thromboplastin and Sepsis

The coagulation system is a part of the mammalian host defense system. Pathogens and pathogen components, such as bacterial lipopolysaccharide (LPS), induce tissue factor (TF) expression in circulating monocytes that then activates the coagulation protease cascade. Formation of a clot limits dissemination of pathogens, enhances the recruitment of immune cells, and facilitates killing of pathogens. However, excessive activation of coagulation can lead to thrombosis. Here, we review studies on the mechanism of LPS induction of TF expression in monocytes and its contribution to thrombosis and disseminated intravascular coagulation. Binding of LPS to Toll-like receptor 4 on monocytes induces a transient expression of TF that involves activation of intracellular signaling pathways and binding of various transcription factors, such as c-rel/p65 and c-Fos/c-Jun, to the TF promoter. Inhibition of TF in endotoxemia and sepsis models reduces activation of coagulation and improves survival. Studies with endotoxemic mice showed that hematopoietic cells and myeloid cells play major roles in the activation of coagulation. Monocyte TF expression is also increased after surgery. Activated monocytes release TF-positive extracellular vesicles (EVs) and levels of circulating TF-positive EVs are increased in endotoxemic mice and in patients with sepsis. More recently, it was shown that inflammasomes contribute to the induction of TF expression and activation of coagulation in endotoxemic mice. Taken together, these studies indicate that monocyte TF plays a major role in activation of coagulation. Selective inhibition of monocyte TF expression may reduce pathologic activation of coagulation in sepsis and other diseases without affecting hemostasis.

Topics: Animals; Humans; Lipopolysaccharides; Mammals; Mice; Monocytes; Sepsis; Thromboplastin; Thrombosis

Tissue factor (TF) is a procoagulant protein released from activated host cells, such as monocytes, and tumor cells on extracellular vesicles (EVs). TF + EVs are observed in the circulation of patients with various types of diseases. In this review, we will summarize the association between TF + EVs and activation of coagulation and survival in different types of diseases, including cancer, sepsis, and infections with different viruses, such as human immunodeficiency virus (HIV), influenza A virus (IAV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We will also discuss the source of TF + EVs in various diseases. EVTF activity is associated with thrombosis in pancreatic cancer patients and coronavirus disease 2019 patients (COVID-19) and with disseminated intravascular coagulation in cancer patients. EVTF activity is also associated with worse survival in patients with cancer and COVID-19. Monocytes are the major sources of TF + EVs in sepsis, and viral infections, such as HIV, Ebola virus, and SARS-CoV-2. In contrast, alveolar epithelial cells are the major source of TF + EVs in bronchoalveolar lavage fluid in COVID-19 and influenza A patients. These studies indicate that EVTF activity could be used as a biomarker to identify patients that have an increased risk of coagulopathy and mortality.

Topics: Biomarkers; COVID-19; Extracellular Vesicles; Humans; Pancreatic Neoplasms; SARS-CoV-2; Sepsis; Thromboplastin; Thrombosis

Disseminated intravascular coagulation (DIC) is a systemic activation of the coagulation system, which results in microvascular thrombosis and, simultaneously, potentially life-threatening haemorrhage attributed to consumption of platelets and coagulation factors. Underlying conditions, e.g. infection, cancer, or obstetrical complications are responsible for the initiation and propagation of the DIC process. This review provides insights into the epidemiology of DIC and the current understanding of its pathophysiology. It details the use of diagnostic biomarkers, current diagnostic recommendations from international medical societies, and it provides an overview of emerging diagnostic and prognostic biomarkers. Last, it provides guidance on management. It is concluded that timely and accurate diagnosis of DIC and its underlying condition is essential for the prognosis. Treatment should primarily focus on the underlying cause of DIC and supportive treatment should be individualised according to the underlying aetiology, patient's symptoms and laboratory records.

Topics: Anticoagulants; Biomarkers; Blood Viscosity; Disease Management; Disseminated Intravascular Coagulation; Endothelium, Vascular; Female; Fibrinolysis; Humans; Male; Neoplasms; Platelet Activation; Pregnancy; Pregnancy Complications, Hematologic; Prevalence; Prognosis; Sepsis; Severity of Illness Index; Thrombin; Thromboembolism; Thromboplastin

Sepsis is a potentially life-threatening, pathological condition caused by a dysregulated host response to infection. Pathologically, systemic inflammation can initiate coagulation activation, leading to organ dysfunction, and ultimately to multiple organ failure and septic death. The inflammasomes are cytosolic multiprotein signaling complexes that control the host response to diverse pathogen-associated molecular patterns (PAMPs) from microorganisms as well as damage-associated molecular patterns (DAMPs) from dead or dying host cells. Recent studies highlight that the activation of canonical and non-canonical inflammasomes not only mediate the maturation and secretion of interleukin-1 (IL1) family cytokines, but also trigger the release of coagulation factor III, tissue factor (F3, best known as TF) in activated macrophages and monocytes. These emerging functions of inflammasomes in immunocoagulation are further positively regulated by stimulator of interferon response cGAMP interactor 1 (STING1, also known as STING or TMEM173, a hub of the innate immune signaling network) and high mobility group box 1 (HMGB1, a nuclear DAMP). This mini-review will discuss the regulation and function of inflammasome-dependent coagulation activation in sepsis.

Topics: Animals; Blood Coagulation; HMGB1 Protein; Humans; Inflammasomes; Macrophage Activation; Macrophages; Membrane Proteins; Monocytes; Sepsis; Thromboplastin

Endothelial cells (ECs) are vascular, nonconventional immune cells that play a major role in the systemic response after bacterial infection to limit its dissemination. Triggered by exposure to pathogens, microbial toxins, or endogenous danger signals, EC responses are polymorphous, heterogeneous, and multifaceted. During sepsis, ECs shift toward a proapoptotic, proinflammatory, proadhesive, and procoagulant phenotype. In addition, glycocalyx damage and vascular tone dysfunction impair microcirculatory blood flow, leading to organ injury and, potentially, life-threatening organ failure. This review aims to cover the current understanding of the EC adaptive or maladaptive response to acute inflammation or bacterial infection based on compelling recent basic research and therapeutic clinical trials targeting microvascular and endothelial alterations during septic shock.

