Ex vivo effects of low-dose rivaroxaban on specific coagulation assays and coagulation factor activities in patients under real life conditions

SummaryGlobal coagulation assays display variable effects at different concentrations of rivaroxaban. The aim of this study is to quantify the ex vivo effects of low-dose rivaroxaban on thrombophilia screening assays and coagulation factor activities based on the administration time, and to show how to mask possible interferences. Plasma samples from 40 patients receiving rivaroxaban 10 mg daily were investigated to measure activities of clotting factor II, V, VII, VIII, IX, XI, XII and XIII; protein C- and protein S-levels; lupus anticoagulants; anticardiolipin IgG and IgM; D-dimer, heparin-platelet factor 4 (HPF4) antibodies and screening tests for von Willebrand disease (VWD). Two hours after rivaroxaban administration, the activities of clotting factors were significantly decreased to different extents, except for factor XIII. Dilution of plasma samples resulted in neutralisation of these interferences. The chromogenic protein C activity assay was not affected by rivaroxaban. Depending on the timing of tablet intake in relation to blood sampling protein S activity was measured falsely high when a clotting assay was used. False-positive results for lupus anticoagulants were observed depending on the assay system used and the administration time of rivaroxaban. ELISA-based assays such as anticardiolipin IgG and IgM, D-dimer, HPF4-antibodies and the turbidimetric assays for VWD were not affected by rivaroxaban. Specific haemostasis clotting tests should be performed directly prior to rivaroxaban intake. Assay optimisation in the presence of rivaroxaban can be achieved by plasma dilution. Immunologic assays are not influenced by rivaroxaban, while chromogenic assays can be used, when they do not depend on factor Xa.

Download Full-text

Ex vivo effects of low-dose rivaroxaban on specific coagulation assays and coagulation factor activities in patients under real life conditions

Thrombosis and Haemostasis ◽

10.1160/th-12-04-0228 ◽

2012 ◽

Vol 109 (1) ◽

Author(s):

H. Mani ◽

C. Hesse ◽

G. Stratmann ◽

E. Lindhoff-Last

Keyword(s):

Low Dose ◽

Ex Vivo ◽

Real Life ◽

Coagulation Factor ◽

Coagulation Assays ◽

Life Conditions

Download Full-text

Coagulation Factors, Postmenopausal Hormone Replacement Therapy and the Risk of Venous Thrombosis: The WHI Clinical Trials of Postmenopausal Hormone Therapy.

Blood ◽

10.1182/blood.v110.11.127.127 ◽

2007 ◽

Vol 110 (11) ◽

pp. 127-127 ◽

Cited By ~ 1

Author(s):

Mary Cushman ◽

Joseph Larson ◽

Frits R. Rosendaal ◽

Lawrence S. Phillips ◽

Barbara V. Howard ◽

...

Keyword(s):

