Extended Data Fig. 1: Administration of zymosan into the air pouch cavity induces neutrophil transcriptional remodeling.
From: Distinct transcription factor networks control neutrophil-driven inflammation
a, Representative H&E staining showing the formation of the granulation tissue (membrane) and infiltration of leukocytes. Representative (left) and zoomed-in (right) images of air pouch cavity from mice subjected to air pouch and zymosan challenge are shown with indicated scale bars. b, Representative image of immunofluorescence imaging on the pouch cavity of Ly6gcre-tdTomato x Cx3cr1GFP illustrates typical neutrophil infiltration. Neutrophils and macrophages appear reddish and green, respectively, and cell nuclei stained with DAPI appear blue. a, b) Representative images from four mice are shown. c, Total number of neutrophils per milliliter of air-pouch exudate (AP) post challenge with zymosan. d, Total number of neutrophils per milligram of air-pouch membrane (MEM) post challenge with zymosan. e, Expression of pro-IL1β, as measured by flow cytometry, in neutrophils from indicated tissues recovered from mice subjected to the air pouch model and zymosan challenge. c, d, e) Results are the means and SD of 15 mice. Significant differences are denoted as: *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001; (DM one-way ANOVA with Dunnett’s multiple comparisons test). f, Gating strategy of neutrophil sorting from Ly6gcre-tdTomato x Cx3cr1GFP mice for subsequent RNA sequencing analysis and ATAC sequencing analysis. g, Gating strategy used to quantify neutrophils under different maturation stages. h, Percentage distribution of neutrophil subsets in BM and blood between steady condition and zymosan-induced inflammation. Data are shown as means and SD from three naïve mice and four mice with zymosan challenge. Significant differences are denoted as: **P < 0.01, ****P < 0.0001 (RM two-way ANOVA with Šidák’s multiple comparisons test). i, Correlation between indicated neutrophil samples with scRNA-seq-defined neutrophil populations reported by Xie et al.11. The mean fraction of indicated scRNA-seq defined clusters (G0-G5) in each group of neutrophil samples. G0-G4: BM neutrophils (G0-CMP, G1-GMP, G2-pre-neutrophils; G3-immature neutrophils; G4-mature neutrophils). G5a, G5b, G5c: peripheral mature neutrophils.
