ATI2341 does not interfere with antimicrobial responses. (a) ROS production measured by Dihydroethidium (DHE) oxidation as determined by flow cytometry in blood neutrophils from vehicle- and ATI2341-treated mice after ex vivo stimulation with vehicle or PMA (left) and in peritoneal neutrophils after in vivo administration of zymosan (right); n = 4–7 (left) and 6–7 (right) mice per group. (b) Expression of IL-1β in blood neutrophils from control and ATI2341-treated mice after ex vivo stimulation with vehicle or PMA as determined by flow cytometry; n = 4 mice per group. (c)Ex vivo NET-formation assay with sorted blood neutrophils at daytime (ZT5) stimulated with PMA from control and ATI2341-treated mice. NETs were quantified (left) by triple colocalization of cit-H3, DNA, and MPO in confocal micrographs (right); n = 7–9 mice per group. (d) Weight loss (up) and survival (bottom) curves of control and neutropenic mice (Mcl1^fl/fl) infected with S. aureus at ZT5 and monitored for 8 days; n = 6 mice per group. (e) Weight loss (up) and survival (bottom) curves of vehicle- and ATI2341-treated mice infected with S. aureus at ZT5 and monitored for 8 days; n = 14–16 mice per group. (f) Survival curves of neutropenic (Mcl1^fl/fl), and vehicle- and ATI2341-treated mice infected with C. albicans at ZT5 and monitored for 5 days; n = 3–10 mice per group. (g) Weight loss curves of vehicle- and ATI2341-treated mice infected with C. albicans at ZT5; n = 10 mice per group. Data in b and d are from single experiments. Data in a, e, f, and g are pooled from two experiments. Data in c are pooled from three experiments. Except for survival curves, data are shown as the mean ± SEM. **P < 0.01; ***P < 0.001; ns, not significant, as determined by two-way ANOVA (a [left], b, d [up], e [up], and g), unpaired t test analysis (a [right], c), and log-rank test (d [bottom], e [bottom], and f).