Figure 3. IRF8 represses the expression of OPN expression in myeloid cells.

(A) RNA was prepared from total spleens of WT (n = 3) and IRF8-KO (n = 3) mice and analyzed by qPCR for OPN mRNA level. (B) Spleen cells of WT (n = 3) and IRF8-KO (n = 3) mice were stained with CD19-, CD3-, CD11b-, and Gr1-specific mAbs, followed by intracellular staining of OPN. The OPN⁺ cells were gated to show the CD11b⁺Gr1⁺ myeloid cells from IRF8-KO mice (left panel). OPN protein level in these CD11b⁺Gr1⁺ myeloid cells is shown in the right panel. (C) The OPN⁺ cells in total spleen cells of WT (n = 3) and IRF8-KO (n = 3) mice as shown in B were quantified. (D) Spleen cells from IRF8-GFP mice were stained with Ly6G- and Ly6C-specific mAbs. Ly6C^hiLy6G⁺ and Ly6C^loLy6G^– cells were overlaid for GFP intensity. Shown are representative plots of 1 of 3 mice (left panel). The GFP⁺ cells of the Ly6C^hiLy6G⁺ and Ly6C^loLy6G^– cells were quantified and are presented in the right panel. (E) The spleen cells were stained with Ly6C- and Ly6G-specific mAbs, followed by intracellular staining with OPN-specific antibody. The Ly6C^hiLy6G⁺ and Ly6C^loLy6G^– cells were gated and analyzed for OPN expression level. Representative plot of 1 of 3 mice is shown. The OPN⁺ cells in Ly6C^hiLy6G⁺ and Ly6C^loLy6G^– cells were quantified and are presented in the right panel.