thromboplastin and Meningococcal-Infections

Neisseria meningitidis causes fulminant meningococcal sepsis with a massive activation of the coagulation and complement cascades. Bacterial cell envelope molecules from N. meningitidis, particularly lipopolysaccharide (LPS), induce tissue factor (TF) expression. In meningococcal sepsis, TF can be detected on circulating monocytes and microparticles (MPs) within the bloodstream. During infection, Nm activates C5 and C5a, which also is able to induce TF. We evaluated the effect of eculizumab, a C5-blocking monoclonal antibodies (mAb), on cell- and MP-associated TF. Using a lepirudin-anticoagulated whole blood model, we activated the coagulation and complement cascades by N. meningitidis, and investigated the interaction between the cascade systems with special focus on cell-associated TF-expression (mRNA and protein) and MP-associated TF-dependent thrombin and fibrin generation in platelet-free plasma. We also examined the ability of TF-positive MPs to support clot formation in whole blood. In addition, the effect of corn trypsin inhibitor and time-dependent changes on MP-associated functional TF activity was examined. Inhibition of C5 reduced cell-associated TF expression at both gene and protein level, and reduced MP-associated TF-dependent thrombin and fibrin generation in platelet-poor plasma, MP-induced TF-dependent clot formation in whole blood, implying that the complement and coagulation cascades are interplayers in N. meningitidis-mediated activation of these cascades.

Topics: Antibodies, Monoclonal, Humanized; Anticoagulants; Blood Coagulation; Blood Platelets; Cell-Derived Microparticles; Complement Activation; Complement C5; Hirudins; Humans; Meningococcal Infections; Neisseria meningitidis; Recombinant Proteins; Sepsis; Thromboplastin

The plasma level of bacterial lipopolysaccharides (LPS) is associated with activation of the coagulation system, inhibition of fibrinolysis and the nature of the clinical presentation and outcome in patients with meningococcal disease. Tissue factor (TF)-bearing microparticles (MPs) appear to contribute to the pathogenesis of disseminated intravascular coagulation (DIC). The aim of this study was to investigate the relationship between MP-associated TF activity and the level of bacterial LPS in plasma from patients with meningococcal septic shock and meningitis.. MPs isolated from citrated plasmas were assessed for TF-dependent activity with both a plasma-based thrombin generation assay (CAT) and whole blood-based thromboelastometry (ROTEM). The LPS level was measured using a chromogenic Limulus amebocyte lysate assay.. MPs obtained from patients with meningococcal septic shock initiated significantly more efficient and TF-dependent thrombin generation in the CAT assay compared to MPs from patients with meningococcal meningitis. Differences in MP-associated TF activity between the septic shock patients and the meningitis patients were also evident when MPs were added to whole blood using ROTEM. The level of plasma LPS in patients with septic shock (range 2-2,100 EU/mL) was correlated with thrombogram parameters in the CAT assay; lagtime (r(s)=-0.84), time to peak (rs=-0.83), peak (r(s)=0.85) and ETP (r(s)=0.83).. MPs obtained from patients with meningococcal septic shock displayed more efficient TF-dependent thrombin generation and clot formation compared to MPs from meningitis patients. MP-associated TF activity was closely associated with plasma LPS levels in the septic shock group.

Topics: Adolescent; Adult; Cell-Derived Microparticles; Child; Child, Preschool; Female; Humans; Lipopolysaccharides; Male; Meningitis, Meningococcal; Meningococcal Infections; Middle Aged; Shock, Septic; Thromboplastin; Young Adult

