格列吡嗪和格列奇特对MIN6细胞的影响

Effects of glipizide and gliclazide on MIN6 cells

目的:研究不同磺脲类药物对胰岛素分泌细胞的影响及机制。方法:不同浓度格列吡嗪和格列奇特干预MIN6细胞一定时间后,MTT法检测细胞活力变化,运用ROS捕获剂双氢溴乙啶(HE)、双氢-乙酰乙酸二氯荧光黄(DCFH-DA)和双氢罗丹明123(DHR123)孵育细胞,用荧光显微镜观察细胞内荧光、流式细胞仪检测MIN6细胞的平均荧光强度(MFI)而测得细胞内ROS水平。用Annexin-vFITC和PI双标记细胞凋亡检测法通过流式细胞仪测定细胞凋亡情况。结果:格列吡嗪干预MIN6细胞后,细胞活力明显被抑制(P<0.05);细胞内的MFI明显增加;细胞凋亡率明显增加;而且,细胞活力下降、MFI和细胞凋亡率与药物作用浓度和时间呈正相关。而格列奇特干预MIN6细胞后,细胞活力无明显抑制(P>0.05);细胞内的MFI无明显增加;细胞凋亡率亦无明显增加。结论:格列吡嗪可诱导MIN6细胞发生氧化应激,诱导MIN6细胞凋亡,说明临床上磺脲类药物治疗糖尿病出现失效可能与磺脲类药物诱发胰岛细胞衰竭相关,然而,格列奇特不诱导MIN6细胞损害,说明并非所有磺脲类药物对β细胞都有损害作用。

Objective： To explore the effects of different sulphonylureas on pancreatic beta-cells. Methods： MIN6 cells were cultured in DMEM and managed with different concentrations of glipizide and gliclazide, and cell viability was detected by M3T. Dihydroethidium （HE）, 2,7-dichlo- rofluorescein diacetate （ DCFH-DA ） or dihydrorhodamine 123 （DHR123） was used as reactive oxygen species （ROS） capture. The mean fluorescent intensity of ethidium, 2 ＇, 7＇-dichlorofluorescein or rhoda- mine 123, product of intracellular oxidation by the above probes, was de- tected by fluorescent microscopy and flow cytometry. Cell apoptosis was analyzed by Annexin-V-FITC. Results：In glipizide treated MIN6 cells, cell viability was inhibited and intracellular fluorescent intensity and cell apoptosis were markedly elevated in a dose-and time-dependent fashion （ P 〈 0.05 ）. No significant changes occurred in gliclazide treated MIN6 cells concerning the cell viability,intracenular fluorescent intensity and cell ap- optosis as compared with the normal control group. Conclusion：Glipizide may induce MIN6 cell oxidative st2yess and cell apoptosis, suggesting that failure of therapeutic response to sulphonylureas documented in clinical trials may be due to the sulphonylurea-induced reduction of beta-cen mass. Contrarily, gliclazide refusing to cause damage to MIN6 cells implies that not individual sulphonylurea compound will result in injury of pancre- atic beta-cells..