Procoagulant State in Current and Former Anabolic Androgenic Steroid Abusers

Background Anabolic androgenic steroid (AAS) abusers are considered at increased risk of cardiovascular morbidity and mortality. We hypothesized that current and former AAS abuse would induce a procoagulant shift in the haemostatic balance. Methods Men 18 to 50 years of age were included as current AAS abusers, former AAS abusers or controls. Morning blood samples were collected after overnight fasting. Thrombin generation (lag time, time to peak, peak height, and endogenous thrombin potential [ETP]) and coagulation factor II (prothrombin), VII and X, antithrombin, protein C, free protein S and tissue factor pathway inhibitor (TFPI) were assessed. Groups were compared by ANOVA or Kruskal–Wallis test and probabilities were corrected for multiple comparisons. Associations were evaluated using linear regression models. Results ETP was increased around 15% in current (n = 37) and former (n = 33) AAS abusers compared with controls (n = 30; p < 0.001). Prothrombin and factor X were increased ≥10% in AAS abusers and prothrombin was a predictor of ETP (p < 0.0005). Lag time and time to peak were increased 10 to 30% in current AAS abusers (p < 0.001) and associated with higher concentrations of TFPI, antithrombin, protein C and protein S (p < 0.0005; = 0.005). Multivariate linear regression, with all coagulation inhibitors as covariates, identified TFPI to be independently associated with lag time and time to peak (p < 0.0005). Conclusion Thrombin generation is augmented in current and former AAS abusers, reflecting a procoagulant state, with altered concentrations of coagulation proteins. Prospective studies are needed to clarify whether these findings translate into an increased thrombotic risk in AAS abusers potentially even after cessation.

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Protein S Regulates Factor IXa in the Absence and Presence of Factor VIIIa Independently of Activated Protein C

Blood ◽

10.1182/blood.v118.21.1197.1197 ◽

2011 ◽

Vol 118 (21) ◽

pp. 1197-1197

Author(s):

Rinku Majumder ◽

Rima Chattopadhyay ◽

Tanusree Sengupta

Keyword(s):

Protein C ◽

Thrombin Generation ◽

Anticoagulant Activity ◽

Activated Protein C ◽

Protein S ◽

Activity Measurement ◽

Clotting Time ◽

Thrombotic Risk ◽

Increased Risk

Abstract Abstract 1197 Coagulation is a finely tuned process. During thrombin formation, several anticoagulant reactions are initiated to prevent systematic activation of coagulation, and impairment of anticoagulant activity causes an increased risk of venous thrombosis. One such anticoagulant factor is protein S, deficiencies of which have been linked to venous and arterial thrombosis. While protein S has been studied for over three decades, the precise role this protein plays in attenuating the hemostatic response is far from clear. Protein S is a vitamin K-dependent plasma protein that functions in feedback regulation of thrombin generation. Protein S was initially identified as a cofactor for activated protein C (APC) but later it was observed that there is only a 3–10 fold increase in APC activity in the presence of protein S. Plasma coagulation assays in the absence of APC suggest that protein S may have other anticoagulant role(s). We report here an anticoagulant activity of Protein S mediated by inhibition of fIXa in the absence and presence of fVIIIa independent of APC. Although an APC-independent anticoagulant activity has been reported for protein S interacting with fVIIIa, no study has shown that the inhibitory effect of protein S is mediated through its interaction with fIXa, thus making our observations novel and significant. Moreover, previous studies that reported an interaction between fVIIIa and protein S were performed with low amounts of phospholipid, a condition that produces activity measurement artifacts due to the presence of active protein S multimers. We used both ex vivo (plasma studies) and in vitro methods at high phospholipid (100–200 micro molar) concentration to determine whether and how the intrinsic pathway of blood coagulation is regulated by protein S. We obtained the following results: 1) activated partial thromboplastin time (aPTT) assays with protein S-supplemented plasma confirmed that protein S prolongs clotting time, and a normal clotting time was restored with addition of anti-protein S antibody, 2) a modified aPPT assay with fIX-deficient plasma confirmed that protein S affects fIX-initiated clotting time, 3) thrombin generation assay through fIXa/fVIIIa pathway, initiated with a limiting amount of tissue factor (TF), was regulated by protein S, 4) in vitro studies with fIXa/fVIIIa and protein S in the presence of phosphatidylserine (PS) vesicles showed ∼40% and ∼65% inhibition in the activity of fIXa in the absence and presence of fVIIIa, respectively, and 5) protein S altered only the KM for fX activation by fIXa but altered both kcat and KM for fX activation by fIXa and fVIIIa. Our findings underscore the central role of protein S in regulation of coagulation. We anticipate these results will unravel important implications for the evaluation of thrombotic risk associated with protein S-deficiency. Disclosures: No relevant conflicts of interest to declare.

