CD4⁺ T cells drive corneal nerve damage but are dispensable for corneal epitheliopathy development in the context of dry eye

Abstract

Purpose : Whether corneal nerve changes in dry eye disease (DED) are a consequence of corneal epithelial damage or directly caused by pathogenic ocular surface CD4⁺ T cells is controversial. We tested the hypothesis that immune-driven corneal neuropathy in DED occurs independently of corneal epitheliopathy.

Methods : DED was induced by extraorbital lacrimal gland excision in wild-type (WT) and T cell-deficient (RAG1KO) mice. Corneal epithelial integrity and nerve function were measured on days 0, 5, and 10 by fluorescein uptake and mechanosensitivity, respectively. Corneal nerve and epithelial morphology were evaluated by whole-mount confocal microscopy while trigeminal ganglion gene expression was analyzed by bulk RNA-seq. Also, CD4⁺ T cells from DED or sham-surgery WT mice were adoptively transferred to RAG1KO mice.

Results : Compared to sham mice, WT and RAG1KO DED mice developed a similar reduction in tear secretion (-37% vs -38%, p=0.85) and conjunctival goblet cell density (-38% vs -47%, p=0.75) and a comparable increase in corneal fluorescein uptake (day 5: 252 vs 200%, p=0.19, day 10: 290% vs 266%, p=0.93). By contrast, corneal mechanosensitivity progressively dropped only in DED WT mice (day 5: -10 vs -0%, p<0.05; day 10: -21 vs +1%, p=0.01) and there was a larger DED-induced decrease in subapical (-54 vs -17%, p<0.001), mid-epithelial (-48 vs -19%, p=0.02), and subbasal (-36 vs -8%, p=0.01) corneal nerve density in WT than in RAG1KO mice. RNA-seq (false discovery rate 5%) of trigeminal ganglia of WT and RAG1KO mice identified 148 (1.6%) and 34 (0.3%) differentially expressed genes (% of total), on which further enrichment analysis found neurodegeneration- and neuropathic pain-associated pathways only in DED WT mice. Compared to sham WT CD4⁺ T cell transfer, DED WT CD4⁺ T cell transfer to naïve RAG1KO mice did not affect corneal epithelial barrier function and proliferation but significantly decreased corneal mechanosensitivity (-21%, p<0.01) and corneal nerve density (p<0.01) at the subapical (-40%), mid-epithelial (-42%), and subbasal (-32%) levels.

Conclusions : Our results indicate that in DED, CD4⁺ T cells drive corneal neurodegeneration but are not required for corneal epithelial damage to develop. This finding holds profound implications in DED therapy.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.

 

Corneal epithelial barrier function and nerve morphology

 

Bulk RNA-seq of trigeminal ganglia of both strains