Neisseria meningitidis causes sepsis with coagulopathy. The present study evaluated the tissue factor (TF)-inducing capacity of bacterial LPS in different presentation forms, i.e. membrane-bound LPS versus purified LPS, and of non-LPS components of N. meningitidis. By using a wild-type N. meningitidis, a mutant N. meningitidis lacking LPS (LPS-deficient N. meningitidis), purified LPS from N. meningitidis and Escherichia coli, we measured TF-expression and TF-activity on human monocytes and microparticles (MPs). The effect of TF-modulators, such as phosphatidylserine (PS), tissue factor pathway inhibitor (TFPI) and recombinant IL-10 (rhIL-10) was investigated. In plasmas from meningococcal patients, fibrinopeptide A (FPA), LPS and IL-10 were quantified. Monocytes and MPs exposed to purified LPS or wild-type N. meningitidis had much higher TF-activity than monocytes and MPs exposed to LPS-deficient N. meningitidis (clot formation assay). Incubation with wild-type N. meningitidis, but also LPS-deficient N. meningitidis, resulted in TF-expression on monocytes (flow cytometry, qRT-PCR). Increased cellular TF-activity is associated with coincident surface-exposure of PS and the number of monocytes positive for both PS and TF was significantly higher for monocytes exposed to wild-type N. meningitidis (7.6%) compared with monocytes exposed to LPS-deficient N. meningitidis (1.8%). Treatment with rhIL-10 reduced monocyte- and MP-associated TF-activity, the number of monocytes positive for both TF and PS, and microvesiculation. Patients with meningococcal septicemia had significantly higher levels of LPS, FPA and IL-10 than patients with distinct meningitis. Our results indicate that LPS from N. meningitidis is crucial for inducing TF-activity, but not for monocyte- and MP-associated TF-expression. TF-activity seems to require coincident expression of TF and PS on monocytes, and LPS induces such double-positive monocytes.

Topics: Blood Coagulation; Cell-Derived Microparticles; Cells, Cultured; Escherichia coli; Escherichia coli Infections; Fibrinopeptide A; Gene Expression Regulation, Bacterial; Humans; Interleukin-10; Lipopolysaccharides; Lipoproteins; Meningitis, Meningococcal; Meningococcal Infections; Monocytes; Neisseria meningitidis, Serogroup B; Phosphatidylserines; Thromboplastin

Disseminated intra-vascular coagulation (DIC) is a serious and frequently fatal condition associated with gross abnormalities of thrombosis, haemostasis, vascular function and inflammation. We measured serial levels of plasma soluble E selectin (marking endothelia damage), interleukin 6 (IL-6, a marker if inflammation) and tissue factor (involved in coagulation) in two young adults with bacterially-induced DIC. Soluble E selectin and IL-6 were grossly elevated on presentation and increased as the DIC progressed. Tissue factor was normal on admission but also increased with the DIC. However, levels remained high in one of the patients who subsequently died, whilst levels resolved in the patient who survived. We suggest that both admission and evolving sE-selectin and IL-6 levels may be useful in predicting outcome, and that measurement of TF has little extra to offer.

Topics: Adult; Biomarkers; Disease Progression; Disseminated Intravascular Coagulation; E-Selectin; Endothelium, Vascular; Female; Humans; Interleukin-6; Male; Meningitis, Meningococcal; Meningococcal Infections; Prognosis; Sepsis; Shock, Septic; Thromboplastin

Patients with meningococcal sepsis generally suffer from disseminated intravascular coagulation (DIC). The aim of this study was to address whether these patients have elevated numbers of circulating microparticles that contribute to the development of DIC. Plasma samples from 5 survivors, 2 nonsurvivors, and 5 healthy volunteers were analyzed for the presence of microparticles by flow cytometry. Ongoing coagulation activation in vivo was quantified by enzyme-linked immunosorbent assay of plasma prothrombin fragment F(1 + 2), and procoagulant properties of microparticles in vitro were estimated by thrombin-generation assay. On admission, all patients had increased numbers of microparticles originating from platelets or granulocytes when compared with controls (P =.004 and P =.008, respectively). Patients had elevated levels of F(1 + 2) (P =.004), and their microparticles supported thrombin generation more strongly in vitro (P =.003) than those of controls. Plasma from the patient with the most fulminant disease course and severe DIC contained microparticles that expressed both CD14 and tissue factor, and these microparticles demonstrated extreme thrombin generation in vitro. We conclude that patients with meningococcal sepsis have elevated numbers of circulating microparticles that are procoagulant. These findings may suggest a novel therapeutic approach to combat clinical conditions with excessive coagulation activation.