Topics: Alarmins; Animals; Apoptosis; Blood Coagulation; Capillary Permeability; Cell Adhesion Molecules; Cytokines; Endothelial Cells; Endothelins; Endothelium, Vascular; Epoprostenol; Fibrinolysis; Glycocalyx; Humans; Inflammation; Microcirculation; Nitric Oxide; Pathogen-Associated Molecular Pattern Molecules; Phenotype; Platelet Adhesiveness; Sepsis; Thromboplastin; Vasoconstriction; Vasodilation

The pathobiology of the septic process includes a complex interrelationship between inflammation and the coagulations system. Antithrombin (AT) and tissue factor are important components of the coagulation system and have potential roles in the production and amplification of sepsis. Sepsis is associated with a decrease in AT levels, and low levels are also associated with the development of multiple organ failure and death. Treatment strategies incorporating AT replacement therapy in sepsis and septic shock have not resulted in an improvement in survival or reversal of disseminated intravascular coagulation.

Topics: Antithrombin III; Critical Illness; Humans; Multiple Organ Failure; Sepsis; Shock, Septic; Thromboplastin

Tissue factor (TF) is the high-affinity receptor and cofactor for factor (F)VII/VIIa. The TF-FVIIa complex is the primary initiator of blood coagulation and plays an essential role in hemostasis. TF is expressed on perivascular cells and epithelial cells at organ and body surfaces where it forms a hemostatic barrier. TF also provides additional hemostatic protection to vital organs, such as the brain, lung, and heart. Under pathological conditions, TF can trigger both arterial and venous thrombosis. For instance, atherosclerotic plaques contain high levels of TF on macrophage foam cells and microvesicles that drives thrombus formation after plaque rupture. In sepsis, inducible TF expression on monocytes leads to disseminated intravascular coagulation. In cancer patients, tumors release TF-positive microvesicles into the circulation that may contribute to venous thrombosis. TF also has nonhemostatic roles. For instance, TF-dependent activation of the coagulation cascade generates coagulation proteases, such as FVIIa, FXa, and thrombin, which induce signaling in a variety of cells by cleavage of protease-activated receptors. This review will focus on the roles of TF in protective hemostasis and pathological thrombosis.

Topics: Animals; Atherosclerosis; Blood Coagulation; Factor IX; Factor VIIa; Factor X; Fibrinolytic Agents; Gene Expression Regulation; Hemostasis; Humans; Neoplasms; Risk Factors; Sepsis; Signal Transduction; Thromboplastin; Thrombosis

Sepsis affects practically all aspects of endothelial cell (EC) function and is thought to be the key factor in the progression from sepsis to organ failure. Endothelial functions affected by sepsis include vasoregulation, barrier function, inflammation, and hemostasis. These are among other mechanisms often mediated by glycocalyx shedding, such as abnormal nitric oxide metabolism, up-regulation of reactive oxygen species generation due to down-regulation of endothelial-associated antioxidant defenses, transcellular communication, proteases, exposure of adhesion molecules, and activation of tissue factor. This review covers current insight in EC-associated hemostatic responses to sepsis and the EC response to inflammation. The endothelial cell lining is highly heterogeneous between different organ systems and consequently also in its response to sepsis. In this context, we discuss the response of the endothelial cell lining to sepsis in the kidney, liver, and lung. Finally, we discuss evidence as to whether the EC response to sepsis is adaptive or maladaptive. This study is a result of an Acute Dialysis Quality Initiative XIV Sepsis Workgroup meeting held in Bogota, Columbia, between October 12 and 15, 2014.

Topics: Animals; Down-Regulation; Endothelial Cells; Endothelium; Glycocalyx; Hemostasis; Humans; Nitric Oxide; Organ Specificity; Sepsis; Thromboplastin; Up-Regulation

Coagulopathy is common in acute sepsis and may range from subclinical activation of blood coagulation (hypercoagulability), which may contribute to venous thromboembolism, to acute disseminated intravascular coagulation, characterized by widespread microvascular thrombosis and consumption of platelets and coagulation proteins, eventually causing bleeding. The key event underlying this life-threatening complication is the overwhelming inflammatory host response to the pathogen leading to the overexpression of inflammatory mediators. The latter, along with the microorganism and its derivatives drive the major changes responsible for massive thrombin formation and fibrin deposition: (1) aberrant expression of tissue factor mainly by monocytes-macrophages, (2) impairment of anticoagulant pathways, orchestrated by dysfunctional endothelial cells (ECs), and (3) suppression of fibrinolysis because of the overproduction of plasminogen activator inhibitor-1 by ECs and thrombin-mediated activation of thrombin-activatable fibrinolysis inhibitor. Neutrophils and other cells, upon activation or death, release nuclear materials (neutrophil extracellular traps and/or their components such as histones, DNA, lysosomal enzymes, and High Mobility Group Box-1), which have toxic, proinflammatory and prothrombotic properties thus contributing to clotting dysregulation. The ensuing microvascular thrombosis-ischemia significantly contributes to tissue injury and multiple organ dysfunction syndromes. These insights into the pathogenesis of sepsis-associated coagulopathy may have implications for the development of new diagnostic and therapeutic tools.

Topics: Animals; Disease Models, Animal; Disseminated Intravascular Coagulation; Endothelium, Vascular; Endotoxemia; Extracellular Traps; Fibrinolysis; Humans; Immunity, Innate; Inflammation; Inflammation Mediators; Macrophages; Models, Biological; Monocytes; Multiple Organ Failure; Neutrophils; Protein C; Sepsis; Thrombophilia; Thromboplastin; Thrombotic Microangiopathies

Activation of the coagulation system is a fundamental host defense mechanism. Microorganisms that have invaded the body are trapped and disposed of in clots. Monocytes/macrophages are widely accepted as the main players in the procoagulant process; however, recent evidence suggests that neutrophils also play important roles. Tissue factor, which initiates the extrinsic coagulation cascade, is reportedly expressed on the surface of neutrophils, as well as on microparticles derived from neutrophils. Neutrophil extracellular traps (NETs) are another source of tissue factor. The components of NETs, such as DNA, histones, and granule proteins, also provide procoagulant activities. For instance, DNA initiates the intrinsic pathway, histones are a strong generator of thrombin, and granule proteins such as neutrophil elastase, cathepsin G and myeloperoxidase contribute to the suppression of the anticoagulation systems. Although understanding of the mechanisms that are involved in coagulation/fibrinolysis in sepsis has gradually progressed, the impact of neutrophils on thrombogenicity during sepsis remains to be addressed. Since the importance of the connection between coagulation and inflammation is advocated nowadays, further research on neutrophils is required.

