CARD8-knockdown facilitates ATP-induced IL-1β secretion. (A) Caspase recruitment domain-containing protein 8 (CARD8) targeting (siCARD8-1, siCARD8-2) or control (siCon) RNA (50 nM) was transfected in human monocyte-derived macrophages (HMDMs). After stimulation of the cells with lipopolysaccharide (LPS), efficiency of the knockdown was evaluated by qPCR. (B) CARD8-knockdown HMDMs were stimulated with LPS and adenosine triphosphate (ATP), and secreted IL-1β was measured by ELISA. (C) CARD8-knockdown HMDMs were stimulated with LPS and ATP, and activation of caspase-1 was determined by flow cytometry using a fluorescence-labeled irreversible caspase-1 substrate (FLICA). The data shown are representative of three independent experiments with similar results. Each column represents the mean ± SD (n = 4), ***P <0.001.