Extended Data Fig. 10: ILC2s are required for the goblet cell adaptation to the microbiota in the middle colon in the steady state.
From: Spatially restricted immune and microbiota-driven adaptation of the gut
a, Representative flow cytometric gating strategy for total ILC2s isolated from mouse MLN. b, Experimental schematic (created using BioRender (credit: H. Yano, https://biorender.com/j03f165; 2024) for ILC2 depletion (top) and gating strategy and summary (bottom) of ILC2s from MLNs in control and Cre+ mice. n = 4 mice per group and summarize 2 independent experiments. Unpaired two-sided t-test. Data are presented as the mean ± s.e.m. c, Data summarizing the number of eosinophils recovered from colon regions A-D gated as in Extended Data Fig. 9c for SPF (purple) and GF (black) mice. n = 22 mice from 7 independent experiments for SPF and n = 9 mice from 3 independent experiments for GF. Box limits are the upper and lower quartiles with the middle line representing the median; and whiskers are 1.5 times the interquartile range from the box. d, Representative histograms showing expression of SIGLEC-F on the surface of eosinophils gated as in Extended Data Fig. 9c for SPF (purple) and GF (black) mice from colon region B. e, Data summarizing MFI of SIGLEC-F on eosinophils recovered from GF colon regions A-D gated as in c in SPF (purple) and GF (black) mice. n = 21 mice from 7 independent experiments for SPF and n = 9 mice from 3 independent experiments for GF. Box limits are the upper and lower quartiles with the middle line representing the median; and whiskers are 1.5 times the interquartile range from the box. f, Experimental schematic (created using BioRender (credit: H. Yano, https://biorender.com/j03f165; 2024) for eosinophil depletion via anti-IL-5 administration (top) and gating strategy and summary (bottom) of eosinophils from MLNs in isotype and anti-IL-5-treated WT SPF mice. n = 4 mice per group; data summarize 2 independent experiments. Unpaired two-sided t-test. Data are presented as the mean ± s.e.m. g, Antibody staining of ANG4 protein (green; Minimum display value = 10 and maximum display value = 30) on colon tissue from isotype and anti-IL-5 treated SPF mice showing enrichment in the C2 region in all conditions. Nuclei are stained with DAPI (gray; Minimum display value = 5 and maximum display value = 50). n = 2 mice per group and images representative of 2 independent experiments. h, Expression of example middle colon genes in Xenium colon samples (n = 2 mice for each condition). i, Difference in cell counts assigned to the Xenium colon samples between DT treated ROSA26LSL-DTR and Nmur1iCre-eGFPROSA26LSL-DTR mice (t statistic with pooled variance). Colon samples were binned into four bins following the way single-cell data were generated along the unrolled X axis. Boxplots display first and third quartiles.