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Thrombin Generation Measurement In Whole Blood As a Control For Vitamin K Antagonist Treatment - Effect Of Thrombomodulin

Blood ◽

10.1182/blood.v122.21.3646.3646 ◽

2013 ◽

Vol 122 (21) ◽

pp. 3646-3646

Author(s):

Saartje Bloemen ◽

Marieke de Laat ◽

Arina ten Cate-Hoek ◽

Hugo ten Cate ◽

Bas De Laat ◽

...

Keyword(s):

Vitamin K ◽

Whole Blood ◽

Protein C ◽

Thrombin Generation ◽

Lag Time ◽

Peak Height ◽

Vitamin K Antagonist ◽

P Value ◽

Time To Peak ◽

20 Nm

Abstract Introduction We tested whether the recently introduced measurement of thrombin generation (TG) in whole blood can be used to evaluate the clotting status of patients on vitamin K antagonist (VKA) prophylaxis. The prothrombin time, and hence the International Normalized Ratio (INR), only evaluates the vitamin K dependent factors II, VII and X but not the anticoagulant factors, protein C and S as well as factor IX. In TG all factors play their role and when thrombomodulin (TM) is added the function of proteins C and S is stressed. The thrombotic tendency in congenital protein C resistance proves the importance of this protein C pathway. Aim To compare the INR in samples from patients under VKA prophylaxis to TG in whole blood and in platelet rich and platelet poor plasma (PRP, PPP) both in the presence and in the absence of TM. Materials & Methods Blood samples were collected from 123 consenting patients on VKA. In two thirds (67%) the indication for prophylaxis was atrial fibrillation. Other indications included prosthetic valves, lung embolisms or thrombosis. The INR was determined in the PPP of the samples and the patients were stratified into 5 groups: INR of 1.0 to 1.5, 1.5 to 2.5, 2.5 to 3.5, 3.5 to 4.5 and higher than 4.5. Thrombin generation (TG) was measured via Calibrated Automated Thrombinography (CAT) in whole blood and in PRP and PPP, with and without 20 nM added TM. From the TG curve lag time and time to peak were obtained as well as the maximal thrombin concentration (peak) and the area under the curve (endogenous thrombin potential: ETP). Also red and white blood cells and platelets were counted. Results With increasing INR values, the ETP and peak height decrease and lag time and time to peak prolong. All TG parameters measured in whole blood were significantly correlated (p-values< 0.01) with the values determined in both PRP and PPP. INR was linearly correlated with lag time and time to peak (p-value< 0.01), whereas for the concentration dependent parameters (ETP and peak height) the correlation with the INR was hyperbolical (p-value< 0.01). In plasma, 20 nM TM causes a diminution of ETP and peak of 50-60 % in normals and in patients in the INR 1 – 1.5 group. At higher INR values inhibition is between 25 and 40%, independent of the INR value. In whole blood, on the contrary, the same concentration of TM causes around 30 % of inhibition in normals and in all patients alike. Conclusions Whole blood TG data correlate well with INR and reflect more of the coagulation mechanism than the INR does. Like the INR it does not reflect the function of the VKAs on the natural anticoagulant factors, however. In PPP and PRP addition of TM shows that VKA treatment induces TM resistance in patients with an INR value higher than 1.5. Disclosures: No relevant conflicts of interest to declare.

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Calibrated automated thrombin generation in normal uncomplicated pregnancy

Thrombosis and Haemostasis ◽

10.1160/th07-05-0359 ◽

2008 ◽

Vol 99 (02) ◽

pp. 331-337 ◽

Cited By ~ 68

Author(s):

Andrea Rosenkranz ◽

Michael Hiden ◽

Bettina Leschnik ◽

Weiss Eva-Christine ◽

Dietmar Schlembach ◽

...

