The impact of rivaroxaban on primary hemostasis in patients with venous thrombosis.

Factor Xa inhibitors are safe and effective alternatives to warfarin, but several studies indicate that rivaroxaban may cause a different risk profile for bleeding. For instance, while the risk of major bleeding in general may be lower with rivaroxaban than for warfarin, the risk of gastrointestinal bleeding or abnormal uterine bleeding may be higher. The underlying mechanisms for these differences are not known, and the effect of rivaroxaban on primary hemostasis is poorly understood. The aim of this study was to investigate the effect of rivaroxaban on platelet function, P-selectin and von Willebrand factor (VWF) antigen and activity. Patients with venous thrombosis assigned to 3 months of treatment due to temporary risk factors were included. Blood was collected both during (on-treatment) and 4-6 weeks after end of treatment (without treatment). The platelet reactivity was assessed by light transmission aggregometry. P-selectin was measured by an enzyme-linked immunosorbent assay and vWF antigen and activity by latex immunoagglutination assays. Platelet reactivity during on-treatment (trough- and peak concentration) was similar to values without treatment. There was a trend toward a reduction of P-selectin during rivaroxaban treatment (peak concentration) compared to value without treatment (p = 0.06). There were no differences in vWF antigen and activity between the different time-points. We found no difference in platelet reactivity or vWF antigen/activity during rivaroxaban treatment compared with values without treatment. Apart from possibly causing a reduction of P-selectin, rivaroxaban seems not to influence primary hemostasis.

The reversal effect of prothrombin complex concentrate (PCC), activated PCC and recombinant activated factor VII against anticoagulation of Xa inhibitor.

BACKGROUND: An increasing number of patients are treated with direct-acting oral anticoagulants (DOACs), but the optimal way to reverse the anticoagulant effect is not known. Specific antidotes are not available and prothrombin complex concentrate (PCC), activated PCC (aPCC) and recombinant factor VIIa (rFVIIa) are variously used as reversal agents in case of a major bleeding. We aimed to determine the most effective haemostatic agent and dose to reverse the effect of rivaroxaban in blood samples from patients taking rivaroxaban for therapeutic reasons. METHODS: Blood samples from rivaroxaban-treated patients (n = 50) were spiked with PCC, aPCC and rFVIIa at concentrations imitating 80%, 100% and 125% of suggested therapeutic doses. The reversal effect was assessed by thromboelastometry in whole blood and a thrombin generation assay (TGA) in platelet-poor plasma. Samples from healthy subjects (n = 40) were included as controls. RESULTS: In thromboelastometry measurements, aPCC and rFVIIa had a superior effect to PCC in reversing the rivaroxaban-induced lenghtening of clotting time (CT). aPCC was the only haemostatic agent that shortened the CT down to below the control level. Compared to healthy controls, patients on rivaroxaban also had a prolonged lag time and decreased peak concentration, velocity index and endogenous thrombin potential (ETP) in platelet-poor plasma. aPCC reversed these parameters more effectively than rFVIIa and PCC. There were no differences in efficacy between 80%, 100% and 125% doses of aPCC. CONCLUSIONS: aPCC seems to reverse the anticoagulant effect of rivaroxaban more effectively than rFVIIa and PCC by evaluation with thromboelastometry and TGA in vitro.

Systemic biomarkers of inflammation and haemostasis in patients with chronic necrotizing pulmonary aspergillosis.

BACKGROUND: The purpose of this study was to investigate mediators of inflammation and haemostasis in patients with chronic necrotizing pulmonary aspergillosis (CNPA), a locally, destructive process of the lung due to invasion by Aspergillus species. METHODS: Measurements of selected biomarkers in 10 patients with CNPA and 19 healthy, matched controls were performed with enzyme-linked immunosorbent assay (ELISA) and multiplex methodology. The gene expressions of relevant biomarkers were analyzed with real-time quantitative RT-PCR. RESULTS: Increased concentrations of circulating mediators of inflammation interleukin (IL)-6, IL-8, RANTES, TNF-α, ICAM-1 and mediators involved in endothelial activation and thrombosis (vWF, TF and PAI-1) were observed in patients with CNPA. The concentration of the anti-inflammatory cytokine IL-10 was increased both in plasma and in PBMC in the patient population. The gene expression of CD40L was decreased in PBMC from the patient group, accompanied by decreased concentrations of soluble (s) CD40L in the circulation. CONCLUSIONS: The proinflammatory response against Aspergillus may be counteracted by reduced CD40L and sCD40L, as well as increased IL-10, which may compromise the immune response against Aspergillus in patients with CNPA.

Thrombin generation and platelet activation related to subintimal percutaneous transluminal angioplasty.

