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C.K. M. Choy, I.F. F. Benzie, P. Cho, T. To; UV–mediated DNA damage in corneal epithelial cells assessed using the comet assay procedure . Invest. Ophthalmol. Vis. Sci. 2004;45(13):1452.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:To develop a comet assay (single cell gel electrophoresis) corneal epithelial cell model for evaluation of DNA damage in single cells, and to assess effect of ultraviolet (UV) irradiation on DNA damage in corneal epithelial cells. Methods: Baseline DNA damage of three types of porcine corneal epithelial cells – ‘superficial–layer’, ‘middle–layer’, and ‘deeper–layer’ cells – were assessed. ‘Deeper–layer’ cells were selected and exposed to ambient room light (control) or an energy–standardized dose (0.216 J/cm2) of UVA, UVB or UVC to determine the effect of UV irradiation. Results: Baseline DNA damage (%DNA in comet tail) in non–irradiated ‘superficial–layer’, ‘middle–layer’ and ‘deeper–layer’ corneal epithelial cells were (Mean±SD) 30.4±3.2%, 10.5±1.0% and 4.0±0.3%, respectively. Exposure to UVA, UVB and UVC caused significant (P<0.001) increases in DNA damage in ‘deeper–layered’ cells: mean±SD %DNA scores (10 gels per treatment, with 100 irradiated cells scored per gel) were 27.4±4.6% for UVB, 10.2±1.4% for UVA, and 14.7±1.8% for UVC (P<0.001), compared to 4.2±0.5% in the control cells. Conclusions:The comet assay can be used successfully with corneal epithelial cells, and will support future studies investigating environmental influences on ocular health and assessment of possible protective strategies. UVB radiation damages DNA in corneal epithelial cells, while UVC causes less damage, probably due to UVC absorption by ascorbic acid in the corneal cells. Results highlight the possible need for additional protection from UVB in high–risk individuals due to either high levels of solar radiation or corneal thinning due to photorefractive keratectomy.
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