The NLRP3 Inflammasome Is Required for Protection Against Pseudomonas Keratitis

PURPOSE: The current study was designed to examine the role of the NLRP3 inflammasome pathway in the clearance of Pseudomonas aeruginosa (PA) infection in mouse corneas. METHODS: Corneas of wild type and NLRP3(−/−) mice were infected with PA. The severity of bacterial keratitis was graded on days 1 and 3 post-infection by slit lamp, and then corneas were harvested for: (i) bacterial enumeration, (ii) immune cell analysis by flow cytometry, (iii) immunoblotting analysis of cleaved caspase-1 and IL-1β, and (iv) IL-1β quantification by ELISA. In parallel experiments, severity of keratitis was examined in the wild-type mice receiving a subconjunctival injection of a highly selective NLRP3 inhibitor immediately prior to infection. RESULTS: Compared to wild type mice, NLRP3(−/−) mice exhibited more severe infection, as indicated by an increase in opacity score and an increase in bacterial load. The hallmark of inflammasome assembly is the activation of proinflammatory caspase-1 and IL-1β by cleavage of their precursors, pro-caspase-1 and pro-IL-1β, respectively. Accordingly, increased severity of infection in the NLRP3(−/−) mice was associated with reduced levels of cleaved forms of caspase-1 and IL-1β and reduced IL-1β(+) neutrophil infiltration in infected corneas. Likewise, corneas of mice receiving subconjunctival injections of NLRP3 inhibitor exhibited increased bacterial load, and reduced IL-1β expression. CONCLUSIONS: Activation of NLRP3 pathway is required for the clearance of PA infection in mouse corneas.