Abstract
Purpose:
Age related macular degeneration (AMD) is the most common form of central retinal blindness globally. Distinct processes of the innate immune system, specifically activation of the NLRP3 inflammasome, have been shown to play a central role in the development of both dry and neovascular (wet) forms of the disease. Autophagy is a homeostatic cellular process that is involved in protein and organelle degradation via the lysosomal pathway, and has been linked with the pathogenesis of AMD in recent times. Autophagy has previously been shown to control IL-1β and IL-18 secretion by targeting the inactive precursor forms of these proteins for degradation in macrophages. As NLRP3 and IL-18 signalling have previously been implicated in RPE cell death associated with dry AMD, we wished to determine the role of autophagy in regulating NLRP3 inflammasome component mediated RPE cell death.
Methods:
We generated adeno-associated-virus (AAV) vectors expressing pro-IL18 cDNA in order to assess the effect of over-expression of this component in retinal pigment epithelial (RPE) cells. Using both in vitro and in vivo techniques, we determined the involvement of autophagy in the increased cell volume observed following over-expression of NLRP3 inflammasome components.
Results:
We observed an increase in the levels of both LC3-I and LC3-II (the cleavage product of LC3-I) in ARPE-19 cells over-expressing pro-IL-18 or pro-IL-1beta. Furthermore, when autophagy was promoted by administration of rapamycin, the cell swelling phenotype induced by over-expression of pro-IL-18 or pro-IL-1beta was inhibited, indicating that autophagy can regulate the pathological RPE cell volume, likely due to the sequestration and degradation of the abundant precursor cytokines. Intriguingly, pro-IL-18 and pro-IL-1beta mediated RPE cell swelling was independent of NLRP3 inflammasome oligomerisation, suggesting that dysregulated autophagic processes may be the the initiator of RPE cell death associated with AMD.
Conclusions:
The mechanisms underlying the increased levels of pro-IL-18 and pro-IL-1beta inducing RPE cell swelling remain unknown; however, it appears that autophagy, as opposed to NLRP3 inflammasome oligomerisation, is a key regulator of the phenotype. A full and comprehensive elucidation of the role played by autophagy in the development of dry AMD may yield novel therapeutic targets for early stage intervention of the condition.
Keywords: 557 inflammation •
695 retinal degenerations: cell biology •
688 retina