饱和脂肪酸对胰岛β细胞凋亡和细胞周期进程的影响

Saturated fatty acids trigger apoptosis and inhibit cell cycle progression in insulinoma cells

目的探讨饱和脂肪酸棕榈酸(pal mitate acids,PA)对MIN6胰岛β细胞生长、增殖及凋亡的影响,研究诱导MIN6细胞凋亡,抑制细胞周期进程的可能细胞分子机制。方法采用MTT法检测细胞活力,流式细胞仪测定细胞周期,Annexin-Ⅴ/FITC/PI双标记流式细胞术检测细胞凋亡。应用Western blot检测细胞周期相关蛋白cyclin D1和CDK4的表达水平,以及抗凋亡蛋白家族(IAP)XIAP和cIAP-2的表达水平。结果和对照组相比:①不同浓度PA显著抑制MIN6细胞增殖(P<0.01);②不同浓度PA作用均呈时间与剂量依赖性诱导MIN6细胞凋亡(P<0.01);③PA亦能显著抑制细胞周期进程,使MIN6细胞周期更多滞留在G0/G1期(P<0.01),G2/M期与S期细胞比例降低(P<0.01);④用Western blot检测发现:和对照组相比,PA降低磷酸化AKT、XIAP和cIAP-2的表达水平(P<0.01),以及降低细胞周期蛋白CDK4、cyclin D1(P<0.01)的表达水平。结论PA能抑制胰岛β细胞的生长、增殖和诱导凋亡,可能是通过降低cyclin D1和CDK4活性,使细胞滞留在G0/G1期,抑制细胞周期进程,影响其复制;同时降低抗凋亡蛋白XIAP和cIAP-2的活性,促使细胞的凋亡。

Objective To investigate the effect of saturated fatty acid palmitate acids （PA） action on viability, proliferation and apoptosis of murine insulinoma MIN6 cells, and the possible intracellular pathways activated by PA. Methods Cell viability was evaluated with MTT reduction conversion assay. The Annexin-V/ FITC cell death detection kit was used to monitor PA-induced apoptosis. （IAP） family （XIAP and cIAP-2）, cyclin D1 and CDK4 were further detected using Western blot. Results （1） PA of different concentration significantly inhibited the proliferation of MIN6 cells （P〈0. 01）. （2） PA increased MIN6 cell apoptosis in time- and concentration-dependent manners （P〈0.01）. （3） The G0/G1 cell cycle arrest was also induced by PA （P〈0.01）, whereas MIN6 cells in S and G2/M phase were decreased （P〈0.01）. （4） Western blot revealed that PA reduced the level of AKT phosphorylation （P〈0.01）, decreased the expression of XIAP （P〈0.01） and cIAP-2 （P〈0.01） compared to BSA-treated control cells. PA also reduced protein expression of cell cycle-related molecules, CDK4 （P〈0.01）, and cyclin D1 （P〈0.01） compared to BSA-treated control cells. Conclusion Our results thus suggest that PA directly affects pancreatic β-cell cell proliferation, possibly by inhibiting Akt kinase and reducing the level of cyclin D1/CDK4 in the cells, which results in arresting in progression through the G1 phase to the S phase of the cell cycle. PA triggers pancreaticβ-cell cell apoptosis, possibly by reducing the levels of XIAP and cIAP-2 in the cells.