Risk Factors ◽

Relative Risk ◽

Venous Thrombosis ◽

Protein C ◽

Coagulation Factor ◽

Protein S ◽

Postmenopausal Hormone ◽

Free Protein ◽

Coagulation Marker ◽

D Dimer

Abstract Background. Postmenopausal estrogen (E) therapy, especially in combination with progestin (P) doubles the relative risk of venous thrombosis (VTE). Risk with hormones is higher with increasing age, obesity and with factor V Leiden. We studied coagulation markers as susceptibility factors for postmenopausal hormone-related VTE. Methods. The Women’s Health Initiative program included two placebo-controlled double-blind randomized trials of two E regimens, E (conjugated equine estrogens) or E+P (E + medroxyprogesterone acetate), in 16,608 postmenopausal women aged 50–79. We performed a nested case control study that measured baseline levels of coagulation markers in 215 women who developed VTE during follow up and 867 age-matched controls. The joint effects of treatment assignment to either E regimen vs placebo and prespecified abnormal levels of each coagulation factor on relative risk of VTE were estimated by logistic regression adjusting for age, race, body-mass index and type of E regimen. Results. Low levels of protein C and free protein S (<5th percentile), high D-dimer (top quartile), and high plasmin antiplasmin complex (PAP) and prothrombin fragment 1–2 (top decile) were all associated with risk of VTE with adjusted odds ratios (95% CI) of 2.0 (1.0–4.1), 2.9 (1.5–5.6), 2.8 (2.0–4.0), 2.5 (1.6–4.0) and 1.9 (1.2–3.1), respectively. Elevated factors II, VIII, IX and fibrinogen were not VTE risk factors. Compared to women with normal coagulation marker levels assigned to placebo, the joint odds of VTE with either E regimen plus an abnormal coagulation marker were more than additive compared to the separate effects of hormones and coagulation abnormalities for low protein C, low free protein S, and elevated D-dimer, PAP and F1–2. The odds ratios of VTE with the combination of an abnormal coagulation factor and assignment to hormones were (in order listed in prior sentence), 4.5 (95% CI 2.0–10.2), 6.7 (3.0–14.5), 6.1 (3.7–10), 5.8 (3.2–10.5) and 4.4 (2.4–7.7). Conclusions. We report new findings of elevated F1-2 and PAP as VTE risk factors in women in this prospective study nested in trials of E or E+P versus placebo. Protein C or S values below the 5th percentile were also clinically relevant even though they do not represent inherited deficiency. Lower protein C and free protein S, and higher D-dimer, F1-2 and PAP all identified women at increased risk of VTE with hormones. If our findings are confirmed in management studies, measurement of these factors might assist women with decision-making on safety of E or E+P.

Download Full-text

APC Resistance and other Haemostatic Variables during Pregnancy and Puerperium

Thrombosis and Haemostasis ◽

10.1055/s-0037-1614518 ◽

1999 ◽

Vol 81 (04) ◽

pp. 527-531 ◽

Cited By ~ 114

Author(s):

U. Kjellberg ◽

N.-E. Andersson ◽

S. Rosén ◽

L. Tengborn ◽

M. Hellgren

Keyword(s):

Factor Viii ◽

Plasminogen Activator ◽

Protein C ◽

Activated Protein C ◽

Plasminogen Activator Inhibitor ◽

Protein S ◽

Reference Level ◽

Soluble Fibrin ◽

Prothrombin Fragment ◽

D Dimer

SummaryForty-eight healthy pregnant women were studied prospectively and longitudinally. Blood sampling was performed at 10-15, 23-25, 32-34 and 38-40 weeks of gestation, within one week and at eight weeks postpartum. Classic and modified activated protein C ratio decreased as pregnancy progressed. In the third trimester 92% of the ratios measured with the classic test were above the lower reference level whereas all modified test ratios were normal. Slight activation of blood coagulation was shown with increased levels of prothrombin fragment 1+2, soluble fibrin and D-dimer. Fibrinogen, factor VIII and plasminogen activator inhibitor type 1 and type 2 increased. Protein S and tissue plasminogen activator activity decreased. Protein C remained unchanged. No correlation was found between the decrease in classic APC ratio and changes in factor VIII, fibrinogen, protein S, prothrombin fragment 1+2 or soluble fibrin, nor between the increase in soluble fibrin and changes in prothrombin fragment 1+2, fibrinogen and D-dimer.

Download Full-text

Increased prothrombin activation in protein S-deficient plasma under flow conditions on endothelial cell matrix: an independent anticoagulant function of protein S in plasma

Blood ◽

10.1182/blood.v85.7.1815.bloodjournal8571815 ◽

1995 ◽

Vol 85 (7) ◽

pp. 1815-1821 ◽

Cited By ~ 16

Author(s):

C van't Veer ◽

TM Hackeng ◽

D Biesbroeck ◽

JJ Sixma ◽

BN Bouma

Keyword(s):

