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的研究(1)
[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。
, http://www.100md.com
[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 signaling pathway， and thus activate the AMPK signaling pathway.
, 百拇医药
[Key words]piperine； AMPK signaling pathway； insulin resistance； leptin； adiponectin
胰島素抵抗是2型糖尿病、高血压和冠心病的生理病理基础[1]。主要表现为胰岛素敏感性降低，不能促进外周靶组织(脂肪组织、肝脏、骨骼肌)的葡萄糖摄取和利用[1]。因此，改善胰岛素抵抗是治疗糖尿病和相关并发症的有效干预措施[2]。AMPK在各组织中分布广泛，是细胞内主要的“能量感受器”，对代谢稳态起着总开关的作用。AMPK调节异常与胰岛素抵抗密切相关[3]。
胡椒碱(piperine，PIP)是胡椒科植物如胡椒、荜茇等植物中一种生物活性较强的生物碱。本课题前期实验研究已证明，不同浓度的荜茇醇提物及有效成分胡椒碱可显著降低IR模型大鼠的FBS水平、血清FFA、血压升高和改善糖耐量异常现象、增强胰岛素敏感指数等关键指标的病理性变化、改善IR模型大鼠血清肿瘤坏死因子-α和瘦素水平异常升高[4-7]。然而其干预作用靶点尚未明确，需进一步研究。本研究进一步在脂肪乳诱导HepG2细胞构建胰岛素抵抗模型上，探讨胡椒碱对AMPK信号通路上游靶点的分子机制，为其治疗代谢性疾病药物开发奠定基础。, 百拇医药(万春平 魏雅改 李晓雪 张丽君 杨瑞 包照日格图)
, http://www.100md.com
[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 signaling pathway， and thus activate the AMPK signaling pathway.
, 百拇医药
[Key words]piperine； AMPK signaling pathway； insulin resistance； leptin； adiponectin
胰島素抵抗是2型糖尿病、高血压和冠心病的生理病理基础[1]。主要表现为胰岛素敏感性降低，不能促进外周靶组织(脂肪组织、肝脏、骨骼肌)的葡萄糖摄取和利用[1]。因此，改善胰岛素抵抗是治疗糖尿病和相关并发症的有效干预措施[2]。AMPK在各组织中分布广泛，是细胞内主要的“能量感受器”，对代谢稳态起着总开关的作用。AMPK调节异常与胰岛素抵抗密切相关[3]。
胡椒碱(piperine，PIP)是胡椒科植物如胡椒、荜茇等植物中一种生物活性较强的生物碱。本课题前期实验研究已证明，不同浓度的荜茇醇提物及有效成分胡椒碱可显著降低IR模型大鼠的FBS水平、血清FFA、血压升高和改善糖耐量异常现象、增强胰岛素敏感指数等关键指标的病理性变化、改善IR模型大鼠血清肿瘤坏死因子-α和瘦素水平异常升高[4-7]。然而其干预作用靶点尚未明确，需进一步研究。本研究进一步在脂肪乳诱导HepG2细胞构建胰岛素抵抗模型上，探讨胡椒碱对AMPK信号通路上游靶点的分子机制，为其治疗代谢性疾病药物开发奠定基础。, 百拇医药(万春平 魏雅改 李晓雪 张丽君 杨瑞 包照日格图)