Abstract
Purpose : RPE cell malfunction plays an important role in AMD pathogenesis. Senescent RPE cells release an assortment of inflammatory, noxious molecules capable of directly or indirectly damaging the macula. We investigated the role of IL1 family cytokines and receptors in RPE senescence and malfunction.
Methods : Four groups of donated human eye sections (geographic atrophy (GA) AMD, neovascular AMD (nAMD), age-matched healthy control and young healthy control) were immunostained for IL1 family cytokines (IL1α, IL1β, IL33, IL36α, IL36β, and IL36γ) and receptors (IL1R1, ST2, IL1RAP, and IL36R) and expression levels in RPE cells were quantified. In vitro, ARPE-19 cells were treated with Doxorubicin (DOX) to induce senescence. The supernatant of the DOX-induced senescent RPE cells was then used to treat healthy ARPE-19 cells with or without blocking antibodies to IL1α/β (Anakinra, rh-IL1Ra), IL33 (anti-IL33R) or IL1RAP (anti-IL1RAP). ARPE-19 phagocytotic ability, cellular senescence, and senescence-associated secretory phenotype (SASP) were examined.
Results : RPE IL1R1 expression was reduced in age-matched healthy eyes compared to young healthy (p<0.05), GA (p<0.01), and nAMD eyes (p<0.01). IL1RAP and IL36R expression were reduced in nAMD eyes compared with age-matched healthy eyes (p<0.05 and p<0.01). RPE cells in GA eyes exhibited lower IL36R expression compared to age-matched healthy eyes (p<0.05). There was no significant change in the expression of IL1α, IL1β, ST2, IL36α, or IL36β in RPE cells between different groups. IL36γ was not detected in all eyes.
Conditioned media from DOX-treated senescent ARPE-19 cells induced senescence in healthy ARPE-19 cells, evidenced by increased β-gal staining (p<0.05). Disruption of IL1 signalling via Anakinra or neutralisation of IL1RAP or IL33R did not affect the percentage of β-gal+ cells, phagocytotic activity, or IL1 family cytokine release (p>0.05) in senescent RPE cells. CCL2, IL6, and IL8, but not IL1 family cytokines, were detected in senescent RPE cell supernatants.
Conclusions : IL1 family cytokine expression in RPE cells did not change during ageing or in AMD. IL1RAP and IL36R expression appears to be reduced in RPE cells in AMD eyes. IL1 family cytokines are not a component of senescent RPE-released SASP, and blocking IL1 family cytokine signalling does not protect healthy RPE cells from senescent cell-mediated damage.
Methods : Four groups of donated human eye sections (geographic atrophy (GA) AMD, neovascular AMD (nAMD), age-matched healthy control and young healthy control) were immunostained for IL1 family cytokines (IL1α, IL1β, IL33, IL36α, IL36β, and IL36γ) and receptors (IL1R1, ST2, IL1RAP, and IL36R) and expression levels in RPE cells were quantified. In vitro, ARPE-19 cells were treated with Doxorubicin (DOX) to induce senescence. The supernatant of the DOX-induced senescent RPE cells was then used to treat healthy ARPE-19 cells with or without blocking antibodies to IL1α/β (Anakinra, rh-IL1Ra), IL33 (anti-IL33R) or IL1RAP (anti-IL1RAP). ARPE-19 phagocytotic ability, cellular senescence, and senescence-associated secretory phenotype (SASP) were examined.
Results : RPE IL1R1 expression was reduced in age-matched healthy eyes compared to young healthy (p<0.05), GA (p<0.01), and nAMD eyes (p<0.01). IL1RAP and IL36R expression were reduced in nAMD eyes compared with age-matched healthy eyes (p<0.05 and p<0.01). RPE cells in GA eyes exhibited lower IL36R expression compared to age-matched healthy eyes (p<0.05). There was no significant change in the expression of IL1α, IL1β, ST2, IL36α, or IL36β in RPE cells between different groups. IL36γ was not detected in all eyes.
Conditioned media from DOX-treated senescent ARPE-19 cells induced senescence in healthy ARPE-19 cells, evidenced by increased β-gal staining (p<0.05). Disruption of IL1 signalling via Anakinra or neutralisation of IL1RAP or IL33R did not affect the percentage of β-gal+ cells, phagocytotic activity, or IL1 family cytokine release (p>0.05) in senescent RPE cells. CCL2, IL6, and IL8, but not IL1 family cytokines, were detected in senescent RPE cell supernatants.
Conclusions : IL1 family cytokine expression in RPE cells did not change during ageing or in AMD. IL1RAP and IL36R expression appears to be reduced in RPE cells in AMD eyes. IL1 family cytokines are not a component of senescent RPE-released SASP, and blocking IL1 family cytokine signalling does not protect healthy RPE cells from senescent cell-mediated damage.
| Original language | English |
|---|---|
| Article number | 771 |
| Journal | Investigative Opthalmology and Visual Science |
| Volume | 65 |
| Issue number | 7 |
| Publication status | Published - 01 Jun 2024 |
| Event | Association for Research in Vision and Ophthalmology Annual Meeting 2024 - Seattle, United States Duration: 05 May 2024 → 09 May 2024 |