Abstract
Purpose :
Topical lubrication is the most common remedy for relieving the signs and symptoms of dry eye. Yet, the therapeutic value of lubricating the ocular surface remains unclear. Here, we explore the restorative properties of lubrication in a mouse model of autoimmune-mediated dry eye.
Methods :
Autoimmune regulator knock-out (Aire KO) female mice were topically treated with PBS 3X daily for 14 days. Age matched untreated Aire KO and wild type (WT) mice served as controls. Alterations in epithelial and stromal cell identities were determined via single nuclei (sn)RNA-seq of 10 corneas per group using 10x genomics pipeline. Cell subpopulations were identified using Seurat and signaling pathways identified using Cellchat and Gene Ontology (GO) analysis. The impact of lubrication on inflammation was assessed in Aire KOs treated with IL1R1 antagonist (IL1RA) 3X for 24 hrs. Changes in corneal epithelial integrity and identity, stromal composition and architecture, keratocyte identity, and altered signaling pathways were validated through a combination of in situ hybridization/immunofluorescence and high-resolution confocal imaging.
Results :
Topical lubrication had no impact on epithelial cell integrity, identity, or proliferation but significantly restored basement membrane components, laminin and perlecan, and collagen fiber alignment essential for maintaining corneal transparency. snRNAseq analyses revealed significant alterations in keratocytes, including an expansion of Col6a1+/Col1a1+ and Ccl8+/Mmp3+ keratocytes indicative of excessive collagen production and ECM remodeling. Strikingly, topical lubrication altered keratocyte cell states resulting in five unique clusters enriched in Thbs1, Ptn, and Igf1, known for their involvement in wound healing.
GO analysis of whole cornea global RNAseq data and snRNAseq stromal data indicated significant enrichment of pathways associated with MAPK activation and increased cellular response to IL-1 exclusively in the KOs, while negative regulation of MAPK was found in the lubricated cornea suggesting lubrication alone reduces MAPK activation in the corneal stroma via inhibition of IL-1 signaling.
Conclusions :
We show that topical lubrication in dry eye promotes regeneration of the stromal compartment, potentially shifting keratocytes into a reparative state and restoring ECM and stromal architecture by inhibiting IL-1/MAPK signaling.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.