miR let-7f对流行性乙型脑炎病毒在神经元细胞中复制的调控作用

miR let-7f inhibited Japanese encephalitis virus replication in neurons

目的探讨miR let-7f对流行性乙型脑炎病毒(Japanese encephalitis virus,JEV)在神经元细胞中复制的影响。方法培养人神经母细胞瘤SH-SY5Y细胞,miRNA芯片检测SH-SY5Y细胞感染JEV后miRNA表达谱,荧光定量RT-PCR检测确认miR let-7f表达情况;实验随机分为miR let-7f mimics组、miR let-7f inhibitor组和miRNA conrol组,分别将miR let-7f mimics、inhibitors和miRNA conrol转染SH-SY5Y细胞后感染JEV,荧光定量RT-PCR和空斑法检测病毒复制变化情况。结果SH-SY5Y细胞感染JEV后差异表达的miRNA有37个,上调的有28个,下调的有9个,其中miR let-7f表达明显升高(3.51倍);SH-SY5Y细胞感染JEV 12h后miR let-7f表达明显上升(3.67倍),与miRNA control组和miR let-7f inhibitor组相比,miR let-7f mimics组细胞培养上清中病毒滴度降低,细胞内JEV RNA也显著减少,差异具有统计学意义(P<0.025)。结论miR let-7f在神经元细胞中能够有效抑制JEV复制,在开发JEV感染的分子治疗方面具有潜在应用价值。

Objective To analyze the role of miR let-7f on Japanese encephalitis virus(JEV)infection and replication in neurons.Methods SH-SY5Y cells were cultured,and miRNA chip was used to detect differentially expressed miRNAs between SH-SY5Y cells after JEV infection and normal cells.The relative expression level of miR let-7f related to JEV infection was determined by qRT-PCR.SH-SY5Y cells were divided into three groups randomly which transfected with miR let-7f mimics,inhibitors,and miRNA conrol respectively,and then infected with JEV.Infected cells and supernatant were harvested and then qRT-PCR and plaque forming test were used to detect the levels of virus replication.Results A total of 28 miRNAs were up-regulated while 9 miRNAs down-regulated in SH-SY5Y cells infected with JEV compared with non-infected cells,of which miR let-7f up-regulated 3.51 folds.miR let-7f was confirmed to be up expressed 3.67 folds in SH-SY5Y cells after JEV infection 12h.After miR let-7f mimics were transfected,JEV RNA in cells and virus titer in supernatant was lower than miR let-7f inhibitors/miRNA conrol groups,and the differences showed statically significant(P<0.025).Conclusion miR let-7f could inhibit JEV replication in neurons which might be as a potential molecular target for novel therapy.