Plasma Concentrations Of Thrombin-antithrombin Complex Research Articles

Extracellular vesicles and coagulation in blood from healthy humans revisited

ABSTRACTBackground: In 2001, we studied the presence and coagulant properties of “microparticles” in the blood of healthy humans. Since then, multiple improvements in detection, isolation and functional characterization of the now called “extracellular vesicles” (EVs) have been made, and shortcomings were identified. Aim: To revisit the presence and function of EVs in blood from healthy humans. Methods: Blood was collected from 20 healthy donors. EV-containing plasma was prepared according to new guidelines, and plasma was diluted to prevent swarm detection. Single EVs were measured by flow cytometry with known sensitivity of fluorescence and light scatter. The haemostatic properties of EVs were measured by thrombin-, fibrin-, and plasmin generation. Plasma concentrations of thrombin-antithrombin complexes and prothrombin fragment 1 + 2 were measured to assess the coagulation status in vivo. Results: Compared to 2001, the total concentrations of detected EVs increased from 190- to 264-fold. In contrast to 2001, however, EVs are non-coagulant which we show can be attributed to improvements in blood collection and plasma preparation. No relation is present between the plasma concentrations of EVs and either TAT or F1 + 2. Finally, we show that EVs support plasmin generation. Discussion: Improvements in blood collection, plasma preparation and detection of EVs reveal that results from earlier studies have to be interpreted with care. Compared to 2001, higher concentrations of EVs are detected in blood of healthy humans which promote fibrinolysis rather than coagulation.

SEPCR Levels in Chronic Myeloproliferative Diseases and Their Association with Thromboembolic Events: A Case-Control Study.

Objective: Venous, arterial, and microcirculatory events are frequently encountered in the clinical course of essential thrombocytosis and polycythemia vera. We aimed to investigate the levels of soluble endothelial protein C receptor (sEPCR) in myeloproliferative diseases to see whether there was a difference between the patients with and without history of thromboembolism. Materials and Methods: The study included patients with polycythemia vera (n=12), patients with essential thrombocytosis (n=13), and controls (n=29). In all groups, we measured proteins C and S, antithrombin and sEPCR levels, and plasma concentrations of thrombin-antithrombin complex, prothrombin fragments 1+2, and D-dimer. Results: Comparing the patients with and without history of thromboembolic attack, statistically significant differences were not observed in terms of sEPCR, D-dimer, thrombin-antithrombin complex, prothrombin fragments 1+2, and hematocrit levels (p=0.318, 0.722, 0.743, 0.324, and 0.065, respectively). Conclusion: Significant increase in the parameters that reflect activation of coagulation, such as sEPCR, thrombin-antithrombin complex, prothrombin fragments 1+2, and D-dimer, reflects the presence of a basal condition that leads to a tendency toward thrombosis development in ET and PV when compared to healthy controls.

In vivo anticoagulant effect of ethyl pyruvate in endotoxemic rats

Introduction Disseminated intravascular coagulation (DIC), a disorder of the blood coagulation function, is a common complication of severe sepsis. Ethyl pyruvate (EP), a derivative of pyruvate, is known for its anti-inflammatory and antioxidant properties. Recently, EP showed anticoagulant effects in vitro. Therefore, the aim of study is to investigate the in vivo anticoagulant effect of EP on a severe DIC model induced by lipopolysaccharide (LPS) in anesthetized Wistar rats. Materials and Methods The animals were intravenously infused with LPS (30 mg/kg) over a period of 4 h. One hour after LPS initiation, rats were treated with EP in three dosages (20, 40 and 60 mg/kg/4 h, i.v.). Results The intermediate and high doses of EP (40 and 60 mg/kg) prevented circulatory failure, improved renal and hepatic function, and reduced the plasma levels of TNF-α and IL-6 after LPS administration. All used doses of EP significantly prevented prothrombin time prolongation, and reduced mRNA expression of tissue factor in lung tissue induced by LPS. The two higher doses of EP attenuated plasma concentrations of thrombin-antithrombin complex. The high dose of EP (60 mg/kg) significantly preserved platelet counts, and improved survival rate. However, EP did not reduce the elevation of plasma plasminogen activator inhibitor-1 during endotoxemia. Conclusion EP demonstrates the in vivo anticoagulant effect and improved organ functions in a severe DIC model in rats, which is likely associated with the inhibitory effect on tissue factor mRNA expression and cytokines release. Its effectiveness in preventing DIC is not mediated by increase in fibrinolysis.

