·糖尿病基础研究·

　　【摘要】 目的 采用过氧化氢(H2O2)制备INS-1氧化应激模型，观察姜黄素对INS-1细胞氧化应激水平及胰岛素分泌的影响及瓣状核酸内切酶(FEN1)和ERK1/2磷酸化的改变在其中的作用。 方法 将INS-1细胞分为阴性对照组，模型组(H2O2处理)，干预组(5、7、10 µM姜黄素+H2O2处理)。采用CCK-8检测细胞增殖，分光光度计法检测细胞丙二醛(MDA)、还原型谷胱甘肽(GSH)水平，酶标仪检测细胞内活性氧(ROS)水平的变化，放射免疫法检测细胞上清液中胰岛素分泌量，Western blot检测FEN1，p-ERK1/2蛋白表达的改变。 结果 (1)模型组MDA,ROS值较对照组明显升高(P<0.05);干预组MDA，ROS较模型组明显降低(P<0.05);模型组GSH较对照组明显降低(P<0.05);干预组GSH较模型组显著升高(P<0.05)。(2)模型组胰岛素分泌水平较对照组明显降低(P<0.05),10 µM姜黄素干预组胰岛素分泌水平较模型组明显升高(P<0.05)。(3)与模型组比较，10 µM姜黄素干预显著提高INS-1细胞FEN1蛋白的表达和ERK1/2的磷酸化，ERK1/2特异性抑制剂U0126可抑制姜黄素引起INS-1细胞FEN1蛋白表达的改变。 结论 姜黄素可以通过pERK1/2-FEN1通路改善INS-1细胞的氧化应激状态以及胰岛素分泌，为姜黄素治疗糖尿病提供了新的作用机制。

　　【关键词】 姜黄素;氧化应激;胰岛素分泌;FEN1;pERK1/2

　　Pilot study on the mechanism and effection of curcumin on oxidative stress and insulin secretion in INS-1 cells ZHANG You-zhi, XU Zi-hui, CHEN Tong, et al. Department of Endocrinology, Xinqiao Hospital, Third Military Medical University, Chongqing 400037, China

　　Corresponding author: XU Zi-hui, E-mail: zihuixu@yahoo.com.cn

　　【Abstract】Objective To observe the effections of curcumin on the oxidative stress and glucose-stimulated insulin secretion (GSIS) and the changes of flap endonuclease 1 (FEN1) and phosphorylation extracellular signal-regulated kinase1/2 (pERK1/2) in this process. Methods INS-1 cells were divided into 3 groups: the negative control group, the model group (treated with H2O2), and the intervention group (treated with curcumin 5, 7, 10µM + H2O2). Proliferation of INS-1 cells was detected through CCK-8 assay. Malondialdehyde (MDA) and glutathione (GSH) levels were detected by spectrophotometer assay. The level of reactive oxygen species (ROS) was observed by enzyme-labeled instrument and insulin secretion stimulated by glucose in the cell supernate was measured by radioimmunoassay. The changes of FEN1 and p-ERK1/2 protein expressions were detected by Western blot assay. Results (1) MDA and ROS levels​​ of the model group were significantly higher than those of the control group (P<0.01); MDA and ROS levels of the intervention group were significantly lower than those of the model group (P<0.05); GSH level of model group was significantly lower than that of the control group (P<0.05); GSH level of intervention group was significantly higher than that of the model group (P<0.05). (2) Insulin secretion of the model group was significantly lower than that of the control group (P<0.05), while the insulin secretion of the 10μM curcumin intervention group was significantly higher than that of the model group (P<0.05). (3) Western blot assay showed that the expressions of Fen1 and pERK1/2 proteins in INS-1 cells were upregulated by 10µM curcumin compared with the model group. U0126, as an ERK1/2 specific inhibitor, could inhibit the expression changes of Fen1 protein which was induced by curcumin in INS-1 cells. Conclusion Curcumin can decrease the oxidative stress damage and has a good effection on insulin secretion through the signal pathway of pERK1/2-FEN1 in INS-1 cells, providing a new mechanism of curcumin treatment on diabetes.

　　【Key words】 Curcumin; Oxidative stress; Glucose-stimulated insulin secretion (GSIS); Flap endonuclease 1 (FEN1); Phosphorylation extracellular signal-regulated kinase1/2 (pERK1/2)