地塞米松诱导L6肌细胞胰岛素抵抗模型的建立及EGCG的保护作用研究

Establishment of insulin-resistant L6 muscle cell model induced by dexamethasone and effect of epigallocatechin gallate on insulin resistance

目的建立地塞米松(DEX)诱导的大鼠骨骼肌细胞胰岛素抵抗模型,并观察表没食子儿茶素没食子酸酯(EGCG)对其的保护作用。方法分化好的L6肌细胞,以1μmol/LDEX诱导L6肌细胞产生胰岛素抵抗,予以不同浓度的EGCG进行干预24h。然后分别用葡萄糖消耗实验、葡萄糖转运实验检测胰岛素刺激下L6细胞对葡萄糖的消耗和利用情况,Westernblot检测细胞胰岛素受体底物-1(IRS-1)的蛋白表达水平。结果与100nmol/L胰岛素组比较,1μmol/LDEX作用24h后胰岛素刺激下细胞上清液中葡萄糖的残存量显著增加和细胞内葡萄糖转运量明显减少(P<0.05),而EGCG作用后明显改善DEX的抑制作用(P<0.05),并呈剂量反应关系。Westernblot结果显示,与胰岛素组相比,DEX增高IRS-1的Ser307磷酸化表达,而EGCG作用后减少IRS-1的Ser307磷酸化表达。结论本研究表明,1μmol/LDEX作用24h能诱导L6肌细胞的胰岛素抵抗,而EGCG能改善这种胰岛素抵抗,可能是通过减少IRS-1的Ser307磷酸化表达来实现的。

Objective To establish a convenient and reliable insulin-resistant cell model of rat skeletal muscle cell treated with dexamethasone （DEX） and to investigate the effects of epigallocatechin gallate （EGCG） on insulin resistance. Methods EGCG was coincubated with DEX-treated rat L6 cells for 24 h. Then 100 nmol/L insulin was added into culture solution of each group. The content of glucose residues of culture solution was measured by the method of glucose oxidizes peroxides （GOD-POD） . We studied the glucose uptake of L6 cells by [ ^3H ] -2-deoxyglucose uptake assay. Insulin receptor substrate-1 （ IRS-1 ） levels were examined by western blotting. Results We found that DEX inhibited insulin-stimulated glucose uptake and increased Set307 phospborylation of IRS-1. However, EGCG treatment improved insulin-stimulated glucose uptake and decreased Ser307 phosphorylation of IRS-1. Conclusion These results suggest that EGCG attenuates DEX induced insulin resistance via decreasing Ser307 pbosphorylation of IRS-1.