Differentiation of rat bone marrow mesenchymal stem cell into mature hepatocyte-like cell in vitro

Objective To establish and optimize the method of isolation and cultivation of rat bone marrow mesenchymal stem cells(MSCs),and to investigate the condition and capability of hepatic differentiation of MSCs.(Methods) Rat bone marrow cells were collected from rats of 5～6 week and mononeuclear cells were isolated by Percoll density gradient centrifugation.MSCs were cultivated and purified by adherence method.Morphology,RT-PCR and immunocytochemistry were used to identify the role of substrates,the kinds of the cytokines and concentrations of cytokines on the hepatic differentiation potential.Results The quality and quantity of the isolated bone marrow mononuclear cells reached the peak by using 57% Percoll density gradient centrifugation.Discarding the suspending cells after 24h incubation and subculturing the cells with low seeding density were helpful for acquiring MSCs with improved reproductive capability and active function.MSCs could be successfully induced under HGF/FGF-4 induction on the substrate of(10 μg/mL) fibronectin.MSCs exhibited round in shape after differentiation,instead of fibroblast-like morphology before.Albumin mRNA and protein were positively expressed in MSCs,withoutdetection of alpha-fetoprotein(AFP).Conclusion The optimization of rats' age,the density of Percoll,the methods of cell cultivation are conducing to obtaining a pure population of MSCs with good differential activity.MSCs are inclined to differentiate into mature hepatocyte-like cells under the induction of HGF/FGF-4.