Role of Rho kinase in regulating thrombus contraction. (A) Lepirudin-anticoagulated human whole blood, spiked with DiIC12-labeled platelets, was preincubated with vehicle (DMSO), H1152 (40 μM), or HA1077 (80 μM), prior to perfusion through collagen-coated microslides at 1800 s−1. The interplatelet distance between firmly adherent platelets was quantified and used as an indirect marker of thrombus contraction. Results represent the mean plus or minus SEM (n = 4; *P < .05; **P < .005; ***P < .001). (B) Thrombus formation in the presence of vehicle (DMSO) or H1152 (40 μM) was recorded in real time, and snapshots of individual thrombi at the indicated times were taken off-line. The original size of the thrombus is outlined by a solid line, while the resultant thrombus size following 2 minutes of flow is outlined by a broken line. These images are taken from 1 representative of 4 independent experiments. (C) To examine the effect of Rho kinase inhibitors on fibrin-dependent clot retraction, citrated PRP was preincubated with vehicle (DMSO), H1152 (40 μM), HA1077 (80 μM), or c7E3 (100 μg/mL), followed by addition of thrombin (1.0 U/mL). The extent of clot retraction was assessed after 30 minutes, as described in “Platelet-mediated fibrin clot retraction.” Results represent the mean plus or minus SEM (n = 3).