Effect of Preoperative Biliary Drainage on Coagulation and Fibrinolysis in Severe Obstructive Cholestasis

To evaluate the function of coagulation and fibrinolysis in cholestatic patients before and after preoperative biliary drainage (PBD). Cholestasis owing to an obstructive biliary malignancy is associated with postoperative complications related to a proinflammatory state, an impaired hepatic synthesis function, and a potential derangement of hemostasis. Hence, PBD is advocated for cholestatic patients undergoing major surgery. Plasma coagulation and fibrinolytic parameters were assessed in 24 cholestatic patients and 10 controls. In 9 cholestatic patients, the parameters were reassessed at least 4 weeks after PBD. Compared with controls, cholestatic patients showed lower concentrations (P<0.001) of plasma vitamin K-dependent factors II and VII, whereas prothrombin time, activated partial thromboplastin time, and factor V were unaltered. Thrombin generation was increased in cholestatic patients, as reflected by higher plasma concentrations of thrombin-antithrombin complexes and D-dimers. Fibrinolysis was significantly impaired as evidenced by low plasminogen activator activity (PAA) owing to an increase in plasminogen activator inhibitor -1). Elevated markers for thrombin generation thrombin-antithrombin decreased after PBD from 10.7±1.2 to 5.7±0.7 ng/mL (P<0.05). Additionally, impairment of fibrinolysis in cholestatic patients resolved after PBD (plasminogen activator inhibitor-1 levels decreased from 19±1 to 10±1 IU/mL and plasminogen activator activity increased from 82±3 to 110±4%, respectively). D-dimers remained unaltered after PBD, likely because of normalization of coagulation and fibrinolytic activity. Obstructive cholestasis is associated with a procoagulant state, despite an impaired vitamin K-dependent coagulation factor synthesis. Virtually all alterations in coagulation and fibrinolysis were reversed by biliary drainage.

Coagulation And Fibrinolytic Responses To Exercise In Hot And Cold Temperatures

Most adverse cardiovascular events are the result of an occlusive blood clot. During exercise, ischemic risk transiently increases, possibly due to exertion-related changes in coagulation and fibrinolytic activity. Previous research suggests ambient temperature affects resting coagulation and fibrinolytic potential, but the effect of heat and cold on hemostasis during exercise is unknown. PURPOSE: To assess changes in coagulation and fibrinolysis during maximal exercise in hot and cold temperatures, and to compare those responses to exercise under normal, temperate conditions. METHODS: Fifteen healthy male subjects (25.3 ± 4.3 years) completed maximal cycle ergometer exercise tests in hot (30°C), temperate (20°C) and cold (5° or 8° C) temperatures. Blood samples were obtained before and immediately after exercise and analyzed by ELISA to determine plasma concentrations of thrombin-antithrombin complex (TAT), tissue plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1). Results were analyzed with a two-factor ANOVA using temperature and time (pre-, post-exercise) as within-subjects factors. Post-hoc pairwise comparisons were made using Fisher‘s LSD method. RESULTS: VO2peak was not different among the three temperatures (temperate = 40.7±8.7, hot = 41.8±9.4, cold = 41.2±9.2 ml/kg/min, p>0.05). A main effect of time was observed for TAT (temperate = 0.45±0.26 - 1.04±0.44 ng/ml, hot = 0.64±0.55 - 1.18±0.95 ng/ml, cold = 0.88±0.57 - 1.34±0.79 ng/ml, PRE to POST, respectively, p<0.01) and tPA activity (temperate = 0.72 ± 0.44 - 2.71±0.55 IU/ml, hot = 0.72±0.38 - 2.64±0.61 IU/ml, cold = 0.86±0.45 - 2.65±0.77 IU/ml, PRE to POST, respectively, p<0.01). A non-significant trend was observed for the PAI-1 response to exercise (temperate = 14.5±23.7 - 12.3±20.2 IU/ml, hot = 15.1±26.5 - 10.0±15.1 IU/ml, cold = 10.5±10.4 - 7.9±9.7 IU/ml, PRE to POST, respectively, p=0.08). Cold temperature resulted in significantly higher TAT concentrations than temperate conditions (p=0.05). No other main effects of temperature or time x temperature interactions were observed. CONCLUSIONS: The potential for clot formation is elevated during exposure to cold temperatures. These data suggest that risk of an ischemic event may be elevated in the cold.

