Abstract
Purpose: :
To study the effects of mild hyperthermia preconditioning on the susceptibility of retinal pigment epithelial (RPE) cells to laser damage at 514 nm.
Methods: :
An existing artificially pigmented in vitro retinal model (Denton et al., 2008, J. Biomed. Opt., 13(5):054014) was used to identify protection or enhancement of laser-induced damage 18 hr after mild hyperthermia preconditioning (PC) treatment. Control and experimental RPE cells were exposed (16 replicates of the same irradiance) to a 0.93-mm diameter laser beam (514-nm line from an argon laser) for 0.25 s. Area of damage was assessed using a live-dead fluorescence indicator method 1-hr post exposure. The hyperthermia pretreatment consisted of a 20-min ramp up from 37 ºC to 42 ºC, an additional 20 min at 42 ºC, followed by a quick ramp down to 37 ºC. In addition to the PC of RPE cells after artificial pigmentation, RPE cells and melanosome particles (MPs) were PC separately and added together 1 hr later. Finally, PC cells without pigment were pigmented with normal MPs. For each exposure, corresponding fluorescence and IR images (high-speed high-magnification) were spatially overlaid with each other to identify thermal history of cells at the boundary of death.
Results: :
Combining the results of all 16 replicates, area of laser damage was summed and compared to control exposures. Cells that were PC after pigmentation had a 5.25 fold increase in damaged area. Cells PC before addition of adding normal MPs had no difference in laser-induced damage area. When cells and MPs were PC separately, but added later, they had markedly less damaged area after laser exposure. In all 4 sample types, the average temperature the cells at the boundary of death experienced was the same, indicating the integrated temperature-time product for cell death was unaltered.
Conclusions: :
The location of MPs during hyperthermia determines whether pigmented RPE cells were resistant or sensitized to subsequent laser damage at 514 nm. The PC appears to alter the photochemistry and/or the absorption properties of the MPs.
Keywords: laser • retinal pigment epithelium • stress response