Generating functional hepatocyte-like cells (HLCs) from mesenchymal stem cells (MSCs) is of great urgency for bio-artificial liver support system (BALSS). Previously, we obtained HLCs from human umbilical cord-derived MSCs by overexpressing seven microRNAs (HLC-7) and characterized their liver functions in vitro and in vivo. Here, we aimed to screen out the optimal miRNA candidates for hepatic differentiation. We sequentially removed individual miRNAs from the pool and examined the effect of transfection with remainder using RT-PCR, periodic acid-Schiff (PAS) staining and low-density lipoprotein (LDL) uptake assays and by assessing their function in liver injury models. Surprisingly, miR-30a and miR-1290 were dispensable for hepatic differentiation. The remaining five miRNAs (miR-122, miR-148a, miR-424, miR-542-5p and miR-1246) are essential for this process, because omitting any one from the five-miRNA combination prevented hepatic trans-differentiation. We found that HLCs trans-differentiated from five microRNAs (HLC-5) expressed high level of hepatic markers and functioned similar to hepatocytes. Intravenous transplantation of HLC-5 into nude mice with CCl4 -induced fulminant liver failure and acute liver injury not only improved serum parameters and their liver histology, but also improved survival rate of mice in severe hepatic failure. These data indicated that HLC-5 functioned similar to HLC-7 in vitro and in vivo, which have been shown to resemble hepatocytes. Instead of using seven-miRNA combination, a simplified five-miRNA combination can be used to obtain functional HLCs in only 7 days. Our study demonstrated an optimized and efficient method for generating functional MSC-derived HLCs that may serve as an attractive cell alternative for BALSS.
Keywords: BALSS; CCl4; hepatic differentiation; mesenchymal stem cells; miRNA.
© 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.