The effect of uremia on platelet contractile force, clot elastic modulus and bleeding time in hemodialysis patients.

INTRODUCTION: Uremic bleeding frequently occurs in dialysis patients. Although its mechanism is not well characterized, acquired platelet dysfunction has been implicated in its pathogenesis. Skin bleeding time has been used to characterize platelet dysfunction in this population. However, the bleeding time is prone to error. The goal of this study was to compare the bleeding time to the novel platelet function parameters platelet contractile force and clot elastic modulus as well as platelet aggregation studies in controls and patients receiving maintenance hemodialysis. MATERIALS AND METHODS: Forty-five subjects completed this study (25 controls, 20 dialysis). All subjects had the Ivy skin bleeding time procedure performed, as well as the collection of whole blood samples for the determination of platelet contractile force, clot elastic modulus, % von Willebrand Factor antigen, and platelet aggregation studies. Pearson's correlation determined the relationships between skin bleeding time and platelet function and clot structure parameters and markers of renal dysfunction. RESULTS: Bleeding time was significantly prolonged in the dialysis group relative to controls. The platelet function parameters were not significantly different between groups. There was a significant relationship between bleeding time and creatinine concentration, however, no relationship existed between bleeding time and platelet function parameters. CONCLUSIONS: Skin bleeding time poorly correlates with measurements of platelet function. There were no significant differences noted in platelet function between the groups despite the prolongations in bleeding time in the dialysis group. These data may suggest that the bleeding time reflects perturbations in platelet adhesion or secretion, and not aggregation. Further study is needed to characterize platelet function in dialysis patients.

Effect of non-heparin thrombin antagonists on thrombin generation, platelet function, and clot structure in whole blood.

Platelet contractile force (PCF), which is absent in blood obtained during cardiopulmonary bypass, significantly recovers after protamine sulfate administration. In vitro studies reveal this effect to be primarily caused by heparin. Because many of heparin's effects are mediated by suppression of thrombin generation and activity, this study assessed the influence of thrombin inhibition on PCF. The effects of natural and synthetic antithrombins were measured. Clots were formed by the addition of batroxobin (0.21 microg/mL) to whole blood (platelet count 200,000/microL). Force development was measured from the moment of batroxobin addition. After 1200 s of clotting, purified antithrombin III (22 microM) reduced PCF by 74%. Thrombomodulin (0.014 microM) reduced PCF by 60%. At 0.040 microM, PCF was reduced by 82% (6.5-1.2 Kdynes). Hirudin decreased PCF in a dose-dependent fashion, with complete suppression at concentrations > or = 0.30 microM. At concentrations between 0.04 and 0.29 microM, Lepirudin (Refludan, a recombinant therapeutic hirudin) produced dose-dependent delay and suppression of PCF. Above 0.29 microM Lepirudin, PCF was totally suppressed. At 1.60 microM, bivalirudin (a synthetic, 20 amino acid peptide) delayed and reduced PCF by 50%. At 6.40 micro;M, PCF was completely suppressed. Although 20 microM of P-PACK II (d-Phenylalanyl-L-Phenylalanylarginine- chloro-methyl ketone 2 HCl) had little effect, 40 microM delayed onset of force development from 300 to 600 s and reduced PCF at 1200 s from 5.2 to 3.3 Kdynes. At 120 microM, force development was totally suppressed. Four micromol Thromstop (BNas-Gly-(pAM)Phe-Pip) delayed force development by greater than 800 s and PCF at 1200 s was reduced by 70%. At 0.20 microM, Argatroban (a synthetic polypeptide direct thrombin antagonist) delayed onset of PCF from 300 to 540 s and decreased PCF by 40%. At a concentration of 0.40 microM and above, Argatroban totally suppressed PCF. These results indicate that some of the antiplatelet effects of heparin are the result of thrombin inhibition and that low-level thrombin generation is essential for clot retraction. The sensitivity of PCF to the presence of thrombin may permit monitoring of antithrombin agents via this assay.

Aprotinin counteracts heparin-induced inhibition of platelet contractile force.

BACKGROUND: Aprotinin interferes with heparin binding to platelets and decreases blood loss during cardiopulmonary bypass (CPB). Heparin abolishes platelet force during CPB, and the extent of platelet force recovery after protamine administration appears to correlate with blood loss. This study assessed the effect of aprotinin on heparin suppression of platelet force. METHODS: Platelet force was measured using the Hemodyne Hemostasis Analyzer. Clots were formed from platelet-rich plasma (PRP) by the addition of batroxobin and 10 mM CaCl(2). Clotting conditions included pH 7.4, ionic strength 0.15 M, fibrinogen level 1 mg/ml and 75,000 platelets/microl. RESULTS: After 1200 s of clotting, force was reduced from 7110+/-1190 to 450+/-450 dyn by 0.2 U/ml of heparin. Platelet force in aprotinin [20 microg/ml (140 KIU/ml)] containing PRP was not suppressed by heparin addition (7480+/-2410 dyn). Aprotinin [40 microg/ml (280 KIU/ml)] addition to previously heparinized plasma counteracted heparin force suppression. Aprotinin (40 microg/ml) increased platelet force from 5630 to 11,138+/-562 in PRP devoid of heparin. Aprotinin did not affect thrombin activity, fibrin structure, platelet aggregation or secretion. CONCLUSIONS: Aprotinin counteracts heparin suppression of platelet force and enhances platelet force in the absence of heparin. Aprotinin-heparin-platelet interactions may help explain aprotinin's ability to reduce blood loss during CPB.

Enhanced platelet force development despite drug-induced inhibition of platelet aggregation in patients with thromboangiitis obliterans--two case reports.

Thromboangiitis obliterans (TAO) is a nonatherosclerotic, nonnecrotizing, nonspecific, segmental inflammatory obliterative vasculitis, characterized by decreased flow to the distal extremities and increased risk of amputation. While smoking cessation is viewed as critical to successful treatment, various therapeutic options have been employed. While many treatment regimens seek to diminish platelet function, there are relatively few studies of platelet function in this disease entity and even fewer that have offered evidence of increased platelet activity. The authors report here 2 cases of TAO in which evaluations for hypercoagulable states and of platelet function were performed. Platelet contractile force (PCF) was found to be 82% higher than a normal control in 1 TAO patient and 340% higher than normal in the second patient. This was true despite the fact that platelet aggregations confirmed suppression of aggregation by antiplatelet medications. Elevated PCF has been seen in a variety of conditions, such as coronary artery disease and diabetes mellitus, in which endothelial function is abnormal. Whether high PCF values play a role in the pathogenesis of these diseases or simply serve as markers of enhanced platelet function and/or endothelial dysfunction awaits additional evaluations.