Platelets promote release of BMDCs during angiogenesis. (A) To induce ischemia, ligation of the femoral artery was performed and the experiment was terminated 14 days later (n = 4). In a separate group of mice, tumor cells were implanted subcutaneously: B16-F10 (2 × 106 cells, n = 8), RM1 (4 × 105 cells, n = 5), or LNCaP-C4–2 (4 × 105 cells, n = 4), and excised after 9 days (B16-F10), 12 days (RM1), or 28 days (LNCaP-C4–2). Blood samples were collected from mice before tumor implantation or ischemic surgery (Initial, white columns) and on experimental termination (Final, black columns). (B-D) After tumor implantation or hindlimb ischemia surgery, 3 × 109 platelets were infused into mice every 5 days by tail vein injection. Control mice were injected with phosphate-buffered saline (PBS). Separately, mice were treated intravenously with 2 μg/g body weight rat anti–mouse GPIbα to deplete platelets or rat IgG as a control. Injections were repeated every 3 days. (B) Whole blood was collected before (Initial, white columns) and 9 days after (Final, black columns) B16-F10 tumor implantation (n = 6). (C-D) Whole blood was collected before ischemic surgery (Initial, white columns), from IgG or PBS-treated mice (Ischemia, gray columns), and from mice after platelet (PLT) infusion or depletion (black columns) (n = 6). Red blood cells were lysed, and the sample was labeled with CXCR4 antibody conjugated with phycoerythrin and analyzed by flow cytometry. Values are mean percentage of CXCR4+ cells ± SEM. *P < .05, **P < .01, and ***P < .005 by Student t test (A-B) or 1-way analysis of variance (C-D) versus initial samples.
