SRC-1基因在小鼠神经干细胞分化过程中的变化

目的观察类固醇受体辅助活化因子-1(SRC-1)基因在小鼠神经干细胞(NSCs)体外培养分化过程中表达的变化。方法用机械分离和酶消化法从1~3 d小鼠大脑皮质获取NSCs原代细胞,体外培养获神经球,传代并消化,经血清诱导神经球细胞分化。形态学、细胞免疫荧光实验观察神经球形态、Nestin表达鉴定NSCs;抗体分别标记神经元、星形胶质细胞,检测细胞分化d 3、9的细胞类型。RT-qPCR和免疫印迹法检测细胞分化d 0、3、9 SRC-1基因mRNA和蛋白的表达。结果新生小鼠大脑皮质获取的细胞,体外培养能扩增形成神经球,Nestin阳性表达。NSCs分化d 3主要为神经元,d 9主要为星形胶质细胞。与NSCs分化d 0相比,SRC-1表达在分化d 3明显升高,在分化d 9明显降低。 结论 成功分离并获取新生小鼠皮质NSCs。在NSCs分化中,SRC-1表达量有明显变化,分布依次为神经元>NSCs>星形胶质细胞。

Changes in Expression of SRC-1 Gene in the Differentiation of Mouse Neural Stem Cells in Vitro

Aim To investigate changes in expression of steroid receptor coactivator-1(SRC-1)gene during the differentiation of mouse Neural Stem Cells(NSCs)in vitro.Methods Primary NSCs were obtained from cerebral cortex of neonatal mouse aged 1~3 days mechanically and enzymatically.The neurospheres were cultured in vitro,The neurosphere cells were induced to differentiate into neurons and astrocytes by serum after primary and passage culture.The cultured NSCs were identified by neurospheric morphology and Nestin immunofluorescence assay;Neurons and astrocytes were recognized respectively by specific antibodies to detect the cell types on days 3 and 9 of differentiation;The expression of SRC-1 gene mRNA and protein were detected by RT-qPCR and Western blotting on days 0,3 and 9 of differentiation.Results Nestin-positive NSCs were observed in cultured neurospheres derived from cerebral cortex of neonatal mouse in vitro.During the process of differentiation,neurons were the main cells on day 3 and astrocytes were the main ones on day 9.Compared with day 0,SRC-1 expression increased on day 3 and decreased on day 9 significantly.Conclusion NSCs could be isolated from cerebral cortex of neonatal mouse and cultured well in vitro.During the differentiation of NSCs,there were significant changes in SRC-1 expression,which was the highest in neurons,followed by NSCs and astrocytes.