Circadian Clock Regulation of Alcoholic Liver Disease

描述（由申请人提供）： 摘要酒精性脂肪肝（ALD）是美国慢性肝病的主要原因之一。特别是，由于肥胖、糖尿病、创伤后应激障碍（PTSD）和药物滥用与酒精消费的增加，ALD正在成为退伍军人中普遍存在的健康问题。昼夜节律紊乱是肝脏和代谢疾病发展的危险因素。然而，肝脏生物钟和ALD之间的关联仍然是一个有趣但未探索的研究领域。此外，最先进的分析工具，如转录组学（RNA-seq）和代谢组学（GC/MS），尚未很好地用于研究ALD。 核受体是分子生物钟机制和代谢疾病之间的关键联系人。我的实验室率先在肝胆汁酸和脂质代谢中的小异二聚体伴侣（SHP，Nrob 2）的体内功能方面取得了重大发现。这项研究是基于我们新发现的SHP在整合生物钟、代谢网络和肝脏ER应激信号中的作用。 总体目标是在酒精介导的内质网应激反应和脂肪变性中建立一个完整的分子昼夜节律网络。核心假设是，酒精狂欢改变了肝脏中有节奏的SHP表达，这反过来又调节了Npas 2和ER应激信号传导和脂肪生成途径中的下游靶点。具体来说，我们将阐明酒精，SHP和Npas 2如何协调控制CHOP和SREBP 1c基因转录，蛋白质稳定性和活性，并建立酒精对整个肝脏转录组和代谢组的调节。目标一：阐明SHP和酒精介导的控制ER应激信号传导的分子昼夜节律途径;目的#2：确定肝脏特异性Npas 2敲低对由SHP和酒精狂欢介导的ER应激反应和脂肪变性的影响;以及目的#3：建立由SHP和酒精狂欢联锁的肝脏昼夜节律转录组和代谢组网络。除了我们独特的体内小鼠（SHP-/-）和体外细胞模型外，还将产生新的肝脏Npas 2敲除小鼠并用于研究。我们的方法充分利用分子生物学、系统生物学和全基因组高通量分析（RNA-seq和GC/MS）来解决与退伍军人健康直接相关的重要科学问题。 本研究在概念上和实验方法的实施上都具有创新性。计划的实验策略是全面的，但可行的。该项目将首次揭示酒精和肝脏昼夜节律钟之间的新型相互作用，以控制ER应激反应和肝脏脂质代谢。这项研究的发现很可能使ALD研究取得实质性进展，使该项目具有很高的临床意义和转化价值。

DESCRIPTION (provided by applicant): SUMMARY Alcoholic fatty liver disease (ALD) is one of the primary causes of chronic liver diseases in the United States. In particular, ALD is becoming a prevalent health issue among veterans due to an increase in obesity, diabetes, post-traumatic stress disorder (PTSD), and drug abuse in conjunction with alcohol consumption. Disturbance of circadian rhythmicity is a risk factor for the development of liver and metabolic diseases. However, the association between the liver circadian clock and ALD remains an intriguing but unexplored research area. In addition, the most advanced analytical tools, such as transcriptomics (RNA- seq) and metabolomics (GC/MS), have not been well utilized to study ALD. Nuclear receptors are key liaisons between the molecular clock machinery and metabolic diseases. My laboratory has pioneered major discoveries in the in vivo function of small heterodimer partner (SHP, Nrob2) in hepatic bile acid and lipid metabolism. The proposed study is based on our newly discovered role of SHP in integrating the circadian clock, metabolic network, and the ER stress signaling in the liver. The overall objective is to establish an integrated molecular circadian network in alcohol mediated ER stress response and steatosis. The central hypothesis is that ethanol-binge alters rhythmic SHP expression in the liver, which in turn modulates Npas2 and the downstream targets in the ER stress signaling and lipogenesis pathways. Specifically, we will elucidate how alcohol, SHP and Npas2 coordinately control CHOP and SREBP1c gene transcription, protein stability, and activity, and establish the modulation by alcohol of the entie liver transcriptome and metabolome. Aim #1: To elucidate a SHP and alcohol mediated molecular circadian pathway that controls ER stress signaling; Aim #2: To determine the effect of liver specific Npas2 knockdown on the ER stress response and steatosis mediated by SHP and ethanol-binge; and Aim #3: To establish hepatic circadian transcriptome and metabolome networks interlocked by SHP and ethanol-binge. In addition to our unique in vivo mouse (SHP-/-) and in vitro cell models, new liver Npas2 knockdown mice will be generated and employed for the study. Our approach makes full use of molecular biology, systems biology, and genome wide high throughput analysis (RNA-seq and GC/MS) to address an important scientific question that is directly relevant to the health of veterans. This study is innovative both conceptually and in the implementation of experimental approaches. The planned experimental strategies are comprehensive, yet feasible. This project will uncover for the first time a novel interplay between alcohol and the liver circadian clock to control the ER stress response and hepatic lipid metabolism. The discoveries made from this study are highly likely to enable substantial advances in ALD research, making this project of high clinical significance and translational value.