Figure 1.

NO^● and O₂^●− donors mimic the respiration profile of 2-signal activated macrophages. Macrophages were activated for 24 h with 40 ng/mL IFNγ plus 10 ng/mL LPS or not (untreated) and subjected to serial injections of 2 μΜ oligomycin (ATP production), 1 μΜ FCCP (Maximal respiration) and 2/2 μΜ antimycin A/rotentone, according to the “mitostress” protocol. A Seahorse XFe96 metabolic analyzer was used to record the changes in the oxygen consumption rate (OCR). The reactive species-donors SIN-1 (NO^● and O₂^●−) or SNAP (NO^●) were added, at the concentration of 3 mM each, prior to the assay in cell-free wells. The last OCR measurement in sequence before addition of oligomycin was used to represent the basal respiration. Two-tailed Mann-Whitney U tests or the Kruskal-Wallis test (with the Benjamini-Krieger-Yekutieli two-stage linear step-up, or with the Dunn’s, procedure) were used for comparisons. ns: not significant, ** p < 0.01, *** p < 0.001. Horizontal intermittent line indicate the basal OCR levels of IFNγ + LPS activated macrophages. One representative experiment is shown, with data presented as mean ± STD of at least three replicates per condition.