Extensively expanded murine‐induced hepatic stem cells maintain high‐efficient hepatic differentiation potential for repopulation of injured livers

Background & aim Shortage of donor hepatocytes limits hepatocyte transplantation for clinical application. Induced hepatic stem cells (iHepSCs) have capacities of self-renewal and bipotential differentiations. Here, we investigated whether iHepSCs could be extensively expanded, and whether they could differentiate into sufficient functional hepatocytes as donors for transplantation therapy after their extensive expansions. Methods Murine extensively expanded iHepSCs (50-55 passages) were induced to differentiate into iHepSC-Heps under a chemically defined condition. iHepSC-Heps were proved for carrying morphological hepatocyte characters and hepatocytic functions including low-density lipoprotein uptake, glycogen storage, CLF secretion, ICG uptake and release, Alb secretion, urea synthesis and metabolism-relative gene expressions respectively. Next, both iHepSCs and iHepSC-Heps were transplanted into Fah-/- mice respectively. Both liver repopulation and alleviation of liver function were compared between two transplantation groups. Results Murine iHepSCs still maintained the capacities of self-renewal and bipotential differentiations after extensive expansion. The efficiency for the functional hepatocyte differentiation from extensively expanded iHepSCs reached to 72.64%. Transplantations of both extensively expanded iHepSCs and iHepSC-Heps resulted in liver engraftment in Fah-/- mice. Survival rate of Fah-/- mice recipients and level of liver repopulation were 50% and 20.32 ± 4.58% respectively in iHepSC-Heps group, while 33% and 10.4 ± 4.3% in iHepSCs group. Conclusions Extensively expanded iHepSCs can efficiently differentiate into hepatocytes in chemical defined medium. Transplantation of iHepSC-Heps was more effective and more efficient than transplantation of iHepSCs in Fah-/- mice. Our results suggested an innovative system to obtain sufficient hepatocytes through hepatic differentiation of iHepSCs generated by lineage reprogramming.