Abstract: Objective To establish an efficient way to induce the mouse embryonic stem cells into homogeneous neural stem cells and to develop culture conditions that support the viability and propagation of stable NSCs in vitro. Methods In the serum-free medium, firstly, we induced ESCs to neuroepithelial progenitor cells (NPCs). Then, NPCs were cultured in a suspension medium supplemented with EGF and FGF2 and aggregates were collected and replate to differentiate to neural stem cells (NSCs) in monolayer. We used 46C for monitoring and quantitating the transition from ESCs to NPCs. Expression of NSCs marker was determined by quantitative PCR and immunoflurescence. Results After 5 days of induction, mESCs differentiated into NPCs with high expression of Sox1+. NPCs can further differentiated in to NSCs after culture in suspension. As determined by real-time quantitative PCR (qPCR), Pax6、Nestin、Mash1、BLBP(FabP7) genes expressed highly in NSCs. The immunoflurescence demonstrated that 90% of NSCs express Nestin, RC2 and Pax6. Conclusion ESCs can successfully be induced to homogeneous NSCs, and NSCs can maintain self-renewal status and propagate in vitro after several passage.

Key words: Embryonic stem cell, Neural differentiation, Neuroepithelial progenitor cells, Neural stem cells