Impact of Decreased Renal Function on Coagulation and Fibrinolysis in Patients With Non-Valvular Atrial Fibrillation

The prothrombotic state in non-valvular atrial fibrillation (NVAF) increases the risk of thromboembolic events. It is also well known that a prothrombotic state exists in renal failure. Hemostatic disorders play a role in thromboembolic mortality in both NVAF and renal failure. However, little is known about the influence of renal function on hemostasis in patients with NVAF. A total of 190 consecutive outpatients with NVAF who were not receiving anticoagulant therapy were enrolled in the present study. Patients were stratified in groups based on estimated glomerular filtration rate (eGFR). Plasma concentrations of thrombin-antithrombin complex (TAT) and D-dimer were measured and the influence of renal function on coagulation and fibrinolysis was assessed. A relatively weak and inverse relationship of eGFR to TAT and D-dimer was observed (r=0.28, P=0.0001; r=0.30, P<0.0001). Both mean TAT and D-dimer concentrations in groups gradually and significantly increased as eGFR decreased (P<0.0001). On multivariate regression analysis, decreased eGFR concentration was a significant predictor for elevation of TAT (P<0.05) and D-dimer (P<0.01) in patients with NVAF. The enhanced coagulation activation appeared to be related to a reduction in residual renal function in patients with NVAF. This suggests that decreased renal function might be a candidate predictor of thromboembolic events in patients with NVAF.

Hyperglycemia enhances coagulation and reduces neutrophil degranulation, whereas hyperinsulinemia inhibits fibrinolysis during human endotoxemia

AbstractType 2 diabetes is associated with altered immune and hemostatic responses. We investigated the selective effects of hyperglycemia and hyperinsulinemia on innate immune, coagulation, and fibrinolytic responses during systemic inflammation. Twenty-four healthy humans were studied for 8 hours during clamp experiments in which either plasma glucose, insulin, both, or none was increased, depending on randomization. Target plasma concentrations were 5 versus 12 mM for glucose, and 100 versus 400 pmol/L for insulin. After 3 hours, 4 ng/kg Escherichia coli endotoxin was injected intravenously to induce a systemic inflammatory and procoagulant response. Endotoxin administration induced cytokine release, activation of neutrophils, endothelium and coagulation, and inhibition of fibrinolysis. Hyperglycemia reduced neutrophil degranulation (plasma elastase levels, P < .001) and exaggerated coagulation (plasma concentrations of thrombin-antithrombin complexes and soluble tissue factor, both P < .001). Hyperinsulinemia attenuated fibrinolytic activity due to elevated plasminogen activator-inhibitor-1 levels (P < .001). Endothelial cell activation markers and cytokine concentrations did not differ between clamps. We conclude that in humans with systemic inflammation induced by intravenous endotoxin administration hyperglycemia impairs neutrophil degranulation and potentiates coagulation, whereas hyperinsulinemia inhibits fibrinolysis. These data suggest that type 2 diabetes patients may be especially vulnerable to prothrombotic events during inflammatory states.

