Elevated BM S1P₁ levels during G-CSF–induced mobilization. (A-E) Mice were injected with PBS (control), G-CSF (5 consecutive subcutaneous injections and evaluation 3-5 hours after the last injection), G-CSF+FTY720 (5 consecutive intraperitoneal injections of both agents), G-CSF+Rapamycin (5 consecutive subcutaneous injections of both agents) or G-CSF+Akt inhibitor (5 consecutive intraperitoneal injections together with G-CSF). A representative histogram (A left) and a total cell analysis (A right) of S1P₁ receptor expression on BM MNC are presented. The percentage of S1P₁⁺/c-Kit⁺/Lin⁻ cells in the BM: (Bi) representative FACS plots of Lin⁻ BM-MNC with the percentage of positive cells from 1 experiment written in each plot and (Bii) total BM-MNC cell analysis are shown. The numbers of WBCs (C), primitive SKL cells (D bars) and CFU-Cs (D dots) per milliliter of blood are shown. BM SKL numbers per 1 × 10⁶ total BM cells (E) are presented. (F-H) WT and S1P₁ conditional deficient mice (S1P₁ KO) were injected with PBS (control), AMD3100 (single subcutaneous injection and evaluation after 1 hour) or G-CSF (5 consecutive subcutaneous injections and evaluation 3-5 hours after the last injection). The numbers of WBCs (F), primitive SKL cells (G), and CFU-Cs (H) per milliliter of blood are shown. Indicated values are presented as mean ± SE in 3 independent experiments, a total of 6 mice in each group (*P < .05, **P < .01).