Extended Data Fig. 4: Tumor cells absorb lipids from lipid-laden lung neutrophils.
From: Lung mesenchymal cells elicit lipid storage in neutrophils that fuel breast cancer lung metastasis
a, Representative images from 2 biologically independent experiments showing lipid transfer from BODIPY FL C16-loaded neutrophils to 4T1-mCherry cells. Arrowheads indicate neutrophils. Scale bars, 20 μm. b, 4T1-mCherry or AT3-mCherry cells were mono-cultured or co-cultured with their respective tumor-bearing mice-derived neutrophils that were pre-loaded with BODIPY FL C16. Then the intensity of BODIPY in tumor cells was examined by flow cytometry. Representative of 3 biologically independent experiments. c, The cellular TG contents of AT3-mCherry cells after monoculture or co-culture with neutrophils. Data are mean ± s.d. from 3 biologically independent cell cultures. d, Measurement of the lipids in early lung-colonizing AT3-mCherry cells in G-CSF-pretreated mice without and with anti-Ly6G-based neutrophil depletion (see Methods) (n = 5). Data are mean ± s.e.m. e, The lipid transfer from BODIPY FL C16-loaded lung neutrophils to the indicated tumor cells in vivo as determined by flow cytometry (see Methods) (n = 4). Representative flow cytometry histograms are shown. f, As depicted in the left panel, AT3-mCherry or 4T1-mCherry cells were mono-cultured or co-cultured with PB or lung neutrophils. In one experimental group, lung neutrophils were pre-activated by phorbol 12-myristate 13-acetate (PMA) and hydrogen peroxide (H2O2). The total lipid levels in tumor cells were determined by BODIPY 493/503 staining and flow cytometry. Data are mean ± s.d. from 5 (4T1-mCherry) or 3 (AT3-mCherry) biologically independent cell cultures. Neutrophils used throughout this figure were isolated from the pre-metastatic stage of 4T1 tumor-bearing mice or AT3-g-csf tumor-bearing mice. n represents biologically independent animals. P values were determined by one-way ANOVA with Tukey’s multiple comparisons test. ns, not significant.
