DOCK5在肝脏脂肪代谢中的作用及信号转导机制研究

The incidence of non-alcoholic fatty liver disease (NAFLD) was in a rapid growth. NAFLD is strongly associated with metabolism syndrome (MS), of which the insulin resistance is their basic characteristics. Published researches and its related data revealed that DOCK5 can regulate the activity of small G protein and interact with PP2A. The data from our latest the experiment confirmed that the DOCK5 can regulate the transduction of insulin signaling pathway and that liver steatosis can be induced by the down-regulation of DOCK5 expression. The relationship between DOCK5 and NAFLD is not reported at present. We hold the hypothesis that DOCK5 is involved in the mechanism of hepatic steatosis through regulating expression and activity of PP2A and/or small G protein. In order to authenticate the hypothesis, the following experiment will be carried out. Cell and animal models is established with different expression of DOCK5. And then, the effects of DOCK5 on the accumulation of hepatic triglycerides were explored in vivo and in vitro. Moreover, the major metabolic pathways of hepatic triglycerides were investigated in above established models. Finally, the mechanism of DOCK5 effects on hepatic lipid metabolism are analyzed through detecting and inhibiting the key member of PP2A, small G protein, Akt, mTORC1 and MAPK signaling pathway. This will provide new experimental data for the prevention and treatment of NAFLD.

非酒精性脂肪性肝病（NAFLD）的发病率呈快速增长趋势，NAFLD的发生与以胰岛素抵抗为特征的代谢综合征密切相关。文献研究及相关数据均提示DOCK5不仅能够调控小G蛋白的活性，还能与PP2A发生相互作用。课题组的前期实验初步证实DOCK5能调控胰岛素信号的转导，DOCK5低表达与肝脏脂肪沉积有关。目前，尚未见DOCK5与NAFLD关系及机制的报道。我们提出科学假设：DOCK5参与肝脏脂肪的代谢，潜在的机制可能涉及对小G蛋白和PP2A表达或活性的调控。本项目建立DOCK5不同表达的细胞和动物模型，首先在体内外进一步证实DOCK5在肝脏脂肪沉积中的作用；然后探索DOCK5表达变化对肝脏脂肪代谢途径的影响；最后通过检测和干预PP2A、小G蛋白、Akt、mTORC1、MAPK等通路关键成员的表达/活性，解析DOCK5参与肝脏脂肪代谢的信号转导机制。研究结果将为NAFLD的防治提供新的理论依据。

胰岛素抵抗是糖尿病、肥胖、非酒精性脂肪性肝病等多种疾病发病的共同病理生理基础。胰岛素抵抗相关的基础研究是探索这些疾病发生发展的重要途径。本项目研究主要探究了DOCK5与胰岛素抵抗的关系，旨在为胰岛素抵抗相关疾病的诊治提供实验理论依据。DOCK5蛋白具有激活小G蛋白Rac的作用，可通过非Rac途径发挥作用，广泛参与细胞内信号系统发挥生理作用，与肥胖和胰岛素抵抗显著相关，高脂饮食可诱导DOCK5表达水平发生显著变化。本项目在前期基础之上进一步观察DOCK5-/-小鼠在高脂和老年因素的诱导下的表型变化，在动物模型中探索了DOCK5小鼠在高脂诱导的肝脏胰岛素信号通路变化，更进一步探讨DOCK5参与胰岛素抵抗及葡萄糖代谢紊乱的分子机制。在高脂诱导及老龄化影响下DOCK5敲除引起小鼠0-12月体重，摄食，血压，血糖增加，呼吸商下降；ITT与GTT实验中血糖水平增加；而12月到16月DOCK5敲除引起小鼠体体重，摄食差异不具有统计学意义。实验结果提示高脂诱导下DOCK5敲除导致葡萄糖代谢障碍引起代谢紊乱。小鼠肝脏组织的raptor/m-TOR及胰岛素信号通路均有明显的变化。提示DOCK5可能通过m-TOR对胰岛素信号通路起调节作用。对raptorloxp/loxp小鼠进行不同的腺病毒转染。观察到高脂诱导下DOCK5敲减引起的胰岛素抵抗导致的血糖调节紊乱在raptor敲减的状态下得到了改善，实验表明DOCK5与raptor直接作用从而介导了胰岛素信号转导及葡萄糖代谢，证实DOCK5/raptor/m-TOR信号转导途径是DOCK5参与代谢紊乱的主要机制之一。