PI3Kβ, and not PI3Kα, is required for mouse and human thrombus stability ex vivo under high shear rate. (A) DIOC₆-labeled platelets in whole blood from p110β^flox/flox control (WT), PF4-Cre/p110β^flox/flox (p110β^null) mice (left) or human preincubated with the PI3Kβ inhibitor (AZD6482) (right) were perfused through a collagen-coated microcapillary at a physiological arterial shear rate of 1500 seconds⁻¹, followed by a high shear rate of 4000 seconds⁻¹. Thrombi volumes (µm³) were analyzed using ImageJ software. Graphs represent the averages of 6 independent experiments and are mean ± SEM; Student t test ***P < .001. (B) Whole blood from p110β^flox/flox control (WT), PF4-Cre/p110β^flox/flox (p110β^null), or PF4-Cre/p110α^flox/flox (p110α^null) mice were perfused through a collagen-coated microcapillary at a physiological shear rate of 500 seconds⁻¹ and then at a high shear rate of 4000 seconds⁻¹. Thrombi volumes (µm³) were analyzed using ImageJ software. Results shown are mean ± SEM of 4 to 5 independent experiments. Student t test, ***P < .001. Representative images of mouse thrombi formed after physiologic shear rate (500 seconds⁻¹) (left) followed by high shear (4000 seconds⁻¹) (right) are shown. Scale bar, 20 µm. (C) DIOC₆-labeled platelets in whole blood from p110β^flox/flox control (WT) or PF4-Cre/p110β^flox/flox (p110β^null) mice were perfused through a collagen-coated microcapillary directly at a high shear rate of 4000 seconds⁻¹ during 3 minutes. Fast Z stack recording was performed every 10 seconds and thrombi volumes (µm³) were analyzed for each time point using ImageJ software. Results shown are mean ± SEM of 4 independent experiments; Student t test *P < .05; ^$P < .001. (D) DIOC₆-labeled whole blood from p110β^flox/flox control (WT) or PF4-Cre/p110β^flox/flox (p110β^null) mice were treated with the α_IIbβ₃ antagonist integrilin and were perfused through a collagen-coated microcapillary at a physiological shear rate of 500 seconds⁻¹ and then at a high shear rate of 4000 seconds⁻¹. Surface covered (%) was analyzed in both conditions. Results shown are mean ± SEM of 3 separate experiments.