Abstract
Purpose :
Variants in genes involved in inflammation and lipid metabolism are known to modulate the risk of age-related macular degeneration (AMD). We aimed to gain insights into their roles in AMD pathogenesis through characterisation of expression profiles in individual retinal cell types by single cell RNA sequencing (scRNA-seq).
Methods :
Wildtype C57BL/6J mouse retina was dissociated and submitted for scRNA-seq. Single cell cDNA libraries were generated with the 10x Chromium platform and sequenced with Illumina NovaSeq. RPE was isolated from posterior eyecups with 0.25% trypsin and submitted for bulk RNA-seq. Data were analysed using Seurat v4 in R.
Results :
Sixteen transcriptionally distinct retinal cell populations were identified on UMAP clustering of scRNA-seq data: rods, cones, microglia, Müller glia, 2 clusters of rod bipolar cells, 2 clusters of amacrine cells, and 8 subtypes of cone bipolar cells. When transcript levels were normalised and compared with reference to a house-keeping gene (Gapdh), complement factor H (Cfh) was expressed significantly by the RPE and microglia. Complement C3 was expressed minimally by RPE while no significant expression of C5 was detected by any retinal cell type. In terms of lipid metabolism genes, apolipoprotein E (Apoe) was abundantly expressed by Müller glia and RPE and, to a lesser degree, by microglia. The cholesterol transporter (Abca1) was minimally expressed by the RPE, while hepatic lipase (Lipc) was not expressed in the retina. Finally, serine protease (Htra1) was expressed in RPE, Müller glia and amacrine cells.
Conclusions :
Single cell retinal transcriptomic profiling implicates specific cell populations in AMD pathogenesis. The results are consistent with ocular complement factors being predominantly blood-derived with selective regulatory components (e.g. Cfh) being produced locally by the RPE and microglia. Apoe stands out as a locally produced lipid transport protein, consistent with its abundance within soft drusen and subretinal drusenoid deposits. Overall, the RPE, Müller glia and microglia appear to be the main modulators of AMD risk.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.