GSDMD is not required for IL-1β secretion induced by PAF. (A) Immunoblot of supernatants (Sup) and cell lysates (Lys) from resting or LPS-primed (300 ng/ml, 3 h) WT BMDMs stimulated with or without PAF (25 µM, 3 h). (B) Immunoblot of Sup or Lys from LPS-primed (300 ng/ml, 3 h) WT BMDMs stimulated with nigericin (10 µM, 3 h) or PAF (25 µM, 3 h). (C) IL-1β ELISA using supernatants from LPS-primed (300 ng/ml, 3 h) WT or Gsdmd^−/− BMDMs stimulated with PAF (25 µM), ATP (10 mM), or silica (300 µg/ml) for the indicated time. (D) Immunoblot of supernatants from LPS-primed (300 ng/ml, 3 h) WT or Gsdmd^−/− BMDMs stimulated with PAF (25 µM) for indicated time. Data are representative of three independent experiments. Data in C are presented as mean ± SD from biological replicates. **, P < 0.01; ***, P < 0.001. ns, not significant. Unpaired Student’s t test (C). FL, full-length. Protein marker size, kilodaltons.