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目的探讨Sox2在神经营养因子3(neurotrophin-3,NT-3)促进原代大鼠神经干细胞(neural stem cells,NSCs)分化中的作用及其机制。方法选取孕14~15 d SD胎鼠,培养原代脑源性NSCs。观察NT-3对NSCs中Sox2表达及NSCs分化的影响:采用不同浓度NT-3(25、50、100 ng/m L)刺激NSCs24 h后,检测Sox2、DCX、Olig2、GFAP蛋白的表达量。探索NT-3对Sox2产生影响的机制:使用PI3K/AKT通路阻断剂(10μmol/L LY294002)选择性阻断NT-3的作用后,检测AKT、p-AKT和Sox2在蛋白水平表达量的变化。结果随着NT-3浓度的升高,NSCs中Sox2蛋白水平的表达明显上调(P<0.05),各NT-3处理组的NSCs向神经元前体细胞分化的比例明显增加(P<0.05),100 ng/m L NT-3组效果最强,呈现出明显的剂量依赖性;与对照组相比,100 ng/m L NT-3组的p-AKT及Sox2蛋白水平显著上调(P<0.05),且LY294002可阻断此作用(P<0.05)。结论 NT-3能够上调NSCs中Sox2的表达,后者进而诱导NSCs向神经元方向分化,其可能是通过激活PI3K/AKT信号通路起作用。
Objective To investigate the role of Sox2 in the differentiation of primary neural stem cells (NSCs) by neurotrophin-3 (NT-3) and its mechanism. Methods Fetal embryos from 14 to 15 days pregnant were selected to culture primary brain derived NSCs. The effect of NT-3 on the expression of Sox2 and the differentiation of NSCs in NSCs: The expression of Sox2, DCX, Olig2 and GFAP protein was detected after 24 h stimulation of NSCs with different concentrations of NT-3 (25, 50 and 100 ng / mL) To investigate the mechanism of NT-3 effect on Sox2: The effect of NT-3 was selectively blocked by PI3K / AKT pathway inhibitor (10μmol / L LY294002) to detect the expression of AKT, p-AKT and Sox2 at protein level Variety. Results The expression of Sox2 protein in NSCs was up-regulated with the increase of NT-3 concentration (P <0.05). The proportion of NSCs differentiated into neuronal precursor cells in each NT-3 group was significantly increased (P <0.05) (P <0.05). Compared with control group, the levels of p-AKT and Sox2 protein in 100 ng / mL NT-3 group were significantly increased (P < 0.05), and LY294002 could block this effect (P <0.05). Conclusion NT-3 can up-regulate the expression of Sox2 in NSCs, which in turn leads to the differentiation of NSCs into neurons. It may be through the activation of PI3K / AKT signaling pathway.