Efficient generation of hepatocyte-like cells from rat bone marrow mesenchymal stem cells in vitro

Abstract

Recent studies indicated that stem-cell–derived hepatocyte-like cells are superior to stem cells in therapy for liver failure. Our aim is to present a modified one-step protocol for high-efficiency in vitro generation of hepatocyte-like cells from rat bone marrow mesenchymal stem cells (rBMSCs). rBMSCs were cultured under optimal differentiation conditions. Hepatic differentiation was evaluated by light microscopy (for morphological analysis), reverse-transcription polymerase chain reaction (for expression of hepatocyte-specific genes), and immunocytochemical and immunofluorescence analyses of hepatic proteins, such as alpha-fetoprotein and albumin. Functional assays comprised periodic acid-Schiff staining, supernatant urea assay, and albumin radioimmunoassay. The results show that differentiated cells exhibited characteristic hepatocyte morphology, expressed hepatocyte-related genes (as shown by reverse-transcription polymerase chain reaction), and displayed antibody-detectable expressions of alpha-fetoprotein (100% at day 8) and albumin (>70% at 2 weeks). Most differentiated hepatocyte-like cells showed evidence of glycogen synthesis and storage, as shown by periodic acid-Schiff staining. Albumin and urea were detected in supernatants. Our one-step protocol induced the efficient differentiation of rBMSCs into functional hepatocyte-like cells in a two-dimensional in vitro model and may be useful for cell transplantation therapy for liver failure.