Development of a Novel Three-Dimensional Biocompatible Nanofibrous Scaffold for the Expansion and Hepatogenic Differentiation of Human Bone Marrow Mesenchymal Stem Cells

In this present study, we examined the differentiation potential of human bone marrow derived mesenchymal stem cells (hBMSCs) into hepatocytes on a three-dimentional (3D) nanofibrous scaffold formed by Poly (ε-caprolactone) (PCL), collagen and polyethersulfone (PES). The nanofiber was prepared by the electrospining technique. HBMSCs were isolated using combining gradient density centrifugation with plastic adherence. Flow cytometric analysis was used to identify the isolated MSCs. The performance of the cells on the scaffold was evaluated by scanning electron microscopy (SEM) and MTT assay. The hBMSCs were then cultured in a hepatic differentiation medium containing hepatocyte growth factor (HGF), oncostatin M (OSM) and dexamethasone (DEX) for up to 21 days. The results showed that the isolated hBMSCs expressed specific markers such as CD44, CD166, CD105 and CD13. The integrity of the MSCs was further confirmed by their differentiation potential to osteogenic and adipogenic lineages. Scanning electron micrographs and MTT analysis revealed that the cells adhered and proliferated well on the nanofibrous hybrid scaffolds. Immunocytochemical analysis of albumin and a-fetoprotein (AFP) showed the accumulation of these markers in the differentiated cells on the scaffold. Hepatocyte differentiation was further confirmed  by showing expression of albumin, AFP and cytokeratin-19 (CK-19) at mRNA levels in differentiated cells. In conclusion, the evidences presented in this study show that the engineered scaffold is promising for maintenance of hepatocyte-like cells suitable for transplantation.