联合应用EGF和NGF对成年大鼠海马神经干细胞分化的影响

Combination of Epidermal Growth Factor and Nerve Growth Factor on the Differentiation of Neural Stem Cells from Hippocampus of Adult Rats

目的探讨联合应用表皮生长因子(EGF)和神经生长因子(NGF)对成年大鼠海马神经干细胞(NSCs)分化的影响。方法用含碱性成纤维生长因子(bFGF)、表皮生长因子、B27的无血清细胞培养技术体外培养成年大鼠海马神经干细胞,单克隆培养细胞行Nestin免疫细胞化学染色,诱导分化1w后细胞行GFAP和NSE免疫细胞化学染色;根据培养基中所加营养因子的不同将第4代细胞分为4组培养:EGF组、EGF+NGF组、NGF组、对照组,此4组细胞培养1w后行NSE免疫细胞化学染色,计数阳性细胞比例后进行统计学分析。结果单克隆培养后克隆球表达Nestin,诱导分化1w后细胞表达NSE、GFAP。与空白对照组相比,EGF组、NGF组和EGF+NGF组细胞分化为神经元的比例较高(P<0.05),其中EGF+NGF组细胞的比例最高。结论单独或联合应用EGF、NGF可以促进成年大鼠海马神经干细胞向神经元分化。

Objective To study the effect of epidermal growth factor（EGF） and nerve growth factor（NGF） on differentiation of neural stem cells（NSCs） from hippocampus of adult rats. Methods The serum-free medium containing bFGF, EGF and B27 was used to culture NSCs isolated from hippocampus of rats. Immunocytochemistry for nestin was used to identify NSCs, for glial fibrillary acidic protein（GFAP） and neurone specific enolase（NSE） to identify astrocytes and neurons respectively one week after NSCs were differentiated. Four passaged NSCs were divided into four groups： EGF, NGF, EGF＋NGF and control groups. The percentage of the positive cells was count and analyzed in statistics. Results The cultured monoclones sphere expressed nestin, and expressed NSE and GFAP one week after induced differentiation. The percentages of NSE positive cells differentiated from NSCs in EGF, NGF and EGF＋NGF groups were much higher than that in normal control,with the highest in EGF＋NGF. Conclusion Alone or combination of EGF and NGF can promote the differentiation of NSCs into neurons in hippocampus of adult rat.