Platelet-Activating Factor Receptor Contributes to Host Defense against Pseudomonas aeruginosa Pneumonia but Is Not Essential for the Accompanying Inflammatory and Procoagulant Response

Pseudomonas aeruginosa is a major cause of nosocomial pneumonia, which is associated with high morbidity and mortality. Because of its ubiquitous nature and its ability to develop resistance to antibiotics, it is a problematic pathogen from a treatment perspective. Platelet-activating factor receptor (PAFR) is involved in phagocytosis of several pathogens. To determine the role of PAFR in the innate immune response to P. aeruginosa pneumonia, pafr gene-deficient (PAFR-/-) mice and normal wild-type (Wt) mice were intranasally inoculated with P. aeruginosa. PAFR deficiency impaired host defense as reflected by increased bacterial outgrowth and dissemination in mice with a targeted deletion of the PAFR gene. PAFR-/- neutrophils showed a diminished phagocytosing capacity of P. aeruginosa in vitro. Relative to Wt mice, PAFR-/- mice demonstrated increased lung inflammation and injury as reflected by histopathology, relative lung weights and total protein concentrations in bronchoalveolar lavage fluid, which was accompanied by higher levels of proinflammatory cytokines in lung homogenates and plasma. In addition, PAFR deficiency was associated with exaggerated local and systemic activation of coagulation as determined by fibrin staining of lung tissue and pulmonary and plasma concentrations of thrombin-antithrombin complexes and D-dimer. These data suggest that PAFR is an essential component of an effective host response to P. aeruginosa pneumonia, at least partly via its contribution to the phagocytic properties of professional granulocytes. Additionally, our results indicate that PAFR signaling is not essential for the induction of a local and systemic inflammatory and procoagulant response to Pseudomonas pneumonia.

Evaluation of coagulation markers in the plasma of healthy cats and cats with asymptomatic hypertrophic cardiomyopathy

Thrombosis and arterial thromboembolism are frequent complications of feline cardiomyopathy, especially when associated with left atrial enlargement. Markers of activated coagulation may be used to evaluate the coagulation status of cats with hypertrophic cardiomyopathy (HCM) in relation to left atrial size. The objective of this study was to compare plasma concentrations of thrombin-antithrombin complex (TAT), D-dimer, and fibrin degradation products (FDP) between clinically healthy cats and cats with HCM. Prothrombin time (PT), activated partial thromboplastin time (aPTT), and antithrombin activity were also compared and the association between left atrial (LA) size and coagulation results in cats with HCM was evaluated. Blood samples from 19 clinically healthy cats and 20 cats with HCM were obtained. All cats with HCM were asymptomatic and had no signs of heart failure. LA diameter and LA to proximal aortic (Ao) diameter ratio (LA:Ao) were determined by echocardiography. Reference intervals for D-dimer and TAT concentrations in plasma of healthy cats were established as 0.09-0.32 microg/mL and 2.0-20.0 microg/L, respectively. TAT, D-dimer, and FDP concentrations were increased in 5, 3, and 2 cats with HCM, respectively. TAT and D-dimer concentrations, and PT and aPTT were not significantly different between groups. Antithrombin activity was significantly decreased in cats with HCM (P=.03) despite marked range overlap. LA and LA:Ao were not correlated with coagulation results. Laboratory evidence of hypercoagulability was found in 45% of cats with HCM. Left atrial size was not associated with laboratory evidence of hypercoagulability. Association between coagulation markers and risk of thrombosis has yet to be evaluated in cats with HCM.

Prednisolone Dose-Dependently Influences Inflammation and Coagulation during Human Endotoxemia

The effects of steroids on the outcome of sepsis are dose dependent. Low doses appear to be beneficial, but high doses do not improve outcome for reasons that are insufficiently understood. The effects of steroids on systemic inflammation as a function of dose have not previously been studied in humans. To determine the effects of increasing doses of prednisolone on inflammation and coagulation in humans exposed to LPS, 32 healthy males received prednisolone orally at doses of 0, 3, 10, or 30 mg (n = 8 per group) at 2 h before i.v. injection of Escherichia coli LPS (4 ng/kg). Prednisolone dose-dependently inhibited the LPS-induced release of cytokines (TNF-alpha and IL-6) and chemokines (IL-8 and MCP-1), while enhancing the release of the anti-inflammatory cytokine IL-10. Prednisolone attenuated neutrophil activation (plasma elastase levels) and endothelial cell activation (von Willebrand factor). Most remarkably, prednisolone did not inhibit LPS-induced coagulation activation, measured by plasma concentrations of thrombin-antithrombin complexes, prothrombin fragment F1+2, and soluble tissue factor. In addition, activation of the fibrinolytic pathway (tissue-type plasminogen activator and plasmin-alpha(2)-antiplasmin complexes) was dose-dependently enhanced by prednisolone. These data indicate that prednisolone dose-dependently and differentially influences the systemic activation of different host response pathways during human endotoxemia.