Topics: Blood Coagulation; Extracellular Traps; Fibrinolysis; Humans; Neutrophils; Sepsis; Thrombin; Thromboplastin

Sepsis, defined as a clinical syndrome brought about by an amplified and dysregulated inflammatory response to infections, is one of the leading causes of death worldwide. Despite persistent attempts to develop treatment strategies to manage sepsis in the clinical setting, the basic elements of treatment have not changed since the 1960s. As such, the development of effective therapies for reducing inflammatory reactions and end-organ dysfunction in critically ill patients with sepsis remains a global priority. Advances in understanding of the immune response to sepsis provide the opportunity to develop more effective pharmaceuticals. This article details current information on the modulation of the lipopolysaccharide (LPS) receptor complex with synthetic Lipid A mimetics. As the initial and most critical event in sepsis pathophysiology, the LPS receptor provides an attractive target for antisepsis agents. One of the well-studied approaches to sepsis therapy involves the use of derivatives of Lipid A, the membrane-anchor portion of an LPS, which is largely responsible for its endotoxic activity. This article describes the structural and conformational requirements influencing the ability of Lipid A analogues to compete with LPS for binding to the LPS receptor complex and to inhibit the induction of the signal transduction pathway by impairing LPS-initiated receptor dimerization.

Topics: Animals; Cytokines; Drug Design; Endotoxins; Humans; Immune System; Inflammation; Lipid A; Lipopolysaccharide Receptors; Lipopolysaccharides; Mice; Molecular Conformation; Phosphorylation; Protein Binding; Sepsis; Signal Transduction; Solubility; Thromboplastin; Tumor Necrosis Factor-alpha

Tissue factor (TF) is a major initiator of extrinsic pathway of blood coagulation. A dual role of TF in the extensive crosstalk between blood coagulation and inflammation has recently become apparent. The majority of the cases of systemic inflammatory response syndrome, disseminated intravascular coagulation, and sepsis are accompanied by hyperactivation of TF in circulating monocytes and damaged tissue. Systemic Gram-negative infection induces expression of TF by vascular cells. In addition to extrinsic coagulation pathway, TF induces proinflammatory signaling cascade originating from activation of protease-activated receptors. Because TF-activated proteolytic cascade is placed in a nexus between coagulation and inflammation, early modulation of TF activity presently becomes a tempting experimental therapeutic strategy in systemic inflammatory response syndrome patients.

Topics: Anti-Bacterial Agents; Blood Coagulation; Disseminated Intravascular Coagulation; Gram-Negative Bacteria; Humans; Lipopolysaccharides; Monocytes; Sepsis; Signal Transduction; Systemic Inflammatory Response Syndrome; Thromboplastin

Binding of factor VIIa (FVIIa) to tissue factor (TF) and the subsequent initiation of the clotting cascade is essential for hemostasis. However, the aberrant expression of FVIIa-TF contributes to thrombosis. Despite the tremendous progress made in the past 25years in understanding the molecular mechanisms involved in the interaction between FVIIa and TF, there is less known about the cell biology of these proteins. Availability of hemophilic mice (by specific knock-out of FVIII or FIX genes) and novel TF transgenic mice has allowed us in recent years to investigate the importance of TF-FVIIa-induced coagulation from wound healing to sepsis. This supplement explores new aspects of TF-FVIIa biology, with a particular focus on structural biology, cell biology and animal models.

Topics: Animals; Cell-Derived Microparticles; Disease Models, Animal; Factor IX; Factor VIIa; Hemophilia A; Hemostasis; Humans; Mice; Mice, Knockout; Sepsis; Thromboplastin; Wound Healing

Sepsis is a systemic host response to infection by pathogenic microorganisms. Activation of the coagulation cascade during endotoxemia and sepsis leads to disseminated intravascular coagulation. This review focuses on tissue factor expression by hematopoietic and non-hematopoietic cells and its contribution to the activation of coagulation during endotoxemia and sepsis.

Topics: Animals; Antithrombins; Blood Coagulation; Endothelial Cells; Endotoxemia; Hematopoiesis; Humans; Lipopolysaccharides; Models, Biological; Sepsis; Thrombin; Thromboplastin

Intravascular and extravascular fibrin formation are characteristic findings in patients with sepsis, suggesting that the activation of coagulation and the inhibiton of fibrinolysis are important in the pathogenesis of sepsis. Activation of coagulation during sepsis is primarily driven by the tissue factor (TF) pathway, while inhibition of fibrinolysis is primarily due to increases in plasminogen activator inhibitor -1(PAI-1). Downregulation of the anticoagulant Protein C pathway also plays an important role in the modulation of coagulation and inflammation in sepsis. Recent advances in the understanding of pathogenetic mechanisms of coagulation and fibrinolysis in sepsis may have therapeutic implications. Recombinant human activated protein C (rhAPC) is currently the only pharmacologic therapy that has been shown to reduce mortality in adults with severe sepsis, highlighting the importance of coagulation and fibrinolysis as a therapeutic target in sepsis. This review summarizes recent basic and clinical findings with regard to the role of the coagulation cascade in sepsis and explores potential therapeutic targets in the coagulation and fibrinolytic pathways in the management of sepsis.

Topics: Animals; Blood Coagulation; Drug Delivery Systems; Fibrin; Fibrinolysis; Humans; Plasminogen Activator Inhibitor 1; Protein C; Recombinant Proteins; Sepsis; Thromboplastin

Sepsis is often associated with systemic intravascular activation of coagulation, potentially leading to widespread microvascular deposits of fibrin, and thereby contributing to multiple organ dysfunction. A complex interaction exists between activation of inflammatory systems and the initiating and regulating pathways of coagulation. A diagnosis of sepsis-associated disseminated intravascular coagulation can be made by a combination of routinely available laboratory tests, for which simple diagnostic algorithms have become available. Strategies to inhibit coagulation activation may theoretically be justified and are being evaluated in clinical studies.

