Mechanisms and metabolic implications of LRP130 in NAFLD

描述（由申请人提供）：非酒精性脂肪性肝病（NAFLD）是美国肝脏疾病的主要原因。它涵盖了从脂肪肝到被称为NASH（非酒精性脂肪性肝炎）的叠加炎症的临床范围。NASH易发生不可逆肝硬化和肝细胞癌，因此需要明确其潜在机制。在NAFLD中，有缺陷的线粒体释放过多的活性氧（ROS）。线粒体ROS引起了人们的兴趣，因为它与胰岛素抵抗和炎症有关，这是NAFLD的两个关键特征。线粒体功能障碍的机制尚不清楚；因此，了解其机制基础可以减轻NAFLD。我们的初步数据表明，LRP130是线粒体功能和ROS的重要调节因子。在NAFLD中，我们观察到LRP130以疾病特异性的方式减弱。值得注意的是，小鼠肝脏中LRP130的缺失会引起胰岛素抵抗和炎症。从机制上讲，LRP130调控整个线粒体基因组。值得注意的是，LRP130深刻影响呼吸链的形成、呼吸和超氧化物的形成。我们假设LRP130是肝脏线粒体缺陷与NAFLD进展之间的分子联系。我们提出三个目标来质疑我们的假设：(1)评估LRP130在线粒体功能和超氧化物形成中的作用。LRP130对线粒体生物能量学的影响将在细胞和小鼠中使用功能丧失和功能获得模型进行评估。我们将使用蓝色天然凝胶电泳来评估呼吸链超配合物。线粒体功能将通过耗氧量、复合体活性、膜电位和超氧化物测量来测量。(2)评价LRP130在非酒精性脂肪性肝病中的作用。用高脂肪饮食刺激小鼠会诱发脂肪肝。肝脏中LRP130缺失小鼠的胰岛素敏感性和炎症将通过一系列试验进行评估：高胰岛素-正糖钳，胰岛素信号的免疫检测，炎症的组织学评估和细胞因子谱。将使用肝脏特异性LRP130转基因模型进行平行研究。(3)评估LRP130的转录和炎症控制。我们假设细胞因子减弱LRP130的转录，并可能将细胞因子与线粒体缺陷和过量超氧化物联系起来。我们将使用基因研究、报告者测定和染色质免疫沉淀来验证这一假设。由于对NAFLD的基本机制了解不足，对ROS的治疗操作受到阻碍。我们关于LRP130的新发现将为线粒体在NAFLD中的作用提供新的范式。

DESCRIPTION (provided by applicant): Non-alcoholic fatty liver disease (NAFLD) is the leading cause of liver disease in the United States. It spans a clinical spectrum from fatty liver to superimposed inflammation called NASH (non-alcoholic steatohepatitis). NASH predisposes to irreversible cirrhosis and hepatocellular carcinoma, which highlights the need to identify underlying mechanisms. In NAFLD, defective mitochondria liberate excess reactive oxygen species (ROS). Mitochondrial ROS is of interest, because it is implicated in insulin resistance and inflammation, two key features of NAFLD. The mechanism for mitochondrial dysfunction is unknown; hence, understanding its mechanistic basis may mitigate NAFLD. Our preliminary data indicate that LRP130 is an important regulator of mitochondrial function and ROS. In NAFLD, we observed that LRP130 is attenuated in a disease specific manner. Notably, depletion of LRP130 in mouse liver induced insulin resistance and inflammation. Mechanistically, LRP130 regulates the entire mitochondrial genome. Notably, LRP130 profoundly influences respiratory chain formation, respiration and superoxide formation. We hypothesize that LRP130 is a molecular link between defective mitochondria in liver and progression of NAFLD. We propose three Aims to query our hypothesis: (1) Evaluate the role of LRP130 in mitochondrial function and superoxide formation. The impact of LRP130 on mitochondrial bioenergetics will be evaluated using loss- and gain-of-function models in cells and mice. We will evaluate respiratory chain supercomplexes using blue native gel electrophoresis. Mitochondrial function will be measured by oxygen consumption, complex activity, membrane potential and superoxide measurements. (2) Evaluate the role of LRP130 in non-alcoholic fatty liver disease. Fatty liver will be induced by challenging mice with a high fat diet. Mice deficient for LRP130 in liver will be evaluated for insulin sensitivity and inflammation using a complement of assays: hyperinsulinemic-euglycemic clamp, immunodetection of insulin signaling, histological evaluation of inflammation and cytokine profiling. Parallel studies will be conducted with a liver specific LRP130 transgenic model. (3) Evaluate transcriptional and inflammatory control of LRP130. We hypothesize that cytokines attenuate transcription of LRP130, and may link cytokines to defective mitochondria and excess superoxide. We will use genetic studies, reporter assays and chromatin immunoprecipitation to test this hypothesis. Therapeutic manipulation of ROS in NAFLD is impeded by an insufficient understanding of basic mechanisms. Our novel findings on LRP130 will advance new paradigms on the role of mitochondria in NAFLD.