Fig. 5

FPN regulates the proliferation and differentiation of RAW264.7 cells. a, b FPN protein levels in RAW264.7 cells after 4 h of treatment with Ctrl OB-CM, TgHamp1-Ocn OB-CM, vehicle or 200 nmol·L⁻¹ hepcidin peptide. FPN levels were analyzed by western blotting. Representative blots are shown in (a). Quantitative data (the mean ± SD of three separate experiments) are presented in (b). *P < 0.05. c, d Western blot analysis of FPN expression in Ctrl and FPN-KD RAW264.7 cells. Quantitative data (the mean ± SD of three separate experiments) are presented in (d). **P < 0.01. e, f Immunostaining analyses of Ki67 in RAW264.7 cells. Ctrl and FPN-KD RAW264.7 cells were treated with vehicle or hepcidin (200 nmol·L⁻¹) for 1 d. Quantitative data are shown in f. The data are presented as the mean ± SD of three separate experiments. **P < 0.01. g, h TRAP staining of RAW264.7 cells. Ctrl and FPN-KD RAW264.7 cells were treated with vehicle or hepcidin (200 nmol·L⁻¹) and 100 ng·mL⁻¹ RANKL for 5 d. Bar, 200 μm. Quantification of TRAP⁺ MNCs is shown in h. The data are presented as the mean ± SD of three separate experiments; **P < 0.01. i, j Immunostaining analyses of Ki67 in RAW264.7 cells. Lv-Ctrl and Lv-FPN-C326S-transfected cells were treated with vehicle or hepcidin (200 nmol·L⁻¹) for 1 d. Quantitative data are shown in j. The data are presented as the mean ± SD of three separate experiments. **P < 0.01. k, l TRAP staining of RAW264.7 cells. Lv-Ctrl and Lv-FPN-C326S-transfected cells were treated with vehicle or hepcidin (200 nmol·L⁻¹) and 100 ng·mL⁻¹ RANKL for 5 d. Bar, 200 μm. The number of TRAP⁺ MNCs is shown in l. The data are presented as the mean ± SD of three separate experiments. *P < 0.05; **P < 0.01