Extended Data Fig. 6: The three mammary gland macrophages respond differently to microbial stimuli.

a-b. Representative flow cytometry plots (a) and violin plots (b) showing the percentage of proIL-1β⁺ or TNF⁺ liMacs (a,b) and F4/80^lo macrophages (Mac) and F4/80^hi macrophages (b) from lactating mammary glands (day 11–14 pp) of wild-type mice after exposure to LPS or zymosan in vitro for 6 h. Data was pooled from two independent experiments, n = 7. Each value is a mean of 3 replicates. Kruskal-Wallis test with Dunn’s multiple comparisons test was performed, *p < 0.05; **p < 0.01; ***p < 0.001. c, Representative flow cytometry plots and violin plots showing the percentage of pHrodo Red⁺ F4/80^lo macrophages, F4/80^hi macrophages and liMacs from the lactating mammary glands (day 10–15 pp) of wild-type mice after exposure in vitro to E. coli, zymosan and S. aureus pHrodo Red bioparticles for 90 minutes. N = 10 for E. coli and zymosan pHrodo Red bioparticles, pooled from 3 independent experiments (day 10–15 pp); n = 7 for S. aureus pHrodo Red bioparticles, pooled from 2 independent experiments (day 12–15 pp). All samples were run in triplicates and the means were calculated. Kruskal-Wallis test with Dunn’s multiple comparisons test was performed, *p < 0.05; **p < 0.01; ***p < 0.001, ns = not significant. d, Violin plots showing total cell counts of other mononuclear phagocytes (MNPs) (CD45⁺CD19^-NK1.1^-CD3^-SiglecF^-Ly6G^- ‘LiMac^-‘), eosinophils, NK cells, B cells and T cells in the lactating mammary glands (day 10–14 pp) of control (untreated) and CSF-1 antibody-treated (on day 7–11 pp) mice, challenged with LPS 18 hours prior to analysis or left untreated. Data (n = 5-6 per group) were pooled from 2 independent experiments. One-way ANOVA test was applied, *p<0.05, **p<0.01, ****p<0.0001, ns = not significant. Related to Fig. 5.