Abstract
Purpose :
Macular degeneration (MD) is a leading cause of irreversible vision loss. The pathogenesis of MD is complex, involving many risk factors, including oxidative stress and a dysregulated complement system. Endoplasmic reticulum (ER) stress has been implicated in MD, contributing to inflammation and cellular dysfunction. This study used an experimental model to examine the effects of oxidative and ER stress on soluble and cell-bound complement proteins.
Methods :
Primary retinal pigment epithelium (RPE) cells were seeded on Millicell®-PCF 0.4-µm inserts for five days. Subsequently, porcine retinal explants were cultured alone and with RPE cells over time (24 - 72 hours). Cultures were exposed to oxidative and ER stress by adding 0.2 mM hydrogen peroxide (H2O2) and 1 µg/mL tunicamycin and cyclic light for eight hours daily. Supernatants were investigated for interleukin 6 (IL-6), interleukin 8 (IL-8), complement component C3, factor H (FH), and factor B (FB) by ELISA or Luminex®. Retinal degenerative injuries and cell-bound complement proteins were investigated by immunocyto- and histochemistry.
Results :
Primary RPE cells secreted IL-6, IL-8, regulatory protein FH, and complement components C3, FB. Oxidative and ER stress upregulated IL-6, IL-8, FH, while the secretion of C3 and FB decreased. The C3a receptor (C3aR) and C5aR showed expression in RPE cells alone in retinas and cocultures. C3aR expression increased in the nerve fiber layer and horizontal cells during culture. C5aR-expression increased in the synaptic and inner nuclear layers following oxidative and ER stress. Over time, in culture, the integrity of the outer plexiform layer (OPL) deteriorated, exhibiting gaps. Opsin and PSD-95 showed colocalization in the OPL in cultured specimens, indicating a potential misrouting of the opsin during the degenerative process. Immunolabeling with glial fibrillary acidic protein (GFAP) showed atrophic Müller cells throughout the cultured retinas.
Conclusions :
The study concludes oxidative and ER stress in RPE cells increased the secretion of inflammatory cytokines IL-6 and IL-8 and modulated the complement system by increasing soluble factor H and decreasing C3 and factor B levels. The experimental degenerative process increased the expression of complement receptors. These findings underscore the intricate relationship between stress responses and complement activation in retinal health and disease.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.