Study On The Anticancer Effect In Vivo And Role Mechanism Of MiRNA-122in Human Hepatocellular Carcinoma

Objective:To investigate anticancer effect in vivo and to validate whether the anti-cancer effect of miR-122on HCC is mediated through regulating Wnt/β-catenin signaling pathways.Methods:HCC xenograft models were established by injecting HepG2-GFP cells to form subcutaneous tumor in nude mice. Sixteen tumor-bearing mice were divided at random into miR-122agomir group and negative control group, each group had8mice. Then5nmol has-miR-122agomir or negative control was respectively injected into subcutaneous tumor every3days. Tumor growth was examined daily for5week. Putative miR-122targets were predicted by algorithms TargetScan, miRanda, and miRBase. Luciferase reporter assay and western blotting were conducted to identify the target of miR-122. The3’-UTR segments of WNT1containing the miR-122 binding sites were amplified by PCR and the luciferase activity in the transfected cells was assayed. WNT1mRNA level was quantified by using RT-PCR. Protein levels of Wntl, β-catenin and TCF-4were detected by using Western blotting respectively.Results:(1) Compared with the negative control group, over-expression miR-122inhibited tumor growth in xenograft models of nude mice (P<0.05).(2) miR-122suppressed the luciferase activity of the pmiR-WNT1-3’UTR-wt by approximately50%compared to the negative control, while mutation or removal of the miR-122binding site using siRNA or miR-122inhibitor blocked the suppressive effect (P<0.05).(3) Over-expressed miR-122down-regulated the protein levels of WNT1, β-catenin and TCF-4(P<0.05).Conclusions:(1) Over expression of miR-122inhibits the growth of of HepG2xenograft models in vivo.(2) WNT1is the target gene of miR-122, miR-122inhibites HCC growth mainlythrough regulating Wnt/β-catenin-TCF signaling pathway.