Proteomic Studies of Age Related Macular Degeneration

描述（由申请人提供）：本研究的长期目标是了解年龄相关性黄斑变性（AMD）中玻璃疣形成和布鲁赫膜增厚的分子机制。统一的假设是，蛋白质修饰是因果关系参与这两个过程。玻璃疣是在视网膜色素上皮和布鲁赫膜之间形成的细胞外沉积物，并且融合性玻璃疣是发展AMD的标志性风险因素。如果可以调节玻璃疣和布鲁赫膜的变化，AMO的进展可能会减慢或停止。拟议的研究将确定来自健康和AMD供体组织的玻璃疣和布鲁赫膜中的蛋白质和脂质，并表征相关的蛋白质修饰和反应性脂质片段。四个具体目标将检验以下假设： (i)玻璃疣蛋白质组在玻璃疣亚型之间不同;（ii）布鲁赫膜蛋白质组随年龄和健康状态而变化;（iii）玻璃疣和布鲁赫膜中的脂质氧化产物随年龄和健康状态而变化;（iv）氧化蛋白质修饰有助于玻璃疣形成和布鲁赫膜增厚。质谱法将用于鉴定和表征蛋白质、磷脂和脂质氧化产物。免疫细胞化学将用于确认蛋白定位于玻璃疣和布鲁赫膜。Western分析、电泳迁移率变化和生物信息学工具将用于检测蛋白质修饰。这些结果将有助于在氧化损伤和AMD的发病机制之间建立牢固的联系，并为开发预防或限制AMD并发症的策略和疗法提供新的机会。

DESCRIPTION (provided by applicant): The long-term goal of this research is to understand the molecular mechanisms of drusen formation and Bruch's membrane thickening in age-related macular degeneration (AMD). The unifying hypothesis is that protein modifications are causally involved in both processes. Drusen are extracellular deposits that form between the retinal pigment epithelium and Bruch's membrane and confluent drusen are the hallmark risk factor for developing AMD. The progression of AMO might be slowed or halted if drusen and Bruch's membrane changes can be modulated. The proposed research will identify proteins and lipids in drusen and Bruch's membrane from healthy and AMD donor tissues and characterize associated protein modifications and reactive lipid fragments. Four specific aims will test the following hypotheses: (i) that the drusen proteome differs among drusen sub-types; (ii) that the Bruch's membrane proteome varies with age and state of health; (iii) that lipid oxidation products in drusen and Bruch's membrane vary with age and state of health; (iv) that oxidative protein modifications contribute to drusen formation and Bruch's membrane thickening. Mass spectrometric methods will be used to identify and characterize proteins, phospholipids and lipid oxidation products. Immunocytochemistry will be used to confirm protein localization to drusen and Bruch's membrane. Western analyses, electrophoretic mobility shifts and bioinformatic tools will be used to detect protein modifications. The results will help establish a firm link between oxidative damage and the pathogenesis of AMD and provide new opportunities for developing strategies and therapies for preventing or limiting the complications of AMD.