Initiating Events in AMD: An Animal Model for the Human Disease

这项提议的广泛、长期的目标是用分子术语定义 外视网膜蛋白质的氧化修饰，免疫系统对它们的识别和 外层视网膜易受攻击，导致老年性黄斑病变 退行性变(AMD)。上一次资助期间完成的工作显示，AMD眼部组织含有 通过加入长链多不饱和的氧化片段而修饰的高水平蛋白质 脂肪酸，二十二碳六烯酸(DHA)。许多被这种加合物和其他加合物修饰的蛋白质被发现 在黄褐斑和布鲁克的膜中。此外，针对这些独特的羧乙基吡咯的自身抗体 AMD患者循环(血浆)中的加合物(CEP)比老年患者更丰富 没有AMD的个体。从这些结果中，出现了以下关于 启动AMD的刺激：(A)。由于光感受器-RPE中DHA的浓度很高 再加上DHA对氧化损伤的脆弱性，CEP加合物的生成速度缓慢。 外视网膜的时间，(B)。这些CEP加合物代表了免疫系统外来的新表位。 导致产生针对CEP的自身抗体。(C)。抗CEP抗体反过来又参与了 Bruch膜-RPE界面补体攻击通路的激活 在外视网膜组织中持续产生CEP表位。为了检验这一假设，我们免疫了 正常小鼠用CEP加成的小鼠血清白蛋白。我们的预测是系统性免疫 CEP将使小鼠对内源性CEP加成蛋白敏感，内源性CEP加成蛋白在 正常的衰老过程。反过来，免疫系统将通过攻击CEP所在的细胞进行反应 表位是最容易产生的。对这些小鼠的分析显示，RPE和 模拟地理萎缩的光感受器，与干性AMD相关的致盲终末期萎缩。 这一新的AMD小鼠模型将在正常小鼠中进一步鉴定，CEP免疫将 扩展到补体途径分子及其调节器存在遗传缺陷的小鼠。一种新的 小鼠AMD模型将是临床前试验的重要新资源 旨在预防或限制AMD进展的治疗方法。

The broad, long-term objective of this proposal is to define in molecular terms the linkage between the oxidative modifications of proteins in the outer retina, their recognition by the immune system and the vulnerability of the outer retina to attack, leading to the disease processes underlying age-related macular degeneration (AMD). Work completed during the last grant period revealed that AMD eye tissues contain high levels of proteins modified by the adduction of oxidation fragments of the long chain polyunsaturated fatty acid, docosahexaenoic acid (DHA). Many of the proteins modified by this and other adducts are found in drusen and Bruch's membrane. Furthermore, autoantibodies against these unique carboxyethylpyrrole adducts (CEP) are more abundant in the circulation (plasma) of individuals with AMD than are found in agematched individuals without AMD. From these results the following hypothesis has emerged regarding an initiating stimulus for AMD: (a). Because of the high concentration of DHA in the photoreceptors-RPE complex coupled with the vulnerability of DHA to oxidative damage, CEP-adducts are slowly generated over time in the outer retina, (b). These CEP-adducts represent new epitopes foreign to the immune system resulting in the generation of autoantibodies against CEP. (c). Anti-CEP-antibodies in turn are involved in activation of the complement attack pathway at the Bruch's membrane-RPE interface in response to the continued generation of CEP epitopes in tissues of the outer retina. To test this hypothesis we immunized normal mice with CEP-adducted mouse serum albumin. Our prediction was that systemic immunization with CEP would sensitize mice to endogenous CEP-adducted proteins generated in the outer retina during the normal course of aging. In turn the immune system would respond by attacking the cells where CEP epitopes are most readily generated. Analysis of these mice demonstrated focal lesions in the RPE and photoreceptors that mimic geographic atrophy, the blinding end-stage atrophy associated with dry AMD. This new mouse model for AMD will be further characterized in normal mice and the CEP-immunizations will be extended to mice with genetic defects in complement pathway molecules and their regulators. A new model for AMD in the mouse will be an important a new resource for use in the preclinical testing of therapeutics designed to prevent or limit the progression of AMD.