Oxidative stress induces mitochondrial dysfunction and autophagy in ARPE‐19 cells

Abstract

Purpose Oxidative stress plays an important role in the pathogenesis of age‐related macular degeneration (AMD), but the exact mechanism of the influence of the stress is not completely clear. The stress can affect both neural retina and retinal pigment epithelium (RPE) cells The purpose of our study was to evaluate changes in some mitochondrial properties and autophagy in the response of RPE cells to oxidative stress. Methods Retinal pigment epithelium ARPE‐19 cells were challenged with 200 mM H2O2 for 2 h. Mitochondrial ROS (mtROS) were assessed by the MitoSOX probe, mitochondrial membrane potential (MMP) with JC‐1, the activity of the complex I of the mitochondrial respiratory chain with MitoCheck. To assess autophagy cells were transfected with the pBABE‐puro mCherry‐EGFP‐LC3B plasmid and autophagic flux was evaluated with confocal fluorescent microscopy; expression of the LC3‐I and LC3‐II proteins were quantified by immunoblotting. Results Hydrogen peroxide induced an increase in mtROS and decrease in MMP in ARPE‐19 cells as well as it inhibited the activity of the complex I. Furthermore, an increase in the LC3‐II/LC3‐I ratio and an increased number of autophagolysosomes was observed in ARPE‐19 cells challenged with hydrogen peroxide. Conclusion Oxidative stress induces detrimental changes in mitochondria of RPE cells that can respond to these changes by increased autophagy, confirming that mitochondrial homeostasis and autophagy may be important in AMD pathogenesis. This work was supported by National Science Centre, Poland grant number 2017/27/B/NZ3/00872.

Anti-Inflammatory and Anti-Oxidative Synergistic Effect of Vitamin D and Nutritional Complex on Retinal Pigment Epithelial and Endothelial Cell Lines against Age-Related Macular Degeneration

“…Earlier studies have effectively described the vital role of oxidative stress by H 2 O 2 in inhibiting cell proliferation in various cells [ 62 ], including RPE in the retina [ 35 , 46 , 63 , 64 ]. The exact mechanisms have not been identified, however, recent studies have revealed the interference of H 2 O 2 in numerous intracellular signaling pathways in retinal epithelial cells, such as the epidermal growth factor receptor/AKT and Notch signaling pathways [ 46 , 64 ], including the regulation of autophagy [ 65 ], kinases, and pro-apoptotic proteins such as caspases [ 66 , 67 , 68 ]. These studies open the way towards future identification of the mechanisms involved in the interplay between vitamin D and retinal cells in restoring cell proliferation and protecting RPE from angiogenesis.…”

Protective Effects of a Lutein Ester Prodrug, Lutein Diglutaric Acid, against H2O2-Induced Oxidative Stress in Human Retinal Pigment Epithelial Cells

“…H 2 O 2 was chosen to induce oxidative stress in ARPE-19 cells because ARPE-19 cells have a high metabolic rate and exist in an environment that is abundant in endogenous ROS, such as O2-, H 2 O 2 and OH-. Long-term accumulation of oxidative damage leads to dysfunction of RPE cells and increases their susceptibility to oxidative stress [ 47 , 48 ]. H 2 O 2 is one of the most common oxidants used in the cellular oxidative stress models for ARPE-19 cells.…”

Potential of Telomerase in Age-Related Macular Degeneration—Involvement of Senescence, DNA Damage Response and Autophagy and a Key Role of PGC-1α

“…Under oxidative stress TERT can shuttle from the nucleus to mitochondria where it exerts a protective action on mitochondrial DNA (mtDNA) against various stresses [42,43]. These activities of telomerase are important in the context of AMD pathogenesis, in which oxidative stress and mitochondria play a crucial role [44].…”

“…In fact, several studies show a greater susceptibility of mtDNA than nDNA in AMD (e.g., [71][72][73][74]). Differences in the susceptibility between mtDNA and nDNA to DNA-damaging factors, especially of endogenous origin, in normal conditions are mainly determined by the differences in their metabolism and DDR mechanisms in mitochondria and the nucleus (reviewed in [44]). Moreover, methods used to measure DNA damage in these two organelles are usually different.…”