May 2003
Volume 44, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2003
DNA Damage in A2E-laden RPE Illuminated with Blue Light
Author Affiliations & Notes
  • J. Zhou
    Ophthalmology, Columbia University, New York, NY, United States
  • B. Cai
    Ophthalmology, Columbia University, New York, NY, United States
  • J.R. Sparrow
    Ophthalmology, Columbia University, New York, NY, United States
  • Footnotes
    Commercial Relationships  J. Zhou, None; B. Cai, None; J.R. Sparrow, None.
  • Footnotes
    Support  NIH EY12951, American Health Assistance Foundation, Research to Prevent Blindness
Investigative Ophthalmology & Visual Science May 2003, Vol.44, 1700. doi:
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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)

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Abstract

Abstract: : 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.

Keywords: retinal pigment epithelium • age-related macular degeneration • cell death/apoptosis 
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