Activated microglia trigger inflammasome activation and lysosomal destabilization in human RPE cells

Activation of the innate immune system plays a major role in retinal degenerative diseases including age-related macular degeneration (AMD). In this study, we investigated whether reactive microglia trigger and sustain NLRP3 inflammasome activation in human retinal pigment epithelium (ARPE-19) cells. Specifically, we analyzed the potential of cell culture supernatants from lipopolysaccharide (LPS)-stimulated human microglia in combination with the lysosomal destabilization agent Leu-Leu-O-Me (LLOMe) to activate the inflammasome in ARPE-19 cells. We found disorganization of ARPE-19 cytoskeletal structure after incubation with conditioned media of LPS-stimulated microglia and LLOMe and accumulation of lipid deposits in these cells using Nile Red staining. LC3-II, the active form of the autophagy marker microtubule-associated protein 1 light chain 3 beta (LC3B), was also elevated in ARPE-19 cells after inducing inflammasome activation. Finally, a significant increase of transcripts for IL-6, IL-8, IL-1ß, GM-CSF and CCL-2 was detected in ARPE-19 stimulated with both microglia-conditioned medium and LLOMe. Our findings highlight a potential role of microglia in RPE inflammasome activation.