Role of ER Stress in the Pathogenesis of Primary Open Angle Glaucoma

摘要 原发性开角型青光眼 (POAG) 是最常见的青光眼形式，通常伴有 由于小梁网 (TM) 无法维持正常水平的眼内压 (IOP) 房水流出。我们最近生成了一种新型转基因小鼠模型（Tg-MYOCY437H） 表达突变型肌纤蛋白（一种已知的人类 POAG 的主要遗传原因），并复制人类 青光眼表型。重要的是，我们将内质网（ER）应激与 Tg-MYOCY437H 小鼠青光眼的发病机制。错误折叠的肌纤蛋白在 ER 中积聚，诱发 ER 压力并激活保护性未折叠蛋白反应（UPR）。随着 UPR 激活，突变肌纤蛋白 还诱导自噬，这是一种已知可降解蛋白质聚集体的溶酶体降解过程。然而， 未能消除肌纤蛋白聚集体可能是由于 UPR 不足和自噬受损，TM 细胞 诱导内质网应激引发的凋亡转录因子 Chop，这可能进一步恶化内质网稳态 并导致 TM 功能障碍/丢失，升高 IOP 并导致 POAG。目前的提案将进一步 研究慢性 ER 应激在肌纤蛋白以及非肌纤蛋白相关发病机制中的作用 波格。在指导阶段，拟议的研究将确定是否未能激活 保护性 UPR (Atf-6¿-/-) 会加剧青光眼表型，而干扰 ER 应激诱导的 细胞凋亡 (Chop-/-) 可预防 Tg-MYOCY437H 小鼠的青光眼。此外，我们将调查 ER 是否 POAG 供体的死后 TM 组织中的应激被激活。在独立阶段，我们将 检查 TM 中 ER 应激的诱导是否与小鼠模型中 IOP 的升高相关 地塞米松诱发的高眼压症。此外，我们将研究自噬在降解中的作用 肌纤蛋白聚集体的变化，并将确定雷帕霉素诱导自噬是否可以挽救青光眼 Tg-MYOCY437H 小鼠。具体来说，考生将使用以下方法学习并生成 ER 压力的新模型 Atf6¿ 和 Chop 敲除小鼠，产生诱导多能干细胞 (iPSC) 衍生的小梁网 - 来自 POAG 患者的类似细胞，并表征了地塞米松诱导的高眼压小鼠模型 在 Val Sheffield 博士的实验室里。此外，在指导阶段，候选人将继续他的 通过咨询委员会的持续指导，实现专业、科学的职业发展。他 将参加科学会议，并与 ER 应激专家 Thomas Rutkowski 博士和青光眼合作 临床专家 Lee Alward 博士。他还将获得教学经验。该项目将有利于继续 对候选人进行技术、智力和专业培训，并协助候选人建立 学术研究所的独立研究实验室。

Abstract Primary open angle glaucoma (POAG), the most common form of glaucoma, is usually accompanied by elevated intraocular pressure (IOP) due to failure of the trabecular meshwork (TM) to maintain normal levels of aqueous humor outflow. We recently generated a novel transgenic murine model (Tg-MYOCY437H) that expresses mutant myocilin, a known leading genetic cause of POAG in humans and replicates human glaucoma phenotypes. Importantly, we have associated endoplasmic reticulum (ER) stress to the pathogenesis of glaucoma in Tg-MYOCY437H mice. Misfolded myocilin accumulates in the ER, induces ER stress and activates a protective unfolded protein response (UPR). Along with UPR activation, mutant myocilin also induces autophagy, a process of lysosomal degradation known to degrade protein aggregates. However, failure to eliminate myocilin aggregates possibly due to insufficient UPR and impaired autophagy, TM cells induce the ER stress-initiated apoptotic transcriptional factor, Chop, which may further worsen ER homeostasis and cause TM dysfunction/loss, elevating IOP and resulting in POAG. The current proposal will further investigate the role of chronic ER stress in the pathogenesis of myocilin as well as non-myocilin associated POAG. During the mentored phase, the proposed studies will determine whether failure to activate the protective UPR (Atf-6¿-/-) exacerbates glaucoma phenotypes, whereas interference with ER stress-induced apoptosis (Chop-/-) prevents glaucoma in Tg-MYOCY437H mice. In addition, we will investigate whether ER stress is activated in port-mortem TM tissues from POAG donors. During the independent phase, we will examine whether induction of ER stress in the TM is associated with elevation of IOP in a mouse model of dexamethasone-induced ocular hypertension. In addition, we will examine the role of autophagy in degradation of myocilin aggregates and will determine whether inducing autophagy by rapamycin will rescue the glaucoma of Tg-MYOCY437H mice. Specifically, the candidate will learn and generate new models of ER stress using Atf6¿ and Chop knockout mice, generate induced pluripotent stem cells (iPSCs)-derived trabecular meshwork- like cells from POAG patients, and characterize a dexamethasone-induced ocular hypertension mouse model in the laboratory of Dr. Val Sheffield. Additionally, during the mentored phase, the candidate will continue his professional and scientific career development through continual guidance from the advisory committee. He will attend scientific conferences, and collaborate with ER stress expert, Dr. Thomas Rutkowski and glaucoma clinical expert, Dr. Lee Alward. He will also acquire teaching experience. This project will facilitate continued technical, intellectual, and professional training of the candidate, and assist the candidate in the establishment of an independent research laboratory at an academic research institute.