ABSTRACT
Objective To establish a satisfactory method of isolating,culturing and determining gut neural crest stem cells,which may provide theoretical basis for clinical application.Methods The guts of embryonic mice removed and dissociated were plated into serum-free DMEM/F12 medium.The mitogen-free DMEM/F12 medium supplementd with 10% fetal bovine serum was used to induce differentiation of GNCSCs.Neurospheres and their derivations were determined with immunocytochemical and immunofluorescent staning.Results Neurospheres were generated in the simplified serum-free medium.The staining results showed that enteric neurospheres were GNCSCs and could differentiate into neurons,glial cells and smooth muscle cells by serum-induction.Conclusion GNCSCs have the capacity of self-renewal and proliferation,and give rise to neurons,glial cells and myofibroblasts.
ABSTRACT
Objective To explore the differences of cultivation and differentiation between cortical neural stem cells(C-NSCs) and mesencephalic neural stem cells(M-NSCs). Methods We dissociated embryonic cortex and ventral mesencephalon respectively,and single cell suspensions were achieved by trypsin digestion and mechanical dissociation in sterile condition.These cells were planted into DMEM/F12 culture medium with B27,bFCF,and then were passaged by mechanical methods and induced to differentiation in DMEM/F12 medium with 10%FBS(without growth factors).For examining the features of C-NSCs and M-NSCs,reverse phasecontrast microscopy,immunocytochemical techniques were applied to detect nestin,NSE,GFAP antigens and the specific antigen(TH) of neurons differentiated from the two kinds of NSCs.We observed and compared the difference of the two kinds of NSCs. Results The NSCs existed both in cortex and ventral mesencephalon of embryonic mice,which expressed nestin and possessed the ability of self-renewing and multipotent, but the density of NSCs in cortex was higher than that in ventral mesencephalon.There were some TH positive cells differentiated from M-NSCs but none came from C-NSCs.Conclusion The proliferative activity of C-NSCs is higher than that of M-NSCs,but more number M-NSCs differentiated into dopaminergic neurons in serum medium.