Drusen and AMD: sub-type isolation and characterization

描述（由申请人提供）：本申请的广泛、长期目标是在分子方面定义黄斑中视网膜色素上皮（RPE）下的软性玻璃疣的积累与发生年龄相关性黄斑变性（AMD）的风险增加之间的联系。黄斑中的软玻璃疣的存在是发展为AMD的标志性风险因素。令人惊讶的是，对玻璃疣的组成或起源知之甚少。为此，已经开发了一种用于玻璃疣分离的新方法，其允许从供体眼组织收集微克量的玻璃疣。在分离时，可以鉴定和分离不同的玻璃疣亚型，用于表征其分子组成的研究。玻璃疣在AMD中的诊断效用可以比作动脉粥样硬化中的血液胆固醇水平。玻璃疣的存在和丰富程度，就像血液中的胆固醇水平一样，表明患者患这种疾病的风险程度。由于玻璃疣和AMD的关系，了解不同玻璃疣亚型的组成将提供重要的信息，可能的途径，是因果关系参与玻璃疣的发展。玻璃疣中存在的新蛋白质或蛋白质的常见修饰应该提供关于治疗AMD的治疗剂的潜在药物靶点的见解。本申请集中于利用这种玻璃疣分离程序来定义正常和AMD组织中玻璃疣亚型的分子组成、分布和细胞起源。三个具体目的是：（1）检验玻璃疣的不同亚群可以从供体眼组织中分离的假设。(hard与软质、中央凹与周边、陈旧与陈旧、球形与扁平、琥珀色与透明、不透明与颗粒状等）。(2)检验玻璃疣亚群的不同结构特征反映不同分子组成的假设（光学显微镜、组织化学、电子显微镜、SDS/PAGE、蛋白质印迹、质谱）。(3)为了检验玻璃疣中存在的一些分子是新的并且在布鲁赫膜中未发现的假设。

DESCRIPTION (provided by applicant): The broad, long-term objective of this application is to define in molecular terms the linkage between the accumulation of soft drusen below the retinal pigment epithelium (RPE) in the macula and the increased risk of developing age-related macular degeneration (AMD). The presence of soft drusen in the macula is the hallmark risk factor for developing AMD. Surprisingly little is known of the composition or origin of drusen. To this end a novel method for drusen isolation has been developed that allows the collection of microgram quantities of drusen from donor eye tissue. At the time of isolation, different drusen sub-types can be identified and separated for use in studies that will characterize their molecular composition. The diagnostic utility of drusen in AMD can be likened to that of blood levels of cholesterol in atherosclerosis. The presence and abundance of drusen, like the level of cholesterol in the blood, indicates the degree to which a patient is at risk for developing the disease. Because of the relationship of drusen and AMD, understanding the composition of different drusen sub-types will provide important information on possible pathways that are causally involved in drusen development. Novel proteins or common modifications of proteins present in drusen, should provide insight as to potential drug targets of therapeutic agents to treat AMD. The current application is focused on exploiting this drusen isolation procedure to define the molecular composition, distribution and cellular origin of drusen sub-types in normal and AMD tissues. The three specific aims are: (1) To test the hypothesis that different sub-populations of drusen can be isolated from donor eye tissue. (hard vs. soft, foveal vs. peripheral, old vs. older, spherical vs. flat, amber vs. clear, opaque vs. granular, etc.). (2) To test the hypothesis that different structural features of drusen sub-populations reflect different molecular composition (light microscopy, histochemistry, electron microscopy, SDS/PAGE, Western blotting, mass spectrometry). (3) To test the hypothesis that some of the molecules present in drusen are novel and are not found in Bruch's membrane.