胡椒碱对HepG2细胞胰岛素抵抗模型糖代谢AMPK信号通路上游靶点干预机制研究.docVIP

胡椒碱对HepG2细胞胰岛素抵抗模型糖代谢AMPK信号通路上游靶点干预机制研究 　　[摘要]探讨胡椒碱对胰岛素抵抗（insulin resistance，IR）细胞模型糖代谢紊乱的作用及干预AMPK信号通路上游靶点的分子机制。通过脂肪乳诱导HepG2细胞构建胰岛素抵抗模型。葡萄糖氧化酶（GOD-POD）法检测各组葡萄糖消耗量；酶联免疫吸附测定（ELISA）法检测上清液中脂联素（adiponectin，APN）、瘦素（leptin，LEP）的水平；荧光定量 PCR法检测APN，LEP mRNA的表达水平，Western blotting法检测AMPKα，р-AMPKα，瘦素受体（LepR），脂联素受体1（AdipoR1）和脂联素受体2（AdipoR2）蛋白的表达。结果显示，胡椒碱与降糖药罗格列酮、AMPK激动剂AICAR均能明显提高胰岛素抵抗细胞模型的葡萄糖消耗量，升高脂联素水平、增加脂联素mRNA、脂联素受体1蛋白的表达，也增加AMPKα mRNA和р-AMPKα蛋白表达；同时降低瘦素mRNA和瘦素受体蛋白的表达。结果表明，胡椒碱能显著改善胰岛素抵抗细胞模型糖代谢紊乱，其作用机制可能通过调控上游脂联素和瘦素的表达，从而激活AMPK信号通路。 　　[关键词]胡椒碱； 磷酸腺苷激活的蛋白激酶； 胰岛素抵抗； 瘦素； 脂联素 　　[Abstract]To investigate the effect of piperine on the disorder of glucose metabolism in the cell model with insulin resistance （IR） and explore the molecules mechanism on intervening the upstream target of AMPK signaling pathway. The insulin resistance models in HepG2 cells were established by fat emulsion stimulation. Then glucose consumption in culture supernatant was detected by GOD-POD method. Enzyme-linked immunosorbent assay（ELISA） was used to measure the levels of leptin（LEP） and adiponectin（APN） in culture supernatant； Real-time quantitative PCR was used to assess the mRNA expression of APN and LEP； and the protein expression levels of LepR， AdipoR1， AdipoR2 and the activation of AMPK signaling pathway were detected by Western blot analysis. The results showed that piperine， rosiglitazone and AMPK agonist AICAR could significantly elevate the glucose consumption in insulin resistance cell models， enhance the level of APN， promote APN mRNA transcripts and increase the protein expression of Adipo receptor. Meanwhile，AMPKα mRNA and р-AMPKα protein expressions were also increased in piperine treated cells， but both LEP mRNA expression and LepR protein expressions were decreased in piperine treated group. The results indicated that piperine could significantly ameliorate the glucose metabolism disorder in insulin resistance cell models through regulating upstream molecules （APN and LEP） of AMPK sig