Postprandial changes in the phospholipid composition of circulating microparticles are not associated with coagulation activation

Introduction

Evidence is present that the phospholipid composition of circulating cell-derived microparticles (MP) affects coagulation in vivo, and that postprandial metabolic alterations may be associated with hypercoagulable state. Our objective was to investigate whether postprandial metabolic responses affect the phospholipid composition of MP, and whether such changes are associated with coagulation activation.

Materials and Methods

Twelve healthy males were studied twice and randomly received two consecutive meals or remained fasted. Blood was collected before and at 2, 4, 6 and 8 h following breakfast. Plasma concentrations of prothrombin-F_1 + 2 and thrombin-antithrombin-complexes were measured. Numbers and cellular origin of MP were determined by flowcytometry. The phospholipid composition of MP was determined by hpTLC. In vitro procoagulant activity of MP was studied by fibrin generation.

Results

During the meal visit, plasma glucose, triglyceride and insulin levels increased, compared to baseline and the fasting visit (all P < 0.05). Postprandially, the total numbers of MP increased in time compared to the fasting visit (P < 0.05). Erythrocyte-derived MP increased (6-fold) during the meal visit, but remained constant on the fasting day (P < 0.001). On the meal versus fasting day circulating MP contained increased phosphatidylcholine (P < 0.05) and decreased sphingomyelin (P < 0.05) amounts. The amount of phosphatidylserine did not change. Concentrations of plasma F_1 + 2 and thrombin-antithrombin were similar on both days, as was the ability of MP to generate fibrin in vitro.

Conclusion

Although numbers, cellular origin and phospholipid composition of MP alter during exposure to two consecutive meals in healthy subjects, this does not lead to changes in the coagulation activation in vivo.     

Comparison of a direct Factor Xa inhibitor, edoxaban, with dalteparin and ximelagatran: A randomised controlled trial in healthy elderly adults

Introduction

Edoxaban (the free base of DU-176b) is a new, oral direct Factor Xa inhibitor. This is the first study to compare the hemostatic response to edoxaban, ximelagatran, and dalteparin in healthy, elderly adults.

Materials and Methods

In this open-label, active-controlled clinical trial, 40 adults (65-75 years), were randomised to: oral edoxaban (60 mg, twice-daily, 7 doses), subcutaneous dalteparin (5000 IU, once-daily, 4 doses), oral ximelagatran (24 mg, twice-daily, 7 doses) or no drug. Blood samples were taken before, and 1.5, 4, 12, 24, 72, 84, 96, 108, 120, and 144 hours after, the first dose. The primary outcomes were changes in thrombin-antithrombin complex, prothrombin fragment 1 + 2 and D-dimer, and adverse events. Additional biomarkers of coagulation, and endothelial cell and platelet activation were compared (ANOVA).

Results

All subjects completed the study. Inhibition of thrombin generation lag time, peak, and constant velocity index were significantly greater, and extended for a longer period of time, following edoxaban administration, compared with dalteparin. We found that the traditional assay for anti-FXa activity was not appropriate for the new anticoagulants. Biomarker changes following edoxaban administration (including prolongation of prothrombin time) reflected inhibition of Factor Xa; there was no effect on platelet, tissue factor or endothelial activation. There were no clinically significant changes in primary outcomes. No serious adverse events were reported.

Conclusion

Oral administration of edoxaban resulted in effective Factor Xa and TG inhibition, and was well-tolerated. Studies are needed to confirm edoxaban (60 mg daily) use in clinical practice.

Sponsorship

Daiichi Sankyo Pharma Development.     

Fibrin clot properties in women heterozygous for factor V Leiden mutation: Effects of oral contraceptives

Introduction

Oral contraceptives (OC) in the presence of factor V Leiden mutation (FVL) markedly increase the risk of venous thromboembolism (VTE). Little is known about the OC and FVL-related alterations in fibrin clot properties.

Subjects and Methods

Plasma fibrin clot permeability (K_s) and efficiency of lysis, reflected by clot lysis time (CLT) and the rate of D-dimer release from clots (D-D_rate) induced by recombinant tissue plasminogen activator (tPA) were determined in 25 women with a family history of VTE who were heterozygous for FVL [FVL(+/−) - twice, on third-generation OC and after their discontinuation. Female non-carriers of FVL, matched for demographics, using OC and after their discontinuation served as controls (n = 25). All participants had no personal history of VTE.

Results

OC discontinuation in FVL(+/−) women resulted in shortened CLT (− 9%), and increased K_s (+ 4%) and D-D_rate (+ 1.4%; all p < 0.01). Alterations in fibrin clot properties were associated with decreased prothrombin fragments 1 + 2 (F1 + 2) (− 8%), plasminogen activator inhibitor-1 (PAI-1) antigen (− 11%), and thrombin activatable fibrinolysis inhibitor (TAFI) activity (− 20%; all p < 0.01). During OC use FVL(+/−) carriers compared with non-carriers had higher platelet count, activity of PAI-1, TAFI, and tPA, as well as prolonged CLT and higher D-D_max, along with lower D-D_rate and K_s. Multiple regression analysis adjusted for fibrinogen and age, showed that PAI-1 antigen and TAFI activity independently predicted CLT in FVL(+/−) women on OC.

Conclusion

FVL(+/−) is associated with hypofibrinolysis in apparently healthy women and third-generation OC administration unfavorably alters plasma clot characteristics in female FVL(+/−) carriers with a family history of thrombotic events.     

