Induction of anemia in adult mice does not result in the expression of Glut1 on differentiating erythroblasts. (A) The spleen of an adult mouse (control) is shown in comparison with those obtained from mice wherein anemia was induced either by phenylhydrazine treatment or phlebotomy of 0.5 mL blood on days 0, 1, and 3. Spleens were obtained from killed mice at day 6 after treatment. (B) Erythroblast differentiation was assessed in freshly isolated splenocytes obtained from control adult mice; 6 days following induction of anemia as indicated, differentiation was assessed by staining with CD71 and Ter119 antibodies. The percentages of cells in each region (regions I to IV) are indicated in the respective dot plot. The relative expression of Glut1 in each of the 4 regions was determined by gating on the identified CD71/Ter119 populations. Shaded histograms show nonspecific staining. (C) Glut1 expression in peripheral red blood cells from newborn mice, adult mice, and adult mice rendered anemic by phenylhydrazine (PHZ) treatment (mice numbered 1 and 2) was assessed by immunoblotting using an anti-Glut1 pAb. Protein loading was monitored by immunoblotting for actin. (D) Glut1 transcripts were assessed in Ter119⁺ splenic erythroid progenitors isolated from newborn, adult, and PHZ-treated anemic mice (numbered 4 to 6; day 6), as indicated, by qRT-PCR. cDNAs were amplified with primers specific for Glut1 and means plus SD of duplicate samples normalized to GAPDH are shown.