Figure 5.

PKC-β recruitment to αIIbβ3 is dependent on AA/thromboxane A₂ generation by cPLA₂α. Where indicated, platelets were treated with 300 μg/mL of human fibrinogen + 1 mM MnCl₂ for 30 minutes at 37°C. Immunoprecipitation was carried out with an anti-β3 antibody or normal rabbit serum (NRS) as a control as indicated in “Methods.” (A) Platelets were collected from 9 mice for each background and resuspended in Walsh buffer at a final concentration of 8 × 10⁸/mL. Platelets were then treated with fibrinogen/MnCl₂ in the presence or absence of 50 μM of pyrrophenone. The experiment was performed 3 times. (B) Washed human platelets were incubated in the presence or absence 50 μM of pyrrophenone and fibrinogen/MnCl₂, fibrinogen/MnCl₂ + 100 μM of AA, or fibrinogen/MnCl₂ + 10 μM of U46619. This experiment was performed twice. (C) Washed human platelets were preincubated with 50 μM of pyrrophenone, 0.18 g/L of aspirin, 10 μM of SQ29,548, 100 μM of BAPTA-AM, 100 μM of 2-APB, or 1 U/mL of apyrase for 15 minutes at room temperature. The results are representative of 4 separate experiments.