Mitochondrial damage and clearance in retinal pigment epithelial cells

Age-related macular degeneration (AMD) is a severe eye condition causing permanent vision loss in the central retina, the macula, significantly impacting patients' quality of life and mortality risk. Despite its severity, effective treatments for dry AMD are lacking. Contributing factors to the disease include genetic predisposition, chronic oxidative stress, mitochondrial damage, protein aggregation, and inflammation. To advance understanding and treatment, our laboratory developed a mouse model lacking peroxisome proliferator-activated receptor gamma coactivator 1-alpha and nuclear factor erythroid 2-related factor 2 (PGC1α/NFE2L2), mimicking dry AMD features such as increased oxidative stress markers, damaged mitochondria, lysosomal lipofuscin accumulation and drusen-like structures in retinal pigment epithelial cells (RPE). Additionally, we established a human RPE cell-based model to study dysfunctional intracellular clearance systems' impact on inflammasome activation. In our investigation, we observed a disturbance in the autolysosomal system responsible for clearing damaged mitochondria in RPE cells of 1-year-old PGC1α/NFE2L2-deficient mice. Confocal immunohistochemical analysis revealed elevated levels of autophagosome marker LC3B and mitophagy markers like PINK1 and PARKIN, indicating mitochondrial damage. However, we did not detect increased autolysosome formation or colocalization of lysosomal and mitochondrial markers. We noted an upregulation of late autolysosomal fusion protein Rab7 and autofluorescent aggregates. Additionally, Toll-like receptors 3 and 9 were increased, while NOD-like receptor 3 levels were decreased in PGC1α/NFE2L2 dKO retinas compared to wild type. There was a trend towards elevated complement component C5a and increased thrombin activity, independent of C3. Furthermore, levels of acute phase protein C-reactive protein and receptor for advanced glycation end products were increased in PGC1α/NFE2L2 dKO retinas. Selective proteasome inhibition with epoxomicin induced NADPH oxidase and mitochondrial-mediated oxidative stress, releasing mitochondrial DNA into the cytosol, activating AIM2 inflammasome, and secreting interleukin-1β in ARPE-19 cells. In summary, our findings suggest a relative decrease in mitophagy, increased C5 and thrombin levels and decreased C3 levels in this dry AMD-like model. Furthermore, selective proteasome inhibition induced mitochondrial damage and AIM2 inflammasome activation in ARPE-19 cells.