P2X₇^−/− BMDCs fail to process and release mature IL-1β. (A) P2X₇^−/− and WT BMDCs were left untreated or were treated with 1 µg/ml LPS and/or 5 mM ATP for 12 h. Western blot analysis was performed to detect pro–IL-1β and mature IL-1β in the culture supernatant. Supernatant from LPS + ATP–stimulated macrophages (MP sup) or recombinant pro–IL-1β was used as positive control. The data are representative of two independent experiments. (B) The same supernatants were used to detect IL-1β secretion by ELISA (***, P < 0.001). (C and D) BMDCs were treated with alum at the indicated concentrations for 12 h in the presence or absence of 0.1 µg/ml LPS. IL-1β secretion was determined by ELISA. (B–D) Data represent the mean of triplicate measurements ± SD and are representative for two (B) or three (C and D) independent experiments.