基于IRS-1/PI3K/AKT/GLUT4通路探讨二苯乙烯类化合物改善胰岛素抵抗作用机制

Mechanism of stilbene compounds on the improvement of insulin resistance by IRS-1/PI3K/AKT/GLUT4 pathway

关键词:  二苯乙烯  脂肪细胞  胰岛素抵抗  IRS-1/PI3K/AKT/GLUT4

Key words:  stilbene  adipocyte  insulin resistance  IRS-1/PI3K/AKT/GLUT4

摘要:

[目的] 探讨3种中药（虎杖、桑白皮、何首乌）来源的二苯乙烯类化合物改善胰岛素抵抗（IR）的作用机制。[方法] 利用肿瘤坏死因子-α（TNF-α）作用于3T3-L1脂肪细胞96 h建立IR细胞模型；采用噻唑蓝（MTT）法检测二苯乙烯类化合物白藜芦醇（RES）、虎杖苷（PD）、桑皮苷A（Mul A）、2，3，5，4’-四羟基二苯乙烯-2-O-β-D-葡糖糖苷（THSG）对脂肪细胞生存率的影响；葡萄糖氧化酶法检测细胞培养基上清液中葡萄糖含量；采用蛋白免疫印迹（Western Blot）法检测胰岛素受体底物-1（IRS-1）、磷酸化IRS-1（p-IRS-1）、磷脂酰肌醇-3激酶（PI3K）、磷酸化PI3K（p-PI3K）、蛋白激酶B（AKT）、磷酸化AKT（p-AKT）、葡萄糖转运体4（GLUT4）的蛋白表达水平；采用实时定量荧光聚合酶链反应（RT-PCR）法检测GLUT4 mRNA表达水平。[结果] 与空白对照组相比，模型组的葡萄糖消耗量显著下降，模型组的p-IRS-1（Ser307）蛋白表达显著增加，p-PI3K、p-AKT（Ser473）、GLUT4蛋白表达显著下降，模型组GLUT4 mRNA表达显著降低；与模型组相比，RES、Mul A、THSG组的葡萄糖消耗量显著增加，PD组的葡萄糖消耗量无明显变化，Mul A、THSG组均能显著逆转p-IRS-1（Ser307）、p-PI3K、p-AKT（Ser473）、GLUT4的蛋白表达，RES仅能够逆转p-IRS-1（Ser307）、GLUT4的蛋白表达；与模型组相比，THSG组GLUT4 mRNA表达显著增加。[结论] Mul A、THSG能够通过IRS-1/PI3K/AKT/GLUT4信号通路改善脂肪细胞IR，而RES则通过激活IRS-1和GLUT4，但不通过PI3K/AKT途径发挥改善脂肪细胞IR的作用。PD无降糖活性。

Abstract:

[Objective] The aim of this study is to investigate the mechanism of three traditional Chinese medicines (Polygoni Cuspidati Rhizoma et Radix,Mori Cortex,Polygoni Multiflori Radix) derived stilbene compounds in improving insulin resistance.[Methods] The insulin resistance model was established by using tumor necrosis factor α (TNF-α) for 96 h in 3T3-L1 adipocytes. Thiazolyl blue tetrazolium bromide (MTT) method was used to detect the survival rates of adipocytes with stilbene compounds,such as resveratrol (RES),polydatin (PD),mulberroside A (Mul A) and 2,3,5,4'-tetrahydroxyl diphenylethylene-2-O-glucoside (THSG). Glucose content in supernatant of cell culture medium was detected by glucose kit. Protein expression levels of insulin receptor substrate-1 (IRS-1),p-IRS-1,phosphatidylinositol3-kinase (PI3K),p-PI3K,protein kinase B (AKT),p-AKT, glucose transporter 4 (GLUT4) were detected by Western Blot,and the mRNA expression of GLUT4 was detected by RT-PCR.[Results] Compared with those in the normal group,the glucose consumption rate of the model group was significantly down-regulated,meanwhile the protein expression of p-IRS-1 (Ser307) was significantly increased,the expressions of p-PI3K and p-AKT (Ser473) and GLUT4 were down-decreased,and the mRNA expression of GLUT4 was significantly decreased. Compared with the model group,RES,Mul A,and THSG could significantly increase the glucose consumption,while PD has no such effect. Moreover,the protein expressions of p-IRS-1 (Ser307) and p-PI3K and p-AKT (Ser473) and GLUT4 were significantly reversed by Mul A and THSG. However,RES only significantly reverse the protein expressions of p-IRS-1 (Ser307) and GLUT4. THSG could significantly increase the mRNA expression of GLUT4.[Conclusion] Mul A and THSG could improve the insulin resistance via IRS-1/PI3K/AKT/GLUT4 signaling pathway in the adipocytes. RES could improve the insulin resistance by activating IRS-1 and GLUT4 but not through the PI3K/AKT pathway in the adipocytes,and PD has no hypoglycemic activity.