Evaluation of conjunctival immunological tolerance in a murine model of evaporative dry eye

GUZMAN M; SABBIONE F; GABELLONI ML; TREVANI AS; CASIRAGHI J; CHIARADIA P; GIORDANO M; GALLETTI J

Los Cocos, Córdoba

Congreso; LXI Reunión Anual de la Sociedad Argentina de Inmunología; 2013

Sociedad Argentina de Inmunología

Introduction: Irrespective of its cause, the dry eye state is characterized by an underlying immune dysfunction in the ocular surface that fuels disease progression. Though yet unexplored, mucosal tolerance (or lack thereof) could play a role in its pathophysiology and even constitute a therapeutic target. Objective: To assess conjunctival immunological tolerance in a murine model of dry eye. Study design: 8- to 12-week-old female Balb/c mice were intraperitoneally injected with saline (Ct) or 0.5 mg scopolamine (SCO) three times daily (9 AM, 1 PM and 5 PM) while exposed to a controlled air draft for 12 h (9 AM to 9 PM) for 5 days (3 mice per group, 3 independent experiments). Tear production was measured with phenol red-impregnated paper threads and expressed in mm. Ovalbumin (OVA) was instilled in both eyes (5 ul per eye, 2 mg/ml) at different time points in order to track the immune response to a harmless antigen. Mice were subcutaneously immunized with OVA in complete Freund?s adjuvant on day 7 and the systemic immune response was measured one week later as the in vitro OVA-induced splenocyte proliferation by [3H]-thymidine incorporation. Stimulation indexes (SI) were calculated as antigen-induced proliferation/basal proliferation. The outcome measurements were tear production and OVA-induced SI for each treatment group. Statistical significance was set at p < 0.05. Results: Relative to pretreatment levels, greater reduction in tear production was observed during treatment (days 1-5) in SCO mice than in Ct mice (52±4% vs 91±4%, p < 0.05). After treatment (days 6-10), tear production returned to pretreatment level in Ct mice (102±11%, p > 0.05) but not in SCO mice (79±14%, p < 0.05). Relative to untreated immunized mice, systemic OVA responses were significantly reduced both in Ct and SCO mice (47±10% and 53±3%, respectively, p < 0.05) if OVA was instilled starting on day 1, whereas when starting on day 4, Ct mice (65±9%, p < 0.05) but not SCO mice (110±20%, p > 0.05) showed a significant decrease in antigen response. Conclusion: SCO treatment for 5 days is sufficient to affect subsequent tear production. Ocular instillation of harmless antigen induces mucosal tolerance when started on the first day, perhaps because there is little ocular surface damage at this timepoint. But if delayed, antigen administration does not induce conjunctival tolerance in SCO mice, probably due to an already established proinflammatory setting at the ocular surface. These results altogether suggests that mucosal immune tolerance could play a role in dry eye pathophysiology.