The Experimental Study Of Inducing Neural Stem Cells Cultured In Vitro To Differentiate Into Dopaminergic Neurons

Objective: To observe the ability of NSCs to differentiate into dopaminergic neurons under different induction protocols, compare and optimize differentiation-induction conditions in order to establish technical basis for using NSCs in the cell transplantation therapy of Parkinson's disease.Methods: Cortex-originated cells from brain of embryonic rats of 13. 5-14. 5 days were obtained, then cultured in serum-free medium with EGF/ bFGF and B27 to isolated NSCs. NSCs of the third passage were induced to differentiate into neural cells by varying protocols: (1) The recombinant plasmids carrying Nurrl gene were introduced into NSCs by method of electroporation, then the positive cell clones were selected by adding hygromycin into the culture medium and the expression of Nurr 1 was identified by immunochemistry.The Nurrl gene modified NSCs were allowed to differentiate. (2)All trans-retinoic acid (ATRA) and extracts of Ginkgo Biloba (Egb) were added into the culture medium to induce NSCs to differentiate. (3) Marrow stromal cells were prepared as the feeder cells and co-cultured with NSCs, then the differentiation-inducing potentiality of the feeder cells were observed. (4) NSCs were induced to differentiate by adding fetal bovine serum into the medium. The transmitter phenotype of the differentiated cells were detected by the method of immunocytochemistry, the TH postive cells in the differentiated neural cells were counted and analyzed to determine the effects of different induction protocols on promoting NSCs to differentiate into dopaminergic...