Fig. 3: Neutrophils and inflammatory monocytes are functionality intact in Il1r1^-/- mice.

A Flow cytometric analysis of intracellular expression of iNOS by monocytes, isolated from kidney and brain of WT and Il1r1^-/- mice, infected with 10⁵ CFU C. albicans, at 3 days p.i.. (n = 5(WT), n = 4(Il1r1^-/-) mice per group). Left: representative dot plot pregated on monocytes (gating is described in Supplementary Fig. 1A). Right: each dot represents a mouse, bars indicate mean values (representative experiment out of 5 repeats is shown), Statistical test: Two-sided unpaired t test per organ. Data are presented as mean values with +/- SD. B Neutrophils were sorted from kidneys or blood of infected WT and Il1r1^-/- mice at 3 days p.i. and co-cultured in-vitro with opsonised C. albicans yeast or hyphae at 5:1 effector:target ratio. Each dot represents mean killing of C. albicans in an independent experiment, data from the same experiment are connected with a line (n = 2/n = 3 mice per group in each experiment). Statistical test: Two-sided paired t test per organ. C In-vitro NET formation by bone marrow neutrophils from WT and Il1r1^-/- mice. Neutrophils were isolated from the bone marrow of uninfected mice, and stimulated for 2.5 h with either PMA, C. albicans (1:1 cell to cell ratio) or C. albicans (1:1) + IL-1b. NET formation was assessed by Sytox green staining (arbitrary units of fluorescence emission at 500-550 nm are used). (n = 3(WT), n = 4(Il1r1^-/-) mice per group, representative of two independent repeats), each dot represents an animal. Plating was performed with 3 technical replicates per condition. Statistical test: Two-sided multiple unpaired t tests, with no correction for multiple comparisons. Data are presented as mean values with +/- SD. D In-vivo NET formation by neutrophils in the brain of Il1r1^+/- and Il1r1^-/- mice, 2 days p.i. with 10⁵ CFU C. albicans. Subsequent slides from brains were stained with either Ly6G or Citrulinated Histone H3 (Cit H3) antibodies. Left panel: Il1r1^+/- mouse, Right panel: Il1r1^-/- mouse. Within each panel: left section: Ly6G immunostaining, right section: Cit H3 immunostaining. (A representative picture from a single experiment with n = 3 mice per group is shown). E–G Analysis of myeloid cells in kidney and brain of infected WT and Il1r1^-/- mice 8, 16 and 24 h after infection with 10⁵ CFU C. albicans. Gating strategy is visualized in supplementary Fig. 1. (E: n = 5(WT), n = 4(Il1r1^-/-) mice per group, F: n = 5(WT), n = 3(Il1r1^-/-) mice per group, G: n = 3(WT), n = 5(Il1r1^-/-) mice per group, all time points are performed as independent experiments), data from representative experiments is displayed. Statistical test: Two sided multiple unpaired t tests, with no correction for multiple comparisons. Data are presented as mean values with +/- SEM. H CXCL1 levels in kidney and brain lysates of Il1r1^+/- and Il1r1^-/- mice 8 h after infection with 10⁵ CFU C. albicans. (n = 3 mice per group). Statistical test: Two-sided multiple unpaired t tests, Two-stage step-up (Benjamini, Krieger, and Yekutieli), Desired FDR 1.00%. Data are presented as mean values with +/- SEM.