Development Of Serum-free Culture System For Embryonic Stem Cells

Embryonic stem cells(ESCs)have potential uses in cell therapy and regenerative medicine because of their unlimited self-renewal capabilities and pluripotency.Nonetheless,successful applications of ESCs will require large numbers of cells with good quality.To address this,the objective of this study was to develop a robust,controllable cuture system for the expansion of ESCs.In this study,serum was replaced with serum replacement and gelatin was applied to replace feeder cells for mouse embryonic stem cells(mESCs).Under these conditions,the fold expasion of mESCs reached 2.95±0.5 and the percentage of SSEA-1+ cells was more than 80%.Based on this,inhibitors of GSK3,MEK and FGFR were selected to investigate whether mESCs could be better maintained in an undifferentiated state in serum-free medium.The reasults showed that the expansion of mESCs was enhaced by BIO,SB-216763,PD184352 and SU5402.A combination of SB-216763,PD0325901 and SU5402 resulted in the expansion fold of mESCs of 318.78±47.95 and the percentage of SSEA-1+ cells remained above 80%,which were both significantly higher than those in the control culture.In addition,the growth characteristics of mESCs on Cytodex 3 in serum-free medium was examined.It was found that the number of mESCs increased by 31,68±7.74 folds within 10 days,which was significantly higher than that in static culture(17.92 ± 9.80 folds)(p<0.05).Moreover,mESCs cultured in the dynamic condition was maintained in an undifferentiated state.In conclusion,this study lays solid foundation for the development of scalable culture systems for ESCs.