Hepcidin-induced ferroportin internalization and hypoferremic response to acute inflammation requires adequate liver iron. Eight-week-old male C57BL/6 mice (n = 24) were fed with an iron-deficient diet for 3 weeks. During the last night, half of the animals were provided a high-iron diet (2% carbonyl iron). On the next day, all mice (n = 6 for each group) were injected with either phosphate-buffered saline or 1 μg/g LPS and sacrificed after 4 hours. (A) Serum iron; (B) transferrin saturation; (C) TIBC; (D) liver iron content; (E) liver Hamp mRNA; (F) serum hepcidin. All data are presented as the mean plus or minus SEM. Dotted lines indicate average values obtained from age-matched male C57BL/6 mice (n = 3) on standard diet. Statistical analysis was performed by 1-way ANOVA. Statistically significant differences (P < .05) across treatments (vs columns a, b, c) are indicated by a, b, c. (G) Immunohistochemical staining of ferroportin in liver sections (original magnification ×20). Arrows indicate ferroportin internalization in Kupffer cells from LPS-treated mice that had overnight access to the high-iron diet.