The Role Of Hbx-Regulated MIR-152-WNT-1in Liver Cancer Proliferation

MicroRNAs (miRNAs) are21-23nt long non-coding RNA sequences, which can regulate gene expression by binding to the3’-untranslated region (UTR) of the target mRNA, miRNAs may involved in physiological processes, like cell proliferation, cell death and host immune response,miRNAs also play a role in viral infections and the development of cancer. MicroRNAs and their target genes could be used as biological markers for clinical diagnosis, and provide potential therapeutic strategies to certain diseases. carcinoma is one of the most common malignant tumors, Hepatitis B virus (HBV) infection constitute the important risk factor during the development of HCC. HBV encode the HBV X protein, which is thought to play a important role in the molecular pathogenesis of initiation, development and metastasis of HBV related HCC, DNMTl is the important target gene of miR-152which could directly targeting the3’untranslated regions of DNMT1, HBX can down regulate the level of miR-152, The down expression of miR-152in liver cell lines resulted in a marked upexpression of DNMTl at both the mRNA and protein levels. Inhibition of miR-152caused global DNA hypermethylation and increased the methylationlevels of two tumor suppressor genes, GSTPland E-cadherin.other research observed that inhibition of miR-152resulted in an increase of the relative percentage of S-phase and G2/M-phase cells. These results suggest that alteration expression of miR-152is important for cell cycle progression.The wnt/β-catenin signaling pathway play a important role in the generation/differentiation of many tissues, Up-regulation of Wnt-1protein has been reported in hepatitis B virus (HBV)-related and hepatitis C virus (HCV)-related hepatocellular carcinoma (HCC) tissues and cell lines. It is known to play a fundamental role insignaling cancer progression, Up-regulation of Wnt-1expression has been shown to correlate with hepatocyte growth stimulated by HCV core protein.Hepatitis B virus X protein is essential for the activation of Wnt/beta-catenin signaling in hepatoma cells, we have found that Wnt-lis the direct target gene of miR-152, alteration expression of miR-152may have the effect on the expression of Wnt-1protein. However, little is known whether upexpression of Wnt-1is regulated by the level of miR-152which is down regulated by HBX and whether there is relationship between the miR-152-WNT-1associated with HBX and hepatic cell apoptosis. These and other data demonstrate that miRNAs play a substantial role in the pathogenesis of cancers and that expression profiling of miRNAs can be used as potential diagnostic/prognostic markers. However, to date very limited information has been published about miRNAs that are specifically involved in the initiation/development/metastasis of HBx related HCCBased on these findings, AdenoVirus which combined with HBx gene was used to transfer HepG2cells, we investigated weather the Up-regulation of Wnt-1expression has been shown to correlate with hepatocyte growth stimulated by miR-152regulated by HBX. We inserted HBX coding domain into adenovirus system. pAd-HBX was transfected into HepG2cells with different concentrations and the transfected time was48h, Real-time PCR and Western-blot was used to verify the expression of HBX; Real-time PCR, Western-blot and FIA was used to analysis the altered expression of miR-152, Wnt-1and cell cycle, real-time PCR,Western-blot and FIA was used to analysis the altered expression of Wnt-1and cell cycle after the infection of miR-152inhibitor.The level of miR-152was down-regulated and the expression of mRNA and protein of Wnt-1was up-regulated via the infection of adv-HBX; the proportion of cells in G1stage decreased with cells in S stage increased after the infection of adv-HBX; miR-152inhibitor up-regulated the expression of mRNA and protein of Wnt-1; miR-152inhibitor also decreased the proportion of cells in G1stage and increased the proportion of cells in S stage.Altered expression of miR-152-Wnt-1mediated by HBx may shed new insight into indicating the molecular mechanism of the development and progression of HBV-HCC.