FIGURE 4.

Increased erythropoiesis in Fgf-23^−/− mice. A, colony-forming assay for erythroid (BFU-E) progenitors (WT, n = 5; Fgf-23^−/−, n = 12). Cells from each mouse were plated in duplicate, and the number of colonies in each plate was counted. B, Epo concentrations measured by ELISA in serum of WT (n = 12) and Fgf-23^−/− mice (n = 8). Samples were measured in duplicate. C, quantitative real time RT-PCR showing changes in Epo mRNA expression in bone marrow (WT, n = 7; Fgf-23^−/−, n = 5), bone (inset) (WT, n = 5; Fgf-23^−/−, n = 6), liver (WT, n = 7; Fgf-23^−/−, n = 7), and kidney (WT, n = 7; Fgf-23^−/−, n = 6) in WT and Fgf-23^−/− mice. Bone graph is amplified in the inset because of small values compared with other tissues. D and E, quantitative real time RT-PCR showing changes in HIF-1α (D) and HIF-2α (E) mRNA expression in bone marrow, bone, liver, and kidney in WT and Fgf-23^−/− mice (WT, n = 7; Fgf-23^−/−, n = 7). F–J, oxygen treatment. WT (n = 4) and Fgf-23^−/− (n = 5) mice were subjected to 100% oxygen at 4 liters/min for 1 h. F, Epo concentrations measured by ELISA in serum of WT (n = 4) and Fgf-23^−/− mice (n = 5). Samples were measured in duplicate. G–I, quantitative real time RT-PCR showing changes in renal HIF-1α (G), renal EPO (H), and BM HIF-1α mRNA expression (I) in WT and Fgf-23^−/− mice (WT, n = 4; Fgf-23^−/−, n = 5). J, flow cytometry analysis of bone marrow from 6-week-old WT and Fgf-23^−/− mice with or without oxygen treatment. A percentage of mature erythroid cells stained positive for Ter119 (WT, n = 4; Fgf-23^−/−, n = 5). The data are represented as mean fold change ± S.E. relative to housekeeping gene HPRT. *, p < 0.05; **, p < 0.01; ***, p < 0.001.