Age-related changes in the photoreactivity of retinal lipofuscin granules: role of chloroform-insoluble components.
OBJECTIVE
Lipofuscin accumulates in human retinal pigment epithelium (RPE) cells with age and may be the main factor responsible for the increasing susceptibility of RPE to photo-oxidation with age. As the composition, absorption, and fluorescence of lipofuscin undergo age-related changes, the purpose of this study was to determine whether photoreactivity of lipofuscin granules also changes with the donor age.


METHODS
To determine whether the photoreactivity of lipofuscin itself is age related, lipofuscin granules were isolated from human RPE and pooled into age groups. Photoreactivity was assessed by measuring action spectra of photo-induced oxygen uptake and photogeneration of reactive oxygen species. Separation of chloroform-soluble (ChS) and -insoluble (ChNS) components by Folch's extraction was used to determine the factors responsible for the age-related increase in lipofuscin photoreactivity.


RESULTS
The observed rates of photo-induced oxygen uptake and photo-induced accumulation of superoxide-derived spin adducts indicated that when normalized to equal numbers of lipofuscin granules, aerobic photoreactivity of lipofuscin increased with age. Both ChS and ChNS mediated photogeneration of singlet oxygen, superoxide radical anion, and photo-oxidation of added lipids and proteins. Although both ChS and ChNS exhibited substantial photoreactivities, neither exhibited significant age-related changes when normalized to equal dry mass. In contrast, ChNS contents in lipofuscin granules significantly increased with aging.


CONCLUSIONS
Aerobic photoreactivity of RPE lipofuscin substantially increases with aging. This effect may be ascribed to the increased content of insoluble components.
Rózanowska
Małgorzata
M
Department of Biophysics, Faculty of Biotechnology, Kraków, Poland. School of Optometry and Vision Sciences, Cardiff University, Cardiff, United Kingdom. rozanowskamb@cf.ac.uk
Pawlak
Anna
A
Rózanowski
Bartosz
B
Skumatz
Christine
C
Zareba
Mariusz
M
Boulton
Mike E
ME
Burke
Janice M
JM
Sarna
Tadeusz
T
Simon
John D
JD
D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000375:Aging / Q000502:physiology*; D002648:Child; D002675:Child, Preschool; D002725:Chloroform / Q000737:chemistry; D004578:Electron Spin Resonance Spectroscopy; D006801:Humans; D007223:Infant; D008062:Lipofuscin / Q000378:metabolism / Q000528:radiation effects*; D008875:Middle Aged; D018384:Oxidative Stress; D010100:Oxygen / Q000378:metabolism; D010857:Pigment Epithelium of Eye / Q000378:metabolism / Q000528:radiation effects*; D017382:Reactive Oxygen Species / Q000378:metabolism; D012995:Solubility; D019097:Spin Trapping; D013481:Superoxides / Q000378:metabolism; D014019:Tissue Donors; D014785:Vision, Ocular
D008062:Lipofuscin; D017382:Reactive Oxygen Species; D013481:Superoxides; D002725:Chloroform; D010100:Oxygen