Hepatic differentiation of mesenchymal stem cells derived from rats in three-dimensional microenvironment formed by hanging drops

AIM: To establish an optimal differentiated three-dimensional microenvironment formed by hanging drops for the high efficiency of hepatic differentiation from rat mesenchymal stem cells(rMSCs).METHODS: rMSCs were cultured in the hanging drops, which provided a three-dimensional microenvironment, for 21 days in the presence of hepatocyte growth factor(HGF, 20 μg/L). The expression of albumin(ALB), alpha-fetoprotein(AFP) and cytokeratin-18(CK-18) was detected by RT-PCR and immunofluorescence staining at 7th, 14th and 21st days. The secretion of albumin in the culture supernatants was measured by ELISA. RESULTS: rMSCs were aggregated in spheroid with a tubiform medium altitude in the center. In rMSCs cultured in the hanging drops with HGF, the expression of albumin, AFP and CK-18 was all detectable by RT-PCR and immunofluorescence staining at 7th day. The production of albumin in the cells cultured in the hanging drops with HGF was 50.25±5.32, 55.03±7.45 and 54.92±3.18(ng·dish^-1·d^-1) at 7th, 14th and 21st days, respectively, significantly higher than that in the cells in the plate cultivation with or without HGF induction at corresponding time points(P<0.01).CONCLUSION: In the presence of HGF, rMSCs are induced to differentiate into hepatocyte-like cells cultured in the hanging drops, where the three-dimensional spheroidal cultures are promising microenvironment for hepatic transdifferentiation of MSCs.