Vagus nerve stimulation inhibits activation of coagulation and fibrinolysis during endotoxemia in rats.

Sepsis and endotoxemia are associated with concurrent activation of inflammation and the hemostatic mechanism, which both contribute to organ dysfunction and death. Electrical vagus nerve stimulation (VNS) has been found to inhibit tumor necrosis factor (TNF)-alpha release during endotoxemia in rodents. To determine the effect of VNS on activation of coagulation and fibrinolysis. Rats received a sublethal i.v. dose of lipopolysaccharide (LPS) after electrical VNS or sham stimulation. Activation of coagulation and fibrinolysis, as well as cytokine release, was measured before LPS injection and 2, 4 and 6 h thereafter. LPS induced activation of the coagulation system (increases in the plasma concentrations of thrombin-antithrombin complexes and D-dimer, and a decrease in antithrombin) and biphasic changes in the fibrinolytic system [early rises of plasminogen activator activity and tissue-type plasminogen activator, followed by a delayed increase in plasminogen activator inhibitor type 1 (PAI-1)]. VNS strongly inhibited all LPS-induced procoagulant responses and more modestly attenuated the fibrinolytic response. In addition, VNS attenuated the LPS-induced increases in plasma and splenic concentrations of the proinflammatory cytokines TNF-alpha and interleukin-6 (IL-6), while not influencing the release of the anti-inflammatory cytokine IL-10. These data illustrate a thus far unrecognized effect of VNS and suggest that the cholinergic anti-inflammatory pathway not only impacts on inflammation but also on the coagulant-anticoagulant balance.

Decreased protein C activation in patients with fulminant hepatic failure

Objective. Abnormalities of the blood coagulation system have an influence on outcome in patients with fulminant hepatic failure (FHF). The protein C (PC) pathway is one of the main modulators of the blood coagulation system. The role of the PC pathway in FHF is not clear. In the present study, we evaluated endothelial cell injury and the grade of activated protein C (APC) generation in FHF patients. Material and methods. The effect of APC on the expression of tumor necrosis factor (TNF)-α and monocyte chemoattractant protein (MCP)-1 from LI90 stellate cells was also evaluated. This study comprised 5 patients with FHF, 6 with acute hepatitis (AH), 12 with chronic hepatitis (CH) and 20 healthy subjects. Results. The plasma concentrations of thrombin-antithrombin complex and thrombomodulin were significantly increased in FHF patients compared with those in AH patients and healthy subjects. The circulating levels of activated protein C-protein C inhibitor (APC-PCI) complex and the APC-PCI/PC ratio were significantly decreased in patients with FHF compared to healthy controls. APC significantly inhibited in vitro the expression of TNFα and MCP-1 from LI90 stellate cells. Conclusions. This study demonstrated enhanced endothelial cell injury in association with decreased PC activation and hypercoagulability in FHF.

Early antithrombotic and anti-inflammatory effects of simvastatin versus fenofibrate in patients with hypercholesterolemia

The aim of the study was to determine whether a short-term treatment with simvastatin or fenofibrate may result in beneficial anti-inflammatory and antithrombotic effects in patients with high risk of coronary artery disease. In a randomized, double-blind study, we compared markers of inflammation, thrombin formation and platelet activation in patients with LDL cholesterol >130 mg/dl assigned to receive simvastatin (40 mg/d; n=20) or micronised fenofibrate (160 mg/d; n=22) for 28 days. Simvastatin, but not fenofibrate, lowered C-reactive protein (CRP) by 32% on day 3 (p<0.001), while both drugs reduced CRP significantly on day 28. Interleukin-6, soluble CD40 ligand, and monocyte chemoattractant protein-1 levels decreased significantly (by 20 to 50%) in both treatment groups on days 3 and 28. Soluble cell adhesion molecules remained unchanged in both groups. Simvastatin and fenofibrate significantly lowered plasma concentrations of thrombin-antithrombin complexes on days 3 and 28, but not platelet beta-thromboglobulin (betaTG) levels. Soluble P-selectin was lowered only in the simvastatin group. The total amount of thrombin generated at the site of microvascular injury also declined (by about 30%) as early as after 3 days of fenofibrate or simvastatin therapy, whereas beta TG release was reduced only in the simvastatin group on days 3 and 28. All the effects were independent of the changes in lipid profiles. Our results suggest that statins and fibrates can exert antithrombotic and anti-inflammatory effects as early as after 3 days of therapy. However, in contrast to statins, fibrates have no influence on platelet function within one month of therapy.

