Characterisation of endogenous IL-33 using a model of Alternaria-induced pulmonary inflammation

Background IL-33 is a potential mediator of chronic inflammatory diseases including asthma. However, endogenous forms of IL-33 are poorly understood. Here we have used a model of Alternaria alternata (ALT) challenge to investigate in vivo mechanisms of IL-33 processing and release. Methods Cytokines were quantified in bronchoalveolar lavage (BAL) from BALB/c mice by MSD assay. Mast cells were differentiated from bone marrow over 5 weeks and stimulated with BAL or recombinant IL-33 for 24 hours. Results Intranasal challenge with 50µg ALT, but not 25µg house dust mite, extract in 50µL to anaesthetised BALB\c mice caused a spike (∼2000pg at 15-30min) of IL-33 in BAL followed by rapid clearance (below detection limit of 2pg within 4-5 hrs). Key downstream effects were an increase in 5 and 24 hours BAL IL-5 (∼400pg and ∼600pg, respectively) and eosinophilia at 24 hours. Dependence on the IL-33-ST2 axis was confirmed in ST2 and IL-33 KO mice which showed no IL-5 or eosinophilia after challenge. IL-33 is considered an alarmin, released on cell damage, yet histological analysis of ALT challenged lungs showed no overt damage to the airway epithelium. The presence of active IL-33 in WT but not IL-33KO BAL was confirmed ex vivo by stimulation of mouse mast cells. Western blot analysis of BAL demonstrated the presence of full length IL-33 (∼31kDa) and a ∼20kDa processed form in WT, but not IL-33KO, ALT-treated mice. Conclusion IL-33 is released rapidly and in large quantities following ALT exposure. This occurs in the absence of overt epithelial injury and results in generation of a ∼20kDa processed form of IL-33 in vivo . These data support the current view of IL-33 as an alarmin-type molecule.