葛根芩连汤含药血清对L6肌细胞胰岛素抵抗模型中PI3K、Akt、GLUT4、AMPK蛋白的影响
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摘要
目的通过观察葛根芩连汤含药血清对L6骨骼肌细胞胰岛素抵抗模型中磷脂酰肌醇-3-激酶(PI3K)、蛋白激酶B(Akt)、AMP依赖的蛋白激酶(AMPK)、葡萄糖转运子4(GLUT4)的影响,探讨葛根芩连汤抗2型糖尿病胰岛素抵抗的信号通路相关机制。方法将诱导分化好的L6肌细胞接种于6孔板内,以1μmol/L DEX诱导L6肌细胞24 h产生胰岛素抵抗,予以不同浓度的葛根芩连汤含药血清(20%、10%和5%),采用western blot法检测L6骨骼肌细胞p-PI3K、Akt、p-Akt、p-AMPK、GLUT4蛋白的表达。另设实验,分组方法如上,采用ELISA法检测膜蛋白GLUT4含量。结果葛根芩连汤含药血清(20%、10%和5%)(高、中和低剂量组)治疗后,p-PI3K、Akt、p-Akt、p-AMPK、GLUT4蛋白的表达明显升高(P<0.05);用葛根芩连汤含药血清(20%、10%和5%)(高、中和低剂量组)治疗后,膜GLUT4蛋白的表达明显升高(P<0.05)。结论葛根芩连汤抗2型糖尿病胰岛素抵抗作用可能与影响PI3K、Akt、GLUT4、AMPK蛋白相关。
Objective To explore the mechanism of Gegenqinlian Decoction on signal transduction of insulin resistance in type 2 diabetes mellitus according to observing the effects of Gegenqinlian Decoction serum on phosphatidylinositol-3-kinase( PI3 K),protein kinase B( Akt),glucose transporter 4( GLUT4) and adenosine 5 '-monophosphate( AMP)-activated protein kinase( AMPK) in insulin resistance model of L6 skeletal muscle cells. Methods The induction of differentiated L6 muscle cell are seeded in 6-well plates. L6 muscle cell were induced with 1 μmol/L DEX for 24 h to produce insulin resistance and were given with different concentrations of Gegenqinlian Decoction-containing serum( 20%,10% and 5%). The expression of p-PI3 K,Akt,p-Akt,p-AMPK and GLUT4 protein in L6 muscle cells were detected by western blot. Set another method,the grouping method as above,using ELISA method to detect the membrane protein GLUT4 content. Results After the treatment of Gegenqinlian Decoction-containing serum( 20%,10% and 5%)( High,medium and low dose groups),the expression of p-PI3 K,Akt,p-Akt,p-AMPK and GLUT4 protein was significantly increased( P < 0. 05),the expression of GLUT4 protein was significantly increased( P < 0. 05). Conclusion The effect of Gegenqinlian Decoction on insulin resistance of type 2 diabetes mellitus may be related to the effects of PI3 K,Akt,GLUT4 and AMPK proteins.
Objective To explore the mechanism of Gegenqinlian Decoction on signal transduction of insulin resistance in type 2 diabetes mellitus according to observing the effects of Gegenqinlian Decoction serum on phosphatidylinositol-3-kinase( PI3 K),protein kinase B( Akt),glucose transporter 4( GLUT4) and adenosine 5 '-monophosphate( AMP)-activated protein kinase( AMPK) in insulin resistance model of L6 skeletal muscle cells. Methods The induction of differentiated L6 muscle cell are seeded in 6-well plates. L6 muscle cell were induced with 1 μmol/L DEX for 24 h to produce insulin resistance and were given with different concentrations of Gegenqinlian Decoction-containing serum( 20%,10% and 5%). The expression of p-PI3 K,Akt,p-Akt,p-AMPK and GLUT4 protein in L6 muscle cells were detected by western blot. Set another method,the grouping method as above,using ELISA method to detect the membrane protein GLUT4 content. Results After the treatment of Gegenqinlian Decoction-containing serum( 20%,10% and 5%)( High,medium and low dose groups),the expression of p-PI3 K,Akt,p-Akt,p-AMPK and GLUT4 protein was significantly increased( P < 0. 05),the expression of GLUT4 protein was significantly increased( P < 0. 05). Conclusion The effect of Gegenqinlian Decoction on insulin resistance of type 2 diabetes mellitus may be related to the effects of PI3 K,Akt,GLUT4 and AMPK proteins.