Keyword(s):

Protein C ◽

Thrombin Generation ◽

Plasminogen Activator Inhibitor ◽

Protein S ◽

Plasminogen Activator Inhibitor 1 ◽

S Protein ◽

Endogenous Thrombin Potential ◽

Time To Peak ◽

Uncomplicated Pregnancy ◽

Pai 1

SummaryPregnancy is associated with substantial changes in the haemostatic system and a six-fold higher incidence of venous thromboembolism. Conventional global tests, such as prothrombin time and activated partial thromboplastin time, do not definitely detect this hypercoagulable condition. We investigated whether the changes in haemostatic system during pregnancy are reflected in the calibrated automated thrombography (CAT). Thrombin generation was measured in platelet-poor plasma (PPP) of 150 healthy pregnant women without any pregnancy associated diseases by means of CAT. In addition, prothrombin (FII), antithrombin (AT), protein S, protein C, tissue factor pathway inhibitor (TFPI), plasminogen activator inhibitor-1 (PAI-1), thrombin-antithrombin complex (TAT), and prothrombin fragments 1+2 (F1+2) were measured. Endogenous thrombin potential (ETP) and peak of thrombin generation increased significantly with gestational weeks, while lag time and time to peak remained unchanged. A significant increase of PAI-1,TFPI,F1+2 and TAT as well as a significant decrease of free protein S, protein S antigen, and protein S activity was observed. Levels of AT and protein C remained stable during pregnancy. Division of population in trimester of pregnancy and analysis of differences between the trimesters showed rather similar results. Our study shows that endogenous thrombin potential does increase with duration of normal uncomplicated pregnancy. Whether parameters of continuous thrombin generation will correlate with thrombembolic disease remains to be shown.

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Microparticle-Associated Thrombin Generation and Procoagulant Activity Is Increased In Patients with Essential Thrombocythemia

Blood ◽

10.1182/blood.v116.21.1985.1985 ◽

2010 ◽

Vol 116 (21) ◽

pp. 1985-1985

Author(s):

Annamaria Leuzzi ◽

Marina Marchetti ◽

Marina Panova-Noeva ◽

Carmen Julia Tartari ◽

Laura Russo ◽

...

Keyword(s):

