NLRP3-IN-57

THP-1 monocytes are differentiated into macrophage-like adherent cells by exposure to phorbol 12-myristate 13-acetate (PMA), a phorbol ester used across published THP-1 macrophage differentiation studies; differentiation is assessed by adherence, macrophage-like morphology, altered macrophage-associated surface markers such as CD11b, CD14, CD36, and CD204, phagocytic capacity, lysosomal/mitochondrial enrichment, cytokine responsiveness, and transcriptomic or proteomic remodeling. Because PMA concentration, exposure duration, and post-PMA resting time change downstream phenotype and immune responses, this protocol treats PMA differentiation as a model-generation step rather than a universal macrophage replacement method; low-dose PMA with a rest period is preferred when subsequent inflammatory or infection assays are planned.

Macrophages are important immune effector cells and play an important role in innate and adaptive immune responses. THP-1 cells are usually induced to differentiate into macrophages with PMA.

Pyroptosis is a lytic inflammatory cell-death pathway executed by gasdermin pores, most classically through inflammasome-mediated activation of caspase-1, cleavage of gasdermin D, membrane pore formation, LDH release, and secretion of IL-1β and IL-18. The canonical pathway is commonly modeled by priming cells with an inflammatory signal such as LPS to induce pro-IL-1β and inflammasome components, followed by an activation signal such as ATP or nigericin to activate NLRP3, ASC speck formation, caspase-1 cleavage, GSDMD cleavage, cytokine release, and pyroptotic membrane rupture. The non-canonical pathway is triggered when cytosolic LPS activates mouse caspase-11 or human caspase-4/5, leading to GSDMD cleavage and pyroptosis, and this can secondarily activate NLRP3-dependent IL-1β release. Pyroptosis is linked to inflammatory injury, infection, cancer, liver disease, ocular disease, placental inflammation, and other disease phenotypes, but unresolved questions include which gasdermin fam