肌肉生长抑制素前肽基因转染对C2C12成肌细胞糖代谢的影响

Effects of myostatin propeptide gene tranfection on glucose metabolism in cultured C2C12 cells

目的探讨肌肉生长抑制素前肽（myostatin propeptide，MPRO）对C2C12成肌细胞葡萄糖摄取、氧化及糖原合成的影响及其作用机制。方法诱导成熟的C2C12成肌细胞分为对照组、胰岛素组、绿色荧光蛋白（GFP）组、胰岛素＋GFP组、MPRO组及胰岛素＋MPRO组，经相应处理后应用2-脱氧-右旋-[1-^14C]葡萄糖检测MPRO对C2C12细胞葡萄糖摄取、氧化及糖原合成的影响，应用Western印迹法检测胰岛素信号通路活性。结果与对照组相比，胰岛素组及胰岛素＋GFP组葡萄糖摄取及糖原合成量明显增加（P〈0．05）；胰岛素＋MPRO组C2C12细胞葡萄糖摄取量及糖原合成较胰岛素组显著增加（P〈0．05）；但MPRO及胰岛素对葡萄糖的氧化无明显影响（P〉0．05）。Western印迹结果显示，与对照组相比，胰岛素组及胰岛素＋GFP组胰岛素信号通路中的信号分子胰岛素受体β（IRβ）、胰岛素受体底物1（IRS-1）、蛋白激酶B（Akt）、糖原合成酶激酶3β（GSK-3β）磷酸化水平及磷脂酰肌醇3激酶（P13K）、细胞膜葡萄糖转运体4（Glut4）的表达显著增高（P〈0．05），转染MPRO后上述基因的蛋白磷酸化和表达水平升高更显著（P〈0．05）。结论MPRO在C2C12可能通过激活IRS-1／P13K／Akt信号通路增加胰岛素刺激的葡萄糖摄取及糖原合成。

Objective To investigate the effects of recombinant adeno-associated virus-mediated myostatin propeptide （MPRO） on uptake and oxidation of glucose, and glycogen synthesis in C2C12 myotubes, as well as the associated molecular mechanism. Methods Mature C2C12 myotubes were assigned to the following 6 groups： control, insulin, green fluorescent protein （GFP） , insulin ＋ GFP, MPRO, and insulin ＋ MPRO groups. Glucose uptake, glucose oxidation, and glycogen synthesis were detected by counting radioactivity of ^14 CO2 or 14C labeled glycogen derived from 2-deoxy-[ 1-^14C ] glucose. The activity of insulin signal pathway was evaluated by Western blot. Results Compared with control group, glucose uptake and glycogen synthesis were significantly increased in insulin and insulin＋GFP groups, and further increased in insulin＋MPRO group as compared with insulin alone （ all P〈 0.05 ）. However, MPRO and insulin had no effect on glucose oxidation. The phosphorylations of insulin receptor （IR） β, insulin receptor substrate 1 （ IRS-1 ） , protein kinase B （Akt） , glycogen synthase kinase-3β （ GSK-3β） , and the expressions of phosphatidylinositol 3-kinase （ PI3K ） and glucose transporter 4 （ Glut4 ） in membrane were significantly increased in insulin and insulin＋GFP groups compared with control group（ all P〈0.05 ）, and were further increased after MPRO transfection （ all P 〈 0.05 ）. Conclusion MPRO may increase insulin-stimulated glucose uptake and glycogen synthesis in C2C12 cells by activating the IRS/PI3K/Akt signal pathway.