Regulation of Hepatic Macronutrient Metabolism by Mitochondrial Citrate Transport

项目总结 在2型糖尿病(T2D)期间，胰岛素敏感性的丧失与肝脏脂肪堆积密切相关，增加了慢性高血糖背后的糖异生。此外，T2D过程中肝脏新生脂肪生成(DNL)的增加被认为是导致胰岛素抵抗和非酒精性脂肪性肝病(NAFLD)的原因。因此，识别直接调节肝脏DNL和糖异生的机制对于理解T2D的病理生理学和治疗机会具有独特的价值。胞浆柠檬酸被认为是肝脏代谢的主要调节剂，通过相互调节糖酵解和糖异生，为DNL提供底物和降低能量。柠檬酸是在线粒体中产生的，需要一种特定的转运体--线粒体柠檬酸盐载体(CIC)--才能到达胞浆。因此，CIC被预测占据了连接肝脏线粒体代谢、DNL、葡萄糖和还原驱动的中心代谢节点。然而，令人惊讶的是，CIC在正常状态和T2D状态下调节肝脏DNL和糖异生的作用在体内仍然鲜见报道。这项应用的总体目标是了解肝脏CIC如何对基础代谢和T2D病理生理学做出贡献。这将通过追求两个具体目标来解决：1)确定肝脏CIC功能如何在T2D状态下调节肝脏DNL；以及2)确定肝脏CIC功能如何导致T2D状态下的高血糖。目标1中的实验将验证这样一个假设，即在T2D状态下扰乱体内肝脏CIC活性会减少作为碳源的柠檬酸盐的供应，并减少可用于延长脂肪链的NAPDH，从而降低DNL。Aim 2中的实验将验证这样的假设：在T2D状态下在体内扰乱肝脏CIC活性可通过减轻肝脏胰岛素抵抗、将肝脏葡萄糖代谢从糖异生转移到糖酵解以及降低小鼠NAFLD进展的相关性来降低高血糖。总体而言，拟议的研究将测试CIC在体内的基本调节作用，并提供关于线粒体柠檬酸盐输出的单一代谢步骤如何有助于肝脏DNL增加和糖异生的核心T2D特征的新的、机械的信息。这项研究意义重大，因为成功完成这项研究将独特地促进对T2D病理生理学的基本理解。这项研究具有创新性，因为它将利用新的体内CIC干扰和代谢组学跟踪模型来测试CIC在T2D中连接线粒体代谢、DNL和糖异生的作用。

PROJECT SUMMARY During type 2 diabetes (T2D), loss of insulin sensitivity strongly associated with hepatic lipid accumulation increases gluconeogenesis underlying chronic hyperglycemia. Furthermore, increased hepatic de novo lipogenesis (DNL) during T2D is thought to drive insulin resistance and non-alcoholic fatty liver disease (NAFLD). Thus, identifying mechanisms directly modulating both hepatic DNL and gluconeogenesis could be uniquely valuable for understanding T2D pathophysiology and therapeutic opportunity. Cytosolic citrate is believed to be a master regulator of hepatic metabolism by reciprocally regulating glycolysis and gluconeogenesis and by supplying substrate and reducing power for DNL. Citrate is produced in the mitochondria and requires a specific transporter, the mitochondrial citrate carrier (CiC), to reach the cytosol. Thus, the CiC is predicted to occupy a central metabolic node linking hepatic mitochondrial metabolism, DNL, glucose, and reductive drive. Yet, surprisingly, the role of the CiC modulating hepatic DNL and gluconeogenesis in normal and T2D states remains sparsely addressed in vivo. The overall goal of this application is to understand how the hepatic CiC contributes to fundamental metabolism and T2D pathophysiology. This will be addressed by pursuing two specific aims: 1) Determine how hepatic CiC function regulates hepatic DNL in T2D states; and 2) Determine how hepatic CiC function contributes to hyperglycemia in T2D states. Experiments in aim 1 will test the hypothesis that disrupting hepatic CiC activity in vivo during T2D states decreases DNL, by decreasing supply of citrate as a carbon source and decreasing NAPDH available for fatty chain elongation. Experiments in aim 2 will test the hypothesis that disrupting hepatic CiC activity in vivo during T2D states decreases hyperglycemia by attenuating liver insulin resistance, shifting hepatic glucose metabolism towards glycolysis away from gluconeogenesis, and decreasing mouse correlates of NAFLD progression. Overall, the proposed investigation will test the fundamental regulatory role of the CiC in vivo and provide novel, mechanistic information on how the single metabolic step of mitochondrial citrate export contributes to the core T2D features of increased hepatic DNL and gluconeogenesis. This research is significant because successful completion will uniquely advance fundamental understanding of T2D pathophysiology. This research is innovative because it will utilize novel in vivo CiC disruption and metabolomic tracing models to test the role of the CiC role linking mitochondrial metabolism, DNL, and gluconeogenesis in T2D.