mTOR复合物在有氧运动改善胰岛素抵抗过程中的作用研究

The Role of m TOR Complex in Attenuating the Development of Insulin Resistance of Mice during Aerobic Exercise

目的:通过研究8周有氧运动对高脂饮食诱导胰岛素抵抗C57BL/6小鼠肝脏m TOR复合物1/2(m TOR Complex 1/2)以及胰岛素信号相关蛋白的影响,分析有氧运动/高脂饮食与m TORC1/C2和胰岛素信号通路蛋白磷酸化活性之间的关系,为全面理解有氧运动改善高脂饮食诱导机体胰岛素抵抗的机制提供理论依据。方法:选取50只4周龄、雄性C57BL/6小鼠随机分为正常饮食组(C组,n=10)和高脂饮食组(H组,n=40)。高脂饮食组小鼠饲以高脂饮食6周以建立胰岛素抵抗模型,6周后通过口服葡萄糖耐量实验(Oral Glucose Tolerance Test,OGTT)鉴定胰岛素抵抗成模小鼠。随后将胰岛素抵抗小鼠再次随机分为安静组(H组,n=10)和运动组(HE组,n=15),此两组小鼠继续饲以高脂饮食,同时对HE组施以8周、强度为75%VO2max的有氧跑台运动干预。实验结束后,采用OGTT检测小鼠葡萄糖耐量,ELISA法测定空腹血清胰岛素水平,Western Blot检测肝脏Akt/m TOR信号通路相关蛋白磷酸化水平,并通过免疫共沉淀方法检测小鼠肝脏Raptor-m TOR及Rictor-m TOR结合水平。结果:与C组相比,H组小鼠体重、血清胰岛素水平均显著升高,口服葡萄糖耐量下降,肝脏胰岛素信号通路相关蛋白p Akt S473、p Akt T308、p IRS1S307、p AMPKαT172磷酸化水平下降,p S6K1T389磷酸化水平升高。免疫共沉淀检测发现Raptor-m TOR结合水平上升,Rictor-m TOR结合水平下降,即m TORC1蛋白表达水平升高,m TORC2蛋白表达水平减低;与H组相比,HE组小鼠体重、血清胰岛素水平均显著降低,口服葡萄糖耐量增高,肝脏胰岛素信号通路相关蛋白p AktS473、p AktT308、p IRS1S307、p AMPKαT172磷酸化水平有所提高,p S6K1T389磷酸化水平降低。Raptor-m TOR结合减少,Rictor-m TOR结合增多,即m TORC1蛋白表达水平降低,m TORC2蛋白表达水平增高。结论:8周有氧运动可改善高脂饮食诱导小鼠胰岛素敏感性,其机制可能与有氧运动激活小鼠肝脏Akt/m TORC2信号通路、抑制Akt/m TORC1信号通路,增强小鼠肝细胞胰岛素信号敏感性、改善胰岛素抵抗有关。

Objective To investigate the effects of 8-week aerobic treadmill exercise on the mammalian target of rapamycin（m TOR）complex 1/2（m TORC1/m TORC2）and insulin signaling related protein expression in the liver of C57BL/6 mice with high-fat-diet induced insulin resistance（IR）and understand the relations of the aerobic exercise/high fat diet to the m TORC1/C2 and activity of insulin signaling pathway protein phosphorylation. Method 4-week old C57BL/6 mice（n=50）were divided into group C（fed with chow diet, n=10）and group H（fed with high fat diet, n =40）. Majority of the mice in group H were proved to have IR symptoms through oral glucose tolerance test（OGTT）,thereafter,the mice with IR symptoms were regrouped into sedentary group RH（n=10）and exercise group RHE（n=15）and continued feeding with high fat diet. Mice in group RHE were forced to run on a treadmill 60 min per day at the intensity of 75% VO2 max,five days per week for 8 weeks. By the end of the experiment,the changes in glucose tolerance,the proteins of the Akt/m TOR signaling pathway,the binding capacity of the regulatory associated protein of m TOR（Raptor）or rapamycin insensitive companion of m TOR（Rictor）in liver,and the serum insulin level were tested. Results Compared with group C, the body weight and fasting serum insulin level increased,and the oral glucose tolerance decreased,the phosphorylation of proteins in liver insulin signaling pathway（p Akt^S473, p Akt^T308,p IRS1^S307, p AMPKα^T172）decreased and the phosphorylation of p S6K1^T389 increased,the binding capacity of Raptor-m TOR enhanced and of Rictor-m TOR reduced significantly in group RH. Compared with group RH,the body weight, fasting serum insulin level decreased,the oral glucose tolerance improved,the phosphorylation of proteins in insulin signaling pathway（p Akt^S473, p Akt^T308, p IRS1^S307, p AMPKα^T172）elevated,the phosphorylation of p S6K1^T389 decreased,the binding capacity of Raptor-m TOR reduced and of Rictor-m TOR improved significantly in group RHE. Conclusion 8-week aerobic exercise resulted in improving insulin sensitivity of mice with high-fat-diet induced IR provably through the activation of Akt/m TORC2 signaling and inhibition of Akt/m TORC1 signaling in liver.