Long-term Use of Diclofenac Exacerbates Immune Driven Dry Eye Disease by Inhibiting Lipoxin A₄ formation in Lymph Nodes and Lacrimal Glands

Abstract

Purpose: Prostaglandins (PG) are lipid mediators (LM) with key roles in many physiologic and pathophysiologic processes; their roles in the eye are poorly defined. Inhibition of PG formation by the NSAID-diclofenac is an important treatment option for inflammatory diseases of the eye, including forms of Dry Eye Syndrome. Clinical data demonstrates that NSAIDs can reduce ocular discomfort and filamentary keratitis in dry eye patients. We have reported significant formation of PG in the eye and important immune regulatory functions of other LM such as LXA₄. Hence, we set out to investigate the role of PG and the impact of long-term diclofenac treatment in immune-driven Dry Eye Disease (DED).

Methods: Mice were treated with diclofenac systemically and topically to globally inhibit PG formation for 10 or 30 days prior to and throughout 10-day desiccating stress. Schirmer’s test, clinical fluorescence scoring and goblet cell density were used to assess dry eye disease severity. CD4⁺ T cells, as marker of T cell activation, were identified and quantified by fluorescence confocal microscopy and flow cytometry. Lipid mediator formation was quantified by LC/MS/MS-based lipidomics, and gene expression by QPCR.

Results: Diclofenac treatment in all groups significantly increased DED, which directly correlated with duration of NSAID treatment. 40-day diclofenac treatment increased CD4⁺ T cell by 86% in limbus, 34% in lacrimal glands and 66% in lymph nodes compared to controls and correlated with marked increased ocular surface defect and decreased goblet cell density. As expected, diclofenac treatment abrogated PGE₂ formation but unexpectedly also inhibited LXA₄ formation in all tissues. Diclofenac-treated mice were treated with LXA₄ systemically and topically, after inducing Dry Eye. LXA₄ treatment markedly increased tear production, decreased corneal defect and significantly reduced CD4⁺ T cells.

Conclusions: The findings provide evidence for a key role of PG in controlling immune-driven DED, which is consistent with recent reports that demonstrate PG regulation of B- and T-lymphocytes. In addition, results suggest that PG regulate the protective and immune regulatory LXA₄ circuit. NSAIDS are widely used systemically and topically, our findings raise questions about their potential side effects in Dry Eye Disease pathogenesis.