CX3CR1 deficiency affects microglial phagocytosis in rd10 retinal degeneration

Abstract

Purpose : Photoreceptor (PR) degeneration in retinitis pigmentosa (RP), while primarily caused by gene mutations in photoreceptors, can be influenced by non-cell autonomous contributions from activated retinal microglia. We investigated the role of CX3CR1, a microglia-specific receptor, in regulating microglial phagocytosis and pro-inflammatory factor production involved in PR degeneration.

Methods : CX3CR1-deficient (CX3CR1^GFP/GFP) and –sufficient (CX3CR1^+/GFP) transgenic mice were crossed into the rd10 background, a mouse model for RP. Microglial physiology and PR degeneration were followed and compared histologically across the time of rod degeneration (P15-P28) to identify the role of CX3CR1 signaling in modulating microglial neurotoxicity in RP. Microglial phagocytosis was monitored by in vitro assays and by ex vivo live-cell imaging. Protein levels of cytokines were measured using a Multiplex assay. Exogenous intravitreal injection of CX3CL1 ligand into rd10 mice to increase CX3CL1-CX3CR1 signaling was evaluated as a means to decrease microglial activation and PR degeneration.

Results : Microglial infiltration into the PR layer was significantly accelerated and increased in rd10;CX3CR1^GFP/GFP relative to rd10;CX3CR1^+/GFP mice, and was associated with increased PR apoptosis (as measured by TUNEL staining) and PR atrophy. CX3CR1 deficiency was associated with increased microglial phagocytosis, as evidenced by (1) increased phagosome number in vivo, (2) increased phagocytosis of fluorescent beads and PR cellular debris in vitro, and (3) increased PR phagocytosis behavior on live cell imaging in retinal explants. CX3CR1 deficiency was also associated with increased protein expression of inflammatory cytokines and increased microglial expression of CD68 and IL1β. Increased CX3CR1 signaling by exogenous intravitreal delivery of CX3CL1 into rd10 animals decreased microglial infiltration, microglial phagocytosis and activation, and photoreceptor atrophy.

Conclusions : CX3CR1 deficiency in retinal microglia increases and accelerates photoreceptor degeneration in a model of RP, likely by increasing microglial phagocytosis of viable photoreceptors. CX3CL1-CX3CR1 signaling in microglia can modulate microglial-mediated degeneration and represent a molecular target in potential therapeutic approaches to RP.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.