Uncoupling caspase 8-mediated-apotosis from caspase 8-mediated-inflammation in glaucoma.

青光眼中一个重要的悬而未决的问题是，视网膜神经节细胞(RGC)和 轴突变性是两个独立的事件，由分子上不同的途径介导，或者无论它们是 通过一条共同的路径链接和触发。回答这个问题是发展有效的关键 保护轴突和RGC SoMAS的神经保护疗法。支持这一想法的有两种截然不同的观点 介导轴突变性和RGC凋亡的途径，研究表明，前-丝裂原的缺失与RGC的凋亡有关。 凋亡的Bax基因阻断小鼠青光眼模型视网膜神经节细胞的凋亡，但不能阻止轴突 退化。越来越多的证据表明，轴突变性是由视神经中的轴突损伤引发的。 神经头，它先于RGC凋亡，与小胶质细胞激活和神经毒性炎症有关。 因此，阻断炎症有可能防止轴突变性和随后的死亡。 RGC，保留了连接轴突变性和RGC的共同途径的可能性 细胞凋亡。支持炎症可能是与轴突变性有关的共同途径的观点 和RGC凋亡，我们的实验室先前证明了阻断Fas/Fas配体(FasL)信号转导 通路预防慢性和可诱导的小鼠轴突变性和RGC细胞凋亡 青光眼和这种保护与抑制小胶质细胞激活和诱导炎症相一致 调解人。然而，由于Fas触发了凋亡和炎症途径，所以问题仍然是 单纯阻断炎症是否能防止轴突变性和随后的RGC凋亡。这个 解决这个问题的挑战是无法将Fas介导的炎症与Fas介导的炎症分开 细胞凋亡。然而，Fas信号需要激活caspase-8以促进细胞凋亡和炎症，并 最近对caspase-8激活分子机制的研究发现了另一种自体- 诱导caspase-8介导的细胞凋亡所必需的但不是诱导所必需的切割步骤 Caspase-8介导的炎症。利用这些信息，在中创建了一个突变小鼠(Casp8DA/DA 其中，自动切割位点的点突变阻止了caspase-8介导的细胞凋亡，但没有阻止。 Caspase-8介导的炎症反应。我们将使用这些小鼠来明确确定caspase-8是如何介导的 炎症和/或细胞凋亡在青光眼中触发轴突变性和RGC凋亡。我们假设 高眼压触发caspase-8介导的炎症，而炎症是触发轴突的共同途径 变性和RGC细胞凋亡。这一假说将在两个目标上得到验证：(1)确定caspase- 8介导的细胞凋亡在微球诱导的小鼠视网膜节细胞轴突变性和死亡中的作用 青光眼模型和(2)Fas介导的小胶质细胞激活介导的神经毒性 微珠诱导的小鼠青光眼模型中视网膜神经节细胞的炎症、轴突变性和死亡。

One important unanswered question in glaucoma is whether apoptosis of retinal ganglion cells (RGCs) and axon degeneration are two separate events mediated by molecularly distinct pathways, or whether they are linked and triggered through a common pathway. Answering this question is critical to developing effective neuroprotective therapies that protect both axons and RGC somas. In support of the idea there are two distinct pathways that mediate axon degeneration and RGC apoptosis, it was demonstrated that deletion of the pro- apoptotic BAX gene blocked apoptosis of RGCs in a mouse model of glaucoma, but did not prevent axon degeneration. There is accumulating evidence that axon degeneration is triggered by axon damage in the optic nerve head, which precedes RGC apoptosis and is linked to microglial activation and neurotoxic inflammation. Therefore, it is possible that blocking inflammation could prevent axon degeneration and subsequent death of RGCs, leaving open the possibility that there is a shared pathway linking axon degeneration and RGC apoptosis. In support of the idea that inflammation may be the shared pathway that links axon degeneration and RGC apoptosis, our laboratory demonstrated previously that blocking the Fas/Fas ligand (FasL) signaling pathway prevents axon degeneration and RGC apoptosis in both inducible and chronic mouse models of glaucoma and this protection coincided with an inhibition of microglia activation and induction of inflammatory mediators. However, since Fas triggers apoptotic and inflammatory pathways, the question remains as to whether blocking inflammation alone would prevent axon degeneration and subsequent RGC apoptosis. The challenge to resolving this issue is the inability to uncouple Fas-mediated inflammation from Fas-mediated apoptosis. However, Fas signaling requires activation of caspase-8 for both apoptosis and inflammation and recent studies examining the molecular mechanism of caspase-8 activation discovered an additional auto- cleavage step that is required for the induction of caspase-8-mediated apoptosis but not required for induction of caspase-8 mediated inflammation. Using this information, a mutant mouse was created (Casp8DA/DA) in which a point mutation in the auto-cleavage site blocked caspase-8-mediated apoptosis, but did not block caspase-8-mediated inflammation. We will use these mice to definitively determine how caspase-8-mediated inflammation and/or apoptosis triggers axon degeneration and RGC apoptosis in glaucoma. We hypothesize that elevated IOP triggers caspase-8-mediated inflammation that acts as a common pathway triggering axon degeneration and RGC apoptosis. This hypothesis will be tested in two aims: (1) Determine whether caspase- 8-mediated apoptosis contributes to axon degeneration and death of RGCs in a microbead-induced mouse model of glaucoma and (2) Demonstrate that Fas-mediated activation of microglia mediates neurotoxic inflammation, axon degeneration, and death of RGCs in the microbead-induced mouse model of glaucoma.