EZH2 suppresses hepatocellular differentiation of mouse bone marrow mesenchymal stem cells.

Our previous studies have indicated that mouse bone marrow mesenchymal stem cells (mBMMSCs) have potential to differentiate into hepatocytes with high efficiency. Our study aimed to evaluate the role of the mouse histone methyltransferase enhancer of zeste homolog 2 gene (EZH2) in the hepatocellular differentiation of mBMMSCs. The mBMMSCs isolated from femurs and tibias were cultured in Iscove's modified Eagle's medium (IMEM) supplemented with 10% fetal bovine serum. Hepatocellular differentiation was induced by 20 ng/mL hepatocyte growth factor and 10 ng/mL fibroblast growth factor 4. The mouse histone methyltransferase EZH2 gene was introduced via PLenti-eGFP-EZH2 or PLenti-eGFP-NEO as a control. Hepatocellular-induced mBMMSCs showed lower expression of EZH2 and lower level of histone H3 lysine 27 trimethylation (H3K27me3) in the AFP and FOXa2 gene promoter regions compared to uninduced mBMMSCs. Introduction of EZH2 inhibited hepatocellular induction, reduced both the mRNA and protein levels of AFP and FOXa2, and increased the level of histone H3K27me3 in the AFP and FOXa2 gene promoter regions. In summary, the mouse histone methyltransferase EZH2 gene could suppress hepatocellular differentiation of mBMMSCs by increasing the level of H3K27me3 in the AFP and FOXa2 gene promoter regions.

FGF4 and HGF promote differentiation of mouse bone marrow mesenchymal stem cells into hepatocytes via the MAPK pathway.

Our research demonstrated the potential for mouse bone marrow mesenchymal stem cells (mBMMSCs) to differentiate into hepatocytes in vitro and in vivo. However, the exact mechanism of this process remains unknown. In this study, we investigated the role of the mitogen-activated protein kinase (MAPK) cell-signaling pathway in the differentiation of mBMMSCs into hepatocytes. mBMMSCs were isolated from femurs and tibias, and hepatic differentiation was induced in Isove's modified Eagle's medium supplemented with 10% fetal bovine serum, containing human growth factor and fibroblast growth factor 4. After seven days of induction, morphological characteristics were examined. For inhibition of signaling molecular activities, the inhibitors p38 (SB203580), ERK1/2 (U0126), and MSK1 (H89) were added to the differentiation medium. Real-time polymerase chain reaction and Western blot analysis were used to evaluate the gene expression profiles and protein expression of several markers, including the early specific markers of hepatocytes (AFP and FOXa2), phosphorylated-p38 (p-p38), phosphorylated-ERK1/2 (p-ERK1/2), and phosphorylated- MSK1 (p-MSK1). Expressions of p-p38, p-ERK1/2, and p-MSK1 were effectively inhibited by their respective inhibitors. Expressions of early specific markers, AFP and FOXa2, in the p38, ERK1/2, and MSK1 inhibitor-treated groups were significantly decreased compared to those of the cytokine-induced control. Notably, the expressions of AFP and FOXa2 in the p38 inhibitor group were more obviously reduced than those in the ERK1/2 inhibitor group. The MAPK signaling pathway, especially p38, is sufficient to drive differentiation of mBMMSCs into hepatocytes. This could increase the efficiency of hepatocyte differentiation, which would benefit clinical applications.