Abstract
Purpose :
Macular degeneration (MD) deteriorates the central macula, leading to irreversible significant vision loss. Risk factors such as dysregulated complement system and light-induced oxidative stress have been associated with MD, but the mechanism behind these events is still under investigation. Herein we used an experimental model for retinal degenerative diseases and investigated soluble and cell-bound complement proteins and the proangiogenic response in an oxidative stressed environment over time.
Methods :
ARPE-19 cells were cultured for one week (young) and six weeks (old) on Millicellâ-PCF 0.4-mm inserts. Young and old ARPE-19 cells were cultured over time (24 - 96 hours) with or without porcine retinal explants. Retinal explants were also cultured alone. Cultures were exposed to oxidative stress by adding 0.2mM hydrogen peroxide (H2O2) and cyclic light eight hours/day. Supernatants were taken daily during the culture period and investigated for complement component C3, factor H (FH), C5a, and vascular endothelial growth factor (VEGF) by ELISA or Luminexâ. Retinal degenerative injuries and cell-bound complement proteins were investigated by immunocyto- and histo-chemistry.
Results :
ARPE-19 cells showed secretion of C3, FH, C5a, and VEGF during the culturing period and, cells were found to secrete increased amounts of C3, FH, and VEGF over time. The in vitro degenerative process induced mislocalization of photoreceptor pigments into the entire photoreceptor cell body and synapse. Locally produced C5a decreased over time from cultured ARPE-19 cells, and the oxidative environment slightly increased the secretion. The C3a and C5a receptors (C3aR) and (C5aR) were expressed in ARPE-19 cells. Neither the aging process nor the oxidative stress changed the expression of C3aR, while C5aR increased over time in ARPE-19 cells. The retinal tissue showed expression of C3aR in inner retina. In contrast, C5aR expression was evident in the inner and outer plexiform layers.
Conclusions :
Oxidative stressed, and aged ARPE-19 secreted increased amounts of complement proteins and the proangiogenic factor VEGF, known to cause injuries in retinal degenerative diseases. Our research suggests that prolonged exposure to an oxidative environment induced locally produced complement components associated with increased VEGF.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.