Confounding effect of therapeutic protamine and heparin levels on routine and special coagulation testing.

: The management of a patient with hemophilia undergoing cardiovascular surgery relies on accurate coagulation test results. Both unfractionated heparin (UFH) and protamine sulfate used during cardiac surgery can interfere with factor and inhibitor assays. Here we describe the effects of UFH and protamine sulfate on routine coagulation, factor activity, and inhibitor assays. Pooled normal plasma (PNP) with UFH, PNP with protamine sulfate, PNP with both protamine sulfate and UFH were tested for the activated partial thromboplastin time (aPTT), prothrombin time (PT), thrombin time (TT), UFH anti-Xa, one-stage factor VIII (FVIII) activity, one-stage factor IX (FIX) activity, and Bethesda inhibitor assays for FVIII and FIX. UFH had a dose-dependent effect with TT, aPTT, and PT. On Bethesda inhibitor testing, FIX inhibition was detected at 1 U/ml UFH and 3 U/ml UFH for FVIII. Increasing protamine sulfate concentration in PNP prolonged the PT and aPTT in a dose-dependent manner, decreased FVIII and FIX activity and did not affect TT or UFH anti-Xa. At protamine sulfate doses of at least 200 µg/ml there was weak FVIII and FIX inhibition detected. At lower ratios of protamine sulfate to UFH (0.6 : 1-0.8 : 1), the aPTT decreased, suggesting reversal of UFH. However, at protamine sulfate to UFH ratios of 1.0 : 1 and higher, aPTT prolongation was observed. Inhibition of FVIII and FIX was detected at low ratios of protamine sulfate to UFH (below 0.4 : 1) and disappeared at higher ratios. UFH and protamine sulfate, alone or in combination, impact factor activity and inhibitor testing for both FVIII and FIX. Hence, factor activity and inhibitor assay results should be interpreted with caution when UFH or protamine sulfate are present.

Treatment and prevention of bleeding in congenital hemophilia A patients with inhibitors.

The treatment of bleeding in hemophilia A patients with persistent inhibitory antibodies to factor VIII is problematic. The current standard hemostatic agents for inhibitor patients are recombinant activated factor VII (rFVIIa) and activated prothrombin complex concentrate (APCC). These "inhibitor bypassing agents" are less reliably effective than are replacement therapies for patients without inhibitors, and there are no validated laboratory assays to monitor their efficacy. Furthermore, only single rFVIIa and APCC products are available worldwide, and their use can be complicated, albeit rarely, by thrombotic events. For all these reasons, new approaches to treat bleeding in inhibitor patients are eagerly awaited. These new approaches include replacement therapy with porcine factor VIII concentrate (currently approved for use in acquired hemophilia patients), bispecific antibodies to simulate the biologic function of factor VIII (already in use in some jurisdictions), pegylated forms of activated factor VII, and strategies targeting the natural anticoagulants TFPI and antithrombin, which create a hypercoagulable phenotype to counterbalance the hypocoagulability imposed by hemophilia.

Current status and future prospects for the prophylactic management of hemophilia patients with inhibitor antibodies.

Inhibitor antibodies to factor VIII arise in a substantial minority of patients with hemophilia A treated with replacement therapy; factor IX inhibitors in treated hemophilia B patients are considerably less common. As replacement therapy is not feasible in most such patients, hemostasis can generally only be achieved with "inhibitor bypassing agents", namely (activated) prothrombin complex concentrates and recombinant factor VIIa. These agents are widely used to treat active bleeding in inhibitor patients but they have been used relatively infrequently as prophylactic agents to prevent bleeding and its consequences, mainly progressive joint damage. This is in contrast to hemophilia patients without inhibitors, in whom prophylactic replacement with concentrates of factor VIII or IX has become widely accepted as the optimal strategy to prevent these adverse outcomes. This review addresses the current experience and evidence and the future prospects regarding prophylaxis in inhibitor patients.

Phosphatidylserine exposure and other apoptotic-like events in Bernard-Soulier syndrome platelets.

In the Bernard-Soulier syndrome (BSS), the giant platelets are said to have increased phosphatidylserine (PS) surface exposure in the resting state and shortened survival in the circulation. When normal platelets are activated, they undergo many biochemical and morphological changes, some of which are apoptotic. Herein, we investigated apoptotic-like events in BSS platelets upon activation, specifically, PS exposure, microparticle (MP) formation, cell shrinkage, and loss of mitochondrial inner membrane potential (DeltaPsi(m)). Platelets from two unrelated BSS patients were examined in whole blood; agonists used were collagen, thrombin, PAR1- or PAR4-activating peptides (APs), or combinations of collagen with thrombin, and the PAR-APs. Flow cytometry was used to measure PS exposure (annexin A5 binding), platelet-derived MPs (forward scatter; events <0.75 microm size), and DeltaPsi(m) (TMRM fluorescence). PS exposure was increased on resting and activated BSS platelets, and this was independent of the platelet size. MP formation by BSS platelets was generally enhanced. Cell shrinkage occurred on activation to form smaller, PS-exposing platelets in BSS and controls. A proportion of PS-exposing BSS and control platelets exhibited DeltaPsi(m) loss, but unlike controls, there was also loss of DeltaPsi(m) in the BSS platelets not exposing PS. Thus, BSS platelets undergo apoptotic-like events upon activation, with PS exposure and MP formation being enhanced. These events may play a role in the shortened survival in BSS, as well as affecting thrombin generation.

