Regulatory functions of neutrophils in the intestine. (A) Experimental strategy and cytometry plots. Dot plots, gated on CD45⁺ CD11b⁺ Ly6G⁺ cells, show neutrophils in tissues of Fut7^RED mice after parabiosis with Fut7^−/− control or WT partners. Numbers indicate mean ± SEM of partner-to-host ratios corrected by ratios in blood and normalized to the Fut7^−/−; Fut7^−/− parabiotic pairs (see Materials and methods). (B) Radial plots showing the relative infiltration of partner-derived and Fut7^−/− neutrophils in different tissues. Polygons show the mean ± SEM ratio of partner to Fut7^RED neutrophils in each tissue, which are shown in the plots in A. Data from four to five pairs per group from three independent experiments. (C) Experimental approach to estimate phagocytosis of partner-derived fluorescent leukocytes by tissue macrophages using flow cytometry. (D) Percentage of macrophages that engulf partner-derived cells in organs of Fut7^−/− parabionts paired with WT^RED or Fut7^RED partners; n = 5–6 mice from two independent experiments. (E) Number of circulating progenitors in Fut7^−/− mice after 1 mo in parabiosis with Fut7^−/− or WT partners or Fut7^−/−; Cd169^DTR mice paired with WT mice after macrophage depletion by treatment with DT; n = 6–12 mice from three independent experiments. (F) Whole-mount staining of the large intestine of Lyz2^GFP mice showing GFP⁺ foci in the mucosal surface. Bar, 1 mm. (G) Detail of an ILF showing the distribution of CD169⁺ macrophages, MRP14⁺ neutrophils, and the microvasculature (laminin; white). Bar, 100 µm (see also Video 9). (H) Representative plot showing intestinal macrophages that phagocytose (red) or not (black) partner-derived DsRed⁺ cells, as in C and D. Bar graph at right shows expression of Il1b and Il23 in phagocytic macrophages relative to non-phagocytic cells; data are from cells sorted from three individual mice. (I) Scheme of the IL-23–IL-17–G-CSF pathway. (J) Number of circulating CFU-C in Fut7^−/− mice after treatment with isotype or blocking antibodies against the cytokines shown in I; n = 5–6 mice from two (left) and one independent experiments. Data shown as mean ± SEM. *, P < 0.05; **, P < 0.001; ***, P < 0.001; n.s., not significant, as determined by unpaired t test analysis (B, D, H, and J) or one-way ANOVA analysis with Tukey’s multigroup test (E).