背景:阻塞性睡眠呼吸暂停(OSA)是一种睡眠呼吸紊乱,其特征是间歇性缺氧(IH),可能导致认知功能障碍。然而,IH对认知功能相关分子过程的影响尚不清楚.
方法:将C57BL/6J小鼠暴露于常氧(对照)或IH中6周。通过羟甲基化DNA免疫沉淀(hMeDIP)测序定量DNA羟甲基化。10-11易位1(Tet1)被慢病毒敲除。具体来说,认知功能通过行为实验进行评估,通过HE染色评估病理特征,通过DNA斑点印迹和免疫组织化学染色检查海马DNA羟甲基化,同时使用qRT-PCR研究了Wnt信号通路及其下游效应,免疫荧光染色,和Luminex液体悬浮芯片分析。
结果:IH小鼠海马出现病理变化和认知功能障碍。与对照组相比,IH小鼠在海马中表现出整体DNA羟基甲基化,3种羟化酶的表达显著增加。Wnt信号通路被激活,以及Wnt3a的mRNA和5hmC水平,Ccnd2和Prickle2显著上调。进一步引起下游神经发生异常和神经炎性激活,表现为IBA1(小胶质细胞的标志物)的表达增加,GFAP(星形胶质细胞的标记),和DCX(未成熟神经元的标记),以及一系列炎症细胞因子(例如TNFa,IL3、IL9和IL17A)。Tet1击倒后,上述指标恢复正常。
结论:海马Tet1激活Wnt信号通路与IH诱导的认知功能障碍有关。
BACKGROUND: Obstructive sleep apnoea (OSA) is a sleep-disordered breathing characterized by intermittent hypoxia (IH) that may cause cognitive dysfunction. However, the impact of IH on molecular processes involved in cognitive function remains unclear.
METHODS: C57BL / 6 J mice were exposed to either normoxia (control) or IH for 6 weeks. DNA hydroxymethylation was quantified by hydroxymethylated DNA immunoprecipitation (hMeDIP) sequencing. ten-eleven translocation 1 (Tet1) was knocked down by lentivirus. Specifically, cognitive function was assessed by behavioral experiments, pathological features were assessed by HE staining, the hippocampal DNA hydroxymethylation was examined by DNA dot blot and immunohistochemical staining, while the Wnt signaling pathway and its downstream effects were studied using qRT-PCR, immunofluorescence staining, and Luminex liquid suspension chip analysis.
RESULTS: IH mice showed pathological changes and cognitive dysfunction in the hippocampus. Compared with the control group, IH mice exhibited global DNA hydroxylmethylation in the hippocampus, and the expression of three hydroxylmethylases increased significantly. The Wnt signaling pathway was activated, and the mRNA and 5hmC levels of Wnt3a, Ccnd2, and Prickle2 were significantly up-regulated. Further caused downstream neurogenesis abnormalities and neuroinflammatory activation, manifested as increased expression of IBA1 (a marker of microglia), GFAP (a marker of astrocytes), and DCX (a marker of immature neurons), as well as a range of inflammatory cytokines (e.g. TNFa, IL3, IL9, and IL17A). After Tet1 knocked down, the above indicators return to normal.
CONCLUSIONS: Activation of Wnt signaling pathway by hippocampal Tet1 is associated with cognitive dysfunction induced by IH.