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目的探讨室管膜前下区(anteriorsubventricularzone,SVZa)神经干细胞体外分化过程中,Nestin和Wnt-1表达的变化,以及Wnt-1对SVZa神经干细胞体外分化的影响。方法以免疫荧光染色方法观察在SVZa神经干细胞体外分化过程中,Nestin和Wnt-1在不同时间点的表达水平,同时以基因转染方法增强SVZa神经干细胞Wnt-1表达,观察Nestin、Wnt-1和Mash-1的表达以及神经元分化比例的改变。结果⑴Nestin主要表达于SVZa神经干细胞分化的早期阶段,随着细胞的分化和成熟,Nestin的表达迅速减弱。⑵SVZa神经干细胞贴壁分化2h即有Wnt-1表达,其表达高峰在细胞分化开始后12h,以后随着细胞分化的进行而迅速降低。⑶SVZa神经干细胞转染Wnt-1基因后,Nestin的表达减弱,Mash-1的表达增强,神经元的分化比例增高,Mash-1的增强与神经元分化之间有很高的一致性。结论⑴Wnt-1表达于SVZa神经干细胞分化过程之中,在细胞分化成熟后则不表达,Wnt-1可能参与调节SVZa神经干细胞的体外分化,并可作为神经干细胞处于分化状态的标志。⑵SVZa神经干细胞转染Wnt-1后表达Mash-1增强,并更多分化为神经元,Wnt-1可能通过Mash-1途径促使SVZa神经干细胞向神经元方向分化。
Objective To investigate the changes of Nestin and Wnt-1 in the differentiation of neural stem cells of anterior subventricular zone (SVZa) in vitro and the effect of Wnt-1 on the differentiation of SVZa neural stem cells in vitro. Methods Immunofluorescence staining was used to observe the expression of Nestin and Wnt-1 at different time points during the differentiation of SVZa neural stem cells in vitro. At the same time, Wnt-1 expression of SVZa neural stem cells was enhanced by gene transfection. The expressions of Nestin, Wnt-1 And Mash-1 expression and neuronal differentiation ratio changes. Results (1) Nestin was mainly expressed in the early stage of SVZa neural stem cell differentiation. With the differentiation and maturation of cells, the expression of Nestin rapidly decreased. ⑵SVZa adherent neural stem cell adherent differentiation that Wnt-1 expression, the peak of expression in the 12h after the start of cell differentiation, and later with the rapid differentiation of cells. (3) After transfection of Wnt-1 gene with SVZa neural stem cells, the expression of Nestin was weakened, the expression of Mash-1 was enhanced, the proportion of neuronal differentiation was increased, and there was a high consistency between Mash-1 enhancement and neuronal differentiation. Conclusion (1) Wnt-1 expression in SVZa neural stem cells in the process of differentiation, after the differentiation does not mature cells, Wnt-1 may be involved in regulating SVZa neural stem cells in vitro differentiation, and can be used as a marker of neural stem cells in a state of differentiation. (2) The expression of Mash-1 in SVZa neural stem cells after Wnt-1 transfection was enhanced and more differentiated into neurons. Wnt-1 might induce SVZa neural stem cells to differentiate into neurons through Mash-1 pathway.