LS174T cell tethering.

(A) LS174T cell adhesion to surface-adherent platelets under conditions of flow (0.8 dyn/cm²)—effects of mAbs and enzymes. Closed bars represent the stable adherent cells (stationary for > 10 seconds) counted at the end of the 10-minute experiment, whereas open bars represent the total interacting cells counted throughout the entire 10-minute experiment. § P < .05 with respect to anti-α_IIbβ₃ sample. Values are mean ± SEM of 3-31 experiments. (B) LS174T cell rolling over purified P-selectin at a wall shear stress of 0.8 dyn/cm². Data are expressed as percentage of untreated (control) LS174T cells that interacted with recombinant P-selectin throughout the 10-minute experiment. *P < .05 with respect to no-treatment control. Values are mean ± SEM of 4-5 experiments. Immobilized platelets were treated with XV454 (blocking α_IIbβ₃ function) during the 10-minute thrombin incubation. LS174T cells were incubated with PL1 (blocking anti-PSGL-1; 10 μg/mL) for 10 minutes before their perfusion over platelet layers. PL1 saturating concentrations were also maintained in the perfusion buffer. Alternatively, LS174T cells were treated with PI-PLC (1 U/mL for 1 hour at 37°C) or neuraminidase (0.1 U/mL for 30 minutes at room temperature) before infusion to the flow chamber. Purified P-selectin was incubated with an anti-P-selectin mAb (AK4) for 10 minutes before LS174T cell perfusion.