波动性高糖对胰岛β细胞增殖及Skp2-p27表达的影响

Effect of intermittent high glucose on INS-1 ceil proliferation and the expression of skp2-p27

目的探讨波动性高糖对INS-1细胞增殖、凋亡及对细胞周期进程的影响，并研究其可能的分子机制。方法采用细胞计数试剂盒（cell counting kit-8）检测细胞增殖活性，流式细胞仪测定细胞周期及细胞ROS水平，Annexin—V／PI双标流式细胞术检测细胞凋亡。应用Western印迹检测细胞周期调控蛋白p27及skp2的表达水平。结果（1）波动性高糖及持续性高糖均明显抑制INS-1细胞的生长，且波动性高糖对细胞增殖的抑制作用更为显著。（2）波动性高糖及持续性高糖均明显增加INS-1细胞的凋亡，且波动性高糖作用更为显著。（3）波动性高糖及持续性高糖能明显抑制细胞周期进程，使INS-1细胞周期更多滞留在G0／G1期，G2／M期与S期细胞比例下降，波动性高糖作用更显著。（4）波动性高糖及持续性高糖均能显著增强细胞周期调控蛋白p27的表达，同时减弱skp2蛋白的表达水平。结论波动性高糖较持续性高糖更能够抑制INS-1细胞的增殖和诱导凋亡，可能是通过减弱Skp2蛋白的表达水平，增加p27蛋白的活性，使细胞阻滞在G0／G1期，抑制细胞周期进程，从而减弱细胞的增殖活性。

Objective To investigate the effect of intermittent high glucose on proliferation, apoptosis, and cell cycle progression of INS-1 cells, and the possible intracellular pathways activated by intermittent high glucose. Methods Cell viability was evaluated by cell counting kit, the cell cycle was determined by flow cytometry, Annexin-V/PI double-labeled cell apoptosis detection kit was used to monitor cell apoptosis. Cell cycle related protein Skp2 and p27 expressions were detected by Western blot. Results （ 1 ） Both intermittent and constant high glucose significantly inhibited the growth of INS-1 cells, and the former effect was more significant. （2） Intermittent and constant high glucose levels significantly increased apoptosis in INS-1 cells, and the former effect was more significant. （3） Intermittent and constant high glucose levels significantly inhibited the cell process, the G0/G1 cell cycle arrest also was induced by intermittent high glucose, resulting in lowered proportion of the G2/M phase and S phase of INS-1 cells. （4） Intermittent and constant high glucose significantly decreased the level of protein Skp2 and increased the level of cell cycle related protein p27. Conclusion Intermittent high glucose levels affect INS-1 cell growth and proliferation, as well as induce cell apoptosis, probably by decreasing the level of protein Skp2 and increasing the level of p27 in the cells, resulting in arrest of progression through the G1 phase to the S phase of INS- 1 ceils, and thus impairment of cell proliferation.