Fig 2: (a) Analysis of differences in tumor purity between three immune subtypes. Tumor purity was importantly more down in Immunity_H group and importantly more excellent in Immunity_L (p < 0.001, Kruskal-Wallis test). (b) Survival analysis of three immune subgroups. The survival curves of Immunity_L, Immunity_M, and Immunity_H subgroups were significantly different (p = 4.396E − 08). It also proved that immune grouping had a good predictive effect on the survival of DLBCL. Patients in Immunity_H had the best prognosis, patients in Immunity_L had got the poorest prognosis, and the Immunity_M was between them. (c–h) The expression of PD-1, PD-L1, CD3D, HIF1A, IRF4, and other genes was meaningfully correlated with the immune subgroup. The expressions of PD-1, PD-L1, CD3D, HIF1A, IRF4, and other genes were meaningfully dissimilar between Immunity_L and Immunity_H (ANOVA text, p < 0.001).

Fig 3: Glucocorticoids promote CD40 signaling and plasmablast-like gene expression ex vivo. (A) Percentage of IgE+ IgG1- B cells (within live B220+) from CD19+ splenic B cells of Stat6 WT, Het, or KO mice cultured with Cont (gray) or DEX (red), as described in Fig. 1 C (n = 2–3 mice/group, representative of two independent experiments). (B) Percentage of IgE+ IgG1- B cells (within live B220+) from WT B cells subjected to IL-4 or aCD40 depletion and treated with Cont (gray) or DEX (red). “Complete” media contained IL-4 and aCD40 (n = 3 mice/group, representative of two independent experiments). (C) Volcano plot showing differentially expressed genes between B cells cultured ex vivo with DEX or Cont. x axis indicates log2(fold change) of counts per million (CPM) and y axis indicates corresponding -log10(adjusted P value) for differential expression, with key B cell or differentially expressed genes highlighted in red (n = 2 mice/group as biological replicates). (D) Effect of DEX on B cell activation of upstream regulators NFKB1 (p105), NFKBIA (I?Ba), and NFKBIB (I?Bb). WT Cont or DEX-treated B cells were analyzed by RNA-seq and pathway analysis for upstream regulators of differentially expressed targets. x axis indicates -log10(P value) for overlap between DEX-induced changes in upstream regulator targets and observed gene expression changes (relative to Cont), by Fisher’s exact test. Numbers and color gradations indicate activation z-score (n = 2 mice/group as biological replicates). (E) Effect of DEX treatment during aCD40 depletion on IRF4 and I?Ba protein abundance by Western blotting. WT B cells were harvested preculture (naive B cells) or after 78 h of complete (lanes 2–3) or aCD40-depleted (lanes 4–5) cultures with Cont or Dex treatment, as similarly described in B (representative of two biological replicates). (F) Heat map showing relative expression of genes associated with plasmablast-like or germinal center and memory B cell–like signatures for Cont vs. DEX-treated IgE+ B cells sorted after culture. Color gradations indicate z-scores by CPM (n = 2 mice/group as biological replicates). (G) Percentage of IgE+ IgG1- cells (within live B220+ CD93+) from Cd19-Cre+/- GR Het vs. cKO B cells cultured with Cont (gray) or DEX (red; n = 5 mice/group, compiled from two independent experiments). Error bars in A–B and G represent SEM. P values <0.05 were considered significant (*, P < 0.05; ****, P < 0.0001; ns, not significant). Ratio-paired Student’s t test was used for A–B and G. Source data are available for this figure: SourceData F2.

Fig 4: Regulation of IRF4 in CD8+ T cells by NAC1. CD8+ T cells were isolated from WT or NAC1−/− mice and analyzed for expression of IRF4. (A) Protein expression by Western blot. For the day 0 sample, T cells were not activated. For the day 3 sample, T cells were activated and cultured for 3 days. (B) mRNA expression by Q-PCR. CD8+ T cells had been cultured for 3 days, and then RNA was extracted, and qPCR was performed (ns, p > 0.05). (C) CHIP-seq analysis. WT CD8+ T cells were isolated from mice and CHIP was performed with anti-NAC1 and anti-IgG Abs (Input control). The sequenced data was visualized by IGV.

Fig 5: T-MPs induce monocytes to upregulate the expression of IRF4. a PKH26-labeled H22-MPs were co-incubated with BALB/c mouse monocytes for 4, 8, or 12 h, and the engulfment of H22-MPs by monocytes was measured by flow cytometry. b, c Monocytes and H22-MPs were co-incubated for 48 h, and the cell morphology was observed under an inverted microscope (b). The percent of CD11c+MHCII+CD64+ moDCs was measured via flow cytometry (c). d Monocytes were co-incubated with H22-MPs. After 24 or 48 h, RNA or protein were collected respectively. Then, the expression of IRF4 was measured at the levels of both gene and protein. e IRF4 mRNA and protein expression of H22-MPs was measured by RT-PCR and western blot. BALB/c mouse bone marrow-derived monocytes co-incubated with H22-MPs were served as positive controls for H22-MPs. f Monocytes transfected with NC siRNA or IRF4 siRNA were cultured with H22-MPs for 48 h. The percent of CD11c+MHCII+CD64+ moDCs was analyzed by flow cytometry. g BALB/c mice were subjected to H22-MPs or PBS via i.m. injection. 24 or 48 h later, IRF4 expression of isolated monocytes from thigh muscles was analyzed at the levels of both gene and protein. Mean ± s.e.m. is represented in the data and two-tailed unpaired Student's t test was used to statistically analyze the P values. **P < 0.01; ***P < 0.001