Thp1 defnlrp3

Human THP-1 cells from the American Type Culture Collection (ATCC, Manassas, VA) were grown at 37°C with 5% CO₂ in RPMI-1640 medium supplemented with 10% FBS (v/v), 100 U/ml-100 µg/ml penicillin-streptomycin, and 50 µM β-mercaptoethanol. The NLRP3-deficient THP-1 cells (THP1-defNLRP3, InvivoGen) were grown at 37°C with 5% CO₂ following InvivoGen’s instruction in RPMI-1640 medium supplemented with 10% FBS (v/v), 200 µg/ml HygroGold, and 100 µg/ml normocin. HEK-Blue IL-1β cells that contain an IL-1β-sensitive reporter were from InvivoGen and grown following InvivoGen’s instruction at 37°C with 5% CO₂ in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 5% FBS (v/v), 4.5 g/l glucose, 2 mM GlutaMAX medium, 100 U/ml penicillin, 100 µg/ml streptomycin, 100 µg/ml zeocin, 200 µg/ml hygromycin, and 100 µg/ml normocin (all from Life Technologies or InvivoGen).The level of secreted embryonic alkaline phosphatase (SEAP) protein, a truncated form of human placental alkaline phosphatase released into the culture medium, was used as a measurement of NF-κB activation through IL-1β stimulation with the QUANTI-Blue™ detection reagent.