Tear hyperosmolarity acts as a disruptor of ocular surface immune tolerance in the context of dry eye disease

Abstract

Purpose : Tear hyperosmolarity is commonly detected in dry eye disease (DED), but its exact role in disease progression is incompletely understood. On the other hand, breakdown of ocular surface tolerance (homeostasis) has been reported in DED models but the trigger remains unknown. The purpose of this work was to evaluate whether hyperosmolar stress could affect ocular surface homeostasis in mice.

Methods : 8-to-12 week-old Balb/c mice were instilled isoosmolar (0.3 Osm) or hyperosmolar (3 Osm) saline on both eyes 3 times daily for 5 days to induce hyperosmolar stress (HS). Ovalbumin (OVA) was also instilled as a surrogate antigen or OVA-pulsed dendritic cells (DCs) were subconjunctivally injected at different time points. Induced T cell responses were measured after subcutaneous immunization with OVA by the delayed-type hypersensitivity (DTH) assay. In addition, T cells in draining lymph nodes (day 5) were evaluated by flow cytometry.

Results : Compared to non-instilled immunized mice, OVA-instilled mice developed reduced DTH responses (i.e. were tolerized), as did their OVA+saline instilled cage mates (p<0.05). By contrast OVA+HS mice exhibited full DTH responses (i.e. were not tolerized). In the latter, T cells in draining lymph nodes showed increased expression of activation (CD69, CD25) and memory (CD44) markers. Local injection of immature dendritic cells (DCs) in HS mice, but not in saline-instilled mice, induced increased T cell proliferation in the draining lymph nodes. Conjunctivae from HS mice showed higher levels of epithelial NF-κB activation and increased numbers of DCs, and conversely, topical NF-κB inhibition prevented ocular surface tolerance disruption in HS mice. Consistently, supernatants from epithelial cells exposed to HS for 4 h, but not to control medium, had higher interleukin-1β and -6 levels and favored DC maturation and T cell proliferation in vitro.

Conclusions : HS is sufficient to abolish conjunctival immune tolerance to a harmless antigen in Balb/c mice, suggesting that HS by itself can skew the immune response at the ocular surface. The HS-induced proinflammatory mucosal environment favors local maturation of DCs and T cell responses in the draining lymph nodes. These results altogether suggest that hyperosmolar stress plays a role in dry eye initiation as an immune disruptor.

This is a 2020 ARVO Annual Meeting abstract.