Essential Thrombocythemia ◽

Thrombin Generation ◽

Lag Time ◽

Procoagulant Activity ◽

Jak2v617f Mutation ◽

Wild Type ◽

Thrombotic Risk ◽

Time To Peak ◽

Mutation Carriers ◽

Free Plasma

Abstract Abstract 1985 Essential Thrombocythemia (ET) is a chronic myeloproliferative neoplasm characterized by an increased thrombotic risk. Numerous quantitative and qualitative abnormalities of platelets and leukocytes, arising from the clonal proliferation of hematopoietic progenitor cells, have been described as responsible of the thrombophilic state in this disease. Recently, a high number of plasma microparticles (MP) of different cellular origin has been described in ET patients. MP are a key component of the hemostatic response and have been found increased in diseases at high thrombotic risk. To explore the contribution of plasma MP to the hypercoagulable state of patients with ET, in this study we aimed to characterize the MP functional procoagulant features. We used two different methods: the calibrated automated thrombogram (CAT), to determine the MP-associated thrombin generation (TG), and the P-PPL/1 assay (Stago R&D) to measure the MP-associated procoagulant activity (PCA). Both assays were performed in platelet free plasma (P-FP) obtained from 69 ET patients (24M/45F; 33 carriers of JAK2V617F mutation) and 67 control subjects (32M/35F). In a subgroup of 23 ET patients and 23 controls MP-associated TG and PCA were also determined in MP-free plasma (MP-FP). PFP was obtained by two serial centrifugations (4,000 rpm for 15 min, then 11,000 rpm for 10 min) and MP-FP by re-centrifuging P-FP at 14,000 rpm for 30 min. MP were isolated from the pellet. For the TG assay, 80 ul of P-FP or MP-FP and 20 ul of buffer were mixed, and TG started by adding CaCl2. The results were expressed as lag-time, peak, area under the curve (ETP), and time-to-peak (ttPeak). For P-PPL/1 assay, 100 ul of P-FP or MP-FP were mixed with 50 ul of phospholipid-depleted plasma. Clotting was started by adding FXa and CaCl2 and results expressed in seconds. The results show that P-FP from ET patients generated significantly higher quantity of thrombin compared to controls, as demonstrated by the shorter lag-time (22.1±8.7 vs 29.5±14.7 min; p=0.004) and time to peak (26.6±8.4 vs 33.3±13.2 min; p=0.004) and the significantly greater peak (52.9±24.9 vs 38.24±20.9 nM; p=0.002) and ETP (765.6±206.4 vs 537.5±295 nM*min; p=0.001). Similarly, the MP-associated PCA was significantly increased in ET patients (79±11 sec) compared to controls (89±11 sec; p<0.05). This increase was due to the presence of MP, as no TG and little PCA was observed in MP-FP from both patients and controls. The addition of isolated MP to autologous MP-FP restored the TG and PCA of the samples to the original values of P-FP for both assays. TG was significantly (p<0.05) increased in the JAK2V617F mutation carriers (lag-time: 19.5±7.4 min; peak: 59.8±26.9 nM; ttpeak: 23.9± 4.9 min) compared to wild-type subjects (lag-time: 24.5±9.1 min, peak: 45.4±20.6 nM, ttpeak: 29.3±10.3 min), while no significant differences were found for PCA. Significant correlations were found between the PCA by PPL-assay and the different parameters of TG assay [lag-time (R2= 0.414), peak (R2= -0.542), ETP (R2= -0.514)]. In conclusion, our results show that MP-associated TG capacity, as well as PCA, are increased in plasma from ET patients. The highest MP-associated TG was found in JAK-2 mutation carriers, who also are at higher risk for thrombosis compared to wild-type subjects. Our data provide evidence for a contribution of MP to the thrombophilic state of these patients and suggest to test MP associated TG and PCA in prospective studies to predict thrombosis in ET patients. Disclosures: No relevant conflicts of interest to declare.

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Effect of oral contraceptives on thrombin generation measured via calibrated automated thrombography

Thrombosis and Haemostasis ◽

10.1160/th07-07-0439 ◽

2007 ◽

Vol 98 (12) ◽

pp. 1350-1356 ◽

Cited By ~ 46

Author(s):

Svetlana chaikovski ◽

Huib van Vliet ◽

M.Christella Thomassen ◽

Rogier Bertina ◽

Frits Rosendaal ◽

...

Keyword(s):

Oral Contraceptives ◽

Protein C ◽

Thrombin Generation ◽

Activated Protein C ◽

Lag Time ◽

Peak Height ◽

Excellent Correlation ◽

Time To Peak ◽

Study Population ◽

Lag Times

SummaryIn a study population consisting of healthy men (n=8), women not using oral contraceptives (OC) (n=28) and women using different kinds of OC (n=187) we used calibrated automated thrombography (CAT) in the absence and presence of added activated protein C (APC) to compare parameters that can be obtained from thrombin generation curves, i.e. lag time, time to peak, peak height and endogenous thrombin potential (ETP). Both with and without APC, plasmas of OC users exhibited the shortest lag time and time to peak, and the highest peak height and ETP. In the absence of APC none of these parameters differed between users of OC containing different progestogens. In contrast, in the presence of APC shorter lag times and time to peak, and higher peak height and ETP were observed in plasma of users of gestodene-,desogestrel-,drospirenone- and cyproterone acetate-containing OC than in plasma of users of levonorgestrel-containing OC. The ETP determined in the absence of APC (ETP-APC) had no predictive value for the APCsr (r=0.11; slope 0.9×10–3; 95%CI: –0.1×10–3 to 2.0×10–3) whereas the ETP measured in the presence of APC (ETP+APC) showed an excellent correlation with the APCsr (r=0.95; slope 6.6×10–3; 95%CI: 6.3×10–3 to 6.9×10–3) indicating that the APCsr is entirely determined by the ETP+APC. In conclusion, OC use increases thrombin generation, but differential effects of second and third generation OCs on the protein C system likely determine the differences in the risk of venous thrombosis between these kinds of OC.

