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

描述（由申请人提供）：原发性开角型青光眼（POAG）是最常见的青光眼形式，由于小梁网（TM）无法维持房水输出的正常水平，通常伴有眼内压（IOP）升高。我们最近建立了一种新的转基因小鼠模型（Tg-MYOCY437H），该模型表达突变心肌蛋白，这是人类POAG的已知主要遗传原因，并复制人类青光眼表型。重要的是，我们已经将内质网应激与Tg-MYOCY437H小鼠青光眼的发病机制联系起来。错误折叠的心肌在内质网中积累，诱导内质网应激并激活保护性未折叠蛋白反应（UPR）。随着UPR的激活，突变型心肌蛋白也会诱导自噬，这是一种溶酶体降解蛋白质聚集体的过程。然而，可能由于UPR不足和自噬受损导致的心肌聚集体无法消除，TM细胞诱导内质网应激启动的凋亡转录因子Chop，这可能进一步恶化内质网稳态，导致TM功能障碍/丧失，IOP升高，导致POAG。本课题将进一步研究慢性内质网应激在心肌及非心肌相关性POAG发病机制中的作用。在指导阶段，拟议的研究将确定未能激活保护性UPR （Atf-6¿-/-）是否会加剧青光眼表型，而干扰内膜应激诱导的凋亡（Chop-/-）是否会预防Tg-MYOCY437H小鼠的青光眼。此外，我们将研究内质网应激是否在POAG供体的死后TM组织中被激活。在独立阶段，我们将在地塞米松诱导的高眼压小鼠模型中检查TM中内质网应激的诱导是否与IOP升高有关。此外，我们将研究自噬在心肌蛋白聚集体降解中的作用，并确定雷帕霉素诱导自噬是否会挽救Tg-MYOCY437H小鼠的青光眼。具体来说，候选人将学习并使用Atf6¿和Chop敲除小鼠产生新的内质网应激模型，从POAG患者中产生诱导多能干细胞（iPSCs）衍生的小骨网样细胞，并在Val Sheffield博士的实验室中表征地塞米松诱导的高眼压小鼠模型。此外，在指导阶段，候选人将通过咨询委员会的持续指导，继续其专业和科学的职业发展。他将参加科学会议，并与ER应激专家Thomas Rutkowski博士和青光眼临床专家Lee Alward博士合作。他还将获得教学经验。该项目将促进候选人持续的技术、智力和专业培训，并协助候选人在学术研究机构建立独立的研究实验室。

DESCRIPTION (provided by applicant): 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, intellectul, and professional training of the candidate, and assist the candidate in the establishment of an independent research laboratory at an academic research institute. PUBLIC HEALTH RELEVANCE: The successful completion of this project will lead to insight into elevated intraocular pressure, a major risk factor of the human blinding disorder, primary open angle glaucoma. The results will potentially improve diagnosis, genetic risk assessment and treatment of this common disorder.