LDN-193189 inhibits short-term IL-6– and turpentine-induced hypoferremia and hepcidin expression. (A) IL-6 (16 μg) or vehicle were injected IV into 10-week-old mice pretreated with or without LDN-193189 (3 mg/kg IP) or vehicle. After 2 hours, livers were harvested, and hepcidin mRNA levels were measured using qRT-PCR (n ≥ 3 each group, 1-way ANOVA P < .005, *P < .05 vs control, †P < .05 vs saline, #P < .05 vs IL-6–stimulated mice). (B-C) Mice were pretreated with LDN-193189 (3 mg/kg IP repeated each 12 hours) or drug vehicle, followed by a single intrascapular injection with turpentine (5 mL/kg) or saline. Twenty-four hours after the turpentine injection, hepcidin mRNA levels were measured by qRT-PCR (B), and serum iron levels were assayed (C; n ≥ 4, *P ≤ .01 vs untreated controls, #P < .05 vs turpentine). (D) Mice were pretreated with ALK3-Fc (2 mg/kg IP) or vehicle, followed by intrascapular injection with turpentine or saline, and serum iron levels measured (n ≥ 5, *P < .000 01 vs untreated control, #P < .05 vs turpentine). Serum IL-6 levels (E) were determined by ELISA before and 6, 12, 24, 48, and 96 hours after turpentine injection with or without LDN-193189 (3 mg/kg) administered every 12 hours (n = 5 mice per time point and treatment). Hepcidin gene expression in the livers of treated mice was measured by qRT-PCR at 0, 24 and 96 hours (F; n = 5, 1-way ANOVA P = .003, *P < .05 vs 0 hours, #P < .01 vs vehicle at 24 hours, †P = .01 vs vehicle at 96 hours). Levels of phosphorylated-SMAD1/5/8 (p-SMAD) and SMAD1 proteins (G) in the livers of mice treated with vehicle, turpentine, or LDN-193189 were measured by immunoblot (n = 4 mice each).