Topics: Anticoagulants; Antithrombin III; Disseminated Intravascular Coagulation; Glycocalyx; Humans; Multiple Organ Failure; Plasma; Platelet Transfusion; Protein C; Sepsis; Thromboplastin

Sepsis is a systemic response to infection, and symptoms are produced by host defense systems rather than by the invading pathogens. Amongst the most prominent features of sepsis, contributing significantly to its outcome, is activation of coagulation with concurrent down-regulation of anticoagulant systems and fibrinolysis. Inflammation-induced coagulation on its turn contributes to inflammation. Another important feature of sepsis, associated with key symptoms such as hypovolemia and hypotension, is endothelial dysfunction. Under normal conditions, the endothelium provides for an anticoagulant surface, a property that is lost in sepsis. In this review, data about the interplay between inflammation and coagulation in sepsis are summarized with a special focus on the influence of the endothelium on inflammation-induced coagulation and vice versa. Possible procoagulant properties of the endothelium are described, such as expression of tissue factor (TF) and von Willebrand factor and interaction with platelets. Possible procoagulant roles of microparticles, circulating endothelial cells and endothelial apoptosis, are also discussed. Moreover, the important roles of the endothelium in down-regulating the anticoagulants TF pathway inhibitor, antithrombin, and the protein C (PC) system and inhibition of fibrinolysis are discussed. The influence of coagulation on its turn on inflammation and the endothelium is described with a special focus on protease-activated receptors (PARs). We conclude that the relationship between endothelium and coagulation in sepsis is tight and that further research is needed, for example, to better understand the role of activated PC signaling via PAR-1, the role of the endothelial PC receptor herein, and the role of the glycocalyx.

Topics: Anticoagulants; Blood Coagulation; Blood Platelets; Endothelium, Vascular; Fibrinolysis; Heparin; Humans; Inflammation; Models, Biological; Sepsis; Thromboplastin; von Willebrand Factor

Tissue factor plays an essential role in the initiation of coagulation in vivo. In severe conditions, including sepsis and acute lung injury, increased expression of tissue factor may induce disseminated intravascular coagulation and fibrin deposition in organs, which are believed to have a determining impact on patient outcome. Tissue factor also acts as a signaling receptor and is involved in the systemic inflammatory response, as in cancer progression and atherosclerosis. Interventions aiming at limiting tissue factor activities have been evaluated in multiple experimental studies and the observed results have supported the potential benefits for coagulation disorders, inflammation, and survival. The effects of the main physiological inhibitor of tissue factor, tissue factor pathway inhibitor, have been evaluated in two large clinical trials in sepsis. Even though they are not associated with an improved outcome, the observed data support further clinical studies.

Topics: Animals; Blood Coagulation; Clinical Trials as Topic; Disease Models, Animal; Disseminated Intravascular Coagulation; Fibrin; Humans; Inflammation; Lipoproteins; Lung; Lung Injury; Sepsis; Signal Transduction; Thromboplastin

Severely septic patients continue to experience excessive morbidity and mortality despite recent advances in critical care. Although significant resources have been invested in new treatments, almost all have failed to improve outcomes. An improved understanding of sepsis pathophysiology, including the complex interactions between inflammatory, coagulation, and fibrinolytic systems, has accelerated the development of novel treatments. Recombinant human activated protein C (rhAPC), or drotrecogin alfa (activated) (DAA), is currently the only US Food and Drug Administration (FDA)-approved medicine for the treatment of severe sepsis, and only in patients with a high risk of death. This review will discuss the treatment of severe sepsis, focusing on recent discoveries and unresolved questions about DAA's optimal use. Increasing pharmacological experience has generated enthusiasm for investigating medicines already approved for other indications as treatments for severe sepsis. Replacement doses of hydrocortisone and vasopressin may reduce mortality and improve hypotension, respectively, in a subgroup of patients with catecholamine-refractory septic shock. In addition to discussing these new indications, this review will detail the provocative preliminary data from four promising treatments, including two novel modalities: antagonizing high mobility group box protein and inhibiting tissue factor (TF). Observational data from the uncontrolled administration of heparin or statins in septic patients will also be reviewed.

Topics: Adrenal Cortex Hormones; Anti-Infective Agents; Anti-Inflammatory Agents; Anticoagulants; Antidiuretic Agents; Drug Administration Schedule; Drug Therapy; Hemorrhage; Heparin; HMGB1 Protein; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Patient Selection; Protein C; Recombinant Proteins; Sepsis; Severity of Illness Index; Thromboplastin; Treatment Outcome; Vasopressins

Tissue factor (TF) is a transmembrane protein that, in complex with factor VIIa (FVIIa), initiates coagulation. It also influences various other physiological and pathological events, such as inflammation, and negatively influences survival during sepsis. TF resembles a conserved class of pro-inflammatory cytokine receptors and activates a set of pro-inflammatory intracellular signal transduction routes. Interestingly, whereas the homology of TF to the class II cytokine receptors is reflected in a similar type of signal transduction, the mechanism by which the signal is transduced across the membrane differs greatly. This review discusses the role of TF and its ligand FVIIa in inflammation, sepsis, and signal transduction, and describes the way in which these processes interact.

Topics: Animals; Cattle; Disseminated Intravascular Coagulation; Factor VIIa; Gene Expression Regulation; Guinea Pigs; Humans; Inflammation; Mice; Protein-Tyrosine Kinases; Rabbits; Rats; Receptors, Cytokine; Sepsis; Sequence Homology; Signal Transduction; Thromboplastin; Trans-Activators

Inhibition of the tissue factor-factor VIIa complex reduces coagulation and inflammation in animal models of endotoxemia and sepsis and in patients with severe sepsis. However, the mechanism by which tissue factor-dependent activation of the coagulation cascade enhances inflammation is not known. We tested the hypothesis that coagulation proteases enhance inflammation during endotoxemia by activating protease-activated receptors (PARs) within the vasculature. We found that genetically modified mice expressing low levels of tissue factor exhibited reduced interleukin-6 expression and increased survival in a mouse model of endotoxemia compared with control mice. In contrast, hirudin inhibition of thrombin or a deficiency in either PAR-1 or PAR-2 did not affect interleukin-6 expression or mortality. However, combining hirudin treatment to inhibit thrombin signaling through PAR-1 and PAR-4 with PAR-2 deficiency reduced lipopolysaccharide-induced interleukin-6 expression and increased survival. Taken together, our results suggest that activation of multiple PARs by coagulation proteases enhances inflammation during endotoxemia.

