Jiao Shujie, Liu Yaling, Yao Yaobing, Teng Junfang
Department of Neurology, the First Affiliated Hospital of Zhengzhou University, 1st of Jianshe East Road, Zhengzhou, 450000 China.
Cell Biosci. 2017 Dec 11;7:68. doi: 10.1186/s13578-017-0194-y. eCollection 2017.
Neural stem cells (NSCs) are able to differentiate into neurons and astroglia. miRNAs have been demonstrated to be involved in NSC self-renewal, proliferation and differentiation. However, the exact role of miR-124 in the development of NSCs and its underlying mechanism remain to be explored.
Primary NSCs were isolated from embryos of Wistar rats. Immunocytochemistry was used to stain purified NSCs. miR-124, Delta-like 4 (DLL4), ki-67, Nestin, β-tubulin III, glial fibrillary acidic protein (GFAP), HES1, HEY2, and cyclin D1 (CCND1) expressions were detected by qRT-PCR and western blot. The interaction between miR-124 and DLL4 was confirmed by luciferase reporter assay. Cell proliferation was assessed by MTT assay.
NSCs could self-proliferate and differentiate into neurons and astrocyte. miR-124 was up-regulated and DLL4 was down-regulated during NSC differentiation. DLL4 was identified as a target of miR-124 in NSCs. Ectopic expression of miR-124 or knockdown of DLL4 promoted the proliferation and the formation of NSCs to neurospheres. Moreover, miR-124 overexpression or DLL4 down-regulation improved β-tubulin III expression but decreased GFAP expression in NSCs. Furthermore, enforced expression of DLL4 partially reversed the effects of miR-124 on NSCs proliferation and differentiation. Elevated expression of miR-124 suppressed the expressions of HES1, HEY2, and CCND1 in NSCs, while these effects were attenuated following the enhancement of DLL4 expression.
miR-124 promoted proliferation and differentiation of NSCs through inactivating Notch pathway.
神经干细胞（NSCs）能够分化为神经元和星形胶质细胞。微小RNA（miRNAs）已被证明参与神经干细胞的自我更新、增殖和分化。然而，miR-124在神经干细胞发育中的具体作用及其潜在机制仍有待探索。
从Wistar大鼠胚胎中分离原代神经干细胞。免疫细胞化学用于对纯化的神经干细胞进行染色。通过qRT-PCR和蛋白质免疫印迹法检测miR-124、Delta样蛋白4（DLL4）、ki-67、巢蛋白、β-微管蛋白III、胶质纤维酸性蛋白（GFAP）、HES1、HEY2和细胞周期蛋白D1（CCND1）的表达。通过荧光素酶报告基因检测法确认miR-124与DLL4之间的相互作用。通过MTT法评估细胞增殖。
神经干细胞能够自我增殖并分化为神经元和星形胶质细胞。在神经干细胞分化过程中，miR-124上调而DLL4下调。DLL4被确定为神经干细胞中miR-124的靶标。miR-124的异位表达或DLL4的敲低促进了神经干细胞的增殖以及向神经球的形成。此外，miR-124过表达或DLL4下调可提高神经干细胞中β-微管蛋白III的表达，但降低GFAP的表达。此外，DLL4的强制表达部分逆转了miR-124对神经干细胞增殖和分化的影响。miR-124表达升高抑制了神经干细胞中HES1、HEY2和CCND1的表达，而在DLL4表达增强后这些作用减弱。
miR-124通过使Notch信号通路失活促进神经干细胞的增殖和分化。
