摘要
目的研究甲基莲心碱对胰岛素抵抗的HepG2细胞降糖的影响。方法用CCK-8法进行细胞毒性实验筛选出合适的甲基莲心碱浓度;取对数生长期的HepG2细胞,胰岛素10^-4 mol/L、10^-5 mol/L、10^-6 mol/L、10^-7 mol/L、10^-8 mol/L 5个浓度干预12 h、24 h、36 h、48 h探索最佳的胰岛素浓度和作用时间;实验分为正常组、模型组、实验组(甲基莲心碱组),正常组不做处理,模型组用10-7 mol/L胰岛素干预36 h,实验组胰岛素干预后给与甲基莲心碱处理。用葡萄糖测定试剂盒(葡萄糖氧化酶-过氧化酶法)检测各组细胞的耗糖量;Western blot法检测各组细胞中哺乳动物雷帕霉素靶蛋白(mTOR、p-mTOR)、自噬微管相关蛋白轻链(LC3)、泛素结合蛋白(p-P62)蛋白的表达。结果给药后培养12 h,当甲基莲心碱的浓度为88.02μmol/L时,HepG2细胞致死率为5%;胰岛素10-7 mol/L干预36 h可成功建立胰岛素抵抗模型;实验组中不同浓度的甲基莲心碱均可促进胰岛素抵抗HepG2细胞葡萄糖消耗量(P<0.05)。与正常组相比,模型组mTOR、p-mTOR、LC3蛋白表达显著升高,P62蛋白表达显著降低(P<0.01)。与模型组比较,实验组的mTOR、p-mTOR、LC3表达不同程度地降低,P62蛋白表达不同程度地升高(P<0.05)。结论甲基莲心碱可增加胰岛素抵抗的HepG2细胞的葡萄糖消耗量,减轻胰岛素抵抗。其可能是通过减少细胞自噬来发挥作用的。
Objective To investigate the effect of neferine on the hypoglycemic effect of insulin resistance HepG2 cells.Methods The appropriate concentration of methyloline was detected by CCK-8 cytotoxicity method.HepG2 cells were in logarithmic growth phase,insulin 10-4,10-5,10-6,10-7,10-8 mol/L 5 concentration intervention for 12,24,36,48 hours to explore optimal insulin concentration and time action.The experiment was divided into blank group,model group,experimental group(neferine group),the blank group was not treated,the model group was treated with 10-7 mol/L for 36 h,and the experimental group was treated with neferine after insulin intervention.The glucose determination kit(glucose oxidation)enzyme-peroxidase method was used to detect the sugar consumption of each group of cells.Western blot detection of breast-fed animal rapamycin target Protein(mTOR,p-mTOR)and autophagy microtubule-associated protein light chain(LC3),ubiquitin-binding protein(P62)protein expression.Results After 12 hours of administration,the lethality of HepG2 cells was 5%when the concentration of neferine was 88.02μmol/L.Insulin resistance model was successfully established after insulin 10-7 mol/L intervention for 36 h.Different concentrations of neferine promoted insulin resistance in HepG2 cells(P<0.05).Western Blot results showed that compared with the normal group,the expression of the model group(mTOR,p-mTOR)and(LC3)protein was significantly increased,and the expression of(P62)protein was significantly decreased(P<0.01).Compared with the model group,the expressions of mTOR,p-mTOR and LC3 in the experimental group were decreased to some extent,and the expression of P62 protein was increased in different degrees(P<0.05).Conclusion Neferine can increase the glucose consumption of insulin-resistant HepG2 cells and reduce insulin resistance.It may work by reducing cellular autophagy.
作者
魏萍
薛春苗
潘霖
雷珍珍
曹俊岭
WEI Ping;XUE Chun-miao;PAN Lin(Department of Pharmacy,College of traditional Chinese Medicine,Beijing University of Chinese Medicine,Beijing 100029,China;Department of Pharmacy,Dongzhimen Hospital,Beijing University of traditional Chinese Medicine,Beijing 100700,China.)
出处
《临床和实验医学杂志》
2020年第12期1283-1286,共4页
Journal of Clinical and Experimental Medicine
基金
北京中医药大学科研课题项目(编号:2014MW12)。
