Macrophage Lcn2 production and antibacterial activity. Macrophages were harvested from WT and Hfe^−/− mice and treated with 100 U/mL IFN-γ or infected with WT S Typhimurium (S Tm) for 24 hours. The production of Lcn2 was investigated in culture supernatants by ELISA (A). Data were log-transformed and compared by ANOVA and are shown as mean ± SEM of at least 3 independent experiments. WT, Hfe^+/−, and Hfe^−/− mice were infected with WT Salmonella. The bacterial burden was determined by gentamicin protection assay (B). In parallel experiments, the acquisition of ⁵⁹Fe by intramacrophage Salmonella was determined (C). In additional experiments, the survival of S Typhimurium within C57BL/6 WT, Hfe^−/−, Lcn2^−/−, and Hfe^−/−Lcn2^−/− macrophages (D) as well as bacterial iron uptake within these cells (E) were determined. These data were analyzed by ANOVA and shown as mean ± SEM of at least 3 independent experiments. *P < .05 compared with the solvent-treated control of the corresponding genotype, #P < .05 for the comparison of cells of different genotype subjected to the same stimulatory regimen.