Abstract
Purpose :
To deduce whether preconditioning of retinal pigment epithelial cells in culture (ARPE19) with long wave red light (625-635nm, 6 W/m2) can significantly blunt an insult of short wave visual blue light (465-475nm, 17.9 W/m2).
Methods :
Human ARPE19 cells were cultured in the dark in DMEM-F12 solution, supplemented with 2% antibiotic penicillin/streptomycin and 10% FBS and in a humidified atmosphere of 5% CO2, at 37oC. Semi-confluent cells in 96 or 48 well plates were either kept for 36 hours in the dark, kept in the dark for 12 hours and then exposed to blue light for 24 hours to blue light or initially exposed to red light for 12 hours, followed by 24 hours of blue light. Cell viability was then assessed by a MTT reduction assay. Mitochondrial status of cells were viewed using JC-1 dye. Dihydroethedium was used for the analysis of ROS. For immunocytochemistry, ARPE19 cell cultures were fixed in cold 4% paraformaldehyde and stained for the localisation of hemoxygenase-1 (HO-1) and NrF2 and NFκB. Cells in 48-well plates were scraped and subjected to western blot analysis for the detection of apoptosis inhibitory factor (AIF), HO-1 and actin.
Results :
Blue light caused loss of ARPE19 cell viability, stimulation of ROS activity and mitochondrial dysfunction as indicated through the use of JC-1. In addition blue light caused AIF to be activated resulting in the upregulation of a 57KDa protein band. Moreover, HO-1 was upregulated by western blot analysis and immunocytochemistry showed it to be translocated into the cytoplasm from mitochondria. Blue light also caused nuclear located Nrf2 to translocate into the cytoplasm. In contrast, blue light caused NFκB to translocate from the cytoplasm into the nucleus of cells. Preconditioning ARPE19 cells with red light as opposed to darkness attenuated all these influences of the blue light effects on the cells. In addition red light on its own seemed to enhance mitochondrial function as indicated by JC-1 staining.
Conclusions :
Evidence is provided that red light has the capacity to stimulated mitochondrial function and in the process upregulate a number of “protective” factors that makes the tissue more resistant to insults, such as blue light. The data also emphasis the potential negative influence of blue light on the retina and possible use of red light non-invasively to treat the retina where oxidative stress (as caused by blue light) participates in a disease process.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.