Endothelial Cell ◽

Tissue Factor ◽

Protein C ◽

Activated Protein C ◽

Coagulation Factor ◽

Protein S ◽

Flow Conditions ◽

Cell Matrix ◽

Anticoagulant Function ◽

Prothrombin Activation

Protein S is a vitamin K-dependent nonenzymatic coagulation factor involved in the regulation of activated protein C (aPC). In this study, we report an aPC-independent anticoagulant function of protein S in plasma under flow conditions. Plasma, anticoagulated with low-molecular-weight heparin allowing tissue factor-dependent prothrombin activation, was perfused at a wall shear rate of 100 s-1 over tissue factor containing matrices of stimulated endothelial cells placed in a perfusion chamber. Fractions were collected in time at the outlet and prothrombin activation was determined by measuring the activation fragment F1+2 of prothrombin. In normal plasma, a time-dependent prothrombin activation was detected by the generation of fragment1+2. Prothrombin activation had ceased after 12 minutes perfusion, independent of the amount of tissue factor present in the matrix. Depletion of protein S from plasma or inhibition of protein S in plasma by monoclonal antibodies induced a 5- to 25-fold increase of prothrombin activation on the procoagulant endothelial cell matrix. A prolonged prothrombin activation was detected in protein S-depleted plasma up to 20 minutes after onset of the thrombin generation. The increased prothrombin activation in protein S-depleted plasma could not be explained by the absence of the cofactor function of protein S for aPC because depletion of protein C from plasma did not result in increased prothrombin activation. These data provide further evidence for a strong anticoagulant function of protein S in plasma independent from activated protein C.

Download Full-text

Prevalence of common thrombophilia markers and risk factors in Indian patients with primary venous thrombosis

Sao Paulo Medical Journal ◽

10.1590/s1516-31802010000500004 ◽

2010 ◽

Vol 128 (5) ◽

pp. 263-267 ◽

Cited By ~ 7

Author(s):

Mahendra Narain Mishra ◽

Varinder Singh Bedi

Keyword(s):

Venous Thrombosis ◽

Protein C ◽

Quantitative Estimation ◽

Protein S ◽

Anticardiolipin Antibodies ◽

Factor V ◽

Cross Sectional ◽

C Protein ◽

Lupus Anticoagulants ◽

Significant Difference

CONTEXT AND OBJECTIVE: Venous thrombosis occurs as a result of interaction of genetic and acquired factors including activated protein C resistance (APC-R), fibrinogen levels, antithrombin, protein C, protein S, lupus anticoagulants and anticardiolipin antibodies. This study was aimed at determining the prevalence of these common thrombophilia markers in Asian Indians with primary venous thrombosis. DESIGN AND SETTING: This was a cross-sectional study carried out in Mumbai. METHODS: Samples from 78 patients with a confirmed diagnosis of venous thrombosis and 50 controls were tested. Semi-quantitative estimation (functional assays) of protein C, protein S and antithrombin was performed. Quantitative estimation of fibrinogen was done using the Clauss method. Lupus anticoagulants were screened using lupus-sensitive activated partial thromboplastin time and β2-glycoprotein-I dependent anticardiolipin antibodies were estimated by ELISA. APC-R was measured using a clotting-based method with factor V deficient plasma and Crotalus viridis venom. Statistical analysis was performed using Epi-info (version 6). RESULTS: The popliteal vein was the most commonly involved site. Forty-four samples (56%) gave abnormal results. The commonest were elevated fibrinogen and APC-R (17.9% each), followed by low protein S (16.6%). CONCLUSIONS: This study confirms the literature findings that fibrinogen level estimation and screening for APC-R are important for the work-up on venous thrombosis patients since these, singly or in combination, may lead to a primary thrombotic episode. The frequency of the other thrombophilia markers was higher among the patients than among the controls, but without statistically significant difference.

Download Full-text

Diabetes does not influence activation of coagulation, fibrinolysis or anticoagulant pathways in Gram-negative sepsis (melioidosis)

Thrombosis and Haemostasis ◽

10.1160/th11-07-0504 ◽

2011 ◽

Vol 106 (12) ◽

pp. 1139-1148 ◽

Cited By ~ 10

Author(s):

Joost Meijers ◽

Rapeephan Maude ◽

Direk Limmathurotsakul ◽

Nicholas Day ◽

Sharon Peacock ◽

...

