Preparation and Treatment of Peritoneal Macrophages and HT-29 Cells

Peritoneal macrophages were prepared as described previously (Zhang et al. 2021b (link)) and were cultured in RPMI 1640 medium supplemented with 10% FBS (SH30071.03, HyClone, USA), 100 units/mL of penicillin, and 100 μg/mL of streptomycin. 2 μg/mL of LT [2 μg/mL of LF (172C, List Biological Labs) and 2 μg/mL of PA (171E, List Biological Labs)] was used to treat peritoneal macrophages for 3 h. HT-29 cells of indicated genotypes were cultured in DMEM supplemented with 10% FBS (10099-141, Gibco, USA), 100 units/mL of penicillin, and 100 μg/mL of streptomycin. 2 μg/mL of LT, 30 ng/mL of human TNF (PHC3011, Gibco), and 10 μmol/L of LCL-161 were used to induce cell death in HT-29 cells. Inhibitors used in this study were 2 μmol/L of SB202190 (S1077, Selleck, USA), 2 μmol/L of SB203580 (559389, Calbiochem), 2 μmol/L of TAK-715 (HY-10456, MedChemExpress), 2 μmol/L of U0126 (S1102, Selleck), 2 μmol/L of GDC-0994 (S7554, Selleck), and 10 μmol/L of SP600125 (S5567, Sigma–Aldrich, USA).