Mechanisms of toll-like receptor-mediated neurotoxicity in the ischemic retina

项目摘要 视网膜组织的缺血性损伤是一种经常导致视觉障碍和失明的临床病症， 影响患者的生活质量和功能状态。但是，更严格地理解的机制， 如果要开发新的治疗视网膜疾病的方法， 开发以前，我们证明了toll样受体4（Tlr 4）信号转导在视网膜神经节细胞中的有害作用。 炎症和损伤引起的局部缺血状况。由于Tlr 4信号传导由两个不同的 信号级联，Myd 88-和Trif依赖，我们分别评估了这些级联的作用， 缺血性视网膜现有的证据似乎表明，Trif和Myd 88信号转导介导的作用， 在缺血视网膜组织中是视网膜特异性和不对称的。而Trif或Myd 88的失活 导致缺血性视网膜组织中的炎症显著减少，Myd 88缺陷型的缺血性视网膜 动物表现出比Trif缺陷动物的缺血性视网膜显著更高的损伤水平。我们 还注意到缺血诱导的Trif信号直接促进坏死的视网膜神经节细胞（RGC）死亡。 此外，我们证明了RGC坏死，这加剧了视网膜损伤，通过促进更多的 炎症，在某些情况下可以调节（称为“坏死性凋亡”）。由于Trif信号可以介导细胞凋亡， 在目的1中，我们假设Trif信号通过以下途径介导缺血诱导的视网膜损伤： 促进RGC坏死性凋亡。此外，Trif依赖性信号传导在以下方面不同于Myd 88信号传导级联： 其激活I型干扰素（IFN）信号传导的能力。以前的研究表明，Trif通过激活IFN， 激活半胱天冬酶-11（Casp 11），后者又可直接和间接（通过NLRP 3）介导细胞死亡 炎性体活性和白细胞介素-1b [IL 1b]释放）。鉴于我们发现高水平的Casp 11和 在目的2中，我们假设缺血诱导的Trif信号转导可能与缺血性RGC中的炎性小体活性有关。 介导Casp 11激活，而Casp 11又直接和间接介导RGC死亡（通过升高的 炎症反应）。最后，由于Müller胶质细胞（MG）活性与神经保护作用有关， 视网膜中的每种病理状况，我们在目标3中假设缺血诱导的MG特异性Tlr 4- 依赖性神经保护活性优于神经毒性活性以促进缺血性RGC的存活。如果 假设被证明是正确的，我们将能够解释Trif的不对称作用， 缺血性视网膜组织中的Myd 88：虽然Myd 88调节神经胶质细胞毒性（星形胶质细胞和小胶质细胞）， 在缺血性视网膜的神经保护（MG）中，Trif激活上述两种现象，同时也直接 介导RGC死亡，从而促进比Myd 88更显著的视网膜损伤。评估我们 假设，我们将使用动物模型，并采用广泛的生物化学，分子和细胞生物学 技术.通过完成这个项目，我们将为新的视网膜特异性 可以更有效地治疗缺血性视网膜疾病的治疗策略。

PROJECT SUMMARY Ischemic injury to retinal tissue is a clinical condition that frequently leads to visual impairments and blindness, affecting patients' quality of life and functional status. But a more rigorous understanding of the mechanisms of ischemia-induced retinal injury will be required if new therapies for the management of retinal disease are to be developed. Previously, we demonstrated the deleterious role of toll-like receptor 4 (Tlr4) signaling in retinal inflammation and damage triggered by ischemic conditions. Since Tlr4 signaling consists of two distinct signaling cascades, Myd88- and Trif-dependent, we individually evaluated the role of these cascades in ischemic retinae. Available evidence appears to indicate that the effects mediated by Trif and Myd88 signaling in ischemic retinal tissue are retina-specific and asymmetric. Whereas inactivation of either Trif or Myd88 resulted in significantly reduced inflammation in ischemic retinal tissue, ischemic retinae of Myd88-deficient animals demonstrated significantly higher levels of damage than ischemic retinae of Trif-deficient animals. We also noted that ischemia-induced Trif signaling directly facilitates necrotic retinal ganglion cell (RGC) death. Furthermore, we demonstrated that RGC necrosis, which exacerbates retinal injury by promoting more inflammation, can be regulated in some cases (termed “necroptosis”). Since Trif signaling can mediate cell necroptosis, we hypothesize in Aim 1 that Trif signaling mediates ischemia-induced retinal damage by promoting RGC necroptosis. In addition, Trif-dependent signaling differs from the Myd88 signaling cascade in its ability to activate type I interferon (IFN) signaling. It was previously shown that Trif, via its activation of IFN, activates caspase-11 (Casp11), which can in turn mediate cell death both directly and indirectly (via NLRP3 inflammasome activity and interleukin-1b [Il1b] release). In light of our finding of high levels of Casp11 and inflammasome activity in ischemic RGCs, we hypothesize in Aim 2 that ischemia-induced Trif signaling mediates Casp11 activation, which in turn mediates RGC death both directly and indirectly (via elevated inflammatory responses). Finally, since Müller glia (MG) activity is associated with neuroprotection in nearly every pathological condition in the retina, we hypothesize in Aim 3 that ischemia-induced, MG-specific, Tlr4- dependent neuroprotective activity prevails over neurotoxic activity to facilitate survival of ischemic RGCs. If the proposed hypotheses are found to be correct, we will be able to explain the asymmetric roles of Trif and Myd88 in ischemic retinal tissue: while Myd88 regulates glial toxicity (astrocytes and microglia) and neuroprotection (MG) in ischemic retinae, Trif activates both aforementioned phenomena while also directly mediating RGC death, thus promoting more significant retinal damage than Myd88. To evaluate our hypotheses, we will use animal models and employ a wide range of biochemical, molecular, and cell biological techniques. By completing this project, we will assemble an intellectual foundation for new retina-specific therapeutic strategies that can more effectively treat ischemic retinal diseases.