Fig 2: The microbiota does not control inflammatory lung phenotypes in epg5-/- mice. (A,B) Flow cytometric analysis was performed on lung tissue from Epg5+/- and epg5-/- SPF, GF, or GF+FT littermates, and the percentage of neutrophils (PTPRC/CD45+ Ly6G+ ITGAM/CD11b+) (A) as well as the ITGAM/CD11b-positivity of alveolar macrophages (pre-gated on PTPRC/CD45+ SIGLECF+ ITGAX/CD11c+) (B) was assessed. (C) Histological analysis of Epg5+/- and epg5-/- SPF, GF, or GF+FT littermate lungs, stained for SQSTM1 with DAPI counterstaining. Representative photomicrographs are shown. (D) QPCR was used to analyze expression of Il1b and Tnf in lungs from these mice. N = 6–8 mice per group over two independent experiments. Statistical analyses performed using Mann-Whitney test.

Fig 3: Inhibition of EZH2 activity promotes MDSC generation from HPCs. a–c Sort-purified HPCs from mouse BM cells were cultured with GM-CSF and IL-6 to allow induction of MDSCs in vitro. a Schematic of in vitro MDSC differentiation from mouse HPCs and experimental design for GSK343 treatment. b Immunoblotting of EZH2 and H3K27Me3 in GM-CSF/IL-6-induced cells at the indicated time points of induction. GAPDH and Histone H3 were used as loading controls (n = 4 per group). The statistical significance of differences is vs. 0 h of cell induction. c Representative flow cytometry plots of MDSCs (CD11b+ Gr-1+) induced by GM-CSF/IL-6. Gated numbers represent the percentage of CD11b+ Gr-1+ cells. d Immunoblots of H3K9Me3, H3K27Me1, H3K27Me2, and H3K27Me3 protein following treatment with GSK343 for 72 h. Histone H3 was used as a loading control (n = 3 per group). e As described in a, MDSCs were induced from mouse HPCs, vehicle (control), or GSK343 was added at different induction times. The total cells were collected at 96 h and representative staining, frequencies, and absolute numbers of developing MDSCs were assessed by flow cytometry. Numbers adjacent to the outlined areas indicated the percentage of CD11b+ Gr-1+ MDSCs (n = 3 per group). The statistical significance of differences was vs. control. f Signaling pathway enrichment analysis was performed using Kyoto Encyclopedia of Genes and Genomes (KEGG). Significantly enriched (nominal P < 0.05) pathways in in vitro-induced HPCs treated with or without GSK343 are plotted by enrichment score (n = 3). g Enrichment plot of the HALLMARK Jak-STAT and TNF signaling pathways for the comparison between in vitro-induced HPCs treated with vehicle (control) and those treated with GSK343. h Heatmap displays the differentially expression of Jak-STAT signaling pathway genes in vitro-induced HPCs treated with vehicle (control) and GSK343. i Heatmap illustrating the differentially expressed genes of TNF signaling pathway in vitro-induced HPCs treated with vehicle (control) and GSK343. Data shown are mean ± SEM and are representative of three independent experiments. *P < 0.05, **P < 0.01 ***P < 0.001, n.s. = not significant (as determined by one-way analysis of variance followed by Bonferroni post test). Source data are provided as a Source Data file

Fig 4: Macrophage CD147 regulates foam cell formation by mediating ox-LDL uptake. (A,B) BMDMs from CD147WT or CD147M–KO mice were examined using either confocal microscopy (A) or flow cytometry (B) after treatment with DiI-oxLDL (20 μg/mL) for 4 h. The scale bar is 20 μm. Right panel: Quantification of MFI of DiI-oxLDL in CD11b+F4/80+ BMDMs. (C,D) BMDMs from CD147WT or CD147M–KI mice were examined using either confocal microscopy (C) or flow cytometry (D) after treatment with DiI-oxLDL (20 μg/mL) for 4 h. The scale bar is 20 μm. Right panel: Quantification of MFI of DiI-oxLDL in CD11b+F4/80+ BMDMs. (E,F) A cholesterol efflux assay kit was used to detect LXR-independent and -dependent cholesterol efflux to HDL or ApoAI in BMDMs isolated from CD147M–KO (E) or CD147M–KI (F) mice and their CD147WT littermates. Data represent the mean ± SEM. of n = 3 biologically independent experiments. ***P < 0.001.

Fig 5: EZH2 inhibition increases colonic myeloid-derived suppressor cells (MDSCs). C57BL/6 mice were exposed to either 2.5% DSS-containing or regular drinking water alone for 5 days. DSS-treated mice were intravenously injected with either vehicle or GSK343 at days 0 and 3. a At day 5, the representative staining, percentage, and absolute number of macrophages (CD11b+ F4/80+), dendritic cells (DCs) (major histocompatibility complex (MHC) class II+ CD11c+), eosinophils (Siglec-F+ Gr-1-), and MDSCs (CD11b+ Gr-1+) in CD45+ cells of the cLP were determined by flow cytometry (n = 5, Water; n = 6 for both DSS and DSS + GSK343). Dot plots are gated on CD45+ cells. Numbers adjacent to the outlined areas indicate the percentage of the gated population in each group. b The expression of Gr-1 (red) in distal colon tissue sections from DSS-exposed mice treated with vehicle or GSK343 was detected by immunofluorescence staining. Magnified views of the marked areas are shown in the adjacent panels. The numbers of Gr-1+ cells per field are enumerated on the right (n = 6 per group). Throughout, data are representative of three independent experiments. The statistical significance of differences was determined by one-way analysis of variance followed by Bonferroni post test (a) and unpaired two-tailed Student’s t-test (b). *P < 0.05, **P < 0.01, ***P < 0.001, n.s. = not significant. Error bars indicate means ± SEM. Source data are provided as a Source Data file