Cytokine induction in early infection does not involve an autocrine mechanism.

<p>(A) After incubation with the indicated amount of recombinant human IL-1Ra for 30 min, miCtrl cells were stimulated with recombinant human IL-1β for 60 min, and the IL-8 secretion was analyzed by ELISA. (B) After incubation with 100 ng/ml IL-1Ra for 30 min, miCtrl cells were infected with <i>S</i>. <i>aureus</i> at an MOI of 4 for the times indicated. IL-8 and TNF-α secretion from each cell line were analyzed by ELISA. (C) After incubation with 20 μM Ac-YVAD-CMK for 30 min, Dox-untreated miCtrl and miNLRP3 cells were infected with <i>S</i>. <i>aureus</i> for 100 min. TNF-α and IL-1β secretion were analyzed by ELISA. (D) THP-1 cells were incubated with 5 μg/ml CHX for 30 min and then infected with <i>S</i>. <i>aureus</i> at an MOI of 4 for the indicated times. TNF-α and IL-1β secretion were analyzed by ELISA. (E) THP-1 cells were treated with CHX and infected as in (D). The total RNA was analyzed for TNF-α and IL-1β mRNA by RT-PCR. (F) ELISA analysis of TNF-α and IL-1β released from THP-1 cells treated with MSU in the presence or absence of 0.63, 1.25, 2.5, 5, and 10 μM Ac-YVAD-CMK for 180 min. (G) ELISA analysis of IL-18 released from THP-1 cells treated as in (C) (left panel) or treated with MSU in the presence or absence of 10 μM of Ac-YVAD-CMK for 20 h (right panel). All results are representative of three independent experiments. ND, not detected.