Abstract
Pancreatic acinar cells AR42J-B13 can transdifferentiate into hepatocyte-like cells permissive for efficient hepatitis B virus (HBV) replication. Here, we profiled miRNAs differentially expressed in AR42J-B13 cells before and after transdifferentiation to hepatocytes, using chip-based microarray. Significant increase of miRNA expression, including miR-21, miR-22, and miR-122a, was confirmed by stem-loop real-time PCR and Northern blot analyses. In contrast, miR-93, miR-130b, and a number of other miRNAs, were significantly reduced after transdifferentiation. To investigate the potential significance of miR-22 in hepatocytes, we generated cell lines stably expressing miR-22. By 2D-DIGE, LC-MS/MS, and Western blot analyses, we identified several potential target genes of miR-22, including parathymosin. In transdifferentiated hepatocytes, miR-22 can inhibit both mRNA and protein expression of parathymosin, probably through a direct and an indirect mechanism. We tested two computer predicted miR-22 target sites at the 3′ UTR of parathymosin, by the 3′ UTR reporter gene assay. Treatment with anti-miR-22 resulted in significant elevation of the reporter activity. In addition, we observed an in vivo inverse correlation between miR-22 and parathymosin mRNA in their tissue distribution in a rat model. The phenomenon that miR-22 can reduce parathymosin protein was also observed in human hepatoma cell lines Huh7 and HepG2. So far, we detected no major effect on several transdifferentiation markers when AR42J-B13 cells were transfected with miR-22, or anti-miR-22, or a parathymosin expression vector, with or without dexamethasone treatment. Therefore, miR-22 appears to be neither necessary nor sufficient for transdifferentiation. We discussed the possibility that altered expression of some other microRNAs could induce cell cycle arrest leading to transdifferentiation.
Highlights
Transdifferentiation from one differentiated cell type to another can occur in vivo and in vitro
AR42J-B13 is a rat pancreatic acinar cell line, which can transdifferentiate into hepatocytes by treatment with dexamethasone (Dex) and oncostatin M (OSM) [3]
We identified miR-22 as a microRNA species highly elevated in transdifferentiated hepatocytes
Summary
Transdifferentiation from one differentiated cell type to another can occur in vivo and in vitro. AR42J-B13 is a rat pancreatic acinar cell line, which can transdifferentiate into hepatocytes by treatment with dexamethasone (Dex) and oncostatin M (OSM) [3]. Transdifferentiated hepatocytes of AR42J-B13 cells expressed several liverspecific markers and transcription factors [3,4]. These rodent hepatocyte-like cells can support very efficient cross-species replication of human hepatitis B virus – a hepatotropic virus [3,4,5]. MicroRNAs can inhibit gene expression through base-pairing of their seed sequences to the 39UTR of target genes, leading to translational suppression or mRNA degradation. Whether miRNAs could play a role in hepatic transdifferentiation remains to be investigated
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