Invitro differentiation of embryonic neural stem cells from rabbit retina and brain

Objective To isolate neural stem cells (NSCs) from rabbit retina and brain,and induce differentiation of those NSCs using different culture media.Methods Single-cell suspensions of retina and cerebral cortex were prepared from rabbit embryo,cultured in 5 types of different media to isolate the NSCs by continual passages.After 3 passages,NSCs were induced to differentiation in 2 types of different media for 8 to 10 days.NSCs and induced-retinal ceils were examined by immunofluorescence and flow cytometry for the expression pattern of some specific antigens.Results Immunofluorescence showed that NSCs from retina and brain,cultured in serum-free media,both expressed Nestin partially.Flow cytometry showed that Nestin positive cells were significantly decreased while the Rhodopsin and Thy1.1 positive cells were increased after induction.Compared with the combined induction of all-trans retinoid acid (ATRA) and serum,5 % FBS (fetal bovine serum) led to higher expression of Rhodopsin (P<0.01),but lower expression of Thy1.1 (P=0.01).Conclusion Serum-free media with N2,EGF,bFGF,LIF is the best for NSCs purification.Both induciton media can induce NSCs to differentiate.Retina NSCs have higher potentials to differentiate into retinal neuro-epithelial cells than brain NSCs. Key words: Stem cells/cytology; Cell culture techniques/methods; Cell differentiation/immunology; Neuroepithelial cells/cytology