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. Author manuscript; available in PMC: 2024 Feb 1.
Published in final edited form as: Nat Immunol. 2023 Jan 5;24(2):239–254. doi: 10.1038/s41590-022-01388-8

Fig. 5 |. Whole beta-glucan particle training increases phagocytic and cytotoxic activity of lung interstitial macrophages.

Fig. 5 |

a, GSEA plot for the regulation of phagocytosis and heat map for the genes related to the phagocytosis regulation pathway. NES, normalized enrichment score. b, Phagocytosis assay was performed with lung AMs and IMs from WGP-trained (n = 4) or PBS control (n = 4) mice. Phagocytosis of pHrodo-green-labeled S. aureus was analyzed by flow cytometry. Representative dot plots and summarized data are shown. c, In vitro cytotoxicity assay using sorted lung IMs from PBS control (n = 3) or WGP-trained (n = 3) mice and co-cultured with LLC cells at different ratios. Cells were cultured for 16 h and cytotoxicity was measured by lactate dehydrogenase (LDH) release assay. d, In vivo cytotoxicity assay. Six-week-old C57BL/6 PBS control (n = 5) and WGP-trained (n = 5) mice were i.v. injected with 1 × 106 LLC-GFP cells and were analyzed for the frequency of LLC-GFP cells in the lungs after 24 h. Representative dot plots and summarized data are shown. e, GSEA plot for ROS biosynthetic process and heat map for the related leading genes in the WGP-trained lung IMs. f, MitoSox Red staining for PBS and WGP-treated peritoneal macrophages. Peritoneal macrophages were treated with PBS (n = 8) or WGP (n = 9) for 24 h and then stained with MitoSox Red and analyzed by flow cytometry. Representative histogram and summarized data from two independent experiments are shown. g, Lung IMs sorted from WGP-trained mice (n = 3) were co-cultured with LLC target cells in the presence or absence of Mito-TEMPO at a 10:1 ratio. Cytotoxicity was measured by the LDH release assay. Data are representative of two independent experiments and presented as the mean ± s.e.m. For a and e, nominal P values were used to determine significance. When the nominal P value is represented as 0, this means P < .0001. The nominal P value was calculated by empirical phenotype-based permutation test. *P < 0.05, **P < 0.01, ****P < 0.0001. P values were derived from an unpaired two-tailed student’s t-test for b, d and f, a two-way ANOVA with Tukey’s multiple-comparison test for c and a one-way ANOVA for g.