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Abstract P683: Thrombin Generation in Plasma of Stroke Patients With Atrial Fibrillation

Stroke ◽

10.1161/str.52.suppl_1.p683 ◽

2021 ◽

Vol 52 (Suppl_1) ◽

Author(s):

Sarina Falcione ◽

Gina Sykes ◽

Joseph Kamtchum Tatuene ◽

Danielle Munsterman ◽

Twinkle Joy ◽

...

Keyword(s):

Atrial Fibrillation ◽

Ischemic Stroke ◽

Thrombin Generation ◽

Thrombus Formation ◽

Lag Time ◽

Thrombin Time ◽

Stroke Patients ◽

Time To Peak ◽

Generation Capacity

Background and Purpose: Thrombus formation is central to pathophysiology of stroke in patients with atrial fibrillation. Whether factors in plasma contribute to thrombus generation in patients with atrial fibrillation remains unclear. In this study we sought to determine whether plasma contributes to thrombin generation in patients with atrial fibrillation. Methods: There were 78 acute ischemic strokes with atrial fibrillation and 37 non-stroke controls. Plasma thrombin generation was measured by thrombin generation assay, resulting lag time, peak thrombin, time to peak and area under the curve was assessed. Thrombin generation capacity was compared in stroke patients with atrial fibrillation to non-stroke controls. The relationship to anticoagulation was assessed. In vitro, the effect of anticoagulation on plasma thrombin generation was determined. Results: Thrombin generation capacity was increased (shorter lag time and time to peak) in ischemic stroke patients with atrial fibrillation compared to non-stroke atrial-fibrillation controls (p<0.05 and p<0.01, respectively). Anticoagulation decreased plasma induced thrombin generation. Ischemic stroke patients with atrial fibrillation treated with anticoagulation (DOAC or warfarin) had lower plasma induced thrombin generation compared to atrial-fibrillation patients not on anticoagulation (p<0.05). Thrombin generation by plasma could be further reduced by DOAC in an in-vitro assay. Conclusions: Stroke patients with atrial fibrillation have a higher plasma induced thrombin generation compared to atrial fibrillation controls. Factors in plasma such as leukocyte derived tissue factor likely contribute to thrombus formation in patients with atrial fibrillation. As such, components in plasma may represent new targets to reduce thrombus formation and stroke risk in patients with atrial fibrillation.

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The APC-independent anticoagulant activity of protein S in plasma is decreased by elevated prothrombin levels due to the prothrombin G20210A mutation

Blood ◽

10.1182/blood-2003-02-0620 ◽

2003 ◽

Vol 102 (5) ◽

pp. 1686-1692 ◽

Cited By ~ 16

Author(s):

Rory R. Koenen ◽

Guido Tans ◽

René van Oerle ◽

Karly Hamulyák ◽

Jan Rosing ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Protein C ◽

Anticoagulant Activity ◽

Factor V Leiden ◽

Protein S ◽

Prothrombin G20210a ◽

Healthy Population ◽

Factor V ◽

Thrombotic Risk ◽

G20210a Mutation

AbstractProtein S exhibits anticoagulant activity independent of activated protein C (APC). An automated factor Xa–based one-stage clotting assay was developed that enables quantification of the APC-independent activity of protein S in plasma from the ratio of clotting times (protein S ratio [pSR]) determined in the absence and presence of neutralizing antibodies against protein S. The pSR was 1.62 ± 0.16 (mean ± SD) in a healthy population (n = 60), independent of plasma levels of factors V, VIII, IX, and X; protein C; and antithrombin, and not affected by the presence of factor V Leiden. The pSR strongly correlates with the plasma level of protein S and is modulated by the plasma prothrombin concentration. In a group of 16 heterozygous protein S–deficient patients, the observed mean pSR (1.31 ± 0.09) was significantly lower than the mean pSR of the healthy population, as was the pSR of plasma from carriers of the prothrombin G20210A mutation (1.47 ± 0.21; n = 46). We propose that the decreased APC-independent anticoagulant activity of protein S in plasma with elevated prothrombin levels may contribute to the thrombotic risk associated with the prothrombin G20210A mutation.

