Abstract
Purpose :
Thyroid hormone (TH) plays an essential role in cell proliferation, differentiation, and metabolism. Recent studies have shown the possible association of TH signaling with age-related macular degeneration. Excessive TH signaling induces cone degeneration while suppression of TH signaling protects cones in mouse models of retinal degeneration. This work investigates the genes/transcriptional networks that might be involved in TH-induced cone degeneration in mice using single-cell RNA sequencing (scRNAseq) analysis.
Methods :
One-month-old C57BL/6 mice received triiodothyronine (T3, 20 µg/ml in drinking water) for 30 days. At the end of the experiment, retinal cells were dissociated using a neural tissue dissociation kit, and cell viability was analyzed before being subjected to 10x Genomics’ scRNAseq. The resulting data were analyzed using the Seurat package and visualized by the Loupe browser. Statistically significant changes in expression of genes in the cone cluster were then submitted to Ingenuity Pathway Analysis (IPA).
Results :
Four individual retinas of untreated and T3-treated mice from both males and females were sequenced and analyzed. The cell viability was in the range of 42-70% after dissociation. Among more than 155,000 single cells, we identified 17 clusters of retinal cells using selected marker genes. We identified 652 genes that statistically significantly changed in cones after T3 treatment. Through IPA analysis, we identified several critical canonical pathways, including phototransduction and oxidative phosphorylation, that were downregulated after T3 treatment. We also detected increased regulation pathways that are involved in sensory neuronal/photoreceptor degeneration and neuronal degeneration, accompanied by inhibition of the upstream regulators Crx (Cone-Rod Homeobox), Nrl (Neural Retina Leucine Zipper), Ppargc1a (PARG Coactivator 1 α), and Rabgef1 (Rab Guanine Nucleotide Exchange Factor 1).
Conclusions :
Our scRNAseq analysis shows that the genes involved in phototransduction and oxidative phosphorylation are inhibited in the mouse cones after T3 treatment, which is concomitant with the downregulation of developmental regulators and the upregulation of the pathways involved in photoreceptor/neuronal degeneration. The data support a broad impact of excessive TH signaling on the function, cellular homeostasis, and cell survival of cones.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.