No abstract

MicroRNAs (miRNAs) are evolutionary conserved small RNAs that post‐transcriptionally regulate the expression of target genes. To date, the role of miRNAs in liver development is not fully understood. By using an experimental model that allows the induced and controlled differentiation of mouse fetal hepatoblasts (MFHs) into mature hepatocytes, we identified miR‐148a as a hepatospecific miRNA highly expressed in adult liver. The main finding of this study revealed that miR‐148a was critical for hepatic differentiation through the direct targeting of DNA methyltransferase (DNMT) 1, a major enzyme responsible for epigenetic silencing, thereby allowing the promotion of the “adult liver” phenotype. It was also confirmed that the reduction of DNMT1 by RNA interference significantly promoted the expression of the major hepatic biomarkers. In addition to the essential role of miR‐148a in hepatocyte maturation, we identified its beneficial effect through the repression of hepatocellular carcinoma (HCC) cell malignancy. miR‐148a expression was frequently down‐regulated in biopsies of HCC patients as well as in mouse and human HCC cell lines. Overexpressing miR‐148a led to an enhancement of albumin production and a drastic inhibition of the invasive properties of HCC cells, whereas miR‐148a silencing had the opposite consequences. Finally, we showed that miR‐148a exerted its tumor‐suppressive effect by regulating the c‐Met oncogene, regardless of the DNMT1 expression level. Conclusion: miR‐148a is essential for the physiology of the liver because it promotes the hepatospecific phenotype and acts as a tumor suppressor. Most important, this report is the first to demonstrate a functional role for a specific miRNA in liver development through regulation of the DNMT1 enzyme. (Hepatology 2013;53:1153–1165)

a b s t r a c tExtracellular vesicles (EVs) contain microRNAs (miRNAs). However, the exact molecular mechanisms of the recruitment of miRNAs in EVs are not well characterized. Based on proteomic analysis, we identified that silencing of Annexin A2 (ANXA2) significantly decreased the amount of miRNAs in EVs. In addition, microarray analysis revealed that ANXA2 regulated the loading of miRNAs into EVs in a sequence independent manner. Lastly, immunoprecipitation analysis confirmed that ANXA2 could bind miRNAs in EVs in the presence of Ca 2+ . These observations demonstrate that ANXA2 plays an important role in the packaging process of miRNAs into EVs.

The MAPK MEK/ERK pathway is often upregulated in cancer cells and represents an attractive target for development of anticancer drugs. Only few data concerning the specific functions of ERK1 and 2 are reported in the literature. In this report, we investigated the specific role of ERK1 and 2 in liver tumor growth both in vitro and in vivo. DNA synthesis and cells in S phase analysed by flow cytometry, correlated with strong inhibition of Cdk1 and cyclin E levels, are strongly reduced after exposure to the MEK inhibitor, U0126. We obtained a significant reduction of colony formation in soft agar assays and a reduction in the size of tumor xenografts in nude mice treated with U0126. Then, we could specifically abolished ERK1 or 2 expression by small-interfering RNA (siRNA) and demonstrated that ERK2 knockdown but not ERK1 interferes with the process of replication. Moreover, we found that colony formation and tumor growth in vivo were significantly inhibited by targeting ERK2 using stable chemically modified siRNA. Taken together, our results emphasize the importance of the MEK/ERK pathway in liver cancer cell growth in vitro and in vivo and argue for a crucial role of ERK2 in this regulation.