IMPACT OF LIPOFUSCIN IN RETINAL PIGMENT EPITHELIAL CELLS

脂褐质对视网膜色素上皮细胞的影响

• 批准号: 6086563
• 负责人: Janet Ruthe Sparrow
• 金额: $ 25.15万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-05-01 至 2003-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6086563
• 关键词: cellular pathology; clinical research; human subject; lipofuscin; macular degeneration; melanins; retinal pigment epithelium

DESCRIPTION (Verbatim from applicant's abstract): A prominent feature of aging and of some inherited retinal degenerations is the accumulation of autofluorescent granules of lipofuscin in retinal pigmented epithelial (RPE) cells. Circumstantial evidence exists for an association between age-related macular degeneration (AMD) and RPE lipofuscin; however, the impact of lipofuscin accumulation on the RPE cell has not been directly tested. Since one of the constituents of RPE lipofuscin, the fluorophore A2E, exhibits structural and photodynamic properties that could be detrimental to cells, it is hypothesized that the accumulation of this fluorophore in RPE cells plays a role in the pathogenesis of AMD. The aim of this work is to understand the mechanisms by which A2E forms in the RPE cell and to determine the impact of its accumulation on the RPE cell. By high performance liquid chromatography (HPLC) analysis, the quantities of A2E in RPE cells isolated from human eyes will be correlated with the age, race and gender of the donors. A2E in eyes from AMD donors will also be quantified. To develop a cell culture model of A2E-containing cells, cultured RPE lacking endogenous A2E will be presented with synthetic A2E in the culture media. Subsequently, the internalization of A2E by the cells will be characterized, as will the intracellular compartmentalization of A2E. To test the hypothesis that A2E, when present at critical concentrations, can have adverse effects on RPE cells, the propensity of intracellular A2E to (a) exhibit detergent-like activity, (b) damage cells as a photosensitizing agent and (c) disrupt lysosomal function, will be evaluated. It will also be determined whether A2E-mediated phototoxicity involves the generation of reactive oxidant species and an apoptotic form of RPE cell death. The ability of melanin pigment to protect against A2E-mediated phototoxicity will also be tested. Finally, in studies related to the biogenesis of A2E, we will obtain evidence for the formation of an intermediate compound (A2-PE) during A2E biosynthesis. We will also address the issue of whether A2-PE/A2E is formed in the photoreceptor outer segment membrane as opposed to the lysosomal compartment of the RPE cell. The long-term goal of these studies is to define conditions that accelerate the formation of A2E in vivo and to evaluate the role of A2E in the causation of AMD. If it can be demonstrated that the amassing of synthetic A2E by RPE cells is significant in terms of RPE cell function, treatments could be aimed at preventing its formation or destroying the formed molecule within the RPE cells.

描述（逐字病申请人的摘要）：衰老的突出特征 在某些继承的视网膜变性中是 视网膜色素上皮（RPE）中脂肪霉素的自荧光颗粒 细胞。存在与年龄相关之间关联的间接证据 黄斑变性（AMD）和RPE脂肪霉素；但是， 脂肪霉素在RPE细胞上的积累尚未直接测试。从一个 RPE脂肪霉素的成分（荧光团A2E）表现出结构性 以及可能对细胞有害的光动力特性，它是 假设该荧光团在RPE细胞中的积累有 在AMD的发病机理中的作用。这项工作的目的是了解 A2E在RPE单元中形成并确定的机制，以确定 它积聚在RPE电池上。高性能液相色谱法 （HPLC）分析，从人眼分离的RPE细胞中A2E的量 将与捐助者的年龄，种族和性别相关。 A2E在眼睛中 从AMD捐助者也将进行量化。开发一个细胞培养模型 含A2E的细胞，将介绍缺乏内源A2E的培养的RPE 与培养基中的合成A2E。随后，内在化 细胞的A2E将被表征，细胞内也会表征 A2E的隔室化。测试A2E的假设，当 临界浓度，可能对RPE细胞产生不利影响，倾向 细胞内A2E至（a）表现出清洁剂样活性，（b）损伤细胞 作为光敏剂，（c）中断溶酶体功能，将是 评估。还将确定A2E介导的光毒性是否 涉及反应性氧化剂物种和凋亡形式的产生 RPE细胞死亡。黑色素颜料预防A2E介导的能力 光毒性也将被测试。最后，在与 A2E的生物发生，我们将获得形成中间体的证据 A2E生物合成期间化合物（A2-PE）。我们还将解决 是否在光感受器外部段膜中形成A2-PE/A2E作为 与RPE细胞的溶酶体隔室相反。长期目标 这些研究是为了定义加速A2E形成的条件 体内并评估A2E在AMD因果关系中的作用。如果可以 证明RPE细胞积聚合成A2E在 RPE细胞功能的术语，治疗可能旨在防止其 形成或破坏RPE细胞内形成的分子。