Role of PI3Kβ and Pyk2 in platelet adhesion and spreading through integrin α2β1. (A) Wild-type and PI3Kβ^KD platelets were allowed to adhere to immobilized monomeric collagen for 10 or 60 minutes. Adherent cells were permeabilized, stained with TRITC-phalloidin, and adhesion (as number of cells/mm²) and spreading (as mean platelet area) were evaluated as described in “Methods.” (i) Representative image of adherent wild-type (WT) and PI3Kβ^KD platelets after 60 minutes (40× amplification). Quantification of adhesion and spreading is reported in panels ii and iii, respectively. Data are the means ± SD of 3 different experiments. (B) Integrin α2β1–mediated adhesion (i) and spreading (ii) of platelets preincubated with wortmannin (100nM, 15 minutes), TGX-221 (0.5μM, 10 minutes), or AS252424 (0.5mM, 10 minutes), as indicated on the right, after 10 or 60 minutes, as indicated on the bottom. Results are expressed as means ± SD of 3 different experiments. (C) Integrin α2β1–mediated adhesion (i) and spreading (ii) of platelets from wild-type (WT) and Pyk2-knockout (Pyk2 KO) after 10, 30, or 60 minutes. Results are expressed as means ± SD of 3 different experiments.