Topics: Animals; Anticoagulants; Blood Coagulation; Endopeptidases; Endotoxemia; Fibrin; Humans; Protein C; Receptors, Proteinase-Activated; Sepsis; Thrombin; Thromboplastin

Severe sepsis and septic shock are among the most common causes of death in noncoronary intensive care units. The incidence of sepsis has been increasing over the past two decades, and is predicted to continue to rise over the next 20 years. While our understanding of the complex pathophysiologic alterations that occur in severe sepsis and septic shock has increased greatly asa result of recent clinical and preclinical studies, mortality associated with the disorder remains unacceptably high. Despite these new insights, the cornerstone of therapy continues to be early recognition, prompt initiation of effective antibiotic therapy, and source control, and goal-directed hemodynamic, ventilatory,and metabolic support as necessary. To date, attempts to reduce mortality with innovative, predominantly anti-inflammatory therapeutic strategies have been extremely disappointing. Observations of improved outcomes with physiologic doses of corticosteroid replacement therapy and activated protein C (drotrecogin alfa[activated]) have provided new adjuvant therapies for severe sepsis and septic shock in selected patients. This article reviews the components of sepsis management and discusses the available evidence in support of these recommendations. In addition, there is a discussion of some promising new strategies.

Topics: Anti-Infective Agents; Anti-Inflammatory Agents; Cause of Death; Critical Care; Evidence-Based Medicine; Fibrinolytic Agents; Fluid Therapy; Hemofiltration; Humans; Incidence; Plasma Exchange; Practice Guidelines as Topic; Prognosis; Protein C; Respiration, Artificial; Risk Factors; Sepsis; Shock, Septic; Survival Rate; Thromboplastin; Treatment Outcome; Tumor Necrosis Factor-alpha

Sepsis is caused by a dysregulated immune response to infection and, without intervention, can lead to septic shock and multiple organ failure. A leading cause of morbidity and mortality in intensive care units worldwide, severe sepsis is also associated with a considerable cost burden that places significant strain on global healthcare budgets. The development of an efficacious and cost-effective treatment strategy is therefore of vital importance to today's intensive care physicians. This paper will examine the pathophysiology of sepsis and multiple organ dysfunction before reviewing trials recently undertaken to investigate three potential anticoagulant therapies: antithrombin III, activated protein C, and tissue factor pathway inhibitor. Finally, other recent developments in the care of sepsis patients will be briefly examined.

Topics: Anticoagulants; Disease Management; Humans; Multiple Organ Failure; Sepsis; Thromboplastin; Treatment Outcome

The transmembrane glycoprotein tissue factor (TF) is the initiator of the coagulation cascade in vivo. When TF is exposed to blood, it forms a high-affinity complex with the coagulation factors factor VII/activated factor VIIa (FVII/VIIa), activating factor IX and factor X, and ultimately leading to the formation of an insoluble fibrin clot. TF plays an essential role in hemostasis by restraining hemorrhage after vessel wall injury. An overview of biological and physiological aspects of TF, covering aspects consequential for thrombosis and hemostasis such as TF cell biology and biochemistry, blood-borne (circulating) TF, TF associated with microparticles, TF encryption-decryption, and regulation of TF activity and expression is presented. However, the emerging role of TF in the pathogenesis of diseases such as sepsis, atherosclerosis, certain cancers and diseases characterized by pathological fibrin deposition such as disseminated intravascular coagulation and thrombosis, has directed attention to the development of novel inhibitors of tissue factor for use as antithrombotic drugs. The main advantage of inhibitors of the TF*FVIIa pathway is that such inhibitors have the potential of inhibiting the coagulation cascade at its earliest stage. Thus, such therapeutics exert minimal disturbance of systemic hemostasis since they act locally at the site of vascular injury.

Topics: Animals; Arteriosclerosis; Blood Coagulation; Blood Coagulation Disorders; Blood Coagulation Factors; Blood Vessels; Disseminated Intravascular Coagulation; Fibrin; Gene Expression Regulation; Humans; Neoplasms; Sepsis; Thromboplastin; Thrombosis

Sepsis, a life-threatening disorder that arises through the body's response to infection, is the leading cause of death and disability for patients in an intensive care unit. Advances in the understanding of the complex biological processes responsible for the clinical syndrome have led to the identification of many promising new therapeutic targets, including bacterial toxins, host-derived mediators, and downstream processes such as coagulation and the endocrine response. Diverse therapies directed against these targets have shown dramatic effects in animal models; however, in humans, their impact has been frustratingly modest, and only one agent--recombinant activated protein C--has achieved regulatory approval. This review summarizes the approaches that have been evaluated in clinical trials, explores the reasons for the discordance between biological promise and clinical reality, and points to approaches that may lead to greater success in the future.

Topics: Adrenal Cortex Hormones; Animals; Clinical Trials as Topic; Endotoxins; Granulocyte Colony-Stimulating Factor; Humans; Interleukin 1 Receptor Antagonist Protein; Nitric Oxide; Platelet Activating Factor; Protein C; Sepsis; Sialoglycoproteins; Thromboplastin; Tumor Necrosis Factor-alpha

To present and discuss the rationale and the results of clinical trials using supplementation with physiologic anticoagulants (Tissue Factor Pathway Inhibitor (TFPI), AntiThrombin (AT), and Protein C (PC) in patients with severe sepsis.. An early activation of the coagulation cascade occurs in severe sepsis. TFPI, AT, and PC are major inhibitors of the coagulation cascade, and additionally modulate inflammatory and vascular reactions. They are consumed or inhibited in the sepsis pathologic process. Therapeutic supplementation with these inhibitors could improve the sepsis-induced organ failures and mortality.. Randomized controlled studies were recently completed. No effect on the mortality rate could be documented after treatment with recombinant TFPI. AT concentrates neither improve mortality, but a biological interaction with heparin therapy could have biased the study results. Treatment with recombinant activated PC (alpha-drotrecogin) was associated with a significant reduction in the mortality rate of severely ill patients and received recently the approval from FDA and EC authorities in this indication. An increase in the rate of hemorrhagic adverse effects has been observed with these compounds, justifying a strict observance of contraindications and of patients selection. PROSPECTIVE: Additional studies are needed to give confirmation of the positive effects of activated PC supplementation in less severely ill patients, children and specific clinical situations. The effects of new anticoagulant compounds are currently evaluated in preclinical studies.

