Immortalized mouse macrophages from wild type (WT) were treated with Kdo₂-Lipid A and Lipid IVa under three experimental conditions: (i) extracellular stimulation to assess TLR4-dependent activation with Kdo2 lipid A or lipid-IVa (Extra_Kdo2/Extra_IVa), (ii) direct cytosolic delivery by transfection to evaluate TLR4-independent sensing with Kdo2 lipidA and lipid IVa (Intra_Kdo2/Intra_IVa), and (iii) double activation consisting of priming with Pam3CSK4 (P3C) for 6 h followed by transfection with either Kdo2 or IVa for 24 h to induce non-canonical inflammasome activation (DA_Kdo2/DA_IVa). We also use extracellular P3C stimulation for 6h for another control of priming. Untreated macrophges were serve as a control. This project also contains the pilot study data of Acyloxyacyl hydrolase (AOAH) knockout under non-canonical inflammasome activation (KO_DA_Kdo2/KO_DA_IVa).