Abstract
Purpose:
To investigate cytokines secreted from cultured human retinal pigment epithelial (RPE) cells under continuous fluorescent lamp illumination.
Methods:
Human RPE line cells (ARPE-19) were cultured to confluence. The cells were incubated in 0.21 mL/cm2 of colorless medium in the dark or under 2000 lx illumination from a daylight-colored (6500 K) fluorescent lamp mimicking ambient light for 24 hours. After the culture, the levels of 27 proinflammatory cytokines in culture supernatants were measured with a multiplex beads array system. To confirm whether the illumination induced oxidative stress, total levels of intracellular reactive oxygen species (ROS) were evaluated by a fluorescent molecular probe using the ROS/RNS Detection Kit. For detection of lipid peroxide and protein oxidization, immunostaining of malondialdehyde (MDA) and western blot of protein carbonyls were performed.
Results:
Fourteen out of 27 cytokines were detected both in the cultures in the dark and under illumination. The levels of IL-1ra, IL-9, IL-15, Il-17, basic FGF and G-CSF were significantly higher in the culture under illumination than in the culture in the dark. The levels of basic FGF increased 3.6-fold. Conversely, the levels of IL-8 and MCP-1 were significantly lower in the culture under illumination than in the culture in the dark. There were not significant differences in the levels of IL-6, IL-7, IL-10, Il-12, IP-10 and VEGF between the cultures in the dark and under illumination. The intensity of ROS signals increased in the culture under illumination compared to the culture in the dark. MDA-positive cells and protein carbonyls with diffuse bands over 240 kDa on the western blot were detected in the cultures under illumination.
Conclusions:
These findings suggest that human RPE cells secrete proinflammatory cytokines, and that photic stimulation which induces oxidative stress modifies production of some of these cytokines by human RPE cells.
Keywords: 412 age-related macular degeneration •
634 oxidation/oxidative or free radical damage •
694 retinal culture