miR-let-7b-5p对急性胰腺炎AR42J细胞凋亡及自噬的作用研究

Effect of miR-let-7b-5p on apoptosis and autophagy of AR42J cells in acute pancreatitis

目的探讨miR-let-7b-5p对急性胰腺炎AR42J细胞凋亡和自噬的影响。方法以未处理的AR42J细胞作为正常对照组,1×10^(-8) mol/L浓度的雨蛙素诱导大鼠胰腺腺泡细胞(AR42J)24h建立体外急性胰腺炎模型,慢病毒携带miR-let-7b-5p的模拟物(miRNA mimic)、抑制物(miRNA inhibitio)和空病毒转染急性胰腺炎模型细胞作为miR-let-7b-5p过表达组、低表达组和阴性对照组,RT-qPCR验证转染效果,检测凋亡基因Caspase-3 mRNA和自噬基因LC3ⅡmRNA的表达量;Western Blot检测Caspase-3及LC3Ⅱ的蛋白表达。结果与正常对照组相比,阴性对照组的miRlet-7b-5p表达量显著降低(P<0.01),凋亡基因Caspase-3mRNA及蛋白表达显著增加(P<0.01),自噬基因LC3ⅡmRNA及蛋白表达增加(P<0.05)。与阴性对照组相比,过表达组miR-let-7b-5p表达量显著增加(P<0.01),凋亡基因Caspase-3mRNA及蛋白表达均减少(P<0.05),LC3ⅡmRNA及蛋白表达均增加(P<0.05);低表达组miR-let-7b-5p表达量显著降低(P<0.01),凋亡基因Caspase-3mRNA及蛋白表达均增加(P<0.05)、自噬基因LC3ⅡmRNA及蛋白表达均增加(P<0.05)。结论miR-let-7b-5p可调控急性胰腺炎AR42J细胞的凋亡基因Caspase-3和自噬基因LC3Ⅱ的表达,从而影响胰腺细胞的凋亡和自噬。

Objective To investigate the effect of miR-let-7b-5p on apoptosis and autophagy of AR42J cells in acute pancreatitis.Methods Untreated AR42J cells were taken as the normal control group.Pancreatic acinar cells(AR42J)were induced by 1×10^(-8) mol/L caerulein for 24 hours to establish the model of acute pancreatitis in vitro.Lentiviruses carrying miR-let-7b-5p mimic(miRNA mimic),those carrying inhibitor(miRNA inhibition)and those carrying empty virus were transfected into the acute pancreatitis model cells as the over-expressed miR-let-7b-5p group,the low-expressed miR-let-7b-5p group and the negative control group.The transfection effect was verified by RT-qPCR.The mRNA expressions of the apoptosis gene Caspase-3 and the autophagy gene LC3Ⅱwere detected.The protein expressions of Caspase-3 and LC3Ⅱwere detected by Western Blot.Results Compared with the normal control group,the negative control group had significantly decreased expression of miR-let-7b-5p(P<0.01)and significantly increased mRNA and protein expressions of Caspase-3(P<0.01)as well as significantly increased mRNA and protein expressions of LC3Ⅱ(P<0.05).Compared with the negative control group,the over-expression group had the significantly increased expression of miR-let-7b-5p(P<0.01),and significantly decreased mRNA and protein expressions of 3(P<0.05)as well as significantly increased mRNA and protein expressions of LC3Ⅱ(P<0.05).The expression of miR-let-7b-5p significantly decreased(P<0.01)and the mRNA and protein expressions of Caspase-3 and autophagy LC3Ⅱsignificantly increased in the low-expression group(P<0.05).Conclusion miR-let-7b-5p can regulate the expression of the apoptotic gene Caspase-3 and the autophagy gene LC3Ⅱin AR42J cells in acute pancreatitis,thus affecting apoptosis and autophagy of pancreatic cells.