Photoreactivity of aged human RPE melanosomes: a comparison with lipofuscin.
OBJECTIVE
To determine whether aging is accompanied by changes in aerobic photoreactivity of retinal pigment epithelial (RPE) melanosomes isolated from human donors of different ages, and to compare the photoreactivity of aged melanosomes with that of RPE lipofuscin.


METHODS
Human RPE pigment granules were isolated from RPE cells pooled into groups according to the age of the donors. Photoreactivity was determined by blue-light-induced oxygen uptake and photogeneration of reactive oxygen species. Short-lived radical intermediates were detected by spin-trapping, hydrogen peroxide by an oxidase electrode, singlet oxygen by cholesterol assay, and lipid hydroperoxides by iodometric assay.


RESULTS
Blue-light photoexcitation of melanosomes resulted in age-related increases in both oxygen uptake and the accumulation of superoxide anion spin adducts. The efficiencies of these processes, however, were still significantly lower than that induced by photoexcited lipofuscin. During irradiation of melanosomes, a substantial amount of oxygen was converted into hydrogen peroxide, whereas for lipofuscin, hydrogen peroxide accounted for not more than 3% of oxygen consumed. In contrast to lipofuscin, photoexcited melanosomes did not substantially increase the rate of oxidative reactions in the presence of polyunsaturated lipids or albumin. However, oxygen uptake was significantly elevated in the presence of ascorbate. Thus, the rate of photo-induced oxygen uptake in samples containing both ascorbate and melanosomes approached that observed in lipofuscin samples.


CONCLUSIONS
Blue-light-induced photoreactivity of melanosomes increases with age, perhaps providing a source of reactive oxygen species and leading to depletion of vital cellular reductants, which, together with lipofuscin, may contribute to cellular dysfunction.
Rózanowska
Małgorzata
M
Department of Biophysics, Institute of Molecular Biology, Jagiellonian University, Kraków, Poland. rozanowskamb@cf.ac.uk
Korytowski
Witold
W
Rózanowski
Bartosz
B
Skumatz
Christine
C
Boulton
Mike E
ME
Burke
Janice M
JM
Sarna
Tadeusz
T
D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000375:Aging / Q000502:physiology*; D001205:Ascorbic Acid / Q000494:pharmacology; D002648:Child; D002675:Child, Preschool; D004578:Electron Spin Resonance Spectroscopy; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D007223:Infant; D008027:Light; D015227:Lipid Peroxidation; D008054:Lipid Peroxides / Q000378:metabolism; D008062:Lipofuscin / Q000378:metabolism / Q000528:radiation effects*; D020460:Melanosomes / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects*; D008875:Middle Aged; D009243:NAD / Q000494:pharmacology; D010101:Oxygen Consumption; D010857:Pigment Epithelium of Eye / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects*; D017382:Reactive Oxygen Species / Q000378:metabolism
D008054:Lipid Peroxides; D008062:Lipofuscin; D017382:Reactive Oxygen Species; D009243:NAD; D006861:Hydrogen Peroxide; D001205:Ascorbic Acid