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

描述（由申请人提供）：本提案的广泛、长期目标是在分子方面定义外层视网膜中蛋白质的氧化修饰、它们被免疫系统识别以及外层视网膜对攻击的脆弱性之间的联系，从而导致年龄相关性黄斑变性（AMD）的疾病过程。在上一次资助期间完成的工作显示，AMD眼组织含有高水平的蛋白质，这些蛋白质通过加合长链多不饱和脂肪酸二十二碳六烯酸（DHA）的氧化片段而被修饰。许多被这种加合物和其他加合物修饰的蛋白质存在于玻璃疣和布鲁赫膜中。此外，针对这些独特的羧乙基吡咯加合物（CEP）的自身抗体在患有AMD的个体的循环（血浆）中比在年龄匹配的没有AMD的个体中发现的更丰富。从这些结果中，已经出现了关于AMD的起始刺激的以下假设：（a）.由于光感受器-RPE复合物中DHA的高浓度以及DHA对氧化损伤的脆弱性，CEP-加合物随时间在外部视网膜中缓慢产生（B）。这些CEP加合物代表免疫系统的新表位，导致产生针对CEP的自身抗体。（c）.抗CEP抗体进而参与响应于外视网膜组织中CEP表位的持续产生而激活布鲁赫膜-RPE界面处的补体攻击途径。为了验证这一假设，我们用CEP加合的小鼠血清白蛋白免疫正常小鼠。我们的预测是，全身免疫与CEP会敏感的小鼠内源性CEP加合物的蛋白质在正常的老化过程中产生的外视网膜。反过来，免疫系统将通过攻击最容易产生CEP表位的细胞来做出反应。对这些小鼠的分析表明，RPE和光感受器中的局灶性病变模拟地图状萎缩，与干性AMD相关的盲性终末期萎缩。这种新的AMD小鼠模型将在正常小鼠中进一步表征，并且CEP免疫将扩展到补体途径分子及其调节剂中具有遗传缺陷的小鼠。小鼠中AMD的新模型将是用于预防或限制AMD进展的疗法的临床前测试的重要新资源。

DESCRIPTION (provided by applicant): 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 age-matched 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.