UCP2 in the pathogenesis of steatohepatitis

描述(由申请人提供) 描述了一个研究解偶联蛋白-2的作用的研究计划。 (UCP2)在脂肪性肝炎(SH)发病机制中的作用。SH从胖子进化而来 肝脏，肥胖、糖尿病和酗酒患者的一种流行疾病 虐待。这种进展的预测因子和生化机制很差。 已定义的，但由活性氧(ROS)和内毒素介导的细胞因子 释放似乎起到了重要作用。UCP2是一种线粒体内膜 作为线粒体ROS产生的潜在调节因子出现的蛋白质。 在肥胖动物模型中，肝脏UCP2的表达显著增加， 酒精摄入和内毒素暴露。我们假设UCP2的诱导 在肝脏中是一种防御机制，它可能涉及到 肝细胞和枯否细胞。UCP2-/-在导师的 实验室将提供一个强有力的工具来检验这一假说。第一, 通过与Ob/Ob小鼠杂交，将在UCP2-/-小鼠中诱导脂肪变性， 高脂饲料和乙醇饲料。UCP2-/-小鼠也将受到挑战 体内应用内毒素、肿瘤坏死因子-α和抗Fas抗体治疗。UCP2-/- 在这种情况下，小鼠预计会出现加速的肝脏损伤。 腺病毒介导的基因转移将用于挽救UCP2功能 活着。UCP2作用的细胞机制将在体外进行研究 培养肝实质细胞和Kupffer细胞。细胞凋亡与ROS 将检测到UCP2-/-和野生型小鼠的肝细胞产生 脂肪酸、乙醇和神经酰胺。模拟超氧化物歧化酶和清道夫化合物将 用于恢复UCP2不存在时的动作。UCP2对多种细胞因子的影响 将研究细胞凋亡的步骤(线粒体通透性转变 开放，投标转移)：库普弗细胞产生RO和 将评估在没有UCP2的情况下释放的细胞因子。最后，Cre/Lox 重组酶系统将被用于创造组织特异性的UCP2基因敲除小鼠。 带有loxP修饰UCP2等位基因的小鼠将被创造出来，并与小鼠交配 表达白蛋白启动子驱动的Cre获得肝细胞特异性UCP2 淘汰赛。用表达溶菌酶的小鼠培育UCP2/loxP小鼠 启动子驱动的Cre将导致Kupffer细胞/巨噬细胞特异性UCP2 淘汰赛。用于组织特异性表达UCP2的Cre/lox系统的构建 将为申请者提供巨大的学习潜力。这项研究 导师实验室的设施将为您提供良好的环境 实现这些目标。生化作用和遗传基因的知识 UCP2在肝脏中的调节将促进我们对其作用的理解 UCP2和SH的病理生理机制。

DESCRIPTION (provided by applicant) A research program is described to study the role of uncoupling protein-2 (UCP2) in the pathogenesis of steatohepatitis (SH). SH evolves from fatty liver, a prevalent condition among patients with obesity, diabetes, and alcohol abuse. The predictors and biochemical mechanisms of this progression are poorly defined, but reactive oxygen species (ROS) and endotoxin-mediated cytokine release appear to have a major role. UCP2 is a mitochondrial inner membrane protein emerging as a potential regulator of mitochondrial ROS production. Expression of liver UCP2 is markedly increased in animal models of obesity, alcohol intake, and endotoxin exposure. We hypothesize that induction of UCP2 in the liver is a defensive mechanism and it may involve both parenchymal hepatocytes and Kupffer cells. UCP2-/- mice generated in the mentor's laboratory will provide a powerful tool to test this hypothesis. First, steatosis will be induced in UCP2-/-mice by crossbreeding with ob/obmice, high-fat feeding, and ethanol feeding. UCP2-/- mice will also be challenged by in vivo treatment with endotoxin, TNF-alpha, and anti-Fas antibody. UCP2-/- mice are expected to show accelerated liver injury under these conditions. Adenovirus-mediated gene transfer will be used to rescue UCP2 functions in vivo. The cellular mechanisms of UCP2 action will then be studied in vitro in cultured parenchymal hepatocytes and Kupffer cells. Apoptosis and ROS production will be detected in hepatocytes from UCP2-/- and wild type mice by fatty acids, ethanol, and ceramide. SOD mimetics and scavenger compounds will be used to revert the action of absent UCP2. The effect of UCP2 on various steps of apoptosis will be investigated (mitochondrial permeability transition opening, Bid translocation): The ability of Kupffer cells to generate ROS and release cytokines in the absence of UCP2 will be assessed. Finally, Cre/lox recombinase system will be used to create tissue-specific UCP2 knockout mice. Mice with a loxP-modified UCP2 allele will be created and bred with mice expressing an albumin promoter-driven Cre to obtain hepatocyte-specific UCP2 knockouts. Breeding of UCP2/loxP mice with mice expressing a lysozyme promoter-driven Cre will result in Kupffer cell/macrophage-specific UCP2 knockouts. Generation of Cre/lox system for tissue-specific UCP2 expression will provide a great learning potential for the applicant. The research facilities of the mentor's laboratory will provide an excellent environment to accomplish these aims. Knowledge of the biochemical actions and genetic regulation of UCP2 in the liver will advance our understanding about the role of UCP2 and the pathophysiology of SH.