鬼针草黄酮对胰岛素抵抗HepG2细胞中胰岛素信号转导的干预机制探讨

Insulin Resistance（IR）is a pathological basis in 2 type diabetes, which cause by the effection of insulin decreased on uptakeing and clearing glucose in peri-constitution. The dysfunction of signal pathway is the important cause of IR,therefore,to enhance the activity of insulin signal pathway is a key strategy to prevent and treat insulin resistance.Our prophase research showed that Bidens Bipinnata L.flavonoids (BBLF) had the effects on improving hypoglycemic and decreasing serum insulin in diabetic mice , so as to improve the sensitivity of insulin of body. But the mechanisms of hypoglycemic and decreasing insulin resistance of BBLF are not clear,especially the mechanisms of signal transduction of insulin. In present studies it was found that there are three signal pathways in insulin transduction,which are Mitogen-activated protein kinase pathway(MAPK pathway), Phosphatidylinositide-3 kinase pathway(PI3K pathway) andβ-arrestin2 pathway.In order to investigate the intervention effects of BBLF on signal pathways of insulin,the HepG2 cells are induced by palmitic acid(PA).Through the glucose consumption and glucose uptake testing in HepG2 cells respectively,the concentration and inducing time of PA are ascertained.The concentration of BBLF is selected by MTT testing.And then HepG2 cells are treated by different groups.At last,RT-PCR and Western Blot testing are adoped to measure relative mRNA expression and protein expression of PI3K,Akt, GLUT4, JNK, P38MAPK,β-arrestin2,Scr,and so on.These proteins exist in signal pathway of insulin respectively. Then BBLF's the molecular mechanism on improving the insulin resistance in HepG2 cell induced by PA is identified.It's a good idea to investigate molecular target of BBLF treatment in insulin resistance.

胰岛素抵抗（IR）是胰岛素在周围组织摄取及清除葡萄糖作用减低。胰岛素信号通路的障碍是发生IR的重要机制。我们在前期研究中发现鬼针草黄酮(BBLF)具有降血糖及减少血清胰岛素作用，从而改善IR。其参与糖代谢和胰岛素信号转导的分子机制尚不明确。目前研究发现，胰岛素信号转导主要通过MAPK、PI3K、β-arrestin2三条通路进行，为明确BBLF在胰岛素信号通路中的作用，本课题拟采用软脂酸（PA）诱导HepG2细胞胰岛素抵抗，运用葡萄糖消耗试验、葡萄糖摄取试验、RT-PCR、Western Blot试验等实验方法，探讨BBLF对胰岛素抵抗HepG2细胞中的PI3K、Akt、GLUT4 、JNK、P38MAPK、β-arrestin2、Scr等相关蛋白及基因表达的影响，明确BBLF参与胰岛素信号转导的信号通路，从而为开发研究BBLF治疗胰岛素抵抗提供新的靶点。

胰岛素抵抗的病理过程是胰岛素在周围组织摄取及清除葡萄糖的作用减低，导致身体糖脂代谢紊乱，它贯穿于2型糖尿病的始终。因此针对胰岛素抵抗的治疗成为治疗糖尿病的关健。调控胰岛素信号通路的活性是目前防治胰岛素抵抗的重要策略，国内外研究的胰岛素信号通路主要有三条：一条是通过Grb2-SOS-Ras-MAPK途经；另一条是通过IRS-PI3K-Akt途经；还有一条是新发现的β-arrestin2- GSK3- GLUT4途经。.本立项课题主要通过软脂酸联合胰岛素作用于HepG2细胞，诱导HepG2细胞胰岛素抵抗模型，并通过Western blot法、RT-PCR法观察鬼针草黄酮干预后， MAPK、PI3K、β-arrestin2三个不同信号通路中各种相关因子的蛋白表达和基因mRNA表达，以初步探讨鬼针草黄酮的改善胰岛素抵抗的作用机制。.本实验结果提示：采用软脂酸+胰岛素建立胰岛素抵抗细胞模型，经鬼针草黄酮干预后，显示： RT-PCR实验结果：与胰岛素抵抗模型组比较，鬼针草黄酮组（80、40mg/L）IRS1、PI3KP85、Glut4mRNA的表达明显增加，差异具有统计学意义（p＜0.05）。Western Blot实验结果：与胰岛素抵抗模型组比较，AKT1、pAKT1蛋白表达明显减少；β-arrestin2、Glut4蛋白表达明显增加，差异具有统计学意义（p＜0.05）。提示：鬼针草黄酮可通过调控IRS1/PI3K/AKT/GLUT4信号通路中信号转导，增加细胞对葡萄糖的利用，从而改善细胞胰岛素抵抗。.本课题的发现为进一步开发广西道地药材提供了理论依据；同时为今后深入研究鬼针草黄酮如何通过调控胰岛素PI3K/AKT/GLUT4信号通路中的靶基因，寻找胰岛素抵抗的靶向治疗药物提供了坚实的研究基础,为开发新型的、有效的、经济的胰岛素抵抗治疗药物具有深远的科学意义。