Activation of coagulation system in preterm labor and preterm premature rupture of membranes

Objective: Thrombin, originally discovered as a coagulation factor, is a multifunctional protease capable of inducing myometrial contractions in vitro and in vivo. This enzyme has been implicated in the mechanisms of premature labor. Plasma concentrations of thrombin-antithrombin (TAT) complexes are an index of in vivo thrombin generation. The purpose of this study was to determine whether patients with premature labor and preterm premature rupture of membranes (PROM) have evidence of increased thrombin generation in maternal blood, as determined by the TAT complex concentrations. Methods: A cross-sectional study was designed to determine plasma concentrations of TAT complexes in 110 women in the following groups: non-pregnant women (n = 20); normal pregnant women (n = 30); women in preterm labor with intact membranes (n = 30); and women with preterm PROM (n = 30). TAT complex concentrations were determined with a sensitive and specific immunoassay. Statistical analysis was conducted with non-parametric statistics. Results: Patients with preterm labor and intact membranes had a significantly higher median plasma TAT complex concentration than normal pregnant women (women in preterm labor, median 19.1 μg/l; range 7.4-406 vs. normal pregnant women, median 15 μg/l; range 6.8-32.5; p = 0.03). Patients with preterm PROM had a higher median TAT complex concentration than normal pregnant women (preterm PROM, median 19.1 μg/l; range 4.7-738.6 vs. normal pregnant women, median 15 μg/l; range 6.8-32.5; p = 0.03). Normal pregnancy was associated with a higher median plasma TAT complex concentration than the non-pregnant state (normal pregnant women, median 15 μg/l; range 6.8-32.5 vs. non-pregnant women, median 2.7 μg/l; range 0.9-14.2; p < 0.001). Conclusion: Preterm labor and preterm PROM are associated with an excess generation of thrombin.

Activation of coagulation system in preterm labor and preterm premature rupture of membranes

Objective: Thrombin, originally discovered as a coagulation factor, is a multifunctional protease capable of inducing myometrial contractions in vitro and in vivo. This enzyme has beenimplicated in the mechanisms of premature labor. Plasma concentrations of thrombin-antithrombin (TAT) complexes are an index of in vivo thrombin generation. The purpose of this study was to determinewhether patients with premature labor and preterm premature rupture of membranes (PROM) have evidence of increased thrombin generation in maternal blood, as determined by the TAT complex concentrations.Methods: A cross-sectional study was designed to determine plasma concentrations of TAT complexes in 110 women in the following groups: non-pregnant women (n = 20); normal pregnant women(n = 30); women in preterm labor with intact membranes (n = 30); and women with preterm PROM (n = 30). TAT complex concentrations were determined with a sensitive and specific immunoassay.Statistical analysis was conducted with non-parametric statistics.Results: Patients with preterm labor and intact membranes had a significantly higher median plasma TAT complex concentrationthan normal pregnant women (women in preterm labor, median 19.1 μg/l; range 7.4-406 vs. normal pregnant women, median 15 μg/l; range 6.8-32.5; p = 0.03). Patients with preterm PROM had a highermedian TAT complex concentration than normal pregnant women (preterm PROM, median 19.1 μg/l; range 4.7-738.6 vs. normal pregnant women, median 15 μg/l; range 6.8-32.5; p = 0.03). Normal pregnancywas associated with a higher median plasma TAT complex concentration than the non-pregnant state (normal pregnant women, median 15 μg/l; range 6.8-32.5 vs. non-pregnant women, median 2.7 μg/l; range0.9-14.2; p < 0.001).Conclusion: Preterm labor and preterm PROM are associated with an excess generation of thrombin.

