Figure 3

iNOS negatively regulates NLRP3 inflammasome activation. (A) Production of cytokines by peritoneal macrophages primed with LPS for different lengths of time in the presence or absence of L-NMMA, followed by stimulation with ATP for 30 min. (B) Immunoblot analysis of caspase-1 and iNOS in lysates of peritoneal macrophages treated as in (A). (C) Production of cytokines by peritoneal macrophages transfected with iNOS-specific (S1 and S2) or control (Sc) siRNA, then primed with LPS for different lengths of time and stimulated with ATP for 30 min. (D) Immunoblot analysis of caspase-1 and iNOS in lysates of peritoneal macrophages treated as in (C). (E) Production of cytokines by peritoneal macrophages from wild-type (WT) and iNOS-knockout (iNOS^−/−) mice primed with LPS for different lengths of time and stimulated with ATP. (F) Immunoblot analysis of caspase-1 and iNOS in lysates of peritoneal macrophages from WT and iNOS^−/− mice treated as in (E). Actin serves as a loading control (B, D and E). ProCasp1, procaspase-1; Casp1 p10, active form of caspase-1. ^**P < 0.01 and ^***P < 0.001 (Student's t-test). The data represent at least three experiments (mean±SD in A, C and E).