小胶质细胞特有质子通道Hv1调控“NOX2/ROS-小胶质细胞炎性激活-CD4+T细胞异常活化”级联反应在EAE中的作用及其机制

Reactive oxygen species (ROS) act as “signaling molecules” in the inflammatory activation of microglial activation，which triggers abnormal CD4+ T-cell activation and finally initiates the pathogenesis of experimental autoimmune encephalomyelitis (EAE) / multiple sclerosis (MS). Therefore, blocking the ROS-mediated pathway above is expected to be an effective strategy for the treatment of EAE/MS. The voltage-gated proton channel Hv1 is a recently cloned ion channel selectively expressed in microglia in the central nervous system and plays an essential role in maintaining the activity of NOX2 and NOX2-dependent ROS production. To date, the role and mechanism of Hv1 in EAE/MS remains unclear. Our latest research results showed that silencing of Hv1 not only significantly reduced the incidence of EAE and the score of clinical neurological function, but also resulted in reduced production of ROS and amelioration of microglial inflammatory activation. Therefore, the aim of this subject is to further explore the role and mechanism of Hv1 in the pathogenesis of EAE. Our findings may supply experimental evidence on the role of Hv1 as a regulator for the "NOX2/ROS-inflammatory activation of microglia-abnormal CD4+ T-cell activation" signaling cascade and utilize Hv1 as a target for the development of therapeutic therapies for EAE/MS.

氧自由基（ROS）可作为“信号分子”不断驱动小胶质细胞炎性激活并进而促进CD4+T细胞的异常活化，而CD4+T细胞所介导的自身免疫反应在EAE/MS病理进程中扮演重要角色，因此，从源头上阻断ROS介导的上述通路有望成为EAE/MS治疗的有效策略。Hv1作为中枢神经系统中特异性表达于小胶质细胞上的一种离子通道，其对于维持NADPH氧化酶2（NOX2）的活性以及NOX2依赖的ROS生成必不可少。目前，Hv1在EAE/MS中的作用及其机制尚不明确。申请者前期预实验结果显示：Hv1沉默后不仅可以明显降低EAE小鼠的发病率并改善其临床症状，还可显著抑制EAE小鼠ROS水平的急剧升高以及小胶质细胞的炎性激活，因此，本项目旨在进一步探索Hv1在EAE中的作用，并试图以“NOX2/ROS-小胶质细胞炎性激活-CD4+T细胞异常活化”级联反应为切入点阐明机制，为MS的防治提供新的治疗靶点和实验依据。

多发性硬化（multiple sclerosis，MS）是一种发生于中枢神经系统的自身免疫性疾病，至今仍无法治愈。活性氧（reactive oxygen species，ROS）和过度激活的小胶质细胞可通过多种途径参与调控CD4+ T细胞的异常活化并进而加剧MS的发生及发展，因此，减缓氧化应激、抑制小胶质细胞炎性活化已成为新型MS治疗药物开发的重要研究方向。电压门控质子通道Hv1对于维持NOX2活性以及NOX2所介导的ROS生成必不可少。在本研究中，我们利用实验性自身免疫性脑脊髓炎（experimental autoimmune encephalomyelitis，EAE）动物模型对Hv1在MS/EAE中的作用及其机制进行系统研究。我们发现：通过Hv1腺相关病毒外源性下调Hv1的表达水平可显著改善EAE小鼠的临床神经功能表现；此外，Hv1下调还可明显缓解EAE小鼠髓鞘脱失、炎性细胞浸润、轴索损伤等典型组织病理学改变；进一步的机制研究表明Hv1下调的保护作用主要是通过抑制EAE小鼠体内NOX2介导的过度积累的ROS以及炎性激活的小胶质细胞，从而减缓CD4+ T细胞的Th1/Th17促炎性分化并促进其Th2抗炎性分化实现的。上述研究结果为MS的防治提供了新的治疗靶点和实验依据。