The differentiation and isolation of mouse embryonic stem cells toward hepatocytes using galactose-carrying substrata

• Published in: Biomaterials 2012-02, Vol.33 (5), p.1414-1427
• Main Authors: Meng, Qingyuan, Haque, Amranul, Hexig, Bayar, Akaike, Toshihiro
• Format: Article
• Language: English
• Subjects: Adsorption - drug effects; Advanced Basic Science; Animals; Asialoglycoprotein Receptor - metabolism; Cadherins - metabolism; Cell Differentiation - drug effects; Cell Separation - methods; Cells, Cultured; Dentistry; Disaccharides - pharmacology; E-cadherin; Embryonic stem cells; Embryonic Stem Cells - cytology; Embryonic Stem Cells - drug effects; Embryonic Stem Cells - metabolism; Endoderm - cytology; Endoderm - drug effects; Endoderm - metabolism; Extracellular matrix; Galactose - pharmacology; Galactose-carrying polymer; Hepatocyte differentiation; Hepatocytes - cytology; Hepatocytes - drug effects; Hepatocytes - metabolism; Immobilized Proteins - metabolism; Mice; Phenotype; Polystyrenes - pharmacology; Receptors, Fc - metabolism; Surface Properties - drug effects; Vinyl Compounds - pharmacology
• ISSN: 0142-9612; 1878-5905
• DOI: 10.1016/j.biomaterials.2011.11.007

Abstract A simple culture system to achieve the differentiation of embryonic stem (ES) cells toward hepatocytes with high efficiency is crucial in providing a cell source for the medical application. In this study, we report the effect of a matrix-dependent enrichment of ES cell-derived hepatocytes using immobilized poly( N-p -vinylbenzyl-4- O -β- d -galactopyranosyl- d -gluconamide) (PVLA) with E-cadherin-IgG Fc (E-cad-Fc) as a galactose-carrying substratum. PVLA and E-cad-Fc were confirmed to be stably co-adsorbed onto polystyrene surface by quartz crystal microbalance (QCM). We showed that the E-cad-Fc/PVLA hybrid substratum was efficient in culturing primary hepatocytes and maintaining liver functions, on which the undifferentiated ES cells also maintained high proliferative capability. Furthermore, ES cell-derived hepatocytes on this hybrid matrix expressed elevated level of liver specific genes and functions together with early expression of definitive hepatocyte marker, asialoglycoprotein receptor (ASGPR). Finally, we isolated a high percentage of cells (about 60%) with ASGPR expression after re-seeding onto PVLA-coated surface, and observed the elimination of the poorly differentiated cells (Gata6+ and Sox17+ ) and the ones toward another cell lineage (brachyury+ and Pdx1+ ). The system uses a glycopolymer as an extracellular substratum for isolation and enrichment of ES cell-derived hepatocytes with adequate homogeneity and functionality.