Caspase-11-dependent IL-1α release boosts Th17 immunity against Paracoccidioides brasiliensis
Fig 2
IFN-β-dependent caspase-11 activation mediates IL-1α production by P. brasiliensis.
(A) BMDMs from WT, Casp11-/- and Gsdmd-/- mice were incubated with 5 yeasts per cell and the supernatant was harvested for IL-1α measurement after 48 hours (B) IL-1β released by WT and Casp11-/- BMDMs after 48 hours of incubation with P. brasiliensis (MOI 5) was analyzed by western blotting and ELISA. As a control of IL-1β p17 subunit secretion, LPS-primed WT BMDMs were stimulated with nigericin (20uM) for 40 minutes. Combined data from three different experiments were plotted. (C) Time-course of IFN-β expression and production in BMDMs after P. brasiliensis infection. (D) Caspase-11 immunoblot and (E) IL-1α quantification in WT BMDMs treated with indicated doses of anti-IFN-β at the time of P. brasiliensis infection. (F) WT BMDMs were pre-treated with or without recombinant IFN-β 2 hours before P. brasiliensis infection. The presence of caspase-11 and (G) IL-1α were assayed by immunoblotting and ELISA after 24 and 48 hours, respectively. Data are representative of three independent experiments expressed as the mean of triplicate wells. Statistical analysis was performed using non-parametric Mann-Whitney U test (A), parametric Student’s t test (B) or one-way ANOVA with Tukey’s multiple comparison test (E and G). Error bars depict ± SD. (*) p < 0.05 versus P. brasiliensis-infected WT mice. (#) p <0.05 compared to infected macrophages that were not subjected to the anti-IFN-β treatment. (&) depicted p < 0.05 comparing P. brasiliensis-infected samples treated with non-treated. NI: non-infected. ns: not significant.