Abstract
Purpose :
Current therapies for proliferative diabetic retinopathy (PDR) do not specifically target many of the VEGF-independent, cell-type specific processes that lead to vision loss. This study aims to identify targetable cell types, and corresponding signaling pathways, by elucidating the single cell landscape of resident cells in the vitreous of patients with PDR.
Methods :
Single cell RNA sequencing (scRNA-seq) was performed on vitreous cells obtained from collection of diluted cassette washings during vitrectomy (n=6 six eyes) and peripheral blood mononuclear cells (PBMCs) (n=5 adults). Droplet-based scRNA-seq was performed using the Chromium 10x platform to obtain single cell transcriptomes. Differences in tissue compartments were analyzed with gene ontology (GO) enrichment of differentially expressed genes and an unbiased ligand-receptor interaction analysis.
Results :
Transcriptomes from 13675 surgically harvested vitreous cells and 22636 PBMCs were included. Clustering revealed four cell states consistently across all batches with representative transcripts for T cells (CD2, CD3D, CD3E, GZMA), B cells (CD79A, IGHM, MS4A1 (CD20), HLA-DRA), myeloid cells (LYZ, CST3, AIF1, IFI30), and neutrophils (BASP1, CXCR2, S100A8, S100A9). Most vitreous cells were T cells (91.6%) compared to the peripheral blood (46.2%) while neutrophils were absent. The full repertoire of adaptive T cells including CD4+, CD8+ and T regulatory cells (Treg) and innate immune system effectors (i.e. Natural Killer T cells) were present in the vitreous. Pathway analysis also demonstrated activation of vitreous CD4+ and CD8+ memory T cells and ligand-receptor interactions unique to the vitreous.
Conclusions :
In the first single-cell transcriptomic characterization of the human vitreous in a disease state, we show the PDR vitreous is primarily composed of T cells, a critical component of adaptive immunity, with activity and proportions distinct from T cells within the peripheral blood, and neutrophils are essentially absent. These results demonstrate the feasibility of liquid vitreous biopsies via collection of diluted cassette washings during vitrectomy to allow mechanistic and therapeutic insights into human retinal and vitreous disease.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.