Evidence of in vivo generation of thrombin in patients with small-for-gestational-age fetuses and pre-eclampsia

Objective: Thrombotic lesions in the maternal or fetal compartments are frequently observed in the placentas of patients with small-for-gestational-age (SGA) fetuses and in pre-eclampsia. The objectiveof this study was to determine whether there was evidence of in vivo generation of thrombin, the ratelimiting enzyme responsible for the formation of fibrin. The plasma concentrations of thrombin-antithrombin(TAT) complexes were used as an index of thrombin generation.Methods: TAT complexes were measured in the plasma from 68 women from the following groups: normal pregnancy (n = 29); pre-eclampsia(n = 26); and SGA (defined as estimated fetal weight below the 10th centile for gestational age, which was confirmed by neonatal birth weight) (n = 13). TAT complex plasma concentrations weredetermined with a specific and sensitive immunoassay. Statistical analysis was performed with non-parametric statistics.Results: The median plasma TAT complex concentrations were significantlyhigher in patients who delivered SGA neonates than in normal pregnant women (SGA, median 24.2 μg/l; range 11.9-788.7 vs. normal pregnancy, median: 14.4 μg/l; range 6.8-26.9; p = 0.001). Patientswith pre-eclampsia had a higher median plasma TAT complex concentration than normal pregnant women (pre-eclampsia, median 18.1 μg/l; range 10.0-75.2 vs. normal pregnancy, median 14.4 μg/l; range 6.8-26.9;p = 0.02).Conclusion: In vivo generation of thrombin, determined by the plasma concentrations of TAT complexes, is higher in patients with SGA fetuses and patients with pre-eclampsiathan in normal pregnancy.

Laboratory signs of activated coagulation are common in Henoch-Sch�nlein purpura

We investigated 17 patients with Henoch-Schönlein purpura (HSP) and describe as yet unreported abnormal results of blood coagulation tests. In parallel to the activity of the disease, D-dimer concentrations in plasma were found to be significantly increased in 15 of the 17 patients; almost 50% of all patients showed values higher than 10 times the upper limit of the normal range. In 11 patients, plasma concentrations of thrombin-antithrombin complex (TAT) and prothrombin fragments 1 and 2 (F1+2) were examined; six of them showed abnormal results. The pathologic values were correlated to the activity of the disease, but abnormalities were also found in milder cases of HSP. These findings probably reflect local reactions within inflamed blood vessels rather than a systemic activation of coagulation and hyperfibrinolysis. Clinicians should be aware of these laboratory findings in order not to confuse common cases of HSP with purpura necroticans, a very severe type of vasculitis in which signs of disseminated intravascular coagulation (DIC) have been reported. Our findings suggest that an activation of coagulation including hyperfibrinolysis secondary to the endothelial damage is a typical feature of the common types of HSP.

Cell-derived Microparticles Circulate in Healthy Humans and Support Low Grade Thrombin Generation

We determined the numbers, cellular origin and thrombin-generating properties of microparticles in healthy individuals (n = 15). Microparticles, isolated from fresh blood samples and identified by flow cytometry, originated from platelets [237 x 10(6)/L (median; range 116-565)], erythrocytes (28 x 10(6)/L; 13-46), granulocytes (46 x 10(6)/L; 16-94) and endothelial cells (64 x 10(6)/L; 16-136). They bound annexin V, indicating surface exposure of phosphatidylserine, and supported coagulation in vitro. Interestingly, coagulation occurred via tissue factor (TF)-independent pathways, because antibodies against TF or factor (F)VII were ineffective. In contrast, in our in vitro experiments coagulation was partially inhibited by antibodies against FXII (12%, p = 0.006), FXI (36%, p <0.001), FIX (28%, p <0.001) or FVIII (32%, p <0.001). Both the number of annexin V-positive microparticles present in plasma and the thrombin-generating capacity inversely correlated to the plasma concentrations of thrombin-antithrombin complex (r = -0.49, p = 0.072 and r = -0.77, p = 0.001, respectively), but did not correlate to prothrombin fragment F1+2 (r = -0.002, p = 0.99). The inverse correlations between the number of microparticles and their thrombin-forming capacity and the levels of thrombin-antithrombin complex in plasma may indicate that microparticles present in the circulation of healthy individuals have an anticoagulant function by promoting the generation of low amounts of thrombin that activate protein C. We conclude that microparticles in blood from healthy individuals support thrombin generation via TF- and FVII-independent pathways, and which may have an anticoagulant function.