Topics: Anticoagulants; Antithrombins; Blood Coagulation Disorders; Drug Evaluation, Preclinical; Drug Interactions; Drug Monitoring; Heparin; Humans; Inflammation; Multiple Organ Failure; Patient Selection; Protein C; Randomized Controlled Trials as Topic; Recombinant Proteins; Sepsis; Severity of Illness Index; Thromboplastin; Treatment Outcome

Topics: Biomarkers; Dehydroepiandrosterone; Humans; Hydrocortisone; Inflammation; Respiration, Artificial; Sepsis; Severe Acute Respiratory Syndrome; Thromboplastin; United Kingdom

Following vessel wall injury, tissue factor (TF) is exposed and forms complexes with already activated factor VII (FVIIa) present in the circulating blood, thereby initiating the hemostatic process. After the first FXa is formed, the TF pathway inhibitor (TFPI) forms a complex with FXa, and a quaternary complex is formed, TF/FVIIa/ FXa/TFPI, which inhibits the first step of the hemostatic pathway. Recombinant activated FVII (rFVIIa) has been developed for use as a hemostatic agent (NovoNordisk A/S, Denmark). Active site-inactivated rFVIIa (rFVIIai) has also been prepared and was shown to have a faster association to and a slower dissociation from TF than rFVIIa, resulting in a lower calculated Kd of rFVIIai compared with rFVIIa. In various animal models rFVIIai has been demonstrated to prevent or diminish immediate thrombus formation at the site of vessel wall injury (athroplasty or other forms of mechanical injury) as well as the development of long-term intima thickening. The inflammatory response following endotoxin-induced sepsis was shown to decrease after administration of rFVIIai. Also, survival increased in the rFVIIai-treated animals in this study. In addition, ischemia-reperfusion injury was mitigated by rFVIIai. In a limited number of patients undergoing percutaneous transluminal coronary angioplasty (PTCA), rFVIIai was observed to allow PTCA to be performed at lower doses of heparin than what has been reported previously.

Topics: Angioplasty, Balloon, Coronary; Animals; Anti-Inflammatory Agents; Arteries; Blood Coagulation; Factor VIIa; Factor Xa; Fibrinolytic Agents; Hemostatics; Humans; Lipoproteins; Myocardial Reperfusion Injury; Recombinant Proteins; Sepsis; Thromboplastin; Thrombosis

To review the experimental and clinical evidence of the emerging role of tissue factor in intravascular thrombosis and to examine evidence supporting the potential use of tissue factor pathway inhibitor as an antithrombotic therapeutic agent.. A PubMed search was conducted encompassing articles in the English language relating to tissue factor and tissue factor pathway inhibitor in intravascular coagulation.. Tissue factor, a membrane-bound procoagulant glycoprotein, is the initiator of the extrinsic clotting cascade, which is the predominant coagulation pathway in vivo. The traditional view localizes tissue factor to extravascular sites, where it remains sequestered from circulating factor VII until vascular integrity is disrupted or until tissue factor expression is induced in endothelial cells or monocytes. This perspective has been challenged since the discovery of tissue factor antigen in plasma, on circulating microparticles, and on leukocytes in whole blood. Recently, the apparent role of tissue factor has expanded with the demonstration that this molecule also functions as a signaling receptor. Recombinant tissue factor pathway inhibitor, an analogue of the physiologic inhibitor of tissue factor, is a potent inhibitor of thrombus formation in experimental models. In summary, the tissue factor pathway initiates thrombosis in vivo. In addition to its classic tissue-bound distribution, recently discovered blood-borne tissue factor may have an important procoagulant function. Despite showing promise in early human studies, a recently completed phase 3 trial of recombinant tissue factor pathway inhibitor in severe sepsis failed to show a reduction in the primary end point of 28-day all-cause mortality. Tissue factor pathway inhibitor, however, remains a plausible therapeutic agent in other conditions of increased thrombogenicity, such as acute coronary syndromes, and further studies to examine this potential are warranted.

Topics: Animals; Anticoagulants; Arteriosclerosis; Hemostatics; Humans; Lipoproteins; Sepsis; Thromboplastin; Thrombosis

In the pathogenesis of disseminated intravascular coagulation, dysfunctional natural anticoagulant pathways appear to play a pivotal role. In this article, we will address the mechanisms that contribute to this defect in the regulation of coagulation activation. Furthermore, we will explore the experimental and clinical evidence that restoration of these anticoagulant pathways results in clinical improvement.. We have searched and reviewed published articles on experimental studies of disseminated intravascular coagulation models in animals and clinical studies in patients with disseminated intravascular coagulation.. All three major anticoagulant pathways, that is, the antithrombin pathway, the protein C system, and tissue factor pathway inhibitor, are defective in sepsis and disseminated intravascular coagulation. Several mechanisms contribute to this defect. Restoration of these pathways, in principle, by administration of coagulation inhibitor concentrates or recombinant anticoagulant factors, appears to ameliorate the coagulation disorder and, more important, result in improvement of clinically relevant outcomes, such as a reduction of organ failure and mortality.. Restoration of disrupted physiologic anticoagulant pathways in disseminated intravascular coagulation is not only a logical point of impact in patients with sepsis and an activated coagulation system, but also is associated with an improved outcome in experimental and (initial) clinical studies.

Topics: Animals; Anticoagulants; Cytokines; Disease Models, Animal; Disseminated Intravascular Coagulation; Humans; Multiple Organ Failure; Protein C; Sepsis; Thrombin; Thromboplastin; Treatment Outcome

Review of primate studies of Escherichia coli sepsis and endotoxemia with a reexamination of the rationale for diagnosis and treatment of these multistage disorders.. Animal research and intensive care units in a university medical school.. Cyanocephalus baboons (E. coli) and normal human subjects (endotoxin).. Baboon studies: anti-tissue factor, protein C, endothelial protein C receptor, and anti-tumor necrosis factor antibodies, and active site inhibited factor recombinant VIIa and factor Xa.. This review concerns the primate microvascular endothelial response to inflammatory and hemostatic stress. Studies of the impact of inflammatory and hemostatic stress on this microvasculature have fallen into four categories. First, studies of pure hemostatic stress using factor Xa phospholipid vesicles showed that blockade of protein C as well as protein C plus tissue plasminogen activator produced a severe but transient consumptive and a lethal thrombotic coagulopathy, respectively. These studies showed that the protein C and fibrinolytic systems can work in tandem to regulate even a severe response if the endothelium is not rendered dysfunctional by metabolic or inflammatory factors. Second, studies of compensated (nonlethal) inflammatory stress using E. coli or endotoxin in baboon and human subjects showed that even under minimal stress in which there is no evidence of overt disseminated intravascular coagulation, injury of the endothelium and activation of neutrophils and hemostatic factors are closely associated. This showed that molecular markers of hemostatic activity could be used to detect microvascular endothelial stress (nonovert disseminated intravascular coagulation) in patients who are compensated but at risk. These studies also showed that the compensated response to inflammatory stress could exhibit two stages, each with its unique inflammatory and hemostatic response signature. The first is driven by vasoactive peptides, cytokines, and thrombin, followed 12 to 14 hrs later by a second stage driven by C-reactive protein/complement complexes, tissue factor, and plasminogen activator inhibitor 1 secondary to oxidative stress after reperfusion. Third, studies of uncompensated (lethal) inflammatory stress using E. coli showed that irreversible thrombosis of the microvasculature was not a link in the lethal chain of events even though inhibition of components of the protein C network (protein C and endothelial protein C receptor) converted compensated responses to sublethal E. coli into uncompensated lethal responses. Fourth, these studies also showed that there were variants of the lethal response ranging from capillary leak and shock to recurrent sustained inflammatory disorders. We believe that each of these variants arises from their sublethal counterparts, depending on underlying or modulating host factors operating at the time of challenge. Such underlying conditions range from preexisting microvascular ischemia, reperfusion, and oxidati

