Photoprotection of chronic macular photochemical injury

老年性黄斑变性(AMD)与白内障、既往白内障手术、累积日光暴露和色素沉着的关系都支持这样的假设，即慢性光化学损伤会导致黄斑随着年龄和AMD的进展而发生变化。随着年龄的增长，脂褐素在视网膜色素上皮(RPE)中积累，并与灵长类视网膜中活性氧中间产物(ROI)的急性光敏作用共存。我们模拟了RPE和Bruch？S膜(BM)复合体中随年龄增长的潜在损害光产物的正常积累，以及到达黄斑的额外光谱过滤所引起的变化。脂褐素颗粒包含至少10种不同的荧光光化学产物，包括A2E(N-视黄醛-N-视黄乙醇胺)、其环氧化物和其他化学上尚未确定的与A2E相关的荧光团。这些荧光团的前体来自视紫红质漂白相关时期(即正常日光)视杆外段(ROS)视盘内全经视网膜的反应。虽然RPE溶酶体处理可以消化掉99%以上的ROS含量，但A2E和相关的荧光团没有被消化，而是集中在脂褐素颗粒中。到60岁时，正常人RPE细胞内A2E的平均浓度达到~400微米。然而，A2E在低得多的浓度下对细胞膜有毒性。我们推测，将A2E分离到脂褐素颗粒中并防止其重新分布到临界膜中是RPE健康所必需的。 我们建立了一个生物物理模型，使用瞳孔大小、晶状体透过率和视杆暗适应时间常数trh的正常值来确定平均视网膜光谱辐照度、全视网膜全视网膜稳态浓度、氧化损伤的全视网膜光敏化、ROS中形成A2E相关物种的全视网膜反应以及RPE脂褐素颗粒内的A2E光氧化作用随年龄和环境光强度的变化。我们的模型预测，作用光谱加权的黄斑短波辐照度每十年下降约三分之一，在前60年，RPE中A2E相关荧光团的产生速度几乎保持不变(此后显着下降)。最近在人的身体眼中也报道了与年龄相关的总脂褐素颗粒体积和每个RPE细胞的总荧光。由于有晶状体眼的脂褐素随年龄增加的速度慢于黄斑短波照度随年龄的下降速度，RPE中的ROI光敏化也应随着年龄的增加而下降。视网膜外部的光氧化应激可能是由于在RPE/BM复合体的关键膜中观察到少量的A2E相关荧光团造成的。然而，如果RPE/BM复合体是导致AMD进展的光氧化损伤的部位，那么在白内障摘除和人工晶状体(IOL)植入后，这种氧化损伤的程度和速率预计将显著增加(未观察到)。 因此，我们提出了一个新的假设，即RPE脂褐素产生的单线态氧允许积累的A2E发生化学变化，从而限制了RPE中A2E([A2E]ss)的稳定水平，限制了A2E在视网膜膜中的重新分布，并限制了A2E的化学毒性。在脂褐素颗粒内以光化学方式产生的单线态氧与其A2E反应生成A2E环氧化物，然后反应形成越来越复杂的交联分子。由于短波长黄斑辐照度随着年龄的增长而下降，A2E光氧化的速率下降了大约20倍，导致正常有晶状体眼的[A2E]ss增加，即使杆状漂白和A2E产生减少。我们的黄斑衰老理论模型再现了脂褐素和A2E的正常年龄依赖性，并提供了一个主要的细胞毒性机制，即一旦A2E在RPE细胞中达到阈值浓度，A2E重新分布到关键膜中会导致损伤，无论是否有额外的光激活。在我们的模型中，主要是随着年龄的增长，晶状体变黄，扭曲了年轻时发现的生产率和光氧化速率之间的原始光谱平衡，并允许[A2E]ss随着年龄的增长而上升。 我们正在评估非侵入性视网膜成像方法，这些方法可能允许临床验证我们对白内障手术后光化学变化的预测，以及我们对特定光谱光保护滤光片的好处的预测。我们提议的特定的光谱选择性？太阳镜？减少棒在明亮环境光下的激活和有毒光产物在RPE中的积累。此外，我们正在寻求通过荧光光谱区分不同的RPE光产物，以评估无创监测此类滤光片对患者分子影响的可能性。与NEI和俄罗斯医学科学院眼科研究所合作，我们正在设计临床研究，研究这种过滤器对白内障手术和人工晶状体植入术后早期和中度AMD进展的影响，以及对年轻时A2E和脂褐素增加的遗传易感性受试者的黄斑变化的影响。

The associations of age-related macular degeneration (AMD) with cataracts, prior cataract surgery, cumulative exposure to sunlight and pigmentation all support the hypothesis that chronic photochemical injury drives macular changes with age and AMD progression. Lipofuscin accumulates with age in the retinal pigment epithelium (RPE) and colocalizes with acute photosensitization of reactive oxygen intermediates (ROI) in the primate retina. We model the normal accumulation of potentially damaging photoproducts with age in the RPE and Bruch?s Membrane (BM) complex as well as changes induced by additional spectral filtering of light reaching the macula. Lipofuscin granules contain at least 10 different fluorescent photochemical products includng A2E (N-retinylidene-N-retinylethanolamine), its epoxides and other as yet chemically unidentified A2E-related fluorophores. The precursors of these fluorophores originate from reactions of all-trans-retinal within the rod outer segment (ROS) discs during periods associated with significant rhodopsin bleaching (i.e., normal daylight). Although RPE lysosomal processing enzymatically digests over 99% of the shed ROS contents, A2E and related fluorophores are not digested, but are concentrated into lipofuscin granules. By age 60 years, the average concentration of A2E within RPE cells reaches ~400 microM in normal eyes. However, A2E is toxic to cellular membranes at much lower concentrations. We hypothesize that segregation of A2E into lipofuscin granules and prevention of its redistribution into critical membranes is required for RPE health. We developed a biophysical model using normal values of pupil size, lens transmission, and rod dark adaptation time constant trh to determine average retinal spectral irradiance, steady-state concentration of all-trans-retinal, all-trans-retinal