Deletion of Socs3 in LysM+ Cells and Cx3cr1 Resulted in Age-dependent Development of Retinal Microgliopathy

Abstract BackgroundWe generated a mouse model of primary microglial dysfunction by deleting two negative immune regulatory genes, Cx3cr1 and Socs3 (in LysM+ cells). This study aimed to understand how primary microglial dysfunction impacts retinal neurons during aging.MethodsThe LysMCre-Socs3fl/flCx3cr1gfp/gfp double knockout (DKO), LysMCre-Socs3fl/fl, Cx3cr1gfp/gfp and Socs3fl/fl mice were maintained up to 12 months. Eyes were collected and processed for immunohistochemistry of IBA-1, cone arrestin, secretagogin, PKCα and GABA. Brain microglia from DKO and WT mice were stimulated with LPS + IFNγ or IL4. The expression of TNFα, IL1β, IL6, iNOS, IL12p40, IL23p19, CCL2, CCL5, CXCL2, IL10, CD206 and Arg1 were examined by real-time RT-PCR and protein production was measured by Luminex assay. Retinal explants from C57BL/6J mice were co-cultured with microglia from DKO or WT mice for 24 h, after which the number of cone arrestin+ cells in retinal flatmount were quantified.ResultsIn 3–5 month old mice, the number of microglia in retinal ganglion cell layer (GCL) and inner plexiform layer (IPL) were comparable in all strains. The DKO mice had a significantly higher number of microglia in the outer plexiform layer (OPL) but significantly lower numbers of cone arrestin+, secretagogin+ and GABA+ cells compared to Socs3fl/fl and single KO mice. During aging, 57% of the DKO mice died before 12 months old. The 10–12 months old DKO mice had significantly highers number of microglia in GCL/IPL and OPL than age-matched Socs3fl/fl and single KO mice. The aged DKO mice developed retinal pigment epithelial (RPE) dysmorphology accompanied by subretinal microglial accumulation. The number of photoreceptors, bipolar cells (Secretagogin+ or PKCα+) and GABA+ amacrine cells was significantly lower in aged DKO mice compared to age-matched Socs3fl/fl and single KO mice. Microglia from DKO mice showed significantly higher levels of phagocytic activity and produced higher levels of TNFα, IL6, CCL2, CCL5, CXCL2 and CXCL10 compared to microglia from Socs3fl/fl mice. Co-culture of retinal explants with DKO microglia (pre-treated with LPS + IFNγ or IL4) significantly reduced cone photoreceptor survival.ConclusionsThe LysMCre-Socs3fl/flCx3cr1gfp/gfp DKO mice displayed primary microglial dysfunction and developed age-related retinal microgliopathy characterized by aggragated microglial activation and multiple retinal neuronal and RPE degeneration.Trial registration: Not applicable. The article does not contain any results from human participants.