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J. Zhou, B. Cai, J.R. Sparrow; DNA Damage in A2E-laden RPE Illuminated with Blue Light . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1700.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: When RPE cells laden with A2E, a lipofuscin fluorophore, are irradiated with blue light, photochemical events are initiated that provoke cell death. We sought to determine whether DNA was a target of the cellular damage. Methods: ARPE-19 cells accumulated A2E before 430 nm illumination. DNA damage was assayed by alkaline comet assay, with and without the addition of the repair enzymes formamidopyrimidine N-glycosylase (Fpg), endonuclease III (endo III) and T4-endonuclease V (T4-Endo V) to characterize DNA lesions. Damage was quantified as tail moment. The base lesion 8-oxo-deoxyguanosine (8-oxo-dG) was detected by immunoperoxidase and histochemical methods.The effects of the singlet oxygen quencher sodium azide was tested and cell viability was quantified. Results: DNA damage was induced in A2E-containing RPE exposed to 430 nm illumination. The extent of damage, measured as tail moment, was proportional to exposure duration and was reduced by pre-incubation with sodium azide. The detection of FPG- and endo III-sensitive DNA lesions revealed the presence of oxidized purine and pyrimidine bases while labeling with specific antibody and binding of fluorescein-labeled avidin indicated that guanine bases were oxidatively modified to 8-oxo-dG. The ability of the cells to repair the DNA damage declined as the severity was increased and kinetic studies disclosed rapid and slow stages of repair. Conclusions: DNA is one of the cellular constitutents that can be damaged by the interaction of A2E and blue light. At least some of the DNA lesions are oxidative base modifications.
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