Topics: Animals; Cytokines; Disease Models, Animal; Endothelium, Vascular; Endotoxemia; Escherichia coli Infections; Factor VIIa; Factor Xa; Hemostasis; Homeostasis; Humans; Inflammation; Microcirculation; Papio; Protein C; Sepsis; Thrombin; Thromboplastin; Tumor Necrosis Factor-alpha

To review the preclinical and clinical evidence that provides the therapeutic rationale for recombinant human tissue factor pathway inhibitor (rTFPI) as a novel treatment for human sepsis.. A summary of published English-language literature regarding preclinical studies and limited information published about three phase II clinical studies for the evaluation of rTFPI safety in sepsis patients.. Tissue factor pathway inhibitor, the physiologic inhibitor of the tissue factor pathway, interrupts activation of coagulation at multiple steps, including tissue factor VIIa activity, Xa activity, prothrombinase complex, and thrombin generation. Recombinant human TFPI exhibits anticoagulant and anti-inflammatory activities in animal models and humans with sepsis. These activities appear to have an important therapeutic role in protecting the microvasculature from injury and preventing multiple organ failure in sepsis.. Tissue factor pathway inhibitor is a potent inhibitor of clotting in the microvasculature, which is thought to protect organs from injury. Recombinant TFPI improved survival of septic animals in multiple models. Recent phase II results suggest that rTFPI is well tolerated, and they show a trend toward reduction in 28-day all-cause mortality in rTFPI-treated patients; in addition, rTFPI demonstrated significant reduction in thrombin generation. These results suggest that a powered study is indicated to further evaluate rTFPI utility for the adjunctive management of severe sepsis.

Topics: Animals; Anticoagulants; Blood Coagulation Disorders; Disease Models, Animal; Drug Evaluation, Preclinical; Fibrinolytic Agents; Humans; Inflammation; Lipoproteins; Sepsis; Survival Analysis; Thromboplastin; Treatment Outcome

Human tissue factor pathway inhibitor (TFPI) is a modular protein comprised of three Kunitz type domains flanked by peptide segments that are less structured. The sequential order of the elements are: an N-terminal acidic region followed by the first Kunitz domain (K1), a linker region, a second Kunitz domain (K2), a second linker region, the third Kunitz domain (K3), and the C-terminal basic region. The K1 domain inhibits factor VIIa complexed to tissue factor (TF) while the K2 domain inhibits factor Xa. No direct protease inhibiting functions have been demonstrated for the K3 domain. Importantly, the Xa-TFPI complex is a much more potent inhibitor of the VIIa-TF than TFPI by itself. Furthermore, the C-terminal basic region of TFPI is required for rapid physiologic inhibition of coagulation and is needed for the inhibition of smooth muscle cell proliferation. Although a number of additional targets for attachment have been reported, the C-terminal basic region appears to play an important role in binding of TFPI to cell surfaces. A primary site of TFPI synthesis is endothelium and the endothelium-bound TFPI contributes to the antithrombotic potential of the vascular endothelium. Further, increased levels of plasma TFPI under septic conditions may represent endothelial dysfunction. We have proposed that the extravascular cells that synthesize TF also synthesize TFPI providing dual components necessary for the regulation of clotting in their microenvironment. Like the TF synthesis in these cells is augmented by serum, so is the case with the TFPI gene expression. TFPI gene knock out mice reveal embryonic lethality suggesting a possible role of this protein in early development. Since TF-induced coagulation is thought to play a significant role in many disease states, including disseminated intravascular clotting, sepsis, acute lung injury and cancer, recombinant TFPI may be a beneficial therapeutic agent in these disease states to attenuate pathologic clotting. The purpose of this review is to outline recent developments in the field related to the structural specificity and biology of TFPI.

Topics: Acute Disease; Amino Acid Sequence; Amino Acids; Antiphospholipid Syndrome; Blood Coagulation; Cardiovascular Diseases; Endothelium, Vascular; Humans; Lipoproteins; Lung Diseases; Models, Biological; Models, Molecular; Molecular Sequence Data; Neoplasm Metastasis; Neoplasms; Protein Conformation; Protein Structure, Tertiary; Sepsis; Sequence Alignment; Sequence Homology, Amino Acid; Structure-Activity Relationship; Thrombophilia; Thromboplastin

Because of its unique position at the convergence point of the intrinsic (contact) and extrinsic (tissue factor/factor VIIa) pathways in the coagulation system, coagulation factor Xa (FXa) has been a theoretically interesting therapeutic target for antithrombotic drugs for many years. More recently, the discovery of naturally occurring FXa inhibitors, such as tick anticoagulant peptide and antistasin, has helped substantiate FXa as a desirable target by demonstrating the efficacy and potential safety advantages of FXa inhibition over conventional antithrombotic therapy. These discoveries led to the design and development of many small-molecule inhibitors of FXa, which have provided potent and selective tools for evaluating the potential role of FXa in various diseases. In addition, these advances have been instrumental in defining the biology of FXa and have aided in the discovery of specific receptors and intracellular signaling pathways for FXa that may be important in the progression of, or the response to, various diseases.

