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Nobuyo Yawata, Anshu ARUNDHATI, Yu-Chi Liu, Jay Jyh-Kuen Siak, Makoto Yawata, Jodhbir S Mehta; Human ocular surface immune profiling in situ in epidemic keratoconjunctivitis. Invest. Ophthalmol. Vis. Sci. 2016;57(12):5863.
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© ARVO (1962-2015); The Authors (2016-present)
Epidemic keratoconjunctivitis (EKC) is the most severe forms of virus-induced ocular surface inflammatory disease; it can impair vision for weeks to years due to subepithelial keratitis (SEK). EKC is highly contagious and affects millions worldwide. Group D human adenoviruses (HAdVs) are the main cause of EKC; however, the mechanisms causing severe inflammation at the ocular surface are unknown. We aimed to study key inflammatory factors of the severe inflammation observed in EKC using ocular surface immune profiling.
Brush cytology was used to collect samples from conjunctiva with acute follicular conjunctivitis over the course of virus infection. High-dimensional flow cytometry analysis was performed to analyse leukocyte subsets in the conjunctiva. The cytokines and chemokines present in tear fluid were analyzed using magnetic bead-based multiplex assays. HAdV infection was confirmed by PCR and HAdV types were determined by direct sequencing of the PCR products using genomic DNA extracted from tear fluid.
Leukocyte frequencies increased nine-fold during the acute phase of conjunctivitis, and HLA-DR+CD20– cells were present at significantly higher frequencies in the conjunctiva with severe inflammation. They consisted of CD14+CD16–, CD14–CD11c+, and CD14–CD11c– subsets, and the majority of the cells expressed CCR2. We found significantly elevated levels of CCL2, a ligand for CCR2, in tear fluid from cases with severe conjunctival inflammation; this corresponded to the increase of CCR2+cells in the conjunctiva. SEK was associated with severe conjunctival inflammation induced by group D HAdVs, where expression of IL-1β and IL-6 was significantly elevated. Increased M-CSF levels were observed in tear fluid with SEK cases.
The ocular surface cellular and molecular immune profiles in EKC suggest that local innate pro-inflammatory cytokines and chemokines, and monocyte/dendritic cell subsets subsequently recruited to the ocular surface play roles in the development of severe conjunctival inflammation and SEK during group D HAdV infection. The approach combining conjunctival brush cytology, high-dimensional flow cytometry, and multiplex bead assays of tear fluid is effective in enabling to understand human ocular surface immune profiles in human-specific ocular diseases.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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