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Liraglutide in polycystic ovary syndrome: a randomized trial, investigating effects on thrombogenic potential

Endocrine Connections ◽

10.1530/ec-16-0113 ◽

2017 ◽

Vol 6 (2) ◽

pp. 89-99 ◽

Cited By ~ 10

Author(s):

Malin Nylander ◽

Signe Frøssing ◽

Caroline Kistorp ◽

Jens Faber ◽

Sven O Skouby

Keyword(s):

Thrombin Generation ◽

Polycystic Ovary ◽

Plasminogen Activator Inhibitor ◽

Lag Time ◽

Plasminogen Activator Inhibitor 1 ◽

Cvd Risk ◽

Ovary Syndrome ◽

Time To Peak ◽

Thrombogenic Potential ◽

Activator Inhibitor

Polycystic ovary syndrome (PCOS) is associated with increased risk of venous thromboembolism (VTE) and cardiovascular disease (CVD) in later life. We aimed to study the effect of liraglutide intervention on markers of VTE and CVD risk, in PCOS. In a double-blind, placebo-controlled, randomized trial, 72 overweight and/or insulin-resistant women with PCOS were randomized, in a 2:1 ratio, to liraglutide or placebo 1.8 mg/day. Endpoints included between-group difference in change (baseline to follow-up) in plasminogen activator inhibitor-1 levels and in thrombin generation test parameters: endogenous thrombin potential, peak thrombin concentration, lag time and time to peak. Mean weight loss was 5.2 kg (95% CI 3.0–7.5 kg, P < 0.001) in the liraglutide group compared with placebo. We detected no effect on endogenous thrombin potential in either group. In the liraglutide group, peak thrombin concentration decreased by 16.71 nmol/L (95% CI 2.32–31.11, P < 0.05) and lag time and time to peak increased by 0.13 min (95% CI 0.01–0.25, P < 0.05) and 0.38 min (95% CI 0.09–0.68, P < 0.05), respectively, but there were no between-group differences. There was a trend toward 12% (95% CI 0–23, P = 0.05) decreased plasminogen activator inhibitor-1 in the liraglutide group, and there was a trend toward 16% (95% CI −4 to 32, P = 0.10) reduction, compared with placebo. In overweight women with PCOS, liraglutide intervention caused an approximate 5% weight loss. In addition, liraglutide affected thrombin generation, although not significantly differently from placebo. A concomitant trend toward improved fibrinolysis indicates a possible reduction of the baseline thrombogenic potential. The findings point toward beneficial effects of liraglutide on markers of VTE and CVD risk, which should be further pursued in larger studies.

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Incidence of VTE in asymptomatic children with deficiencies of antithrombin, protein C, and protein S: a prospective cohort study

Blood Advances ◽

10.1182/bloodadvances.2020002781 ◽

2020 ◽

Vol 4 (21) ◽

pp. 5442-5448

Author(s):

Daniela Tormene ◽

Elena Campello ◽

Chiara Simion ◽

Giacomo Turatti ◽

Michelangelo Marobin ◽

...

Keyword(s):

Cohort Study ◽

Confidence Interval ◽

Prospective Cohort Study ◽

Prospective Cohort ◽

Protein C ◽

Protein S ◽

Healthy Children ◽

Thrombotic Risk ◽

Asymptomatic Children ◽

Risk Period

Abstract Although antithrombin, protein C, and protein S defects are well-recognized inherited risk factors for venous thromboembolism (VTE) in adults, whether they predispose children to these vascular disorders as well is undefined. In a prospective cohort study, we assessed the incidence of spontaneous and risk period–related VTE in children who were family members of adults who, after an episode of symptomatic VTE, had then been identified as carriers of these abnormalities. A total of 134 children from 87 families were enrolled. Seventy (51.5%) of these children were carriers of an inherited defect, and the remaining 64 were not; the mean observation period was 4 years (range, 1-16 years) and 3.9 years (range, 1-13), respectively. Sixteen risk periods were experienced by carriers, and 9 by noncarriers. Six VTE occurred in the 70 carriers during 287 observation-years, accounting for an annual incidence of 2.09% patient-years (95% confidence interval, 0.8-4.5), compared with none in the 64 noncarriers during 248 observation-years. Of the 14 children with thrombophilia who experienced a risk period for thrombosis, 4 (28.6%) developed a VTE episode. The overall incidence of risk-related VTE was 25% per risk period (95% confidence interval, 6.8-64). In conclusion, the thrombotic risk in otherwise healthy children with severe inherited thrombophilia does not seem to differ from that reported for adults with the same defects. Screening for thrombophilia in children who belong to families with these defects seems justified to identify those who may benefit from thromboprophylaxis during risk periods for thrombosis.

