Inflammation triggers a potentially toxic iron sequestration response in the retina

Abstract

Purpose : Inflammation and retinal iron are believed to play a role in AMD pathogenesis. Pro-inflammatory cytokines can induce iron sequestration in the liver, heart, and brain, starving extracellular pathogens of the iron necessary for survival. We have evaluated the ability of several pro-inflammatory cytokines known to be upregulated in AMD to trigger the retinal cellular iron sequestration response (CISR).

Methods : Müller and RPE cell lines and primary cultures were treated with IL-1β, IL-1α, or TNF-α for 24 hours at 0.1-100ng/mL (n=3). Gene expression changes in the iron importers, Tfrc, Dmt1, and Zip14, and the iron exporter, Fpn, were analyzed by qRT-PCR. Statistical significance was determined by one-way ANOVA with Bonferroni correction. Mice expressing IL-6 in retinal astrocytes, driven by a GFAP-IL-6 transgene, were tested indirectly for retinal iron accumulation by immunolabeling for the iron-induced protein L-ferritin.

Results : Expression of the iron importers, Zip14 and Dmt1, was significantly elevated in Müller cells exposed to IL-1β or IL-1α and RPE cells exposed to IL-1β. Cytokine stimulation of Müller cells did not increase Tfrc, but IL-1β increased Tfrc in RPE cells. Fpn expression was significantly decreased in response to IL-1β, IL-1α and TNF-α in Müller cells. GFAP-IL-6 transgenic mice exhibited increased L-ferritin in the inner retina compared to age/strain matched controls.

Conclusions : Our data suggest that pro-inflammatory cytokines trigger changes in the iron transport transcriptome in vitro and increase intracellular iron burden in vivo, a response which we term the cellular iron sequestration response (CISR). IL-1β was a particularly potent activator of the CISR, triggering increased expression of iron importers (Tfrc, Dmt1 and Zip14) and decreased expression of the iron exporter, Fpn. IL-1β was sufficient to elevate expression of Tfrc in ARPE-19 cells, but failed to induce a similar change in M1 Müller cells suggesting differential regulation of the CISR among retinal cell types. Increased retinal L-ferritin in GFAP-IL-6 transgenic mice indicates that CISR occurs in vivo as well as in vitro and can be triggered by IL-6. This study suggests that a vicious cycle of inflammation and iron sequestration-induced oxidative damage may exacerbate retinal disease.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.

 

mRNA levels in Müller and RPE cells after 24h of 1ng/ml IL-1β, IL-1α, or TNF-α. * p<0.05. For Fpn, p<0.001 for all cytokines.