MAC激活NLRP3炎症小体通路促进近视发生发展的分子机制

Myopia is a public health issue worldwide which can cause visual impairment and blindness, however, its pathogenic mechanism has not been fully clarified. Our previous studies and published literatures provided evidence that inflammation plays a crucial role in the development of myopia. As an important biological system, complement is responsible for the central point of inflammation through its activation. Based on our study investigating the pathogenic and regulatory mechanism of complement on pathological myopia, we currently point our research to the terminal product of complement activation—membrane attack complex (MAC) and NOD-like receptor 3(NLRP3) inflammasome pathway. Our working hypothesis is that, the environmental factor responsible for myopia activates local complement system, which leads to the over deposition of MAC, and consequently trigger NLRP3 inflammasome pathway, which in turn promote the development of myopia through regulating the signals involved in the progression of myopia. We propose that (1) to investigate the ordered molecular mechanism of MAC-NLRP3 inflammasome pathway in the myopic animal model and cultured cells by utilizing MAC or NLRP3 deficient mouse and siRNA targeting MAC or NLRP3; (2) to investigate whether the progression of myopia could be blocked by CD59, a MAC inhibitor through breaking MAC-NLRP3 inflammasome pathway. Our study aimed to provide new insight on the mechanism responsible for myopia and the potential therapeutic strategies of inhibitor targeting MAC on myopia progression.

近视是导致视力受损和盲的全球性健康问题，发生机制未明。课题组前期研究和相关报道显示炎症在近视发生中的重要作用。补体系统是机体重要的反应体系，其活化是炎症反应的中心环节。本课题在“补体对病理性近视的致病作用及其调控机制”基础上，锁定补体活化的终末产物—膜攻击复合物（MAC），选择NLRP3炎症小体通路，提出假设：机体在环境因素作用下，局部补体活化，MAC过度沉积，激活NLRP3炎症小体通路，通过调控“近视信使”促进近视。拟通过（1）利用MAC和NLRP3基因缺失小鼠以及靶向MAC和NLRP3的siRNA，从动物实验和细胞实验分割性探讨MAC-NLRP3炎症小体通路促进小鼠形觉剥夺性近视形成的有序性分子机制；（2）借助靶向MAC的抑制剂CD59，探讨抑制MAC形成，阻断MAC-NLRP3炎症小体通路是否可以阻止近视进展。本课题旨在研究近视发病机制的新切入点和靶向MAC抑制剂的潜在近视防控策略。

近视是导致视功能损伤的全球性健康问题，发生机制未明。本课题在“补体对病理性近视的致病作用及其调控机制”研究基础上，锁定补体活化的终末产物—膜攻击复合物（MAC），选择NLRP3炎症小体通路，立项研究：MAC激活NLRP3炎症小体通路促进近视进展的分子机制。采用MAC和NLRP3基因缺失小鼠以及靶向MAC和NLRP3的siRNA，从动物水平和细胞水平分割性探讨MAC-NLRP3炎症小体通路促进小鼠形觉剥夺性近视形成的有序性分子机制。动物研究显示：野生型小鼠形觉剥夺眼后巩膜NLRP3炎症小体通路及其下游促炎因子白细胞介素-1β（IL-1β）表达水平显著高于对照眼，伴随基质金属蛋白酶-2（MMP-2）表达上调，并与近视度增加和眼轴延长存在显著正相关性，去除近视诱导因素后，上述变化可有所逆转。NLRP3-/-小鼠形觉剥夺眼后巩膜NLRP3通路表达被抑制，MMP-2表达降低，近视漂移和眼轴延长程度减少。NLRP3的特异性抑制剂MCC950可有效抑制野生型小鼠形觉剥夺性近视的进展。野生型小鼠后巩膜MAC表达与近视偏移和眼轴延长呈正相关。抑制补体6（C6，MAC形成的必要组成部分）可有效减少MAC形成，C6-/-小鼠剥夺眼的近视偏移和眼轴延长程度均低于野生型，且MMP-2升高程度和胶原蛋白-1（Collagen-1）的下降趋势有所减弱，伴随NLRP3炎症小体通路表达下降。细胞研究显示：补体3a（C3a，强效补体激活蛋白）干预人视网膜色素上皮细胞（hRPEs）和人巩膜成纤维细胞（hSFs）后，补体和NLRP3炎症小体通路活化，MAC合成增加，NLRP3炎症小体和IL-1β表达增高。同时，C3a显著抑制hSFs细胞增殖，促进细胞凋亡，伴随MMP-2表达升高和Collagen-1表达下降。靶向CD59（MAC体内抑制剂）siRNA可显著促进hSFs的NLRP3表达，而NLRP3-siRNA则可下调hSFs的C5b-9水平，验证MAC-NLRP3炎症小体二者的密切关联。本课题采用正反两方面研究，设置关键节点，结果支持假设：在近视诱导中，后巩膜MAC过度组装，细胞破坏增加，NLRP3炎症小体活化，“近视信使”释放增加，使细胞外基质代谢失衡，进而促进近视进展，解析了形觉剥夺-MAC-NLRP3炎症小体-“近视信使”-近视这一有序性信号通路的分子机制，为未来建立近视防控新靶点提供依据。