Keyword(s):

Protein C ◽

Burkholderia Pseudomallei ◽

Blood Donors ◽

Protein S ◽

Admission Diagnosis ◽

Procoagulant Effect ◽

Multiple Abnormalities ◽

The Impact ◽

D Dimer

SummaryDiabetes is associated with a disturbance of the haemostatic balance and is an important risk factor for sepsis, but the influence of diabetes on the pathogenesis of sepsis remains unclear. Melioidosis (Burkholderia pseudomallei infection) is a common cause of community-acquired sepsis in Southeast Asia and northern Australia. We sought to investigate the impact of pre-existing diabetes on the coagulation and fibrinolytic systems during sepsis caused by B. pseudomallei. We recruited a cohort of 44 patients (34 with diabetes and 10 without diabetes) with culture-proven melioidosis. Diabetes was defined as a pre-admission diagnosis of diabetes or an HbA1c>7.8% at enrolment. Thirty healthy blood donors and 52 otherwise healthy diabetes patients served as controls. Citrated plasma was collected from all subjects; additionally in melioidosis patients follow-up specimens were collected seven and ≥28 days after enrolment where possible. Relative to uninfected healthy controls, diabetes per se (i.e. in the absence of infection) was Characterised by a procoagulant effect. Melioidosis was associated with activation of coagulation (thrombin-antithrombin complexes (TAT), prothrombin fragment F1+2 and fibrinogen concentrations were elevated; PT and PTT prolonged), suppression of anti-coagulation (antithrombin, protein C, total and free protein S levels were depressed) and abnormalities of fibrinolysis (D-dimer and plasmin-antiplasmin complex [PAP] were elevated). Remarkably, none of these haemostatic alterations were influenced by pre-existing diabetes. In conclusion, although diabetes is associated with multiple abnormalities of coagulation, anticoagulation and fibrinolysis, these changes are not detectable when superimposed on the background of larger abnormalities attributable to B. pseudomallei sepsis.

Download Full-text

Acquired activated protein C resistance is associated with lupus anticoagulants and thrombotic events in pediatric patients with systemic lupus erythematosus

Blood ◽

10.1182/blood.v97.4.844 ◽

2001 ◽

Vol 97 (4) ◽

pp. 844-849 ◽

Cited By ~ 55

Author(s):

Christoph Male ◽

Lesley Mitchell ◽

James Julian ◽

Patricia Vegh ◽

Penny Joshua ◽

...

Keyword(s):

Systemic Lupus Erythematosus ◽

Lupus Erythematosus ◽

Protein C ◽

Pediatric Patients ◽

Activated Protein C ◽

Factor V Leiden ◽

Protein S ◽

Activated Protein C Resistance ◽

Systemic Lupus ◽

Lupus Anticoagulants

Abstract Acquired activated protein C resistance (APCR) has been hypothesized as a possible mechanism by which antiphospholipid antibodies (APLAs) cause thrombotic events (TEs). However, available evidence for an association of acquired APCR with APLAs is limited. More importantly, an association of acquired APCR with TEs has not been demonstrated. The objective of the study was to determine, in pediatric patients with systemic lupus erythematosus (SLE), whether (1) acquired APCR is associated with the presence of APLAs, (2) APCR is associated with TEs, and (3) there is an interaction between APCR and APLAs in association with TEs. A cross-sectional cohort study of 59 consecutive, nonselected children with SLE was conducted. Primary clinical outcomes were symptomatic TEs, confirmed by objective radiographic tests. Laboratory testing included lupus anticoagulants (LAs), anticardiolipin antibodies (ACLAs), APC ratio, protein S, protein C, and factor V Leiden. The results revealed that TEs occurred in 10 (17%) of 59 patients. Acquired APCR was present in 18 (31%) of 58 patients. Acquired APCR was significantly associated with the presence of LAs but not ACLAs. Acquired APCR was also significantly associated with TEs. There was significant interaction between APCR and LAs in the association with TEs. Presence of both APCR and LAs was associated with the highest risk of a TE. Protein S and protein C concentrations were not associated with the presence of APLAs, APCR, or TEs. Presence of acquired APCR is a marker identifying LA-positive patients at high risk of TEs. Acquired APCR may reflect interference of LAs with the protein C pathway that may represent a mechanism of LA-associated TEs.

