Single Nucleus Multi-modal Analysis of the Aging Mouse Retina

Abstract

Purpose : To identify cell type-specific and age-related changes in gene expression and gene regulation in the mouse retina.

Methods : Retina and RPE tissue were isolated from 2.5 month-old (n=3) and 21 month-old (n=3) mice and analyzed via single-nucleus RNA-seq and single-nucleus ATAC-seq.

Results : Cluster analysis of the snRNA-seq data (~62,000 nuclei) revealed >30 clusters encompassing known retinal cell-types such as photoreceptors, horizontal cells, RPE, and RGCs. To identify differences in gene expression in the retina between young and old mice, we performed differential expression (2.5 months vs. 21 months) for each cell cluster. Overall, we identified several age-related gene expression patterns across retinal cells. To further investigate these changes, we compared age-related retinal genes to those that have previously been reported in the brain. Strikingly, 168 genes that were identified in retinal tissue were also reported in the brain. 67 of these genes followed the same trends of being up or down regulated with age in both the mouse retina and brain tissue, although there was variation in expression among cell types in both tissues. Among the 67 genes in common between the retina and brain, 10 are known to be associated with retinal disease. Notably, Ahi1, Son, Ubb, Cst3, and Gngt2 were all significantly up-regulated (p < 0.01, Wilcoxon Rank-Sum) with aging in the retina via snRNA-seq. These findings were further confirmed by qRT-PCR. Ahi1 was up-regulated in RGCs, (p=8.92E-09) and Gngt2 was up-regulated in cones (p=5.49E-94). Son and Cst3 were both up regulated in all cell types with age with p-values ranging from 1.32E-84- 0.015 and 3.12E-132-1, respectively. Ubb was up regulated in all cell types except for rods (P = 1.25E-75 to 1). Out of these 5 genes, only Cst3 did not show the same pattern of expression in the retina and brain.

Conclusions : In the retina, horizontal cells, RPE, and RGC cells showed most of the age-related differences. Several genes that matched the aging pattern in both mouse brain and retina are found to have a significant role in apoptosis and regulation of the apoptotic pathway. Analysis of additional genes/pathways and cell types as well as snATAC-seq data is in progress. This study provides key insight into genes that show altered regulation with age in retinal tissue and contribute to aging.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.