Klf6在少突胶质细胞发育以及髓鞘损伤后修复过程中的作用研究

Myelin sheath formed by mature oligodendrocytes is an essential structure to maintain the proper physiological function of nervous system. Injury or abnormal oligodendrocytes development usually leads to demyelination disease. Until now, the molecular basis of demyelination disease remains unclear. In our preliminary study, we firstly performed genome-wide H3K27Ac ChIP-Seq analysis on oligodendrocytes treated with a demyelinating compound LPC, to identify the genes whose expression were mostly changed after oligodendrocytes injury. Among the identified genes, a transcription factor Klf6, whose biological function in oligodendrocytes has not been studied was chosen for further study in our project. According to our obtained data, Klf6 expressed in oligodendrocytes, and its expression level decreased during the oligodendrocytes maturation process. Similarly, expression of Klf6 decreased in mice at adulthood. In cultured primary oligodendrocytes, although overexpression of Klf6 had no effect on cell proliferation, it inhibited the differentiation and maturation of oligodendrocytes. The inhibitory effect of Klf6 on oligodendrocytes maturation resulted from its inductive effect on the expression of Hes5, a key downstream factor of Notch signaling pathway. Meanwhile, overexpression of Klf6 reduced the expression level of cell apoptotic genes in oligodendrocytes, suggesting that Klf6 might protect mature oligodendrocytes from apoptosis. Basing on our findings, further experiments will be performed in mice using retro-virus system to overexpress or knock-down endogenous Klf6 level to elucidate the effects of Klf6 in oligodendrocytes and myelin development, as well as the underpinning molecular basis, aiming to provide new therapeutic targets for demyelinating disease in clinic.

髓鞘是维持神经系统正常生理功能的重要组成部分。少突胶质细胞损伤及发育异常可导致脱髓鞘疾病。目前，脱髓鞘疾病形成的机理尚未明确。我们首次利用全基因组H3K27Ac ChIP-Seq对溶血磷脂酰胆碱诱导少突胶质细胞损伤相关基因进行了分析筛选，并选取了转录因子Klf6进行研究。我们发现Klf6在少突胶质细胞中表达水平随着细胞分化而减少，而在小鼠体内则随着年龄增长而降低。过表达Klf6对少突胶质细胞增殖无影响，但能够抑制其分化成熟。此外，Klf6还可以上调少突胶质细胞中Notch信号通路下游分子Hes5的表达，并且可能通过下调凋亡相关基因的表达而抑制其凋亡。在此基础上，我们拟通过慢病毒系统在小鼠模型中分别过表达及抑制表达Klf6，深入研究Klf6在少突胶质细胞发育、髓鞘发育以及髓鞘损伤后修复等过程中的作用，阐明其作用机制，也为后续临床防治脱髓鞘疾病提供新的靶点。

髓鞘是维持神经系统正常功能的重要组成部分。少突胶质细胞是髓鞘的主要成分，其发育成熟受到多种信号通路及转录因子的综合调控。在本课题中，我们发现并证明了KLF6是少突胶质细胞发育过程中的重要调控因子之一。KLF6在少突胶质细胞中表达，在细胞分化前期表达水平较高。在少突胶质细胞中过表达KLF6可诱导细胞分化成熟并抑制细胞凋亡，但对细胞增殖水平无显著影响。KLF6对少突胶质细胞凋亡的影响可能通过其对细胞内氧化应激反应的调节而实现。过表达KLF6抑制细胞内ROS浓度，而使用ROS清除剂NAC处理可逆转敲低KLF6所引起的细胞凋亡。为了阐明KLF6调节少突胶质细胞分化的机理，我们进行了RNA-Seq和ChIP-Seq分析。结果显示，Gpr37是KLF6的下游基因，其表达水平受KLF6所调控。过表达KLF6诱导Gpr37表达，而敲低KLF6则抑制其表达水平。在少突胶质细胞中过表达Gpr37能够逆转敲低KLF6所引起的细胞分化水平下降，证实Gpr37介导了KLF6对少突胶质细胞分化的诱导作用。本课题初步阐明了KLF6调控少突胶质细胞发育的功能和机理，为探讨脱髓鞘疾病的治疗靶点提供了新的参考。