photosensitization of oxidative damage, all-trans-retinal reactions to form A2E-related species in the ROS, and A2E photo-oxidation within RPE lipofuscin granules as a function of age and ambient light intensity. Our model predicts a decline of about one third in the action spectra-weighted short-wavelength macular irradiance with each decade and a nearly constant production rate of A2E-related fluorophores in the RPE during the first 60 years (falling significantly thereafter). A similar age dependence of total lipofuscin granule volume and total fluorescence per RPE cell was reported recently in human cadaver eyes. Since the rates of lipofuscin increase with age are slower than the rate of decrease in short-wavelength macular irradiance in the phakic eye with age, ROI photosensitization in the RPE should also fall with increasing age. Photo-oxidative stress in the outer retina might arise from the smaller amounts of A2E-related fluorophores observed in critical membranes of the RPE/BM complex. However, if the RPE/BM complex were the site of photo-oxidative injury driving AMD progression, the magnitude and rate of this oxidative injury would be expected to increase dramatically (not observed) following cataract removal and intraocular lens (IOL) implantation. Consequently, we propose a novel hypothesis that singlet oxygen generation by RPE lipofuscin allows the chemical alteration of accumulating A2E, thereby limiting the steady-state levels of A2E ([A2E]ss) in the RPE, the redistribution of A2E into retinal membranes, and A2E chemical toxicity. Singlet oxygen generated photochemically within the lipofuscin granule reacts with its A2E to form A2E epoxides which then react to form increasingly complex cross-linked molecules. As short-wavelength macular irradiance falls with age, the rate of A2E photo-oxidation falls approximately up to 20-fold, causing [A2E]ss in the normal phakic eye to increase even as rod bleaching and A2E production decrease. Our theoretical model of macular aging reproduces the normal age dependence of lipofuscin and A2E and provides a primary cytotoxic mechanism in which, once A2E reaches a threshold concentration in the RPE cell, A2E redistribution into critical membranes causes damage with or without additional photo-activation. In our model, it is primarily the yellowing of the lens with age that distorts the original spectral balance between rate of production and rate of photo-oxidation found in youth, and allows the [A2E]ss to rise with age. We are evaluating noninvasive retinal imaging methods that might permit clinical validation of our predictions of photochemical changes following cataract surgery and our predictions of the benefits of specific spectral photo-protective filters. Our proposed specific spectrally selective ?sunglasses? reduce both rod activation in bright ambient light and the accumulation of toxic photoproducts in the RPE. In addition, we are seeking to distinguish different RPE photoproducts by their fluorescence spectra to evaluate the potential for noninvasive monitoring of the molecular effects of such filters in patients. In collaboration with the NEI and the Eye Institute of the Russian Academy of Medicine, we are designing clinical studies of the effects of such filters on progression of both early and moderate AMD following cataract surgery and IOL implantation and macular changes in subjects with genetic predisposition to generation of increased A2E and lipofuscin at younger ages.