Zhang Zhen, Gao Feng, Kang Xiaokui, Li Jia, Zhang Litong, Dong Wentao, Jin Zhangning, Li Fan, Gao Nannan, Cai Xinwang, Yang Shuyuan, Zhang Jianning, Ren Xinliang, Yang Xinyu
Department of Neurosurgery, Tianjin Medical University General Hospital, Tianjin 300052, PR China; Tianjin Neurological Institute, Tianjin Key Laboratory of Injuries, Variations and Regeneration of Nervous System, 154 Anshan Road, Heping District, Tianjin 300052, PR China.
Department of Neurosurgery, Liaocheng People's Hospital, 67 Dongchang West Road, Liaocheng 252000, PR China.
Brain Res Bull. 2015 Apr;113:8-16. doi: 10.1016/j.brainresbull.2015.02.003. Epub 2015 Feb 17.
The Notch pathway is a highly conserved pathway that regulates hippocampal neurogenesis during embryonic development and adulthood. It has become apparent that intracellular epigenetic modification including DNA methylation is deeply involved in fate specification of neural stem cells (NSCs). However, it is still unclear whether the Notch pathway regulates hippocampal neurogenesis by changing the Notch genes' DNA methylation status. Here, we present the evidence from DNA methylation profiling of Notch1, Hes1 and Ngn2 promoters during neurogenesis in the dentate gyrus (DG) of postnatal, adult and traumatic brains. We observed the expression of Notch1, Hes1 and Ngn2 in hippocampal DG with qPCR, Western blot and immunofluorescence staining. In addition, we investigated the methylation status of Notch pathway genes using the bisulfite sequencing PCR (BSP) method. The number of Notch1 or Hes1 (+) and BrdU (+) cells decreased in the subgranular zone (SGZ) of the DG in the hippocampus following TBI. Nevertheless, the number of Ngn2-positive cells in the DG of injured mice was markedly higher than in the DG of non-TBI mice. Accordingly, the DNA methylation level of the three gene promoters changed with their expression in the DG. These findings suggest that the strict spatio-temporal expression of Notch effector genes plays an important role during hippocampal neurogenesis and suggests the possibility that Notch1, Hes1 and Ngn2 were regulated by changing some specific CpG sites of their promoters to further orchestrate neurogenesis in vivo.
Notch信号通路是一条高度保守的信号通路，在胚胎发育和成年期均参与调节海马神经发生。目前已经明确，包括DNA甲基化在内的细胞内表观遗传修饰深度参与神经干细胞（NSCs）的命运决定。然而，Notch信号通路是否通过改变Notch基因的DNA甲基化状态来调节海马神经发生仍不清楚。在此，我们展示了来自出生后、成年和创伤性脑的齿状回（DG）神经发生过程中Notch1、Hes1和Ngn2启动子DNA甲基化谱分析的证据。我们通过qPCR、蛋白质免疫印迹和免疫荧光染色观察了Notch1、Hes1和Ngn2在海马DG中的表达。此外，我们使用亚硫酸氢盐测序PCR（BSP）方法研究了Notch信号通路基因的甲基化状态。创伤性脑损伤（TBI）后，海马DG颗粒下区（SGZ）中Notch1或Hes1（+）和BrdU（+）细胞的数量减少。然而，受伤小鼠DG中Ngn2阳性细胞的数量明显高于非TBI小鼠的DG。相应地，三个基因启动子的DNA甲基化水平随其在DG中的表达而变化。这些发现表明，Notch效应基因严格的时空表达在海马神经发生过程中起重要作用，并提示Notch1、Hes1和Ngn2可能通过改变其启动子的某些特定CpG位点来进行调控，从而在体内进一步协调神经发生。
