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M. L. Denton, K. J. Schuster, M. S. Foltz, L. E. Estlack, R. J. Thomas; A Search for Preconditioning Laser Irradiances Capable of Producing a Protective Adaptive Response in RPE Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):2536.
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To search for photochemical preconditioning conditions (laser irradiances) in RPE cells that lead to protection against subsequent photochemical damage.
Pigmented hTERT-RPE1 cells were used to determine an ED50 irradiance threshold for damage by a photochemical laser exposure (0.3 mm, 100 s, 413 nm). Binary (yes/no) damage endpoints, based on a fluorescent dye assay (calcein-AM and ethidium homodimer1) one hr post-exposure, were used as input to the Probit computer program. This threshold value served as a reference point for damaging exposures (challenge) in preconditioning experiments. Sublethal preconditioning exposures (2.5 mm, 100s, 413 nm) were applied to RPE cells at predetermined time intervals prior to a suprathreshold challenge exposure (0.3 mm, 100 s, 413 nm). Two types of damage assessment were used to determine if preconditioning caused a protective adaptive response.
Threshold ED50 values for 0.3 mm and 2.5 mm laser exposures (100 s at 413 nm) were 2.3 fold different from each other. The use of a large beam for preconditioning and a small beam for challenging, along with meticulous alignment procedures, reduced uncertainty regarding challenge exposures occurring outside of preconditioned regions in culture dishes. Using both a Probit and a Fractional Damage Area approach to damage assessment, no measurable protective effect was found over a broad range of preconditioning (more than 4 orders of magnitude) and challenging (3 fold at, or above ED50) irradiances.
Preconditioning with photochemical laser exposure is either insufficient in producing a measurable protective response against photochemical damage in pigmented RPE cells, or the effective range of preconditioning laser irradiances is very narrow or extraordinarily low.
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