Airway allergic inflammation induced by subtilisin. (A) Experimental protocol for the development of allergic airway inflammation. C57BL/6 mice were s.c. sensitized with subtilisin or papain with or without alum on days 0 and 7 and i.n. challenged with the respective enzyme on days 14 and 21. Control groups received saline and samples were harvested on day 22, 24 h after the last enzyme challenge. (B) Total serum IgE and (C) subtilisin-specific IgG1. BAL (D) total and (E) differential cell counts. (F) Representative lung sections (200×) showing mucus production and peribronchovascular cellular infiltrates. (G) Remaining specific enzymatic activities of subtilisin (left) and papain (right) after incubation with different concentrations of alum. Error bars show ***, P ≤ 0.0001; **, P ≤ 0.001; *, P ≤ 0.01 for significant differences to control group or • to protease without alum group. Data are mean ± SEM and are representative of three independent experiments (n=5) (B-E) or are representative of, at least, five experiments (F).