The Culturing,Identification Of Neural Stem Cells Of Embryonic Rat In Vitro And The Effect Of HGF On Differentiation Of Them

Objective: To establish the methods of culturing, identification of neural stem cells of embryonic rat in vitro and to explore the effect of HGF on the differentiation of them.Methods: Neocortex was dissected out from E14 embryonic rat and made into single cell suspension, which were cultured by the technique of serum free cultivation combined with growth factors (EGF and bFGF). The characteristics of NSCs were detected in different growing phases. NSCs were subcultured continuously. After 3 passages, adding variant concentrations of HGF to investigate the differentiation of NSCs. Immunofluorescence was performed to identify NSCs and the differentiation result of them . All the data were analyzed by statistics.Results: NSCs were isolated from the E14 embryonic rat's neocortex continuously proliferated and formed neurospheres. The results of immunofluorescence showed: both the primary and the passaged neurospheres were nestin positive. The rate of the nestin positive cells in the the primary and the passaged neurospheres were same (P>0.05). The differentiated cells were GFAP or NSE positive.Then we revealed EGF and bFGF ,when HGF was added into the NSCs with different concentration to induce them differentiation . Immunoflurescene was used to examine those differentiated cells. The NSE and GFAP positive rate of the cells were calculated. With the concentraton of HGF increased, the NSE positive rate was also increased.