Topics: Adolescent; Adult; Biomarkers; Blood Coagulation Factors; Blood Platelets; Child; Child, Preschool; Disseminated Intravascular Coagulation; Female; Granulocytes; Humans; Infant; Lipopolysaccharide Receptors; Male; Meningococcal Infections; Particle Size; Sepsis; Survivors; Thrombin; Thrombophilia; Thromboplastin

We have evaluated the quantitative relationship between lipopolysaccharide (LPS, endotoxin), fibrinopeptide A (FPA), antithrombin (AT), protein C (PC) and extrinsic pathway inhibitor (EPI) in plasma from 39 consecutively admitted patients with systemic meningococcal disease (SMD). The most severely ill patients with fulminant meningococcal septicemia (n = 13, 6 dead) had significantly (p less than 0.01) higher plasma levels of LPS and FPA and lower levels of PC and AT on admission as compared with the less severe clinical presentations (n = 26, 1 dead). The levels of EPI on admission were significantly (p less than 0.05) higher in nonsurvivors vs survivors with fulminant septicemia. As the disease progressed, the levels of LPS, FPA, AT and PC declined, while the levels of EPI increased. Three of six nonsurviving septicemic patients had levels of EPI greater than 200% within 16 hours of admission vs two of 30 survivors (p = 0.02). The results suggest that increasing levels of LPS in SMD elicit increasing consumption coagulopathy, contributing to the organ pathophysiology. The kinetics of EPI, inhibiting the thromboplastin-FVIIa-FXa complex, differs markedly from the kinetics of AT and PC i.e. increases as opposed to decreases.

Topics: Antithrombin III; Disseminated Intravascular Coagulation; Endotoxins; Factor VII; Fibrinogen; Fibrinopeptide A; Humans; Lipopolysaccharides; Lipoproteins; Meningitis, Meningococcal; Meningococcal Infections; Multiple Organ Failure; Neisseria meningitidis; Protein C; Sepsis; Thromboplastin

In 16 patients, 13 with meningococcal infection and 3 suspected to have this infection, 8 patients were found to possess significant higher level of tissue thromboplastin activity of their monocytes isolated from the blood at the admission to the hospital than normal. Five of those 8 patients had an extremely high concentration, greater than 60-300 fold increase, and all these patients died. The exposed tissue thromboplastin activity on the surface of the endotoxin stimulated monocytes is probably the direct inducer of disseminated intravascular coagulation (DIC) in meningococcal infection.

Topics: Adolescent; Adult; Blood Cell Count; Child; Child, Preschool; Female; Humans; Infant; Male; Meningococcal Infections; Monocytes; Prognosis; Thromboplastin; Time Factors

Topics: Antithrombin III; Binding Sites; Endotoxins; Factor VII; Humans; Meningococcal Infections; Monocytes; Thromboplastin

Topics: Blood Coagulation Tests; Blood Platelets; Child; Child, Preschool; Disseminated Intravascular Coagulation; Fibrin; Fibrinogen; Heparin; Humans; Infant; Meningococcal Infections; Prothrombin Time; Thromboplastin

Topics: Adolescent; Adult; Aged; Anticoagulants; Blood Coagulation Disorders; Blood Coagulation Factors; Child; Child, Preschool; Female; Fibrinogen; Histocytochemistry; Humans; Infant; Male; Meningitis; Meningococcal Infections; Middle Aged; Neisseria meningitidis; Prothrombin Time; Sepsis; Shock, Septic; Skin Manifestations; Thrombocytopenia; Thromboplastin