The Retinal Environment Controls the Transcription Pattern of Resident and Recruited Mononuclear Cells

Abstract

Purpose : Previously, we reported that dendritic cells (DC) increase in the retina following sterile injury while the number of microglia (MG) remains constant. To further characterize the nature and function of DC and MG in the retina, we analyzed the transcription of neurotrophic factors and genes associated with immune mononuclear cell function in DC and MG isolated from naïve and injured retinas.

Methods : CDG mice, which expressed green fluorescent protein (GFP) and diphtheria toxin under control of the CD11c promoter, were used so that MG (CD45⁺CD11b⁺GFP^lo) could be distinguished from DC (CD45⁺CD11b⁺GFP^hi). CD45 levels were used to determine retinal mononuclear cells recently recruited from the circulation (CD45^hi) versus those derived from resident progenitors or had established residence in the retina (CD45^med). Retinal injury was induced by optic nerve crush (ONC) with analysis done ten days post-ONC. Retinal mononuclear cells were isolated by tissue digestion followed by fluorescence-activated cell sorting (FACS). Transcriptional analysis was done on mRNA from explanted retinas and on pools of defined, sorted retinal mononuclear cells by RT-qPCR. Single cells from defined mononuclear cell populations were analyzed by RT-PCR.

Results : Our previous studies suggested that the majority of DC in the retina after ONC were derived from circulating precursors cells. However, analysis of thirty-eight genes showed that MG (CD45^medGFP^lo) and DC (CD45^medGFP^hi) from ONC treated mice were very similar in their transcription of chemokine receptors, chemokine ligands, neurotrophic factors, and markers of DC development and function. MG and DC from naïve mice were also similar in their transcription pattern to each other and to those from ONC mice. Significantly different expression in CD45^med populations was observed only with CCR2, CCL2, CD135 and GDNF. Analysis of CD45^hi cells showed more transcriptional differences compared to CD45^med cells.

Conclusions : The results suggest that the retinal environment imparts significant control upon the phenotype and function of mononuclear cells. Although retinal injury or stimulation is efficient at recruiting immune mononuclear cells from the circulation, continued residency and function within the retina requires that mononuclear cells adopt the phenotype and function imposed by the retinal environment.

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