Abstract
Purpose :
We aimed to investigate naïve murine cornea through single-cell RNA sequencing, identify populations of non-immune cells, and explore the expression of proinflammatory mediators in these cell groups.
Methods :
Single-cell gene expression libraries were prepared from adult 6–8-week-old naïve C57BL/6 mice (n=20 male and female each) using the Chromium Single Cell Gene Expression 3’v3.1 kit (10X Genomics). The gene expression matrix obtained was preprocessed and explored using Loupe Browser 7.0.1. Only non-immune cells were considered for analysis.
Results :
Totally, 5184 cells passed quality control, with 11 non-immune cell clusters identified by their expression of standard markers as stromal keratocytes-Mmp3hi (484, 9%) and Mmp3lo (272, 5%), stromal stem cells or SSCs (198, 4%), epithelial cells (1988, 38%), melanocytes (6, 0.1%), transit amplifying cells or TACs (107, 2%), endothelial progenitor cells or EPCs (522, 10%), vascular/lymphatic endothelial cells (32, 1%), Schwann cells (19, 0.3%), and limbal (691, 13%) and epithelial (788, 15%) stem cells. Of note, stromal keratocytes Mmp3hi and Mmp3lo expressed interleukin Il15, cyclooxygenase-2 (Ptgs2/Cox2), and prostaglandin E synthase (Ptges). SSCs expressed Il15, Il16 and Ptges. EPCs expressed pro-inflammatory calcium binding protein s100a9, tumor necrosis factor (Tnf), neurotoxin-1 (Lynx1), and interleukins (Il1α, Il18). TACs, limbal, and epithelial stem cells expressed Il18 and Lynx1. Activated leukocyte cell adhesion molecule or Alcam (Cd166) was expressed by many non-immune cell clusters. While melanocytes expressed Ptges, no expression of proinflammatory mediators was seen in mature epithelial cells. Schwann cells expressed nerve growth factor (Ngf), Alcam, and s100b, a pro-inflammatory calcium binding protein. Among these genes, Ngf and Lynx1 show both pro- and anti-inflammatory activity. No sex differences were observed in the gene expression patterns among cell clusters.
Conclusions :
Non-immune cells in the naïve cornea, including stromal cells, endothelial progenitor cells, transient amplifying cells, Schwann cells, limbal and epithelial stem cells express a host of pro-inflammatory mediators such as CD166, tumor necrosis factor, interleukins, prostaglandin E2, s100 proteins, neurotoxin-1, and nerve growth factor. Thus, non-immune cells in both sexes may play a significant role in inflammatory processes of the cornea.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.