VEGFR1 and VEGFR2 are expressed by Sca1⁺endothelial cells in the BM. BM cells were extracted from the endosteal region, enriched in stromal cells by magnetic depletion of CD45⁺ leukocytes and Ter119⁺ erythroid cells, and stained for hematopoietic, endothelial, and mesenchymal markers together with anti-VEGFR1 and anti-VEGFR2 fluorescent antibodies. (A) Gating strategy to identify endothelial and mesenchymal cells. Following the exclusion of residual CD45⁺ and Ter119⁺ leukocytes and erythroid cells, nonhematopoietic CD45⁻Ter119⁻ cells were gated as CD31⁺Sca1⁻ endothelial cells and CD31⁺Sca1⁺ endothelial cells. Remaining CD45⁻Ter119⁻CD31⁻ mesenchymal cells were gated as PDGFRα⁻Sca1⁻, PDGFRα⁺Sca1⁻, PDGFRα⁺Sca1⁺, and PDGFRα⁻Sca1⁺ cells. (B) Typical dot-plots showing VEGFR1 and VEGFR2 expression on previously defined endothelial and mesenchymal populations. (C) Percentage of endothelial and mesenchymal BM cells expressing VEGFR1 and/or VEGFR2. Bar graphs show mean ± standard deviation of 4 mice. (D) Kdr (VEGFR2) mRNA expression by qRT-PCR on CD11b⁺F4/80⁺VCAM1⁺CD169⁺ macrophages (CD11b+Mφ), CD11b⁻F4/80⁺VCAM1⁺CD169⁺ macrophages (CD11b-Mφ), CD11b⁺F4/80⁺VCAM1⁻CD169⁻ monocytes (MO), CD11b⁺F4/80⁻Ly6G⁺ granulocytes (Gran), CD45⁻Ter119⁻CD31⁺ endothelial cells (EC), CD45⁻Ter119⁻CD31⁻PDGFRα^+/−Sca1⁺ mesenchymal progenitor cells (P±S+), PDGFRα⁺Sca1⁻ mesenchymal progenitor cells (P+S-), and CD45⁻Ter119⁻CD31⁻PDGFRα⁻Sca1⁻ stromal cells (P-S-) sorted from the BM of mice treated with saline (C), FG-4497 alone (F), G-CSF (G), or G-CSF plus FG-4497 (G+F) for 3 days. Each dot represents a separate mouse and separate sort. Data are relative to Hprt mRNA. The P values were calculated using ANOVA.