Differentiation of neural stem cells from the anterior subventricular zone into dopaminergic neurons induced by astrocyte-conditioned medium in vitro
- VernacularTitle:星形胶质细胞条件培养液体外诱导胎鼠室管膜前下区神经干细胞向多巴胺能神经元的分化
- Author:
Zhicheng XIN
;
Zheng ZHOU
;
Jiwen YAN
- Publication Type:Journal Article
- From:
Chinese Journal of Tissue Engineering Research
2007;0(06):-
- CountryChina
- Language:Chinese
-
Abstract:
BACKGROUND: Effective proliferation in vitro of neural stem cells and directional induction and differentiation of dopaminergic neurons play a key role in neural stem cell transplantation for treatment of degeneration of the nervous system and repair of injured nervous system. OBJECTIVE: To explore the differentiation of neural stem cells from anterior subventricular zone into dopaminergic neurons in astrocyte-conditioned medium. DESIGN, TIME AND SETTING: The in vitro controlled cytology experiment was performed at the Experimental Center of Xinqiao Hospital of Third Military Medical University of Chinese PLA from December 2006 to August 2007. MATERIALS: A total of 15 neonatal KM mice aged 2 days and 40 pregnant 16 days Wistar rats were provided by the Experiment Animal Center of Third Military Medical University of Chinese PLA. METHODS: Astrocytcs of KM mice were isolated and purified by a standard shaking method and differential adhesion. Astrocyte-conditioned medium was collected when astrocytcs at passage 3 were cultured for 5 days maintained at –20 ℃ for use. Neural stem cells from the anterior subventricular zone of Wistar rats were isolated and cultured in serum-free DMEM/F12, supplemented with B27 and basic fibroblast growth factor for primary culture. Neural stem cells at passage 3 were collected and incubated at 0.5?108/L. Cells in the control group were incubated in the DMEM/F12 medium containing fetal bovine serum of 0.1 volume fraction. Cells in the experimental group were incubated in the astrocyte-conditioned medium. MAIN OUTCOME MEASURES: Neural stem cells in the anterior subventricular zone were determined by the immunocytochemical technique. Positive rate of the neural stem cell differentiation into dopaminergic neurons in the anterior subventricular zone was detected by the flow cytometry. RESULTS: Cells in neurospheres expressed nestin and differentiated into neuro-specific enolase and glial fibrillary acidic protein. Tyroxine hydroxylase positive cells were found in the experimental group, showing round or ellipse. Tyroxine hydroxylase was seen in cytoplasm and cluster-shape processes. Tyroxine hydroxylase positive cells in control group were different from that in the experimental group in amount and appearance. Positive cell rate of tyroxine hydroxylase was significantly higher in the experimental group than in the control group (t =35.296, P
