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，通常伴有由于小梁网（TM）无法维持正常水平的房水流出而导致的眼内压（IOP）升高。我们最近产生了一种新的转基因小鼠模型（Tg-MYOCY 437 H），表达突变型肌球蛋白，一个已知的主要遗传原因POAG在人类和复制人类青光眼表型。重要的是，我们已经将内质网（ER）应激与Tg-MYOCY 437 H小鼠青光眼的发病机制联系起来。错误折叠的myocilin在ER中积累，诱导ER应激并激活保护性未折叠蛋白反应（UPR）。沿着UPR激活，突变型肌球蛋白还诱导自噬，这是一种已知降解蛋白质聚集体的溶酶体降解过程。然而，可能由于UPR不足和自噬受损而未能消除肌钙蛋白聚集体，TM细胞诱导ER应激启动的凋亡转录因子Chop，这可能进一步恶化ER稳态并导致TM功能障碍/丧失，升高IOP并导致POAG。目前的建议将进一步研究慢性内质网应激的作用，在发病机制中的myocilin以及非myocilin相关的POAG。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ER应激诱导的细胞凋亡（Chop-/-）是否会预防Tg-MYOCY 437 H小鼠的青光眼。此外，我们将研究ER应激是否在POAG供体的尸检TM组织中被激活。在独立阶段，我们将研究地塞米松诱导的高眼压小鼠模型中TM中ER应激的诱导是否与IOP升高相关。此外，我们将研究自噬在肌球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-MYOCY 437 H小鼠的青光眼。具体而言，候选人将使用Atf 6和Chop敲除小鼠学习和生成新的ER应激模型，从POAG患者中生成诱导多能干细胞（iPSC）衍生的小梁网样细胞，并在瓦尔谢菲尔德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，在辅导阶段，候选人将通过咨询委员会的持续指导继续其专业和科学职业发展。他将参加科学会议，并与ER压力专家托马斯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的持续技术，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.