摘要
目的大黄酸改善糖代谢的作用机制尚不明确,文章旨在探讨大黄酸对C2C12细胞胰岛素信号通路的影响。方法测定大黄酸抑制蛋白酪氨酸磷酸酶1B(PTP1B)的最适底物、最佳酶浓度,计算大黄酸对PTP1B的半抑制浓度(IC50)。将C2C12细胞诱导分化成熟,分为大黄酸组[分别给予不同浓度大黄酸(0.1、1、10、100μmol/L)处理];阳性对照组(给予10nmol/L胰岛素刺激0.5 h),空白对照组(加入等体积溶剂处理)。Western blot方法检测各组细胞胰岛素信号通路相关蛋白的变化,免疫共沉淀法检测PTP1B的活性变化。结果大黄酸对PTP1B的半抑制浓度(IC50)为80.5μmol/L。C2C12肌细胞经不同浓度大黄酸处理后,随大黄酸浓度的增加,细胞内总蛋白酪氨酸磷酸化水平略增加;经胰岛素处理后,细胞内总蛋白酪氨酸磷酸化水平有一定增加。C2C12细胞经胰岛素处理后,IRβ、IRS1的相对磷酸化水平以及Glut4的蛋白表达水平升高;空白对照组IRβ、IRS1、Glut4蛋白相对表达量分别为0.37、0.39、0.15,经不同浓度大黄酸处理,IRβ的相对磷酸化水平在0.1~10μmol/L处理时无明显变化(0.37、0.31、0.32),100μmol/L处理时升高至0.67;IRS1的相对磷酸化水平在0.1~100μmol/L浓度时升高至0.49、0.50、0.41、0.59;Glut4的蛋白相对表达水平在0.1~100μmol/L处理时升高至0.37、0.48、0.90、0.88。在C2C12细胞中,与0μmol/L大黄酸处理组比较,10 nmol胰岛素处理组及0.1~10μmol/L大黄酸处理组细胞中PTP1B活性无明显变化,100μmol/L大黄酸处理组细胞中PTP1B活性降低。结论大黄酸能降低PTP1B活性并增强胰岛素信号传递,推测胰岛素信号传递增强可能与骨骼肌细胞PTP1B活性降低有关。
Objective Recent studies have shown that rhein could improve glucose metabolism,while the specific mechanism is not yet clear,the aim of this study is to explore the effects of rhein on the insulin signaling pathway in C2 C12 cells. Methods To measure the optimum substrate and optimum enzyme concentrations of rhein and the protein tyrosine phosphatase-1 B（PTP1 B） and calculate the half maximal inhibitory concentration of rhein on PTP1 B（IC50）. Differentiated and maturated C2 C12 myotubes were divided into two groups,the rhein group and the control group. The rhein groups were treated with different rhein concentrations（0.1,1,10,100 μmol/L）, the positive control group was given 10 nmol/L insulin stimulation for 0. 5 hours and the blank control group was treated with equal volume solvent. Related proteins in the insulin signaling pathway were detected by Western blotting and PTP-1 B activity was measured by immunoprecipitation. Results The IC50 of rhein to PTP1 B was 80.5 μmol/L,and when C2 C12 myotubes were treated with 100 μmol/L rhein,the activity of PTP1 B decreased significantly.Compared with the blank control group,the rhein groups＇ phosphorylation levels in insulin receptor and insulin receptor substrates were enhanced and the levels of GLUT4 were obviously improved in C2 C12 cells. Moreover,the protein levels of insulin receptor and insulin receptor substrates were not affected significantly.Conclusion Rhein could reduce the activity of PTP1 B and enhance insulin signaling transduction. Therefore,we speculated that the enhancement of insulin signaling may be related to the decrease of PTP1 B activity in skeletal muscle cells.
作者
李柯
卢斌
王魏
邵加庆
LI Ke;LU Bin;WANG Wei;SHAO Jia-qing(Department of Endocrinology,Jinling Hospital,Southern Medical University/Nanjing General Hospital of Nanjing Military Region,PLA,Nanjing 210002,Jiangsu,China)
出处
《医学研究生学报》
CAS
北大核心
2018年第7期709-713,共5页
Journal of Medical Postgraduates
基金
国家自然科学基金(81471018)
江苏省博士后基金(1501120C)
