Fig. 3: Functionally reprogrammed hepatocytes orchestrate Egr1⁺ neutrophil positioning at the tumor–liver interface via the SAA2–FPR2 axis.

A Flow cytometric quantification of Egr1⁺ neutrophils across five tissue compartments (tumor, LN, blood, spleen, liver) on day 14 (n = 3 mice, Student’s t test). B Time-course analysis of Egr1⁺ neutrophil frequency in the liver over 18 days (n = 3 mice per time point, two-way ANOVA). C UMAP visualization and re-clustering of hepatocytes into 13 subpopulations (Hepa-C0 to Hepa-C12). Bar plot shows proportional distribution. (n = 4 mice). D Pseudotime enrichment analysis of hepatocyte subpopulations (n = 4 mice). E. Spatial transcriptomic maps of two CRLM patients showing the localization of Hepa-C11 (functionally reprogrammed hepatocytes) and EMT signatures. Right: Spatial proximity (k-distance) analysis of Hepa-C11 to tumor borders, EMT regions, and Egr1⁺ neutrophils. F mIHC showing co-localization of SAA2 (cyan), EGR1 (red), and LY6G (green) in the liver of KIAA1199-high mice (n = 2 mice). Yellow arrowheads: Egr1⁺ neutrophils. Scale bars: 100 µm. G mIHC showing spatial distribution of Egr1⁺ neutrophils, SAA2, and CD31⁺ vasculature in human CRLM samples across tumor core, transitory, and leading edge regions (n = 2 patients). Scale bars: 100 µm. H–J In vivo validation. Mice bearing orthotopic tumors were treated with WRW4 (FPR2 antagonist, 2 mg/kg/day) or recombinant SAA2 (1 μg/kg/day). Livers were harvested on day 14 for neutrophil isolation. I Western blot of isolated neutrophils (n = 3 independent experiments). (J) Flow cytometry of hepatic Egr1⁺ neutrophils (n = 3 mice, Student’s t test). K Schematic of therapeutic intervention in the dual-tumor model. Mice received WRW4 (2 mg/kg/day) treatment. L Representative gross liver and H&E images at day 21 (n = 4 mice). Scale bars: 2 mm (gross) and 2 mm (histology). M, N Quantification of liver metastatic area (M) and nodule number (N) (n = 4 mice, Student’s t test). O. Kaplan–Meier survival curves over 60 days (n = 10 mice, log-rank test). FACS sequential gating strategies are shown in Supplementary Fig. 5G. Where applicable, all statistical tests are two-sided. Data are presented as mean ± s.d. Source data and exact p values are provided as a Source Data file.