NLRP3信号通路介导Th细胞调节在糖尿病视网膜病变发病中的作用机制研究

Diabetic retinopathy （DR）is the most common complication of diabetes and the leading cause of blindness in the world. Immunological mechanisms is believed to contribute actively to DR associated damage to the retinal vasculature and retinal neovascularization. Our preliminary studies have provided evidences suggesting DR patients have an increase in the proinflammatory Th17/Th22 axis. We also found that IL-1β and IL-18 ,which can enhance Th17/Th22 cell differentiation, were significantly elevated in patients with DR ; NLRP3 was proved to be associated with autoinflammatory disorders characterized with excessive production of IL-1βand IL-18.Based on these results, we hypothesize that NLRP3 plays a a critical role in the induction of retinal inflammation in diabetic retinopathy, likely through effects on caspase-1-dependent cytokines which then influence Th17/Th22.To test this hypothesis, we propose the following studies: 1) to elucidate that NLRP3 activation increases caspase-1- IL-1β/IL-18 pathway activity in T2DM rat model and in the primary culture of retinal capillary endothelial cells, pericytes and müller cells incubated by high glucose;2) To explore whether NLRP3 pathway regulates the Th17/Th22 cell differentiation by evaluate the mechanisms of the effect of TXNIP and siRNA-NLRP3. These studies will establish the NLRP3 pathway as a novel pathogenic pathway in diabetic retinopathy and provide scientific theoretical basis for the prevention and treatment of DR.

糖尿病视网膜病变（DR）是由高血糖引起的致盲性视网膜血管病变，与免疫应答紊乱所致的炎症反应密切相关。申请人前期研究发现DR患者外周循环及房水中Th17、Th22细胞优势性表达，可调控其分化的IL-1β及IL-18同时上调。NLRP3炎症小体是IL-1β及IL-18活化的关键因子，其信号通路可能通过调节Th17、Th22细胞分化在DR炎症损伤中发挥重要作用。本课题拟从Th17、Th22的上游调控机制着手，通过体内外实验在大鼠DR模型及高糖培养视网膜内皮细胞、周细胞、müller细胞中检测NLRP3信号通路各分子表达水平，采用NLRP3天然配体TXNIP及RNA干扰技术了解上调和抑制其信号通路对Th17、Th22细胞分化及相关炎症损伤因子表达的调节作用，从正反两方面探讨NLRP3介导Th17、Th22细胞调节在DR发病中的作用，明确其促进DR炎症损伤的分子机理，有望为DR防治提供新的干预路径。

目前，大量研究已证实炎症小体在多种免疫与炎症相关疾病中发挥重要作用，然而其在糖尿病视网膜病变(DR)中的作用却仍不清楚。本研究通过体内外实验从分子水平探讨NLRP3炎症小体及其信号传导通路在DR发病中的作用机制，为DR的防治研究提供新的途径。实验结果表明：（1）T2DM大鼠视网膜和高糖培养视网膜内皮细胞胞中NLRP3—ASC—caspase-1—IL-1β轴各分子的表达上调，明确NLRP3炎症小体确实在DR发病过程中发挥作用；（2）同时通过NLRP3配体TXNIP诱导NLRP3炎症小体及其信号通路活化可导致Th17细胞表达增加，促进视网膜炎症损伤；（3）通过RNA干扰技术抑制 NLRP3炎症小体及其信号通路活化，可导致Th17细胞表达下降，减轻视网膜炎症损伤的影响，进一步阐明NLPR3炎症小体信号通路介导Th17细胞调节失衡可引起DR炎症损伤，从而揭示NLPR3炎症小体在DR发病中的作用机制，为DR的防治研究提供新的思路和理论依据。