lipin-1和DGKδ共调控骨骼肌糖脂代谢和胰岛素敏感性机制研究

Insulin resistance plays a key role in the development of type 2 diabetes. Skeletal muscle is the biggest target of insulin. Efforts to identify mechanisms of the cause and cure of insulin resistance in skeletal muscle are very important to the discovery of new drug for diabetes.Diacylglycerol (DAG) is identified as a potential mediator of insulin resistance in skeletal muscle. DAG can be phosphorylated to phosphatidic acid (PA) by diacylglycerol kinase (DGK), whereas lipin-1 could catalyze the dephosphorylation of PA to form DAG by its phosphatidate phosphatise (PAP) enzymatic activity. The previous study showed that down regulation of the DGK isoform delta (DGKδ) contributed to hyperglycemia -induced insulin resistance. However, it is unknown whether lipin-1 is involved in the regulation of glucose and lipid metabolism by its opposite role of DGKδ in skeletal muscle. Moreover, lipin-1 has been reported as a transcriptional coactivator which involved in the regulation of glucose and lipid metabolism in adipose tissue and liver. However, its role in skeletal muscle is also unknown. Therefore, in the present study, we will elucidate the involvement of Lipin-1 and DGKδ in the regulation of glucose and lipid metabolism and development of insulin resistance in skeletal muscle, and analyze the possibility that therapeutic approaches to target DAG and PA metabolism by regulation of DGKδ and lipin-1 may control and prevent insulin resistance in type 2 diabetes mellitus. Our study will offer molecular insight into the role of DGKδ and lipin in the development of skeletal muscle insulin resistance and contribute to the new drug research for type 2 diabetes.

胰岛素抵抗在2型糖尿病中起重要作用，骨骼肌是胰岛素作用的最大靶组织，研究骨骼肌中胰岛素抵抗发生机制对抗糖尿病创新药物发现具有重要意义。骨骼肌中甘油二酯(DAG)增加是胰岛素抵抗产生的重要诱因，DGKδ和lipin-1是调控细胞内DAG和磷脂酸(PA)平衡的重要蛋白，DGKδ催化DAG磷酸化产生PA，而lipin-1则通过其磷脂酸磷酸酶活性催化PA转变为DAG。骨骼肌内DGKδ下调参与胰岛素抵抗发生，但lipin-1的作用尚不清楚，此外，lipin-1可作为转录协同因子调节相关基因的表达而参与脂肪和肝脏糖脂代谢的调控，但其在骨骼肌中作用则研究很少。本项目将系统研究lipin-1和DGKδ对骨骼肌糖脂代谢的调控作用及其分子机理，探讨其成为抗糖尿病药物研发新靶标的可能性，并尝试寻找以DGKδ或lipin1为靶标的抗糖尿病活性先导化合物，为治疗糖尿病创新药物研究提供新思路和理论依据。

本课题系统研究了Lipin-1和DGKδ对骨骼肌糖脂代谢相关信号通路及功能的调控作用及其分子机制，其中重点研究了Lipin-1和DGKδ如何共同调控细胞内二酰基甘油（DAG）/磷脂酸（PA）平衡以及Lipin-1如何通过其磷脂酸磷酸酶（PAP）活性和转录协同因子双重活性而参与骨骼肌细胞内能量代谢平衡的调节。研究结果显示，降低C2C12细胞内DGK活性或DGKδ蛋白表达可增加细胞内DAG水平，降低PA水平、削弱胰岛素信号通路并抑制胰岛素刺激的葡萄糖摄取；Lipin-1过表达可增加C2C12细胞内DAG含量，削弱胰岛素信号通路并抑制胰岛素刺激的葡萄糖摄取；在Lipin-1过表达的同时过表达DGKδ可使细胞内DAG的增加回复至正常水平，并可逆转Lipin-1过表达引起的胰岛素信号通路削弱以及胰岛素刺激的葡萄糖摄取能力降低。小鼠C2C12骨骼肌细胞中Lipin-1表达水平特异性下调后可造成胰岛素抵抗，具体表现为胰岛素刺激条件下Akt蛋白磷酸化水平的下降和葡萄糖摄取及糖原合成功能的削弱。同时，在棕榈酸诱导的小鼠C2C12骨骼肌细胞胰岛素抵抗后，Lipin-1的蛋白表达也明显下调，提示Lipin-1可能参与糖尿病的发生及发展过程。进一步的研究发现，小鼠C2C12骨骼肌细胞中lipin-1表达下调可导致TAG合成减少和线粒体功能紊乱，使得脂质更多流向神经酰胺合成，细胞内神经酰胺含量的增加激活PP2A，从而造成细胞胰岛素抵抗。因此，Lipin-1和DGKδ均参与骨骼肌糖脂代谢功能以及胰岛素敏感性的调控，且二者之间密切相关。改变骨骼肌细胞中Lipin-1和DGKδ的活性或者表达水平均可以影响细胞内DAG/PA平衡、胰岛素信号通路以及葡萄糖摄取功能，从而调节骨骼肌的胰岛素敏感性。