Optimal concentration of brain-derived neurotrophic factors for rat hippocampus neural stem cell differentiation into neurons in the medium containing epidermal growth factors
- VernacularTitle:表皮生长因子培养条件下脑源性神经生长因子诱导大鼠海马神经干细胞向神经元分化的最佳浓度
- Author:
Zhenyu WANG
;
Lei TONG
;
Lili JI
;
Jiuhong ZHAO
;
Yuanyuan TANG
- Publication Type:Journal Article
- From:
Chinese Journal of Tissue Engineering Research
2008;12(16):3171-3174
- CountryChina
- Language:Chinese
-
Abstract:
BACKGROUND:The differentiation of neural stem cells (NSCs) is a hot research. Whether brain-derived neurotrophic factors (BDNF) can induce the differentiation of NSCs into neurons has not been detailed studied. OBJECTIVE:To investigate the effect of different concentrations of BDNF on the differentiation of NSCs from adult rat hippocampus into neurons in the medium with epidermal growth factor. DESIGN:A controlled study. SETTING:Department of Human Anatomy, China Medical University. MATERIALS:The experiment was performed at the Neurotomia Laboratory of China Medical University in August 2007. NSCs isolated from hippocampus of adult SD rats were inoculated in a serum-free medium. Three clean adult SD rats (200-250 g each) were provided by Laboratory Animal Department of China Medical University. Dispositions to the rats were consistent with ethical standards of animals. Epidermal growth factor (EGF) and BDNF were bought from R&D Company. METHODS:①NSCs obtained from rat hippocampus were cultured in the serum-free medium containing basic fibroblast growth factor (bFGF), EGF and B27. ②Single cell clone was obtained via limited dilution for the fourth passage of cells. Identification of NSCs for the passage of cells from monoclonal spheres was performed by Nestin immunocytochemistry. Neurons and astroglial cells were identified by immunocytochemistry for neuron specific enolase (NSE) and glial fibrillary acidic protein (GFAP) a week after differentiation. ③Differentiation for the monoclonal spheres was done to appraisal the proportion of neurons. NSCs were divided for several groups containing 0 μg/L, 50 μg/L, 100 μg/L, 150 μg/L, and 200 μg/L groups according to the doses of BDNF. EGF was added in the media of each group. Immunocytochemistry for NSE was done a week later to count the positive cells. MAIN OUTCOME MEASURES:Proportion of neurons differentiated from NSCs.RESULTS:①NSCs immunocytochemical staining showed that Nestin was positive in monoclonal spheres, and NSE and GFAP were positive in differentiated cells. ②The proportion of differentiation from NSCs into neurons was significantly higher in groups treated with 50 μg/L and 100 μg/L BDNF compared to other groups（t=2.502-5.025, P < 0.05）, but there was no significant difference between 50 μg/L and 100 μg/L BDNF groups（P > 0.05）. Moreover, no significant difference was detected among 0 μg/L, 150 μg/L and 200 μg/L BDNF groups (P > 0.05). CONCLUSION:The optimal concentration of BDNF is 50 μg/L for differentiation from NSCs into neurons while the concentration of EGF is 20 μg/L.
