Accumulating evidence supports an important role for the hepatitis B virus x protein (HBx) in the pathogenesis of hepatitis B virus (HBV)-induced hepatocellular carcinoma (HCC), but the underlying mechanisms are not entirely clear. Here, we identified a novel long noncoding RNA (lncRNA) DBH-AS1 involved in the HBx-mediated hepatocarcinogenesis. The levels of DBH-AS1 were positively correlated with hepatitis B surface antigen (HBsAg) and tumor size in HCC tissues. Functionally, transgenic expression of DBH-AS1 significantly enhanced cell proliferation and tumorigenesis, whereas short hairpin RNA knockdown of DBH-AS1 caused an inhibition of cell proliferation. Mechanistically, overexpression of DBH-AS1 induced cell cycle progression by accelerating G1/S and G2/M transition concomitantly with upregulation of CDK6, CCND1, CCNE1 and downregulation of p16, p21 and p27. We also found that enhanced DBH-AS1 expression inhibited serum starvation-induced apoptosis of HCC cells. In contrast, suppressed DBH-AS1 expression had opposite effects. Furthermore, DBH-AS1 was shown to activate MAPK pathway. We also provide evidence that DBH-AS1 could be significantly induced by HBx protein and markedly down-regulated by p53. Thus, we concluded that DBH-AS1 can be induced by HBx and inactivated by p53, and consequently promote cell proliferation and cell survival through activation of MAPK signaling in HCC. Our study suggests that DBH-AS1 acts as an oncogene for HCC.

BackgroundHepatocyte nuclear factor-3β (HNF-3β) plays a critical role in hepatocyte differentiation and controls liver-specific gene expression during the development of hepatocellular carcinoma (HCC), but the molecular basis of this process has not been fully elucidated. microRNAs (miRNAs) are powerful, post-transcriptional regulators of gene expression. Whether miRNAs can impact the effects of HNF-3β in HCC is still unknown.MethodsHNF-3β and miR-141 expression levels were detected in HepG2 cells, using real-time quantitative RT-PCR (qRT-PCR). Luciferase reporter assays and Western blots were used to validate HNF-3β as a direct target gene of miR-141. Cell proliferation, invasion, and apoptosis were also examined to confirm whether miR-141 could impact on HNF-3β in HCC.ResultsIn this study, we found that HNF-3β protein levels were consistently upregulated in HCC clinical tissues compared with matched normal adjacent tissues. However, the mRNA levels of HNF-3β varied in random tissues, suggesting that a post-transcriptional mechanism was involved in its regulation. We used bioinformatic analyses to search for miRNAs that could potentially target HNF-3β, and identified specific targeting sites for miR-141 in the 3′-untranslated region (3′-UTR) of the HNF-3β gene. By overexpressing miR-141 in HepG2 cells, we experimentally validated that miR-141 directly regulated HNF-3β expression. Furthermore, the biological consequences of targeting HNF-3β by miR-141 were examined using cell proliferation, invasion and apoptosis assays in vitro. We demonstrated that the repression of HNF-3β by miR-141 suppressed the proliferation and invasion and promoted the apoptosis of HepG2 cells.ConclusionsmiR-141 functions as a tumor suppressor in HCC cells through the inhibition of HNF-3β translation.

Non-alcoholic fatty liver disease (NAFLD) affects obesity-associated metabolic syndrome, which exhibits hepatic steatosis, insulin insensitivity and glucose intolerance. Previous studies indicated that hepatic microRNAs (miRs) play critical roles in the development of NAFLD. In this study, we aim to explore the pathophysiological role of miR-194 in obesity-mediated metabolic dysfunction. Our findings show that the high fat diet or palmitic acid treatment significantly increase hepatic miR-194 levels in vivo and in vitro. Silence of miR-194 protects palmitic acid-induced inflammatory response in cultured hepatocytes, and attenuates structural disorders, lipid deposits and inflammatory response in fatty liver. MiR-194 inhibitor also improves glucose and insulin intolerance in obese mice. Through dual luciferase assay, we demonstrate that miR-194 directly binds to FXR/Nr1h4 3'-UTR, and inhibits gene expression of FXR/Nr1h4. Furthermore, overexpression of miR-194 downregulates FXR/Nr1h4 in cultured hepatocytes, but miR-194 inhibitor reversely increases FXR/Nr1h4 expression in obese mouse liver tissues. On the contrast, silence of FXR/Nr1h4 abolishes the hepatic benefits in obese mice treated with miR-194 inhibitor. Present study provides a novel finding that suppression of miR-194 attenuates dietary-induced NAFLD via upregulation of FXR/Nr1h4. The findings suggest miR-194/FXR are potential diagnostic markers and therapeutic targets for NAFLD.