Fig. 5.

Inhibition of MIF prevents NLRP3 inflammasome activation. ASC-cerulean macrophages were primed with LPS (10 ng ml⁻¹) overnight. The following day cells were treated with COR123625 (50 µM) for 2 h before activation of the inflammasome with a nigericin (10 µM) (1 h) or b silica (150 µg ml⁻¹) (4 h). Confocal images are representative of three independent experiments. c, d Data are presented as the percentage of ASC-speck-positive cells. Data shown are mean ± SEM of three independent experiments. e WT BMDMs were left untreated, treated with LPS alone (100 ng ml⁻¹) for 5 h, treated with COR123625 for 1 h prior to the addition of LPS, primed with LPS before inflammasome activation with nigericin (5 µM) for 1 h, or treated with COR123625 before LPS and nigericin stimulation. Levels of LDH release were quantified using the Promega cytotoxicity assay. f WT or Mif^−/− BMDMs were treated with LPS (10 ng ml⁻¹) + nigericin (5 μM) and lysates and supernatants analyzed by Western blot for caspase-1 and IL-1β. Images are representative of >3 mice. g BMDMs from WT and Mif^−/− mice were treated with (10 ng ml⁻¹) + nigericin (5 μM), fixed and stained for ASC, and analyzed by confocal microscopy. Images are z projections of multiple z-stacks. h Quantitation of ASC specks in g, n = 3 mice per group. Data are expressed as percentage increase of mean ± SEM from three mice. *P < 0.05, **P < 0.01, ***P < 0.005, or ****P < 0.001, one-way ANOVA with a correction provided by the Tukey's multiple comparisons test