Self-renewal and Proliferation of Murine Embryonic Stem Cells: A Study of Glycosaminoglycans Effect on Feeder-Free Cultures

The self-renewal and proliferation of murine embryonic stem (ES) cells can be preserved indefinitely in the presence of the mouse embryonic fibroblast (MEF) feeder layer. Since the feeder layer has several drawbacks, including viral contamination and large-scale production, gelatin solutions are the most prominent alternative to replacing it. In this investigation, self-renewal and proliferation of ES cells was carried out by supplementing the gelatin solution with glycosaminoglycan components, such as chondroitin sulfate and heparin. The methylcellulose-based embryoid body (EB) assay was used to evaluate the self-renewal ability of the ES cells. Chondroitin sulfate and heparin were mixed with 0.1% gelatin solution in 0:100, 50:50 and 100:0 (heparin/chondroitin sulfate) ratios and compared with 0.1% gelatin (positive control) and MEF (negative control) in three independent parallel experiments. Chondroitin sulfate addition to 0.1% gelatin enhanced the cell population and the number of EBs by 57 and 32%, respectively.