过表达microRNA-126诱导INS-1细胞凋亡

Over-expression of MicroRNA-126 Induces Apoptosis of Cultured INS-1 Cells

目的研究过表达microRNA-126对INS-1细胞(大鼠胰岛β细胞株)增殖和凋亡的影响。方法将INS-1细胞分为4组:阴性对照(NC)组、阳性对照(miR-375)组、miR-126过表达(miR-126m)组、miR-126低表达(miR-126in)组,分别转染nonsense segment、miR-375mimic、miR-126mimic、miR-126inhibitor,48h后予胰岛素刺激细胞12h。用实时定量RT-PCR方法检测转染效果;MTS法检测INS-1细胞增殖情况;电子显微镜下观察细胞超微结构的变化。结果 miR-126m组INS-1细胞的miR-126水平较NC组升高(P<0.01),miR-126in组miR-126水平较NC组降低(P<0.05);miR-126m组及miR-375组INS-1细胞活力较NC组及miR-126in组降低(P<0.05);电镜下可见miR-126m组及miR-375组INS-1细胞大量凋亡,细胞核折缝样改变,染色质高度浓缩、边集;NC组及miR-126in组偶见凋亡细胞。结论过表达microRNA-126可以显著降低INS-1细胞活力,诱导细胞凋亡,提示microRNA-126可能与糖尿病的发生有关。

Objective To observe the effect of over- expression of microRNA- 126 on apoptosis and proliferation of INS- 1 cells( rat pancreatic β cell line). Methods INS- 1 cells were divided into four groups:negative control( NC) group,positive control( miR- 375) group,miR- 126 high- expression( miR- 126m)group and miR- 126 low- expression( miR- 126in) group. After transfection with nonsense segment,miR-375 mimic,miR- 126 mimic and miR- 126 inhibitor respectively for 48 h,cells in the four groups were stimulated with synthetic insulin for 12 h. The transfection efficiency was detected by real- time qRT- PCR; proliferation of INS- 1 cells was detected by MTS; and ultrastructural changes were observed by electron microscopy. Results Compared with NC group,miR- 126 level in miR- 126 m group significantly increased( P <0. 01),while it significantly decreased in miR- 126 in group( P < 0. 05). Compared with NC group and miR- 126 in group,cell viability in miR- 126 m group and miR- 375 group was significantly reduced( P <0. 05). There were large numbers of apoptotic cells in miR- 126 m and miR- 375 groups,where cell nuclei became creased or rippled,and chromatin was concentrated and peripherally aggregated under electron microscopy. However,there was only a small number of apoptotic cells in NC and miR- 126 in groups. Conclusion Over- expression of microRNA- 126 could significantly reduce the viability of INS- 1 cells and induce cell apoptosis,suggesting that microRNA- 126 may be associated with the pathogenesis of diabetes mellitus.