）治疗。该项目将通过使用肝细胞生物钟无功能的遗传小鼠模型，即肝细胞特异性BMAL 1敲除小鼠来促进。总之，这些创新的研究将提供关于生物钟调节细胞生物能量稳态的机制的新见解，并揭示慢性酒精介导的生物钟破坏导致肝脏线粒体功能障碍。此外，这项工作可能会为未来针对治疗ALD患者的时钟的药理学研究奠定基础。

 DESCRIPTION (provided by applicant): Alcoholic liver disease (ALD) is a major cause of morbidity and mortality from chronic alcohol consumption. Despite increased medical and scientific knowledge of ALD, the molecular events responsible for ALD remain poorly understood. Previous studies from our labs have shown that mitochondrial bioenergetic function is severely compromised in the liver following chronic alcohol consumption and is a significant causative factor for hepatocyte necrosis. However, the mechanisms responsible for mitochondrial damage remain unknown. We propose that disruption in the hepatocyte molecular circadian clock underpins chronic alcohol-induced mitochondrial dysfunction. Published studies have shown that the clock influences hepatic oxidative metabolism (β-oxidation) and that key regulators of mitochondrial biogenesis (Pgc1a) exhibit diurnal rhythms in peripheral tissues. Importantly, we have found that rhythms in Pgc1a, Pgc1b, and Pdk4 are significantly dampened in livers of alcohol-fed mice, suggesting that the ability of mitochondria to be metabolically flexible and responsive to time-of-day dependent fluctuations in energetic supply and demand is severely compromised in the chronic alcohol-exposed liver. In line with this, we observed that the transcription factor, nuclear respiratory factor 1 (Nrf1), which is essential for integration o nuclear and mitochondrial encoded gene transcription exhibits a diurnal rhythm, that is abolished by genetic disruption of the circadian clock in the liver. Moreover, we found that the activity of cytochrome c oxidase, the terminal and rate-limiting enzyme of the mitochondrial respiratory chain, displays a robust diurnal rhythm in liver mitochondria from control mice that is significantly depressed by chronic alcohol. Taken together, these new data strongly suggest that the circadian clock regulates mitochondrial function over the course of the day, and that disruption in these rhythms by chronic alcohol impairs hepatic bioenergetic function. Accordingly, our long-term goal is to test the hypothesis that chronic alcohol-mediated disruption in the liver clock is a critical mechanism underpinning hepatic mitochondrial dysfunction in the chronic alcohol consumer. To test this hypothesis we propose the following Aims. In Aim 1, we will determine whether the hepatocyte clock directly regulates mitochondrial bioenergetics and influences chronic alcohol-induced mitochondrial dysfunction in the liver. In Aim 2, we will determine whether alcohol-mediated alterations in the hepatocyte clock and impairments in liver mitochondrial bioenergetics can be treated with a drug (an ROR ligand) targeting the clock. This project will be facilitated by using a genetic mouse model in which the hepatocyte circadian clock is nonfunctional, namely the hepatocyte- specific BMAL1 knockout mouse. In summary, these innovative studies will provide new insight regarding the mechanisms by which the circadian clock regulates cellular bioenergetic homeostasis and reveal that chronic alcohol-mediated disruption to the clock contributes to mitochondrial dysfunction in the liver. Moreover, this work will likely lay a foundation for future pharmacological studies targeting the clock in treating ALD patients.