Engagement of PECAM-1 leads to Akt activation.

(A) Akt activation evaluated by Western blot with mAbs recognizing unphosphorylated (Akt; bottom) or phosphorylated Akt (P-Akt; top) in purified CD14⁺CD34⁺ cells after 2 minutes (lanes 2 and 4) or 4 minutes (lane 3) upon PECAM-1 ligation (incubation with 5 μg/mL of the specific anti–PECAM-1 mAb followed by 10 μg/mL GAM). Lane 1 shows untreated cells (exposed to GAM alone); lane 4, cells exposed to the PI-3K inhibitor LY294002. (B) Akt activation in cell lysates of CD14⁺CD34⁺ cells assessed with the Akt assay kit, using the specific substrate and³²γATP, after immunoprecipitation with anti–Akt antibody. CTR represents untreated cells. Ig+GAM, cells incubated with murine Ig plus GAM. PECAM-1 ligation was obtained as indicated above in the absence (nil) or in the presence of LY294002, wortmannin, rapamycin, or PD98059. Results are expressed as cpm ×10⁻³, means ± SDs of 4 independent experiments.