高浓度葡萄糖对体外培养小鼠胰岛细胞NF-κB的表达及细胞凋亡的影响

Effects of high glucose on expression of NF-κB in mouse islet cells and apoptosis in vitro

目的:探讨核因子κB(NF-κB)在高浓度葡萄糖诱导胰岛细胞凋亡中的作用。方法:对分离纯化的小鼠胰腺胰岛细胞进行体外培养,按DMEM培养液里葡萄糖浓度不同,分为6组:G1组(5.6mmol·L-1葡萄糖)、G2组(7.8mmol·L-1)、G3组(11.1mmol·L-1)、G4组(16.7mmol·L-1)、G5组(22.2mmol·L-1)及G6组(27.6mmol·L-1)。采用放射免疫法检测各组胰岛细胞胰岛素分泌水平;免疫组织化学染色法检测NF-κB活性;免疫荧光法检测细胞色素C释放和细胞核染色检测细胞凋亡。结果:当葡萄糖浓度5.6～11.1mmol·L-1时,随着葡萄糖浓度升高,胰岛细胞胰岛素分泌逐渐增加(P<0.05),NF-κB的活性逐渐增加(P<0.05),但此时细胞凋亡未见显著的变化(P>0.05),细胞色素C释放的组间比较差异无显著性(P>0.05)。当葡萄糖浓度≥16.7mmol·L-1时,与葡萄糖浓度≤11.1mmol·L-1各组比较,NF-κB表达明显增加,细胞色素C的释放也逐渐增强(P<0.05),胰岛细胞凋亡百分率同时也呈明显增加趋势(P<0.05)。结论:高浓度葡萄糖可以诱导胰岛细胞NF-κB表达增加,细胞色素C释放增强,同时诱导胰岛细胞凋亡,提示NF-κB的激活与胰岛细胞凋亡可能存在必然的联系,抑制NF-κB活性可能对于保护胰岛细胞有重要作用。

Objective To study the role of NF-kappaB （NF-κB） on islet cell apoptosis in high concentration glucose in vitro. Methods Pancreatic islet cells were isolated from Kunming mice and these cells were cultivated in medium with different eoncerntrationsofglucose： G1 （5.6mmol·L^-1）, G2 （7.8mmol·L^-1）, G3 （11.1mmol·L^-1）, G4 （16.7mmol·L^-1）. G5 （22.2mmol·L^-1）, G6（27.6 mmol·L^-1）. After the islet cells were cultivated for 72 h, insulin secretion was evaluated by radio-immunity technique. The NF-κB expression was detected by immunocytochemistry and cytochrome C （ Cyt. C ） release was detected by immunofluorescenee technique.Apoptosis of islet cells was determined by Hoechst33342 assay. Results Insulin secretion was enhanced and NF-κB activity was increased （P〈0.05） when the concentrations of glucose were 5.6-11.1mmol·L^-1, but Cyt. C release and apoptosis didn＇t change. When the concentration of glucose exceeded 16.7mmol·L^-1, the apoptotic percentages of islet cells were increased signficantty accompanying with the glucose concentration elevation compared with G1 , G2 and G3 groups （P〈0.05）, the NF-κB activity and Cyt. C relesae were significantly increased （P〈0.05）.Conclusion High glucose can induce the apoptosis of islet cells and decrease insulin secretion, enhance the activity of NF-κB and Cyt C release. It suggests that the activation of NF-κB may be related to apoptosis of islet cells, and the inhibition of NF-κB may protect islet cells.