Effects of N omega-nitro-L-arginine methyl ester on the endotoxin-induced disseminated intravascular coagulation in porcine septic shock.

Nitric oxide is known to prevent platelet aggregation and clot formation. Inhibitors of nitric oxide synthase might promote or enhance endotoxin disseminated intravascular coagulation. The present study was designed to evaluate the effects of the arginine analog, N omega-nitro-L-arginine methyl ester (L-NAME), on the endotoxin-induced disseminated intravascular coagulation in a porcine model of septic shock. Prospective, comparative, experimental study. Laboratory at a large university hospital. Sixteen female piglets, weighing 20 to 28 kg. Three groups of animals were studied: a control group (n = 6); a lipopolysaccharide (LPS)-treated group (n = 5) receiving Escherichia coli endotoxin (5 micrograms/kg/min over 30 mins); and an LPS + L-NAME group (n = 5) receiving endotoxin and, 1 hr after, a bolus of L-NAME (25 mg/kg). Hemodynamic changes, usual coagulation parameters, and plasma concentrations of thrombin-antithrombin complexes, antithrombin III activity (At III), tissue plasminogen activator, plasminogen activator inhibitor type 1, and von Willebrand factor were measured at baseline, and at 30, 60, 90, 120, 180, 240, and 300 mins. After euthanasia or death, lungs and kidneys were withdrawn for histologic study. The extent of microvascular thrombosis was assessed by a semiquantitative disseminated intravascular coagulation score. In both septic endotoxin group, administration of LPS resulted in hemodynamic changes typical of severe septic shock, with disseminated intravascular coagulation and histologic changes characterized by adult respiratory distress syndrome and kidney microthrombosis. L-NAME administration normalized mean arterial pressure with a dramatic increase in systemic vascular resistances and a marked decrease in cardiac index. The changes in usual coagulation parameters, AT III, tissue plasminogen activator, and plasminogen-activator inhibitor type 1 concentrations were not different between both septic groups. However, in the LPS + L-NAME group, thrombin-antithrombin complexes and von Willebrand factor were higher and associated with a higher histologic disseminated intravascular coagulation score. In this model of endotoxin septic shock, L-NAME administration resulted in histologic and coagulation changes consistent with an increased activation of intravascular coagulation.

Gemfibrozil Reduces Thrombin Generation in Patients with Combined Hyperlipidaemia, without Influencing Plasma Fibrinogen, Fibrin Gel Structure or Coagulation Factor VII

A double-blind, placebo-controlled, cross-over study was conducted in 21 men with combined hyperlipoproteinaemia to examine if lipid-lowering treatment with gemfibrozil (10-12 weeks) affects blood coagulation and fibrin gel structure at rest or during mental stress. Gemfibrozil lowered plasma triglycerides by 57 +/- 4%, whereas high density lipoprotein (HDL) cholesterol increased by 22 +/- 5%. Gemfibrozil lowered the triglyceride content of low density lipoprotein (LDL). Gemfibrozil reduced the plasma concentrations of thrombin-antithrombin complex (TAT) and prothrombin fragment F1 + 2 (F1 + 2), both at rest and during mental stress. However, there were no effects of gemfibrozil treatment on the plasma concentrations of fibrinogen, factor VII antigen, activated factor VII (VIIa) or activated factor XII (XIIa), or on fibrin gel structure. Acute mental stress per se did not influence coagulation factors, reaction products or fibrin gel structure, or their responses to the study drug. Thus, gemfibrozil reduced thrombin generation in men with combined hyperlipoproteinaemia, without influencing the plasma levels of fibrinogen, VIIa and XIIa, or fibrin gel structure. Attenuation of thrombin generation may contribute to the primary-preventive effects of gemfibrozil on coronary heart disease.