LA ameliorates iron burden in Hbb^th3/+ mice. Four-week-old Hbb^th3/+ mice were treated with intravenous LA (150 mg/g body weight) or vehicle for 4 weeks (n = 4-5 mice per group). (A-C) Serum hepcidin levels (A) and serum (B) and hepatic tissue (C) iron contents of wild-type (WT) and Hbb^th3/+ mice in response to LA administration. (D) Immunohistochemical staining of FPN on duodenum sections (×100) from Hbb^th3/+ mice after LA administration, with quantitative analysis in the right panel. (E) Enhanced DAB iron staining of liver (×200) and duodenum (×200) sections from Hbb^th3/+ mice with LA treatment. Brown represents iron deposition (denoted by yellow arrows). (F) Western blotting of Smad1, P-Smad1/5/8, CREB1, P-CREB1, FPN, and FTL in liver specimens from LA-treated mice relative to untreated control, with quantitative analysis in the right panel. (G) Spleen morphology (the upper panel) and spleen weight per body weight (the lower panel) of WT and Hbb^th3/+ mice treated with LA or solvent. (H) Blood smears stained with Wright-Giemsa stain, with black arrows indicating damaged or deformed erythrocytes (×1000) and quantification of normal cells. (I) Reticulocyte counts were performed using new methylene blue staining with red arrows indicating positive cell (×1000) and quantification of positive cells. (J) Representative erythropoiesis profiles of bone marrow from Hbb^th3/+ mice treated with LA or PBS. (K) Index of splenic erythropoiesis (as calculated by spleen weight × % spleen erythroid cells determined in flow cytometry analysis). ∗P < .05 and ^#P < .001, relative to untreated WT mice or as indicated. Data are represented as mean ± standard deviation. Baso, basophilic erythroblasts; Ortho, orthochromatic erythroblasts; Poly, polychromatic erythroblasts; Pro, proerythroblasts; Retic, reticulocytes.