Generation and validation of IL-6 expressing EL4 tumor in vitro and in vivo. (A) Sort-purified MDPs were cultured with Flt3L with or without 25 ng/ml IL-6 and analyzed by FACS after 7 d. Shown is FACS analysis of MerTK⁻ B220⁻ SiglecH⁻ cells. Data is representative of four independent experiments. (B) EL4-empty and EL4-IL-6 cell lines were treated with brefeldin A (Sigma-Aldrich) for 3 h and analyzed by FACS for intracellular staining of IL-6 and EGFP. Rat IgG1 antibody was used for isotype control (ctl) staining. (C) Sort-purified cKit^hi BM progenitors were cultured for 9 d with Flt3L and the indicated concentration of conditioned media (CM) from EL4-empty or EL4-IL-6 cells or recombinant murine IL-6. (D and F) Related to Fig. 1 E, the bar-scatter graphs show the average (D) frequencies (%) and (F) absolute number of cDC in the spleen ± SD. (E and G) Related to Fig. 1 F, the bar-scatter graphs show the average (E) frequencies (%) and (G) absolute number of cDC in the MLNs ± SD. cDC total (gray and/or white), cDC1 (red), and cDC2 (blue). The numbers of mice for each experimental group were indicated as dots in the bar graphs, PBS (n = 7), EL4-empty (n = 9), EL4-IL-6 (n = 11). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (Student’s t test).