Phototoxic Blue-Violet Wavelengths Determined by Tissue Culture Model of Aging RPE Cells Exposed to Sunlight-Normalized Irradiance Levels
Narrative Responses:
Purpose
While there is considerable evidence that blue light may damage the retina, its action spectrum has not been precisely defined. This study aimed to identify the specific hazardous wavelengths within the blue-violet spectrum (380 to 500 nm) using an in vitro model of retinal pigmented epithelium (RPE) cell aging.
Methods
Primary cultures of swine RPE cells accumulated A2E from serum-free media that contained 12.5, 20, or 40 μM concentrations of A2E. Levels of internalized A2E were quantified by ultra-performance liquid chromatography. RPE cells with A2E were exposed to 10-nm illumination bands across the range from 390 to 520 nm for 18 hours. The illumination bands were generated by a custom-made, computer-controlled LED-based illumination system, with irradiance levels normalized to mimic normal sunlight retinal exposure. To quantify phototoxicity, the ApoTox-Glo™ Triplex assay was performed 6 hours after exposure to assess cell viability, caspase-3/7 activation (to determine apoptosis), and necrosis.
Results
The greatest photodamage to the A2E-containing RPE cells, including morphological changes and decreased cell viability, followed exposure to the 10-nm illumination bands between 415 and 455 nm. A significant increase in caspase-3/7 activity (P < 0.001) in this blue range indicated apoptotic rather than necrotic cell death. Blue light phototoxicity increased as a function of the concentration of A2E used to load the RPE cells before illumination but was not correlated to irradiance levels. Blue light exposure did not induce RPE cell necrosis, regardless of A2E concentration. Illumination with 630-nm light had no effect on apoptosis or necrosis.
Conclusion
A2E confers susceptibility to blue light toxicity in RPE cells. The hazardous portion of the blue spectrum is the narrow band between 415 and 455 nm. This finding may prove critical in developing selective filters to provide protection from blue light hazards without disrupting the retina’s visual and nonvisual functions.