OBJECTIVE: The purpose of this study was to measure the in vivo platelet activation and thrombin generation in arterial blood after passing a subintimal conduit. METHODS: Subintimal percutaneous transluminal angioplasty (SPTA) is a technique where a subintimal channel is created, allowing recanalization of long peripheral arterial occlusion. From 10 patients with intermittent claudication, undergoing successful SPTA for femoropopliteal occlusive disease, we collected antecubital venous blood samples immediately before treatment, preprocedural arterial blood samples taken at the entry level proximal to the vessel occlusion, and subsequently at the reentry level after successful recanalization. Venous follow-up blood samples were taken after 24 hours. Plasma concentrations of ß-thromboglobulin (ß-TG), RANTES, and Prothrombin fragment (F1 + 2), were determined by immunoassay. Fibrinogen binding to platelets, leukocyte-platelet adhesion, and P-selectin were determined by flow cytometry. RESULTS: We found a statistically significant transluminal increase in the plasma concentrations of RANTES, ß-TG and F1 + 2 (p = 0.002, 0.001 and 0.001 respectively), which all normalized within 24 hours. Platelet-leukocyte aggregates significantly decreased after 24 hours compared with preprocedural and preentry levels (3.26% versus 5.26 %, p = 0.017). P-selectin expression on circulating platelets was statistically significantly increased in the blood sample taken at the re-entry level compared with the pre-procedural and pre-entry level (p = 0.007). After 24 hours there was no statistically significant difference to pre-procedural levels. There was no significant change in platelet fibrinogen binding at any levels. CONCLUSION: When passing a subintimal conduit, in vivo sampled blood demonstrated an extremely rapid and substantial uniform platelet activation and thrombin generation.

The effect of emicizumab and bypassing agents in patients with hemophilia - An in vitro study.

BACKGROUND: Emicizumab is a nonfactor replacement therapy for hemophilia A (HA) and is a bispecific monoclonal antibody mimicking factor VIII by binding both factors IXa and X. Although it reduces the frequency of bleeding episodes, there is still need for bypassing agents in case of breakthrough bleeds or need for surgery. The HAVEN-1 study showed an increased risk of thrombotic events and episodes of thrombotic microangiopathic hemolytic anemia with simultaneous treatment with emicizumab and activated prothrombin complex concentrate (aPCC) in high doses (>100 U/kg daily) for more than 1 day, and it is suspected that these drugs have a synergistic hemostatic effect. OBJECTIVES: To evaluate and compare the hemostatic effect of bypassing agents in vitro in people with HA before and after starting treatment with emicizumab to investigate if dosing should be adjusted to optimize treatment. PATIENTS/METHODS: Blood collected before and after start of treatment with emicizumab was spiked with aPCC and recombinant factor VIIa (rFVIIa) at different concentrations. The effect of aPCC and rFVIIa was assessed by thrombin generation assay and thromboelastometry. RESULTS: Six people with HA were included. The response to aPCC in thrombin generation after starting emicizumab was significantly stronger than before. This synergistic effect was less pronounced for emicizumab and rFVIIa. Furthermore, aPCC shortened thromboelastometry clotting time more effectively after starting emicizumab than before starting this treatment. CONCLUSIONS: We demonstrated a strong synergistic effect of emicizumab and aPCC and a similar but less pronounced effect of rFVIIa in people treated with emicizumab.

Variability in aggregometry response before and after initiation of clopidogrel therapy.

BACKGROUND: Evaluation of clopidogrel therapy by in vitro methods has limitations which may be of clinical importance. We wanted to explore the variability in aggregometry response in aspirin sensitive patients before and after initiation of clopidogrel therapy. METHODS: ADP 9.37 microM, AA 1.2mM and TRAP 25 mM stimulated light transmissions aggregometry (LTA) were performed twice before (Exams 1 and 2; 3 weeks apart)-and within one year after-initiation of clopidogrel therapy (Exam 3) in 79 patients treated with PCI. Repeated ADP aggregometry was also performed in 16 healthy volunteers in order to estimate LTA measurement error. RESULT: Inter-individual differences in ADP aggregation e.g. at Exam 1 were substantial (range 17-77%, SD 15.8%). Intra-individual changes between Exams 1 and 2 were significant (-27 to +36%, SD 14.6%, p<0.05). Inter-individual differences at Exam 3 (on clopidogrel treatment) were larger than expected from Exams 1 and 2 (p<0.01). AA aggregation was the same before and during clopidogrel treatment. In controls, inter-individual differences were smaller at ADP 10 than at ADP 5 microM. CONCLUSIONS: Inter-individual differences in ADP aggregation were significant both before and during clopidogrel therapy, and there were significant intra-individual variations over time. Therefore, prediction of aggregometry response before or during clopidogrel therapy based on single tests may be unreliable. Inter-individual differences in healthy controls are smaller at high concentrations of ADP, and comparisons of aggregometry response should be performed with caution unless ADP concentrations are standardized.

The reversal effect of prothrombin complex concentrate (PCC), activated PCC and recombinant activated factor VII in apixaban-treated patients in vitro.

