Abstract
Retinal lipofuscin accumulates with age in the retinal pigment epithelium (RPE), where its fluorescence properties are used to assess retinal health. It was observed that there is a decrease in lipofuscin fluorescence above the age of 75 years and in the early stages of age-related macular degeneration (AMD). The purpose of this study was to investigate the response of lipofuscin isolated from human RPE and lipofuscin-laden cells to visible light, and to determine whether an abundant component of lipofuscin, docosahexaenoate (DHA), can contribute to lipofuscin fluorescence upon oxidation. Exposure of lipofuscin to visible light leads to a decrease in its long-wavelength fluorescence at about 610 nm, with a concomitant increase in the short-wavelength fluorescence. The emission spectrum of photodegraded lipofuscin exhibits similarity with that of oxidized DHA. Exposure of lipofuscin-laden cells to light leads to a loss of lipofuscin granules from cells, while retaining cell viability. The spectral changes in fluorescence in lipofuscin-laden cells resemble those seen during photodegradation of isolated lipofuscin. Our results demonstrate that fluorescence emission spectra, together with quantitation of the intensity of long-wavelength fluorescence, can serve as a marker useful for lipofuscin quantification and for monitoring its oxidation, and hence useful for screening the retina for increased oxidative damage and early AMD-related changes.
Highlights
Retinal pigment epithelium (RPE) is a monolayer of neuroepithelial cells separating the retina from its choroidal blood supply
RPE cells gradually accumulate a product of incomplete lysosomal digestion of phagocytosed photoreceptor outer segments with age, as well as some autophagocytosed material known as age pigment or lipofuscin [1,2,3]
We have shown that oxidation of DHA leads to the formation of products absorbing visible light with more potent photosensitizing properties than the lipophilic extract of lipofuscin, suggesting that even if they contribute only a small percentage to the light absorption of the lipofuscin, they could still be responsible for the majority of its photosensitizing properties [60]
Summary
Retinal pigment epithelium (RPE) is a monolayer of neuroepithelial cells separating the retina from its choroidal blood supply. RPE cells gradually accumulate a product of incomplete lysosomal digestion of phagocytosed photoreceptor outer segments with age, as well as some autophagocytosed material known as age pigment or lipofuscin [1,2,3]. RPE lipofuscin emits a characteristic golden-yellow fluorescence with a broad maximum at about 600 nm and large variations between individual granules (the maxima vary from 550 to 645 nm) [3,10,11,12,13]. It has been determined that, when excited with blue light, the fluorescence is emitted mostly by yellow-emitting fluorophores [12]. A considerable part of that emission originates not from a direct
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