Thioesterase-mediated lipotoxicity in liver and thermogenic adipose tissue

项目摘要/摘要 肥胖引起的脂毒性是易患非酒精性脂肪肝的主要病理生理缺陷 疾病(NAFLD)。由于目前的管理选择仍然有限，因此确定新的监管 管理对营养过剩的不良适应反应的机制将有助于确定 药物干预。酰基辅酶A硫代酯酶(ACoT)通过以下途径控制细胞对脂肪酸的利用 将酰基辅酶A水解成非酯化脂肪酸。我们的长期目标是定义异常的ACoT活动 可用于治疗目的。这项研究的目的是通过以下方式确定其作用机制 其中Acot9介导的乙酰辅酶A的水解会导致代谢性疾病。我们的初步数据显示 Acot9位于肝脏线粒体的内膜(IM)，在那里它运输乙酰辅酶A 走向柠檬酸(TCA)循环。这导致肝脏葡萄糖产生(HGP)增加和从头开始 活性氧(ROS)和肝脏胰岛素证明的脂肪生成(DNL)和脂毒性 抵抗。除了TCA循环，Acot9还增加了蛋白质赖氨酸乙酰化的乙酰辅酶A供应(ACK) 通过控制乙酰辅酶A的生物利用度和抑制脱乙酰酶sirtuin 3(SIRT3)，从而减少ROS 通过抑制ACK。与肝脏相反，在生热性脂肪组织(BAT)中，冷诱导的Acot9易位 从IM进入线粒体基质，通过将乙酰辅酶A从TCA循环转移到线粒体基质中来抑制产热。 我们的中心假设是肥胖诱导的Acot9的激活通过促进 肝脏中的脂肪毒性和限制蝙蝠的产热作用。其基本原理是，乙酰基的机理是- 通过Acot9转运CoA将揭示预防脂毒性及其病理生理的新靶点 后果。中心假设将在三个具体目标中进行检验：1)确定分子 肝脏Acot9促进肝脏脂毒性的机制；2)阐明Acot9促进肝脏脂毒性的机制 Acot9在蝙蝠中限制产热；以及3)确定Acot9控制ack和 肝脏中的ROS。在目标1中，将阐明Acot9诱导的脂毒性、HGP和DNL的机制。 使用脂质组学和代谢组学对Acot9(Acot9LKO)进行肝脏特异性消融的小鼠。对胰岛素的影响 信号和脂毒途径将在小鼠和原代肝细胞中确定。AIM 2将使用小鼠 用于气候控制笼子中间接量热法测量的Acot9的BAT特定消融。夹子， 组织组织学和代谢组学将评估Acot9在BAT中的代谢功能。养殖褐变 脂肪细胞和重组Acot9将被用来确定Acot9易位到 线粒体基质。AIM 3将使用Acot9LKO/SIRT3-/-双基因敲除小鼠来确定SIRT3在 Acot9介导的肝脏ACK和ROS的调节。总体而言，这项提议将阐明新的机制 硫代酯酶介导的乙酰辅酶A利用的控制。这一点很重要，因为乙酰辅酶A是 几乎所有膳食脂肪和糖类的常见分解最终产物。这些研究预计将 将Acot9确立为管理NAFLD的一个易处理的目标。

PROJECT SUMMARY/ABSTRACT Obesity-induced lipotoxicity is the primary pathophysiological defect that predisposes to non-alcoholic fatty liver disease (NAFLD). Because current management options remain limited, identification of new regulatory mechanisms that govern the maladaptive response to overnutrition will serve to identify novel opportunities for pharmacologic intervention. Acyl-CoA thioesterases (Acots) control the cellular utilization of fatty acids by hydrolyzing acyl-CoA into non-esterified fatty acids. Our long-term goal is to define how aberrant Acot activity can be leveraged for therapeutic purposes. The objective of this research is to determine the mechanisms by which Acot9-mediated hydrolysis of acetyl-CoA culminates in metabolic disease. Our preliminary data indicates that Acot9 locates to the inner membrane (IM) of the mitochondria in the liver where it traffics acetyl-CoA towards the citric acid (TCA) cycle. This results in increased hepatic glucose production (HGP) and de novo lipogenesis (DNL) as well as lipotoxicity as evidenced by reactive oxygen species (ROS) and hepatic insulin resistance. In addition to TCA cycle, Acot9 increased acetyl-CoA supply for lysine acetylation of proteins (AcK) by controlling acetyl-CoA bioavailability and by inhibiting the deacetylase sirtuin 3 (Sirt3), which reduces ROS by inhibiting AcK. In contrast to liver, in thermogenic adipose tissue (BAT), cold-induced translocation of Acot9 from IM into mitochondrial matrix suppressed thermogenesis by trafficking acetyl-CoA away from TCA cycle. Our central hypothesis is that obesity-induced activation of Acot9 impairs nutrient homeostasis by promoting lipotoxicity in the liver and by limiting thermogenesis in BAT. The rationale is that the mechanisms of acetyl- CoA trafficking by Acot9 will reveal novel targets for the prevention of lipotoxicity and its pathophysiological consequences. The central hypothesis will be tested in three specific aims: 1) To identify the molecular mechanisms by which hepatic Acot9 promotes hepatic lipotoxicity; 2) To elucidate the mechanisms by which Acot9 in BAT limits thermogenesis; and 3) To determine the mechanisms by which Acot9 controls AcK and ROS in the liver. In Aim 1, the mechanisms of Acot9-induced lipotoxicity, HGP and DNL will be elucidated in mice with liver-specific ablation of Acot9 (Acot9LKO) using lipidomics and metabolomics. Impact on insulin signaling and lipotoxic pathways will be determined in mice and primary hepatocytes. Aim 2 will use mice with BAT-specific ablation of Acot9 for the indirect calorimetry measurements in climate-controlled cages. Clamp, tissue histology and metabolomics will assess the metabolic function of Acot9 in BAT. Cultured brown adipocytes and recombinant Acot9 will be used to determine the mechanism of Acot9 translocation into mitochondrial matrix. Aim 3 will use Acot9LKO/Sirt3–/– double knockout mice to determine the role of Sirt3 in Acot9-mediated regulation of AcK and ROS in the liver. Overall, this proposal will elucidate new mechanisms of thioesterase-mediated control of acetyl-CoA utilization. This is significant because acetyl-CoA is the common breakdown end-product of nearly all dietary lipids and sugars. These studies are expected to establish Acot9 as a tractable target for the management of NAFLD.