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目的通过观察地塞米松(Dex)对胰岛β细胞NADPH氧化酶表达水平和细胞凋亡的影响,探讨氧化应激在地塞米松诱导胰岛β细胞凋亡中的作用。方法将大鼠胰岛细胞β系INS-1细胞分为正常对照组、不同浓度的地塞米松组(0.01μΜ-Dex组、0.1μΜ-Dex组、1μΜ-Dex组)。培养48h后MTT法检测细胞增殖情况;Annexin V-FITC/PI双染经流式细胞仪检测细胞凋亡情况;Realtime-PCR检测Nox1、Nox3、Nox4、p47phox mRNA表达水平;荧光法检测ROS释放量;Weastern-blot检测Nox4蛋白表达情况。结果地塞米松诱导INS-1细胞凋亡呈现时间剂量依赖性,并且在此凋亡过程中Nox1、Nox3、Nox4、p47phox mRNA表达上调,Nox4蛋白表达上调,ROS释放量增加,差异均具有统计学意义(P<0.05)。结论地塞米松可诱导INS-1细胞凋亡,其机制可能与上调Nox1、Nox3、Nox4、p47phox表达,激活氧化应激反应有关。
Objective To observe the effect of dexamethasone (Dex) on the expression of NADPH oxidase and apoptosis in pancreatic β cells and to explore the role of oxidative stress in the induction of pancreatic β-cell apoptosis by dexamethasone. Methods The rat pancreatic islet β-cell INS-1 cells were divided into normal control group, dexamethasone group (0.01μM-Dex group, 0.1μM-Dex group and 1μM-Dex group). The cell proliferation was detected by MTT assay after 48 hours of culture; the apoptosis of cells was detected by flow cytometry with Annexin V-FITC / PI double staining; the expression of Nox1, Nox3, Nox4 and p47phox mRNA was detected by Realtime-PCR; Weastern-blot detection of Nox4 protein expression. Results Dexamethasone induced the apoptosis of INS-1 cells in a dose-and time-dependent manner. In the process of apoptosis, Nox1, Nox3, Nox4 and p47phox mRNA expressions were up-regulated, Nox4 protein expression was up-regulated and ROS release was increased with statistical difference Significance (P <0.05). Conclusion Dexamethasone can induce apoptosis of INS-1 cells. The mechanism may be related to the up-regulation of Nox1, Nox3, Nox4 and p47phox expression and activation of oxidative stress.