Ghrelin提高L6大鼠成肌细胞胰岛素敏感性及其机理

Ghrelin Enhances The Sensitivity of Insulin in L6 Rat Skeletal Muscle Via PI3K/Akt/GSK-3β Signaling Pathway

目的研究Ghrelin对L6大鼠成肌细胞在棕榈酸诱导下的葡萄糖代谢和胰岛素敏感性的影响,并探讨其可能的机理。方法培养L6大鼠成肌细胞,诱导分化后利用0.3 mmol/L的棕榈酸作用16 h,实验组加入不同剂量的Ghrelin(1、10和100 nmol/L)作用8 h,采用葡萄糖氧化酶-过氧化物酶法(GOD-POD)检测骨骼肌细胞对葡萄糖的摄取,并在荧光共聚焦显微镜下观察细胞膜葡萄糖转运因子-4(GLUT-4)蛋白染色;采用免疫印迹(Western blot)法检测骨骼肌细胞中总蛋白激酶B(Akt)、磷酸化蛋白激酶B(pAkt)、总糖原合成酶激酶-3β(GSK-3β)和磷酸化糖原合成酶激酶-3β(pGSK-3β)蛋白的表达。结果 Ghrelin使胰岛素抵抗之L6大鼠成肌细胞葡萄糖摄取量增加,细胞膜GLUT-4染色加深,pAkt及pGSK-3β蛋白表达增加(磷酸化表达增加),而总Akt和总GSK-3β蛋白无明显变化,这种作用可被PI3K阻断剂(LY294002)消除。结论 Ghrelin通过磷酯酰肌醇-3激酶/蛋白激酶B/糖原合成酶激酶-3β(PI3K/Akt/GSK-3β)信号途径促进L6大鼠成肌细胞葡萄糖摄取量,从而提高L6大鼠成肌细胞胰岛素的敏感性。

Objective To study the effects of Ghrelin for glucose metabolism and insulin sensitivity of L6 rat myoblasts in palmitic acid induced， and to explore its possible mechanisms. Methods The L6 rat myoblasts were cultured until differentiation， then using palmitic acid （0.3 mmol/L） for 16 hours. The experimental group was treated with different doses of Ghrelin （1， 10， and 100 nmol/L） for 8 hours， then the glucose uptake was detected by using glucose oxidase peroxidase method （GOD-POD）， the cell membrane glucose transporter 4 （GLUT-4） protein staining was observated under confocal microscopy， and the expressions of total protein kinase B （Akt）， phosphorylated protein kinase B （pAkt）， total glycogen synthase kinase-3β （GSK-3β）， and phosphorylated glycogen synthase kinase-3β （pGSK-3β） were detected by using immunoblotting （Western blot）.Results Ghrelin enhanced the glucose uptake of L6 rat myoblasts with insulin resistance， the cell membrane Glut-4 stain was deepen， the expressions of pAkt and pGSK-3β protein increased， and this effect could be PI3K blocker （LY294002） eliminated. Conclusion Ghrelin promotes the glucose uptake of L6 rat myoblasts through PI3K/Akt/GSK-3β signaling pathway， so as to improve the sensitivity of insulin in L6 rats muscle cells.