Role of Alpha 2 Collagen VIII in Fuchs Corneal Dystrophy

描述（由申请人提供）：本提案的总体目标是为首席研究员（PI）提供必要的经验和技能，使其成为研究角膜疾病基本机制的独立研究人员。本提案的科学重点是更好地理解由VIII型胶原突变引起的细胞和细胞外基质（ECM）蛋白异常，从而导致Fuchs内皮营养不良（FED）患者角膜内皮细胞（CEC）丢失。胶原VIII （COL8）在CECs特化基底膜Descemet membrane （DM）的形成中起核心作用，编码胶原VIII (COLSA2) α 2链的基因突变导致一些患者发生FED。在人类角膜移植中，FED已经发展了几十年，占角膜移植的29%。该疾病的早期阶段是无症状的，没有轻度受影响的组织。因此，几乎没有关于导致FED的早期细胞和ECM变化的信息。通过开发可靠的FED动物模型以及对致病性COL8A2突变对胶原VIII功能的详细生化分析，可以大大增加对这些早期致病事件的了解。该提案的基本假设是COL8A2基因突变产生细胞和生化异常，导致FED中CEC丢失。识别这些异常应该为FED的发病机制提供重要的见解，这可能为这一重要角膜疾病的合理治疗提供建议。为了解决这一假设，提出了两个具体目标：目标1：利用基因靶向技术引入R155Q COL8A2突变，建立并表征小鼠FED模型。目的2：研究已知引起FED的COLSA2突变对α - 1胶原VIII（另一个COL8亚链与COL8A2在体内相互作用）的同三聚体和异三聚体形成的影响。在拟议的研究和选定的教学活动过程中，PI将获得宝贵的培训经验，并在开发眼部疾病的动物模型和ECM蛋白的生化分析方面获得指导。在这些领域的专业知识被认为是宝贵的PI谁渴望开发一个独立的研究计划，研究角膜疾病的基本机制。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to provide the principal investigator (PI) with the experiences and skills necessary to become an independent researcher studying basic mechanisms of corneal diseases. The scientific focus of this proposal is to gain a better understanding of the cellular and extracellular matrix (ECM) protein abnormalities caused by mutations in collagen VIII which lead to corneal endothelial cell (CEC) loss in Fuchs endothelial dystrophy (FED). Collagen VIII (COL8) plays a central role in the formation of Descemet membrane (DM), the specialized basement membrane of CECs, and mutations in the gene encoding the alpha2 chain of collagen VIII (COLSA2) cause FED in some patients. FED progresses over decades in humans and accounts for up to 29% of corneal transplants. Early stages of the disease are asymptomatic, and mildly affected tissues are not available. Thus, virtually no information exists about the early cellular and ECM changes leading to FED. Understanding these early pathogenic events could be increased greatly through the development of a reliable animal model of FED as well as detailed biochemical analysis of pathogenic COL8A2 mutations on collagen VIII function. The underlying hypothesis of this proposal is that mutations in the COL8A2 gene produce cellular and biochemical abnormalities which cause CEC loss in FED. Identifying these abnormalities should provide important insights into the pathogenesis of FED, which may suggest rational therapies for this important corneal disease. To address this hypothesis, two specific aims are proposed: Aim 1: To develop and characterize a mouse model of FED by introducing the R155Q COL8A2 mutation using gene targeting techniques. Aim 2: To investigate the effects of COLSA2 mutations known to cause FED on homotrimer and heterotrimer formation with alpha1 collagen VIII, the other COL8 subchain interacting with COL8A2 in vivo. In the course of the proposed research and selected didactic activities, the PI will gain invaluable training experience and mentoring in developing animal models of ocular disease and biochemical analyis of ECM proteins. Expertise in these areas is deemed invaluable for the PI who aspires to develop an independent research program studying basic mechanisms of corneal diseases.