Download Full-text

Complex formation between urokinase and plasma protein C inhibitor in vitro and in vivo

Blood ◽

10.1182/blood.v74.2.722.722 ◽

1989 ◽

Vol 74 (2) ◽

pp. 722-728 ◽

Cited By ~ 55

Author(s):

M Geiger ◽

K Huber ◽

J Wojta ◽

L Stingl ◽

F Espana ◽

...

Keyword(s):

Complex Formation ◽

Protein C ◽

Specific Activity ◽

Ex Vivo ◽

Plasminogen Activator Inhibitor ◽

Enzyme Linked Immunosorbent Assay ◽

Plasma Samples ◽

Protein C Inhibitor

Abstract Protein C inhibitor (PCI) and plasminogen activator inhibitor 3 (PAI-3; urinary urokinase inhibitor) are immunologically identical. The role of PCI for urokinase (uPA) inhibition in vivo was investigated. We therefore developed an enzyme-linked immunosorbent assay (ELISA) specific for uPA-PCI complexes: Rabbit anti-PCI IgG was immobilized on a microtiter plate and following incubation with uPA-PCI complex- containing samples, bound uPA-PCI complexes were quantified with a horseradish-peroxidase-linked monoclonal antibody (MoAb) to uPA. Using this assay, time, dose, and heparin-dependent complexes were detected when uPA was incubated with normal plasma or purified urinary PCI, whereas no complexes were measurable using PCI-immunodepleted plasma. Plasma samples (containing 20 mmol/L benzamidine to prevent complex formation ex vivo) from patients undergoing systemic urokinase therapy (1 x 10(6) IU/60 min intravenously [IV]) after myocardial infarction were also studied. uPA present in these plasma samples (up to 1,200 ng/mL) had only 43% to 70% of the specific activity of purified 2-chain uPA, suggesting that a major portion of uPA is complexed to inhibitors. In these plasma samples uPA-PCI complexes were present in a concentration corresponding to 21% to 25% of inactive uPA antigen. These data suggest that at high uPA concentrations, such as during uPA therapy, plasma PCI might contribute significantly to uPA inhibition in vivo.

Download Full-text

β2-Glycoprotein I: Target Antigen for Autoantibodies in the ‘Antiphospholipid Syndrome’

Lupus ◽

10.1177/096120339600500509 ◽

1996 ◽

Vol 5 (5) ◽

pp. 381-385 ◽

Cited By ~ 14

Author(s):

DA Kandiah ◽

YH Sheng ◽

SA Krilis

Keyword(s):

Clinical Significance ◽

Protein C ◽

Binding Proteins ◽

Protein S ◽

Target Antigen ◽

Major Protein ◽

Neurological Sequelae ◽

Foetal Loss ◽

Lupus Anticoagulants ◽

Glycoprotein I

‘Antiphospholipid’ (aPL) antibodies are of important clinical significance because of their association with thrombosis both arterial and venous, recurrent foetal loss, specific neurological sequelae like seizures and chorea, cardiac valvular abnormalities and thrombocytopenia.1 Traditionally these autoantibodies have been assayed using phospholipid (PL) dependent tests and are classified as lupus anticoagulants (LA) and anticardiolipin (aCL) antibodies based on the method of detection.2 The antibodies thus, had been thought to bind PLs but it has now become clear that the true antigens are PL-binding proteins. The major protein consistently found as the target antigen for these autoantibodies is β2-glycoprotein I (β2-GPI).3 Other candidate PL-binding proteins have also been investigated including prothrombin, protein C and protein S4 but thus far appear to play less important roles in the binding of these antibodies.

Download Full-text