QUANTITATIVE ANAYLSIS OF AGING RETINA

年龄相关性黄斑病变（ARM）是发达国家老年人无法治愈的视力丧失的主要原因，其治疗方法的改进在于更好地了解早期的ARM。基于我们的初步数据，我们假设早期黄斑病变是眼部动脉粥样硬化的一种形式。根据这一假说，ARM的病理生理涉及两种机制，类似于大动脉内壁衰老和动脉粥样硬化过程中发生的机制：根据这一假说，大动脉内壁：与细胞外基质蛋白相关的血清脂蛋白衍生的酯化胆固醇和未酯化胆固醇（EC和UC）的年龄相关沉积；以及在局部细胞的参与下，与疾病相关的富含uc的物质沉积。利用一种独特的人类供体眼睛资源和一种成熟的实验性动脉粥样硬化动物模型，我们提出了三个目标来验证这一假设的几个预测。1)在人眼中，我们将利用filipin荧光和数码显微镜测定Bruch膜EC和UC含量随年龄和视网膜位置的变化。我们将通过确定溶剂如何影响filipin荧光和超结构来识别布鲁赫膜中富含ec的颗粒。我们将使用酶分析和气相色谱-质谱法表征分离的布鲁赫膜的脂质组成。2)在人眼中，我们将确定发生在视网膜周围的膜状碎片的形态。我们将使用荧光、数码显微镜和超微结构立体学来测定膜碎片和光感受器外段的UC含量。3)在兔眼实验中，我们将利用放射自显影技术在组织切片中定位全身注射放射性标记的酪胺-纤维素-低密度脂蛋白，确定是否长期补充胆固醇会导致布鲁氏膜EC沉积。这些目标的结果对于评估ARM和动脉粥样硬化在细胞外胆固醇积累和来源方面的共同发病机制的程度是有价值的。为了确定是否应该考虑对动脉粥样硬化进行有效的治疗来治疗早期ARM，需要这些信息。

Improved treatments for age-related maculopathy (ARM), the leading cause of untreatable vision loss among the elderly in the developed world, lie in better understanding of early ARM. On the basis of our preliminary data, we hypothesize that early age-related maculopathy is an ocular form of atherosclerosis. According to this hypothesis, the pathophysiology of ARM involves two mechanisms similar to those that occur during aging and atherosclerosis in the inner wall of the large arteries: According to this hypothesis, the inner wall of the large arteries: an age-related deposition of serum-lipoprotein-derived esterified cholesterol and unesterified cholesterol (EC and UC) in association with extracellular matrix proteins; and a disease-related deposition of a UC-rich material with the participation of local cells. Using a unique resource for human donor eyes and a well-established animal model for experimental atherosclerosis, we propose three aims to test several predictions of this hypothesis. 1) In human eyes, we will determine how the EC and UC content of Bruch's membrane varies with age and retinal location, using filipin fluorescence and digital microscopy. We will identify EC-rich particles in Bruch's membrane by determining how solvents affect filipin fluorescence and ultra-structure. We will characterize the lipid composition of isolated Bruch's membrane using an enzymatic assay and gas-chromatography-mass spectrometry. 2) In human eyes, we will determine the morphology of membranous debris occurs in peripheral retina. We will determine the UC content of membranous debris and photoreceptor outer segments using filipin fluorescence, digital microscopy, and ultrastructural stereology. 3) In rabbit eyes, we will determine if an extended diet of cholesterol supplementation results in deposition of EC in Bruch's membrane EC, using autoradiography to localized systemically injected radiolabeled tyramine-cellubiose-low density lipoprotein in tissue sections. Results from these aims will be valuable in assessing the extent to which ARM and atherosclerosis share pathogenetic mechanisms with regard to the accumulation and source of extracellular cholesterol. This information is required in order to determine if effective treatments for atherosclerosis should be considered for the treatment of early ARM.