Abstract
Purpose :
The blood-retinal barrier regulates immune cell entry into the retina and restricts potentially vision-threatening inflammatory responses. Understanding gene expression changes in cells that maintain the blood-retinal barrier during retinal inflammation (in particular, the retinal pigment epithelium and Müller glia) may help identify novel therapeutic targets.
Methods :
Using experimental autoimmune uveitis (EAU) in mice as a model for retinal inflammation, we performed transcriptomic analyses to compare gene expression profiles in individual retinal cell types between healthy and inflamed states. EAU was induced in 6-10 week old C57BL/6J mice by immunization with human IRBP peptide emulsified in complete Freund’s adjuvant. One healthy and two EAU retinae 21 days post-immunization were dissociated and single cell RNA-sequencing performed with 10X Chromium and Illumina NovaSeq. Naïve (n=3) and EAU (n=3) RPE were isolated from posterior eyecups with 0.25% trypsin and processed for bulk RNA-seq. Data analysis was performed in R using Seurat and DEseq2 packages.
Results :
Differential expression analysis revealed a significant shift in gene expression in Müller glia during retinal inflammation with marked enrichment of interferon-γ stimulated genes. This was associated with alterations in adhesion molecule expression, MHC class II upregulation, chemokine and cytokine expression profiles and immune checkpoint ligands. The RPE in inflamed state also displayed wide-ranging changes in gene expression, including similar upregulation of MHC-II and cytokine/chemokine profiles, and downregulation of several tight junction molecules. Infiltrating leukocytes captured in the EAU scRNA-seq dataset expressed receptors for several genes upregulated by blood-retinal barrier cells during active inflammation.
Conclusions :
Blood-retinal barrier cells undergo significant gene expression changes during autoimmune retinal inflammation, with complex pro- and anti-inflammatory gene expression programmes being implicated. Upregulation of genes associated with immune cell recruitment and activation (e.g. leukocyte adhesion molecules, co-stimulatory ligands and MHC-II) highlights extensive blood-retinal barrier-leukocyte cross-talk. Thus, targeted interference of blood-retinal barrier-leukocyte interactions represents an attractive strategy for local and specific therapeutic intervention in retinal inflammation and non-infectious uveitis.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.