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目的观察碱性成纤维生长因子(bFGF)和表皮生长因子(EGF)对胚胎脊髓神经干细胞(NSC)增殖与分化的影响。方法从14 d胚胎大鼠的脊髓组织中分离培养脊髓NSC,并随机分为3组:EGF组、bFGF组和bFGF+EGF组。通过光镜观察不同时间点各组脊髓NSC克隆细胞团数量及直径大小,并采用免疫荧光染色检测各组脊髓NSC向神经元和星形胶质细胞分化的情况。结果①EGF组培养1、3、7 d后NSC克隆细胞团数量和直径均少于bFGF和bFGF+EGF组,差异有统计学意义(P<0.05)。而bFGF+EGF组仅在培养1 d时克隆细胞团数量多于bFGF组,在培养3、7 d时差异无统计学意义。②EGF组分化细胞中神经元比例显著少于bFGF和bFGF+EGF组,星形胶质细胞数量明显大于bFGF和bFGF+EGF组,差异有统计学意义(P<0.05)。而bFGF和bFGF+EGF组组间差异无统计学意义。结论 EGF对脊髓NSC克隆形成有一定作用,而bFGF能较好地促进克隆细胞团的形成及生长,两者联合应用在培养早期可显著促进克隆细胞团形成。EGF可诱导脊髓NSC更多分化为星形胶质细胞,而bFGF则可促进脊髓NSC向神经元分化。
Objective To observe the effect of basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) on the proliferation and differentiation of embryonic spinal cord neural stem cells (NSC). Methods Spinal cord NSCs were isolated and cultured from spinal cord tissue of 14-day-old embryonic rats and randomly divided into three groups: EGF group, bFGF group and bFGF + EGF group. The numbers and diameters of NSC clonal cell clusters in each group at different time points were observed with light microscope. The differentiation of NSCs into neurons and astrocytes in each group was detected by immunofluorescence staining. Results ① The numbers and diameters of NSC clone cells in the EGF group were less than those in the bFGF and bFGF + EGF groups after 1, 3 and 7 days of culture. The difference was statistically significant (P <0.05). However, in the bFGF + EGF group, the number of clonal cell clusters was more than that of the bFGF group only at 1 d of culture, and there was no significant difference at 3 and 7 d of culture. ② The percentage of neurons in EGF-positive cells was significantly lower than that in bFGF and bFGF + EGF groups. The number of astrocytes was significantly higher than that in bFGF and bFGF + EGF groups (P <0.05). There was no significant difference between bFGF and bFGF + EGF groups. Conclusion EGF can promote the formation of NSC clones in spinal cord, while bFGF can promote the formation and growth of clonal cell clusters. The combination of the two can promote the formation of clonal cell mass in the early stage of culture. EGF can induce more differentiation of spinal cord NSCs into astrocytes, while bFGF can promote the differentiation of spinal cord NSCs into neurons.