Coagulation activation and ultrasound characteristics in patients with carotid artery disease

Introduction

Elevated levels of markers for thrombin activation are associated with plaque echogenicity and degree of stenosis in patients with carotid artery stenosis. The Activated Protein C-Protein C Inhibitor (APC-PCI) complex reflects activation of the Protein C system and is a measure of thrombin generation. The aim of the present study was to examine APC-PCI complex in patients undergoing thrombendartherectomy for carotid artery stenosis, and to relate the findings to clinical characteristics and plaque morphology as determined by ultrasound.

Materials and Methods

Blood was obtained from 125 patients (39 female, median age 71 years) with carotid artery stenosis admitted from September 2005 to May 2007. The APC-PCI complex was measured using a sandwich immunofluorometric method and compared to an age- and sex-matched healthy control-group. Clinical and demographic characteristics, routine laboratory markers and ultrasound characteristics were analysed using univariate and multivariate analysis.

Results

APC-PCI complex concentration was significantly increased in patients with carotid artery stenosis (median 0.21 µg/L; 10th to 90th percentile 0.15-0.36) compared to a healthy control-group (0.19 µg/L; 0.11-0.31; P = .009). There was no significant difference in APC-PCI-values between asymptomatic (n = 48) and symptomatic (n = 77) patients with carotid artery stenosis (0.22 vs. 0.20 µg/L; p = 0.626). Patients with minor stroke (n = 31) had a higher median APC-PCI-concentration (0.27 µg/L; 0.15-0.63) than patients with amaurosis fugax (0.19 µg/L; 0.15-0.36) or transient ischemic attack (0.21 µg/L; 0.12-0.36) (p = 0.016). No association was found between APC-PCI-values and the degrees of carotid artery stenosis or the time from the latest neurological symptoms to blood sampling. Patients with echolucent plaques had significantly lower APC-PCI concentrations (0.20 µg/L; 0.14-0.35 vs. 0.24 µg/L; 0.15-0.60; p = 0.043), according to the Gray-Weale classification.

Conclusions

Patients with carotid artery disease exhibit increased concentrations of APC-PCI compared to a healthy control-group, particularly those patients with echogenic plaques, who have significantly higher APC-PCI levels than patients with echolucent plaques.     

Effects of raloxifene and continuous combined hormone therapy on haemostasis variables: a multicenter, randomized, double-blind study

INTRODUCTION: Hormone replacement therapy is known to increase the risk of thromboembolic events. We compared the effects of HRT and raloxifene on some haemostasis variables. MATERIALS AND METHODS: In a multicenter, double-blind study, 54 healthy postmenopausal women were randomized to receive either continuous treatment with 2 mg 17beta-estradiol plus 1 mg norethisterone acetate (n=30) or 60 mg raloxifene (n=24) daily for 12 months. Blood samples were collected at baseline and at 3, 6 and 12 months to evaluate therapy effects on some haemostasis variables (factor VII, factor VIII, prothrombin fragments 1 and 2, protein C, protein C activity, protein S, thrombin-antithrombin complex, D-dimer, antithrombin, fibrinogen and plasminogen activator inhibitor). RESULTS: Both raloxifene and continuous combined hormone therapy modified the haemostasis variables toward a more prothrombotic profile. Factor VIII (p<0.01) and fibrinogen (p<0.05) plasma levels significantly increased at 6 months, prothrombin fragments 1 and 2 (p<0.05) significantly increased at 12 months, whereas protein C activity (p<0.001) and antithrombin (p<0.01) significantly decreased at 12 months in both groups. CONCLUSIONS: Our results demonstrate that raloxifene and continuous combined hormone therapy exhibit the same prothrombotic profile. Both treatments induced an increase in procoagulant parameters at 6 months and a decrease in anticoagulant parameters at 12 months.     

Short-term effects of estrogen, tamoxifen and raloxifene on hemostasis: a randomized-controlled study and review of the literature

Introduction

Estrogen therapy (ET), tamoxifen and raloxifene are associated with a two- to three-fold increased risk of venous thrombosis (VT); however, the mechanisms by which each drug increases venous thrombosis propensity are not fully understood. The objectives of this investigation were to compare the effects of these three treatments on hemostasis in a head to head randomized placebo-controlled trial.

Patients/methods

Ninety-four postmenopausal women were assigned to receive oral estrogen (conjugated equine estrogen [CEE] 0.625 mg, n=23), tamoxifen 20 mg (n=24), raloxifene 60 mg (n=24) or placebo (n=23) daily for 6 months. Blood samples were analyzed for procoagulant factors (prothrombin, factors VII [fVII], VIII [fVIII], IX [fIX] and XI [fXI], D-dimer and von Willebrand factor [vWf]), anticoagulant factors (antithrombin [AT], total and free protein S, protein C and activated protein C [APC] resistance) and fibrinolytic factors (thrombin activatable fibrinolysis inhibitor [TAFI] and plasminogen activator inhibitor-1 [PAI-1]), at baseline and at 6 months of treatment.

Results

Estrogen increased factor VII and D-dimer, and decreased antithrombin, total and free protein S and PAI-1. Changes with tamoxifen were distinct from estrogen with increases in factors VIII, IX, vWf and free protein S, and decreases in AT, total protein S, protein C and plasminogen activator inhibitor-1. Raloxifene produced similar effects as tamoxifen, but did not increase factor IX or decrease protein C.

Conclusions

Estrogen, tamoxifen and raloxifene affected hemostasis favoring procoagulation and impairing anticoagulation. The biochemical effects of the selective estrogen receptor modulators (SERMs) were distinct from those of estrogen and differed only subtly from each other.