RIP1/RIP3信号通路调控程序性坏死在视网膜炎症中的作用及机制研究

Retinal inflammation, a common feature of multiple retinal diseases, could promote degeneration and death of neurons and lead to blindness. In recent years, great progress has been made for the treatment of inflammatory diseases by modulating immune cell responses. However, the exact role of microglia, the resident immune cells in the retina, in the process of retinal inflammation largely remains unknown. Necroptosis is a newly discovered pathway of regulated cell death and provide a novel concept of necroptosis-mediated inflammation in macrophage. Interestingly, our previous work found elevated levels of the necroptosis-specific kinases RIP1 and RIP3 in retinal microglia of rd1 mice, indicating that microglia underwent necroptosis in retinal degeneration. Whether the necroptotic microglia contribute to the retinal inflammation needs to be elucidated. Based on our previous work and recent findings of others, we hypothesize that microglia would undergo RIP1-RIP3-dependent necroptosis after TLR4 activation, and release the inflammatory cytokines, aggravating the retinal degeneration. In this study, we are trying to elucidate the role of microglia necroptosis in retinal inflammation and the underlying molecular mechanisms by loss-and gain-of-function strategies. In addition, the effect of necroptosis blockade by Nec-1 in the diseased mice will be identified. The results thus may provide new evidence on the mechanism of microglia necroptosis in retinal inflammation and also present an attractive new strategy for the treatment of retinal inflammatory diseases such as retinal degeneration.

炎症反应普遍存在于视网膜变性等难治性致盲眼病中。调控免疫细胞治疗炎症性疾病研究近年取得了突破，但视网膜小胶质细胞这一特殊免疫细胞的功能特性及其在视网膜炎症中的作用和机制尚未明确。我们前期研究发现视网膜变性小鼠的小胶质细胞高表达RIP1和RIP3，提示其发生程序性坏死，这一新型死亡方式可能是小胶质细胞发挥促炎功能的新形式，是调控视网膜炎症的新靶点。由此我们提出小胶质细胞程序性坏死是否及如何介导炎症促进视网膜变性的科学问题。本项目将利用基因敲除鼠、siRNA等技术在体内外水平研究：1)视网膜变性疾病中小胶质细胞程序性坏死介导视网膜炎症的作用；2)小胶质细胞经TLR4激活RIP1/3通路进而释放炎症因子的分子机制；3)以RIP1为靶点抑制程序性坏死缓解视网膜变性的可能性和有效性。研究成果将从特殊免疫细胞死亡的新角度揭示视网膜炎症的细胞和分子机制，为调控小胶质细胞干预视网膜变性疾病提供依据。

视网膜炎症是视网膜变性等难治性致盲眼病的重要病理基础。调控免疫细胞被认为是近年来炎症性疾病治疗研究的新方向。本项目基于前期发现视网膜变性小鼠的小胶质细胞可高表达RIP1和RIP3，创新性提出视网膜小胶质细胞可能通过程序性死亡发挥介导炎症功能的新观点。本项目主要围绕RIP1/RIP3信号通路调控程序性坏死在视网膜炎症中的作用及机制展开系列研究。首先在视网膜多种炎症疾病体内外模型中明确小胶质细胞发生了程序性坏死，并可高表达TNF-α、CCL2等促炎因子和趋化因子，从而促进视网膜炎症。进一步研究小胶质细胞发生程序性坏死的上游机制，利用TLR4基因敲除小鼠发现TLR4活化介导了RIP1和RIP3通路激活，及TNF-a、CCL2的释放，提示TLR4信号通路介导了小胶质细胞的程序性坏死，参与视网膜炎症反应。另外，本项目还以细胞死亡及调控为切入点，明确调控小胶质细胞活化状态及抑制程序性坏死，缓解视网膜变性的有效性。发现以RIP1为靶点应用程序性坏死抑制剂Necrostatin-1及AAT调控小胶质细胞极化，可有效缓解神经炎症，减轻视网膜变性。这些研究结果揭示了小胶质细胞活化和程序性坏死可能是小胶质细胞发挥促炎功能的新形式，可作为调控视网膜炎症的新靶点。该项目研究成果目前已发表SCI论文4篇，相关结果在眼科学国际大会上交流。本项目从动物模型、细胞水平及分子机制不同层次开展研究，研究成果从特殊免疫细胞死亡的角度，揭示了视网膜炎症的新细胞和分子机制，为调控小胶质细胞干预视网膜变性疾病提供依据，为治疗视网膜变性疾病提供新思路和新策略。