Vascular injury and thrombotic potential: a note of caution about recombinant factor VIIa.

Postoperative hemorrhage following cardiac surgery increases morbidity, mortality, and costs. Several case reports have described the successful use of recombinant factor VIIa to decrease or stop bleeding in patients undergoing cardiac surgery. The mechanism of action of recombinant factor VIIa is thought to be increased site-specific thrombin generation by tissue factor-mediated activation of coagulation or from activated platelets. However, there have also been many reports of thrombotic complications after recombinant factor VIIa administration. Randomized clinical trials and further laboratory studies should help better clarify the efficacy, safety, cost-effectiveness, and optimal dosing of recombinant factor VIIa in the cardiac surgical setting.

Thr325Ile polymorphism of the TAFI gene is related to TAFI antigen plasma levels and angiographic restenosis after percutaneous coronary interventions.

BACKGROUND: The fibrinolytic system is closely related to several processes that are involved in restenosis. We have previously shown that high pre-procedural plasma levels of thrombin-activatable fibrinolysis inhibitor (TAFI) antigen predicted angiographic restenosis. The aims of this study were to evaluate the relationship between Thr325Ile polymorphisms, plasma levels of TAFI antigen, and late angiographic restenosis after percutaneous coronary intervention (PCI). METHODS: We prospectively studied 159 patients with stable angina who underwent successful elective angioplasty or stenting of de novo native coronary artery lesions. Blood samples were drawn before the procedure. TAFI antigen levels were measured, as well as the presence of TAFI Ile325Thr genetic variation. Follow-up coronary angiography was performed in 90% of patients. RESULTS: The genotypes based on Ile325Thr substitution had significantly different TAFI antigen levels: genotype C/C>C/T>T/T (111+/-29%, 87+/-24%, and 59+/-22%, respectively, p<0.006). T/T genotype was associated with lower rates of restenosis compared to C/T and C/C genotypes (25% versus 37% and 33%, respectively, p<0.05). CONCLUSIONS: These data suggest that plasma TAFI antigen levels are genetically controlled. The T/T Thr325Ile polymorphism of the TAFI gene is associated with lower plasma levels of TAFI antigen and lower restenosis rates after PCI.

Thrombin-activatable fibrinolysis inhibitor (TAFI): a novel predictor of angiographic coronary restenosis.

The fibrinolytic system is closely related to several processes that are involved in restenosis. We previously showed that low PAI-1 plasma levels predicted restenosis. Recently, a different fibrinolytic inhibitor, TAFI, has been described. The aims of this study were to evaluate the relationship between pre-procedural plasma levels of TAFI and late angiographic restenosis and the interaction between TAFI and PAI-1.We prospectively studied 159 patients with stable angina who underwent successful elective angioplasty or stenting of de novo native coronary artery lesions. TAFI and PAI-1 antigen levels were measured in plasma samples drawn before the procedure. Follow-up coronary angiography was performed in 92% of patients. There was a significant correlation between pre-procedural TAFI levels and 6-month % diameter stenosis (DS) (r = 0.21; p = 0.013). The overall angiographic restenosis rate (DS>50%) was 31%. Pre-procedural TAFI levels were significantly higher in patients with restenosis (108 +/- 33% versus 94+/-30%, p = 0.011). Restenosis rates for patients in the upper tertile of TAFI levels were 2-fold higher than for those in the lowest tertile (45% versus 22%; p = 0.016). A combination of high TAFI and low PAI-1 levels identified patients at the highest risk of restenosis (53%) compared to 14% in patients with low TAFI and high PAI-1 levels; p = 0.027. In conclusion, pre-procedural plasma TAFI antigen levels identify patients at increased risk for restenosis after PCI.

Successful treatment using recombinant factor VIIa for severe bleeding post cardiopulmonary bypass.

PURPOSE: To describe a case of persistent and excessive bleeding following an aortic valve and ascending aorta replacement that was successfully managed with recombinant factor VIIa (rFVIIa). The postulated mechanisms for rFVIIa are discussed. CLINICAL FEATURES: A 75-yr-old female with no preoperative coagulopathy underwent a tissue aortic valve replacement and supracoronary ascending aorta replacement for severe aortic stenosis and an ascending aortic aneurysm. Following surgery, she bled in excess of 200 mL x hr(-1) despite a nearly normal platelet count and nearly normal coagulation parameters. The patient was surgically re-explored twice in seven hours, and despite the presence of near normal in vitro coagulation parameters, the patient continued to bleed. Multiple units of fresh frozen plasma, platelets and cryoprecipitate were administered empirically. We then administered a single 6-mg (107 microg x kg(-1)) iv dose of rFVIIa. Following the administration of rFVIIa, blood loss decreased to a total of 440 mL over the next 12 hr. CONCLUSIONS: This case describes the use of rFVIIa for intractable bleeding postcardiovascular surgery in the presence of nearly normal laboratory markers of coagulation. Further controlled laboratory and clinical studies are required to define the role of rFVIIa in patients undergoing cardiovascular surgery.