BACKGROUND: The number of patients under treatment with FXa inhibitors is increasing, but there is no concensus on how to reverse their anticoagulant effect in case of a life-threatening bleeding. A specific antidote is not yet commercially available. Prothrombin complex concentrate (PCC), activated PCC (aPCC) and recombinant factor VIIa (rFVIIa) are suggested available reversal agents. OBJECTIVES: To find the most effective reversal agent to apixaban and to determine the optimal dose. PATIENTS/METHODS: PCC, aPCC, and rFVIIa at concentrations imitating 80%, 100%, and 125% of suggested therapeutic doses were added to blood drawn from apixaban-treated patients (n=30). aPCC was also tested in a 50% dose. Samples from healthy subjects (n=40) were used as controls. Thromboelastometry in whole blood (WB) and thrombin generation in platelet-poor plasma (PPP) were measured to assess the reversal effect. RESULTS: aPCC shortened clotting time (CT) in WB, and increased the peak thrombin concentration and velocity index in PPP to a greater extent than PCC and rFVIIa. No significant differences were seen between rFVIIa and aPCC on thrombin generation lag time, or between PCC and aPCC on endogenous thrombin potential (ETP). The 50% dose of aPCC had a slightly inferior effect, but was comparable to the other reversal agents. CONCLUSIONS: In this in vitro study the 80% dose of aPCC (40 IU/kg) reversed the anticoagulant effect of apixaban more effectively than the corresponding dose of rFVIIa and PCC both in WB (CT) and PPP (peak, ETP).

Intermittent saline flushes during haemodialysis do not alleviate coagulation and clot formation in stable patients receiving reduced doses of dalteparin.

BACKGROUND: Heparin-free haemodialysis (HD) with intermittent saline flushes (ISF) in patients with bleeding risk is widely used. The aim of this study was to investigate if ISF reduce coagulation and clotting in stable patients receiving reduced doses of dalteparin. METHODS: Inclusion criteria were stable chronic HD patients >or=18 years of age and haemoglobin >or=11 g/dl. Exclusion criteria were use of warfarin and acetylsalicylic acid. Six HD sessions were evaluated per patient. Dalteparin was given as one bolus dose at start of HD (50% of the conventional dose). In HD number 1, 3 and 5, 100 ml saline solution was flushed through the filter each 30 min. In HD 2, 4 and 6, no ISF were given. Potential clotting in the bubble trap was visually observed each hour and graded on a 4-point scale: 1 = normal, 2 = fibrinous ring, 3 = clot formation and 4 = coagulated system. The dialyser was visually inspected at the end of each session: 1 = normal, 2 = a few blood stripes (affecting less than 5% of the surface fibres), 3 = many blood stripes (more than 5% of the fibres) and 4 = coagulated filter. The coagulation marker PF1+2, the platelet activation marker beta-TG and anti-FXa activity were repeatedly measured during HD. RESULTS: Six men and two women were included. In four cases (four different patients), HD was stopped due to a coagulated system, all cases on days with ISF performed. Multiple linear regression analyses with repeated measurements showed that ISF adjusted for dalteparin dose/kg significantly increased mean clot in the bubble trap, estimate (B) = 0.717, P = 0.0001 and also showed that ISF increased PF1+2, B = 0.16, P = 0.001 when adjusted for anti-FXa activity and hours of dialysis, whereas beta-TG was only borderline increased, B = 0.09, P = 0.055. CONCLUSIONS: ISF during HD does not alleviate visible clotting or intravascular coagulation activity in stable patients receiving reduced doses of dalteparin and polysulphone dialysers. Whether this applies to unstable patients with increased bleeding risk not receiving any anticoagulation remains to be shown.

Tranexamic acid given into the wound reduces postoperative blood loss by half in major orthopaedic surgery.

OBJECTIVE: To evaluate the effect of locally applied tranexamic acid on postoperative blood loss and measures of fibrinolysis in drained blood. DESIGN: Prospective study. SETTING: University hospital, Norway. PATIENTS: 30 patients operated on for low back pain by screw fixation of the lumbar spine, 16 of who were randomised to be given topical tranexamic acid. MAIN OUTCOME MEASURES: Postoperative blood loss after 18 hours. Concentrations of plasmin/alpha2-antiplasmin (PAP) and D-dimer in arterial and drained blood at the time of wound closure and in drained blood after 1 hour. RESULTS: In the tranexamic group median (interquartile) blood loss was reduced by half from 525 (325-750) ml to 252 (127-465) ml, p = 0.02. In drained blood after one hour the increase in the concentration of PAP was 150 (109-170)% and D-dimer 150 (107-272)% in the tranexamic group compared with the control group where the increase in PAP was 320 (140-540)% and D-dimer 260 (161-670)%. CONCLUSION: Tranexamic acid applied in the wound inhibits blood loss by up to a half in major orthopaedic surgery probably because it prevents excessive fibrinolysis.