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主要研究者（PI）提供成为研究角膜疾病基本机制的独立研究者所需的经验和技能。该提案的科学重点是更好地了解由胶原蛋白VIII突变引起的细胞和细胞外基质（ECM）蛋白异常，这些突变导致Fuchs内皮营养不良（FED）中的角膜内皮细胞（CEC）损失。胶原蛋白VIII（COL8）在后弹力膜（DM）（CEC的特化基底膜）的形成中起核心作用，并且编码胶原蛋白VIII的α 2链（COLSA2）的基因中的突变在一些患者中引起FED。 FED在人类中发展了几十年，占角膜移植的29%。疾病的早期阶段是无症状的，并且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突变对胶原蛋白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。 该建议的基本假设是，COL8A2基因突变产生细胞和生化异常，导致FED中CEC损失。识别这些异常应提供重要的见解FED的发病机制，这可能会建议合理的治疗这一重要的角膜疾病。 为了解决这一假设，提出了两个具体的目标：目标1：通过使用基因打靶技术引入R155Q COL8A2突变来开发和表征FED小鼠模型。 目标二：研究已知导致FED的COLSA 2突变对与α 1胶原VIII形成同源三聚体和异源三聚体的影响，α 1胶原VIII是另一种在体内与COLSA 8A2相互作用的COLSA 8亚链。 在拟议的研究和选定的教学活动的过程中，PI将获得宝贵的培训经验和指导，在开发眼部疾病的动物模型和ECM蛋白的生化分析。 这些领域的专业知识被认为是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是无价的。

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.