Abstract
Purpose :
Age-related macular degeneration (AMD) is a degenerative disease of the macula, leading cause of blindness in the elderly population, and it is caused by an interplay of diverse risk factors (genetic predisposition, age and lifestyle habits). One of the main genetic risks is the Y402H polymorphism in the Complement Factor H gene (CFH/FH), an inhibitor of complement system activation. In our previous work, we showed that FH loss in RPE cells disturbs cell homeostasis. In this study, we investigate the impact of the AMD risk polymorphism Y402H on iPSC-RPE with focus on the endoplasmic reticulum (ER) stress response.
Methods :
Following differentiation of iPS cells into mature RPE cells, we investigated the phenotype of iPSC-RPE cells carrying CFH 402Y (low risk) or 402H (high risk) variants. RPE cells were treated with Hydroquinone (HQ) to induce oxidative damage, and Tunicamycin (TM) to directly induce ER stress. The ER stress response was assessed by protein expression analyses via Western Blot of ER stress sensor protein Binding immunoglobulin protein (Bip/GRP78) and unfolded protein response (UPR) effectors PKR-like ER kinase (PERK) and inositol-requiring transmembrane kinase/endoribonuclease 1α (IRE1α). Data are shown for n≥3 (mean ± SEM).
Results :
CFH 402Y and CFH 402H RPE cells did not show differences in the basal levels of ER stress sensor and UPR effector proteins Bip, PERK or IRE1α. CFH 402H RPE cells showed increased levels of Bip compared to CFH 402Y when exposed to oxidative stress condition in response to HQ treatment (1.2± 0.4 vs 4.2± 1.7, p=0.01). As a consequence, UPR effector IRE1α was also upregulated in CFH 402H RPE cells (0.95± 0.06 vs 1.6± 0.1, p<0.001). In response to TM treatment, ER stress sensor Bip was upregulated in CFH 402H RPE cells compared to CFH 402Y cells (7.2± 2.1 vs 12.2± 2.2, p=0.04).
Conclusions :
Our data support the hypothesis that FH exerts non-canonical functions in RPE cells and CFH 402H affects cellular homeostasis. Indeed, CFH 402H RPE cells are unable to properly respond to stress insult. Here, we demonstrated that increased ER stress may play a role in the pathology of RPE cells carrying the CFH Y402H polymorphism associated with AMD. These findings help to elucidate the function of FH in the RPE and AMD pathology.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.