Modulation of Neuroinflammation in Glaucoma

项目摘要 来自人类青光眼研究和实验模型的越来越多的信息支持胶质细胞驱动的青光眼。 神经炎症对视网膜神经节细胞（RGC）胞体、轴突和轴突产生额外的破坏力， 突触，其治疗调制可以提供广泛的神经保护。我们积累的数据来自 最近关于青光眼神经炎症的研究导致了我们的新假设，我们的目标是在这个项目中进行测试。 显然，cFLIP作为半胱天冬酶-8介导的细胞死亡/存活/炎症之间的分子开关起作用 通过TNFRs，TLR和炎性小体的通路，我们的第一个特定目的将测试条件性缺失是否 星形胶质细胞cFLIP的表达可抑制caspase-8/cFLIP相互作用的炎症结局， 实验性青光眼炎症/神经毒性星形胶质细胞表型我们预计，cFLIP的删除， 星形胶质细胞将选择性地靶向炎性/神经毒性（A1）星形胶质细胞（因为这些细胞被启动以 炎性caspase-8通过TNF-α/TNFR、Fas/FasL、TLR和炎性小体发挥作用），并将提供 免疫调节（而不危及神经胶质神经支持功能）。除了cFLIP，我们将有选择地针对 cFLIPL还可以确定炎症结果和细胞命运决定中的亚型特异性差异。我们将 通过形态学和分子生物学方法分析对星形胶质细胞cFLIP（或cFLIPL）缺失的空间和时间响应 视网膜和视神经星形胶质细胞的分析，包括炎症反应，A1/A2星形胶质细胞分布，细胞 死亡（凋亡/坏死性凋亡）和神经支持功能。消除星形胶质细胞驱动的炎症可以 在实验性青光眼中保护RGCs免受炎性损伤。我们的具体目标2将检验这一假设 通过分析神经元存活（通过RGC轴突/索马计数）和功能（通过PERG）。星形胶质细胞和小胶质细胞 它们密切相互作用并调节彼此的表型和免疫功能。我们最近的研究结果也 支持星形胶质细胞的细胞因子反应可能塑造实验性青光眼中的小胶质细胞反应。我们 因此，特定目标3被设计为测试cFLIP（或cFLIPL）的胶质细胞亚型特异性缺失是否可以影响cFLIP（或cFLIPL）的表达。 个体的贡献和神经炎症过程中星形胶质细胞和小胶质细胞之间的相互作用。我们将 分析小胶质细胞对星形胶质细胞缺失的反应（包括M1/M2转换），以及星形胶质细胞的反应 （包括A1/A2分布）与不同位点/时间点的小胶质细胞缺失的关系。纵向分析还将 包括少突胶质细胞和Müller神经胶质反应。通过分析细胞类型特异性反应（包括 分离的胶质细胞亚型的分子反应）胶质细胞亚型靶向小鼠模型，该项目将价值 胶质细胞cFLIP作为免疫调节治疗靶点预防/逆转神经退行性炎症 实验性青光眼该项目的成果预计也将广泛用于其他研究。 眼/神经变性/神经炎性疾病、衰老、干细胞/轴突再生和癌症研究。

PROJECT SUMMARY Growing information from the studies of human glaucoma and experimental models supports that glia-driven neuroinflammation creates an additional damaging force on retinal ganglion cell (RGC) somas, axons, and synapses, and its therapeutic modulation can provide widespread neuroprotection. Our accumulated data from recent studies of neuroinflammation in glaucoma led to our new hypotheses that we aim to test in this project. Evidently, cFLIP functions as a molecular switch between caspase-8-mediated cell death/survival/inflammation pathways through TNFRs, TLRs, and inflammasome, Our first Specific Aim will test whether conditional deletion of astroglial cFLIP can inhibit the inflammatory outcomes caspase-8/cFLIP interaction and prevent/reverse the inflammatory/neurotoxic astrocyte phenotype in experimental glaucoma. We expect that deletion of cFLIP in astroglia will selectively target inflammatory/neurotoxic (A1) astrocytes (because these cells are primed for inflammatory caspase-8 functions through TNF-α/TNFR, Fas/FasL, TLRs, and inflammasome) and will provide immunomodulation (without risking the glial neurosupport functions). Besides cFLIP, we will selectively target cFLIPL to also determine isoform-specific differences in inflammatory outcomes and cell fate decisions. We will analyze spatial and temporal responses to astroglial cFLIP (or cFLIPL) deletion by morphological and molecular analysis of retina and optic nerve astroglia, including inflammatory responses, A1/A2 astrocyte distribution, cell death (apoptosis/necroptosis), and neurosupport functions. Elimination of astroglia-driven inflammation can protect RGCs against inflammatory injury in experimental glaucoma. Our Specific Aim 2 will test this hypothesis by analysis of neuron survival (by RGC axon/soma counting) and function (by PERG). Astroglia and microglia closely interplay and regulate each other’s phenotype and immune functions. Our recent research findings also supported that astroglial cytokine responses may shape microglia responses in experimental glaucoma. Our Specific Aim 3 is therefore designed to test whether glial subtype-specific deletion of cFLIP (or cFLIPL) can impact individual contributions and the cross-talk between astroglia and microglia during neuroinflammation. We will analyze microglia responses (including M1/M2 conversion) to astroglial deletions, and and astroglia responses (including A1/A2 distribution) to microglial deletions at different sites/time points. Longitudinal analysis will also include oligodendrocyte and Müller glia responses. Through the analysis of cell type-specific responses (including the molecular responses of isolated glial subtypes) to glial subtype-targeting mouse models, this project will value glial cFLIP as an immunomodulatory treatment target to prevent/reverse neurodegenerative inflammation in experimental glaucoma. Outcomes of this project are also expected to be widely useful in studies of other ocular/neurodegenerative/neuroinflammatory diseases, aging, stem cell/axon regeneration, and cancer research.