引文
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[12]章常华,盛军庆,熊淑平,等.葛根芩连汤及其主要成分对L6骨骼肌细胞糖代谢的调节作用[J].时珍国医国药,2016,27(4):797.
[13]张旭,陈俊国,吴鸣,等.细胞因子信号转导抑制因子3对糖尿病大鼠胰岛素受体底物1及其丝氨酸磷酸化水平的影响[J].中华全科医学,2016,14(7):1095.
[14]Lim CT,Kola B,Korbonits M.AMPK as a mediator of hormonal signaling[J].J Mol Endocrinol,2010,44(2):87.
[15]曹永红,王长江.葡萄糖转运蛋白4表达调节及与糖尿病的关系[J].中国临床保健杂志,2007,10(1):93.
[2]Zhao L.The gut microbiota and obesity:from correlation to causality[J].Nat Rev Microbiol,2013,11(9):639.
[3]Zhu SM,Sun F,Li WJ,et al.Apelin stimulates glucose uptake through the PI3K/Akt pathway and improves insulin resistance in 3T3-L1 adipocytes[J].Mol Cell Biochem,2011,353:305.
[4]Saltiel AR,Kahn CR.Insulin signalling and the regulation of glucose and lipid metabolism[J].nature,2001,414:799.
[5]Saltiel AR,Pessin JF.Insulin signaling pathways in time and space[J].Cell Biol,2002,12(2):65.
[6]Jaiswal N,Yadav PP,Maurya R,et al.Karanjin from Pongamia pinnata induces GLUT4 translocation in skeletal muscle cells in a phosphatidylinositol-3-kinase-independent manner[J].Eur J Pharmacol,2011,670:22.
[7]吴瑛.二氢杨梅素通过PPARγ/FGF21/AMPK信号通路改善胰岛素抵抗的机制研究[D].第三军医大学硕士学位论文,2015.
[8]Zhang C H,Xu G L,Liu Y H,et al.Anti-diabetic activities of Gegen Qinlian Decoction in high-fat diet combined with streptozotocininduced diabetic rats and in 3T3-L1 adipocytes[J].Phytomedicine,2013,20(3-4):221.
[9]熊淑平,章常华,邓永兵,等.葛根芩连汤改善KK-Ay糖尿病小鼠糖脂代谢作用的实验研究[J].时珍国医国药,2016,27(9):2090.
[10]章常华,涂秀英,熊淑平,等.基于蛋白质组学技术筛选葛根芩连汤防治胰岛素抵抗作用的血清差异表达蛋白[J].中药新药与临床药理,2016,27(1):81.
[11]章常华,邓可众,于梅,等.葛根芩连汤含药血清对Hep G2肝细胞胰岛素抵抗模型糖代谢的调节作用[J].中国实验方剂学杂志,2015,21(5):120.
[12]章常华,盛军庆,熊淑平,等.葛根芩连汤及其主要成分对L6骨骼肌细胞糖代谢的调节作用[J].时珍国医国药,2016,27(4):797.
[13]张旭,陈俊国,吴鸣,等.细胞因子信号转导抑制因子3对糖尿病大鼠胰岛素受体底物1及其丝氨酸磷酸化水平的影响[J].中华全科医学,2016,14(7):1095.
[14]Lim CT,Kola B,Korbonits M.AMPK as a mediator of hormonal signaling[J].J Mol Endocrinol,2010,44(2):87.
[15]曹永红,王长江.葡萄糖转运蛋白4表达调节及与糖尿病的关系[J].中国临床保健杂志,2007,10(1):93.