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Factor VIII Efficacy and Tissue Factor Levels in Hemophilia A Patients Undergoing Total Knee Replacement.

Blood ◽

10.1182/blood.v104.11.4029.4029 ◽

2004 ◽

Vol 104 (11) ◽

pp. 4029-4029

Author(s):

Wolfgang Wegert ◽

Manuela Krause ◽

Inge Scharrer ◽

Ulla Stumpf ◽

Andreas Kurth ◽

...

Keyword(s):

Total Knee Replacement ◽

Plasma Levels ◽

Thrombin Generation ◽

Hemophilia A ◽

Lag Time ◽

Major Surgery ◽

Plasma Samples ◽

Time To Peak ◽

Thrombin Generation Assay ◽

Total Knee

Abstract The changes of tissue factor (TF) blood levels in patients undergoing major surgery has been reported presenting controversial data. Whether this TF is hemostatically active or if it interacts with other coagulation factors, e.g. FVIII, is still unclear, making thrombotic risk and complications assessment for even more difficult. We analyzed plasma samples from four male patients aged 27–55 with severe hemophilia A without inhibitory antibodies, undergoing total knee replacement, which all gave informed consent. Initial FVIII doses before intervention was 75–80 U/kg. Treatment following intervention was targeted at 100 % FVIII serum levels. None received heparin. No bleeding events occurred during the observation period. The samples were taken at these timepoints (TP): 1. before preoperative FVIII substitution, 2. at the time of first incision (intervention start), 3. at circulation arrest release + 90 s after prosthesis implantation, 4. final suture (intervention end), 5. 24 h and 6. 48 h after intervention to assay procedurally induced TF production. Coagulation analyses were carried out using a fluorometric thrombin generation assay (TGA) in platelet rich plasma (PRP), RoTEG (rotation thrombelastography) in whole blood and a TF ELISA for the plasma samples’ TF levels. Both clotting function tests were started using TF diluted 1:100.000 and calcium chloride 16,7 mM (final conc.). TGA parameters were ETP, PEAK (maximum thrombin generation velocity), TIME TO PEAK, LAG TIME. TGA parameters directly related to thrombin activity (ETP; PEAK) showed no change during the intervention, but a sharp decrease 24 h later with a partial recovery 48 h later. TGA time marks (TIME TO PEAK, LAG TIME) changed in an inverse way, except for the difference from LAG TIME and TIME TO PEAK, which shortened continously after circulation arrest removal. RoTEG was characterized using 4 parameters: clotting time (CT), clot formation time (CFT), maximum clot firmness (MCF) and clot formation rate (CFR). After preoperative FVIII substitution, CT decreased by 10 % and CFT by 50 % in 48 h. MCF stayed unchanged during the intervention and the following 24 h, but increased by 20 % at 48 h. CFR increased by 10 % during intervention, and by 20 % from 24 to 48 h. TF ELISA showed preoperative TF plasma levels from 11 to 271 pg/ml. After release of circulation arrest (TP 3) TF concentration increased sharply (4 times the initial value), which was not detectable in the samples taken at TPs 2 and 4. TF levels further increased at TPs 5 and 6 to 170 % and 317 % resp. Altogether, TF plasma levels elevated after major surgery seem to correspond to a potential risk factor for postoperative thrombosis, especially when elevation is induced after intervention. However, functional coagulation assays do not change uniformly, as the thrombin generation assay reflects no marked changes under intervention, but in the period after(24–48 h). Changes in the RoTEG whole blood clotting assay are not dramatic but indicate a thrombophilic shift in coagulation balance also pronouned at 24–48h, too. These results demonstrate that increased coagulability after orthopedic surgery detected using functional clotting assays correlates with increased TF levels, but further studies must be performed to prove this relation in healthy individuals.

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