Fig 1: Activation of the NLRP3 inflammasome in vitro. (A) IL1 beta expression in supernatant at different time points upon cholesterol monohydrate and calcium bilirubinate crystal stimulation of M1 macrophages. (B) Comparison of cholesterol monohydrate and calcium bilirubinate crystal‐induced IL1 beta expression at 24 h in M0 (non‐differentiated) and M1 (differentiated, pro‐inflammatory) macrophages, measured by ELISA. (C) Immunoblot analysis showing procaspase 1 and caspase one expression in M1 macrophages after 24 h of cholesterol monohydrate and calcium bilirubinate crystals stimulation. (D) Light microscopy images of M1 macrophages stimulated with cholesterol monohydrate and calcium bilirubinate crystals in different treatment groups, indicating merged immunofluorescence staining of NLRP3 (orange), ASC (green), and nuclei (DAPI, blue) (Brightfield; magnification, x100; scale bar, 100 μm). Data points and mean ± SEM from three independent experiments are shown.