Foxo1 modulates hepcidin expression in hepatocytes. (A-B) Primary hepatocytes treated with the Foxo1 specific inhibitor AS1842856; (A) hepcidin (Hamp1) mRNA expression (A) and pSmad1/5/8 (B) were measured under basal or BMP6 treated condition (AS1842856, 10 μM, 7 hours; BMP6, 20 ng/mL, 6 hours; cells were treated with AS1842856 1 hour before the addition of BMP6). (C-D) HepG2 cells were treated with the Foxo1 specific inhibitor AS1842856; hepcidin (HAMP) mRNA (C) and pSmad1/5/8 (D) were measured at indicated time duration (left) and quantified (right). Cells were treated with 10 μM AS1842856 1 hour before the addition of 20 ng/mL BMP6. (E) Primary hepatocytes were treated with the Foxo1-specific inhibitor AS1842856; holo-Tf-induced Hamp1 mRNA expression was measured (AS1842856, 10 μM, 7 hours; holo-Tf, 30 μM, 6 hours; cells were treated with AS1842856 1 hour before the addition of holo-Tf). (F-G) Primary hepatocytes were transfected with the pCDNA3.1 vector containing human wild-type FOXO1 or the constitutively active form of FOXO1 (FOXO1-AAA) for 24 hours; Hamp1 expression (F) and pSmad1/5/8 (G) were measured under basal and BMP6 treated conditions. Cells were treated with 20 ng/mL BMP6 for 6 hours before the end of plasmids transfection. Primary hepatocytes were isolated from 8-week-old male wild-type 129S2/SvPasCrl mice. Results from qPCR were obtained from 3 biological replicates. Quantification of western blot was obtained from 3 independent experiments. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001. The P value was calculated by the 1-way ANOVA with the Sidak multiple comparison test.
Figure 2.

Foxo1 modulates hepcidin expression in hepatocytes. (A-B) Primary hepatocytes treated with the Foxo1 specific inhibitor AS1842856; (A) hepcidin (Hamp1) mRNA expression (A) and pSmad1/5/8 (B) were measured under basal or BMP6 treated condition (AS1842856, 10 μM, 7 hours; BMP6, 20 ng/mL, 6 hours; cells were treated with AS1842856 1 hour before the addition of BMP6). (C-D) HepG2 cells were treated with the Foxo1 specific inhibitor AS1842856; hepcidin (HAMP) mRNA (C) and pSmad1/5/8 (D) were measured at indicated time duration (left) and quantified (right). Cells were treated with 10 μM AS1842856 1 hour before the addition of 20 ng/mL BMP6. (E) Primary hepatocytes were treated with the Foxo1-specific inhibitor AS1842856; holo-Tf-induced Hamp1 mRNA expression was measured (AS1842856, 10 μM, 7 hours; holo-Tf, 30 μM, 6 hours; cells were treated with AS1842856 1 hour before the addition of holo-Tf). (F-G) Primary hepatocytes were transfected with the pCDNA3.1 vector containing human wild-type FOXO1 or the constitutively active form of FOXO1 (FOXO1-AAA) for 24 hours; Hamp1 expression (F) and pSmad1/5/8 (G) were measured under basal and BMP6 treated conditions. Cells were treated with 20 ng/mL BMP6 for 6 hours before the end of plasmids transfection. Primary hepatocytes were isolated from 8-week-old male wild-type 129S2/SvPasCrl mice. Results from qPCR were obtained from 3 biological replicates. Quantification of western blot was obtained from 3 independent experiments. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001. The P value was calculated by the 1-way ANOVA with the Sidak multiple comparison test.

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