Topics: Animals; Cell Adhesion; Clinical Trials as Topic; Coronary Restenosis; Cysteine Endopeptidases; Cytokines; Factor Xa; Factor Xa Inhibitors; Graft Occlusion, Vascular; Humans; Hyperplasia; Inflammation; MAP Kinase Signaling System; Neoplasm Proteins; Neoplasms; Platelet-Derived Growth Factor; Receptor, PAR-2; Receptors, Platelet-Derived Growth Factor; Receptors, Thrombin; Sepsis; Thromboplastin; Up-Regulation

Tissue factor, a 47 kDa membrane glycoprotein, lies at the basis of the extrinsic pathway of the coagulation cascade. Interaction of TF with factor VIIa results in the formation of fibrin from fibrinogen, thereby inducing the formation of a blood clot. In addition to this well-established role in blood coagulation, TF is associated with various other physiological processes such as sepsis, inflammation, angiogenesis, metastasis and atherosclerosis. The molecular basis of the latter events is slowly emerging. It has become clear that TF-FVIIa interaction elicits a variety of intracellular signalling events that may be implicated in these actions. These events include the sequential activation of Src-like kinases, MAP kinases, small GTPases and calcium signalling. How this progress in the understanding of TF associated signal transduction may generate answers as to the mechanism through which TF exerts it pleiotropic effects will be focus of this review.

Topics: Animals; Arteriosclerosis; Blood Coagulation; Calcium Signaling; Cytokines; Disseminated Intravascular Coagulation; Factor VIIa; Fibroblasts; Gene Expression Regulation; GTP Phosphohydrolases; Humans; Inflammation; Lipoproteins; Lipoproteins, LDL; MAP Kinase Signaling System; Mice; Mice, Knockout; Muscle, Smooth, Vascular; Neoplasm Metastasis; Neoplasms; Neovascularization, Physiologic; Papio; Phagocytes; Sepsis; Signal Transduction; src-Family Kinases; Thromboplastin

To review the role of the tissue factor pathway of coagulation in experimental sepsis.. Studies published in biomedical journals.. Studies on the role of the tissue factor pathway in animal or human models for sepsis.. Variables reflecting tissue factor pathway activation in the various models are discussed; the effects of administration of tissue factor pathway inhibitors on these and inflammatory variables, as well as on the course and outcome, are analyzed.. Activation of coagulation during experimental sepsis occurs mainly, if not exclusively, via the tissue factor pathway; inhibitors of this pathway improve mortality, presumably by a combined attenuating effect on coagulative and inflammatory responses.

Topics: Animals; Blood Coagulation; Humans; Sepsis; Thromboplastin

Tissue factor mediated pathways leading to microvascular thromboses and endothelial activation appear to play an important role in the development of multiple organ failure associated with severe sepsis. Tissue factor pathway inhibitor (TFPI) is an endogenous inhibitor of tissue factor associated coagulation cascades. In experimental models of severe sepsis, treatment with TFPI results in significant reduction in mortality. Similarly, a recently completed Phase II 210-patient study comparing placebo and infusions of TFPI showed trends toward a relative reduction in day 28 all-cause mortality in TFPI treated patients. These data suggest that coagulation cascades involving tissue factor contribute to organ dysfunction in critically ill septic patients. TFPI may be a useful therapy in improving outcome of severe sepsis.

Topics: Animals; Clinical Trials as Topic; Disseminated Intravascular Coagulation; Lipoproteins; Recombinant Proteins; Sepsis; Thromboplastin

In sepsis, levels of the endogenous coagulation inhibitors antithrombin III and protein C are lowered as a result of complex formation with multiple activated clotting factors. In addition, their activity can further be curtailed by proteolytic inactivation. Loss of antithrombin III and protein C activity blocks the endogenous control mechanism for thrombin generation resulting in a state of systemic activation of coagulation and inflammatory processes. Levels of tissue factor pathway inhibitor, a third endogenous coagulation inhibitor, are increased in sepsis rather than decreased, probably reflecting a depletion of the endothelial cell bound tissue factor pathway inhibitor pool with loss of its endothelial protective function. Administration of any of these three inhibitors in various animal species and sepsis models reduces morbidity and mortality. In addition to their anticoagulant effects, these inhibitors also have various anti-inflammatory activities that may contribute to their protective effects. Phase II studies in patients with severe sepsis using coagulation inhibitors have indicated that this therapeutic approach may be useful. Large-scale phase III trials will ultimately decide whether adjunctive coagulation inhibitor replacement will have a place in the treatment of patients with severe sepsis.

Topics: Animals; Anticoagulants; Antithrombin III; Clinical Trials as Topic; Humans; Protein C; Sepsis; Thromboplastin

The expression of tissue factor (TF) by monocytes/macrophages leads to thrombin generation and contributes to their physiological and pathophysiological roles in wound repair, disseminated intravascular coagulation linked to sepsis, postoperative thrombosis, unstable angina, atherosclerosis, chronic inflammation and cancer. Regulation of TF expression in monocytes is controlled by the transcription factors NF-kappaB and AP-1. In whole blood, the activation of the transcription factors is mediated through the phospholipase A2 pathway. Platelets play a crucial role in the expression of TF activity in monocytes, and granulocytes are mandatory in provoking the platelet effect in a P-selectin-dependent reaction. Although all induced or constitutive TF is expressed on the surface of monocytes, its catalytic activity is only about 10% compared to the activity of lysed cells. This phenomenon has been attributed to the increased availability of anionic phospholipid (phosphatidylserine) after cell lysis. At the surface of viable cells, the transmembrane phospholipid distribution and its regulation may be important for the expression of the catalytic activity of the complex of TF and activated factor VII. Phosphatidylserine pathophysiologically exposed at the outer surface of monocytes may, similar to that for platelet membranes, provide a strong stimulus for thrombin generation.

Topics: Blood Platelets; Cells, Cultured; Granulocytes; Humans; Macrophages; Monocytes; Sepsis; Thrombin; Thromboplastin

Topics: Animals; Blood Coagulation; Blood Coagulation Factors; Cell Membrane; Cytokines; Disseminated Intravascular Coagulation; Endothelium, Vascular; Endotoxins; Gene Expression Regulation; Growth Substances; Humans; Mice; Neoplasms; Organ Specificity; Protein Processing, Post-Translational; Sepsis; Thromboplastin

This review presents the rationale for and main results of coagulation inhibitor substitution during experimental and human sepsis. Activation of the contact system induces activation of the classical complement pathway with generation of anaphylatoxins, of the kinins pathway and of fibrinolysis. Physiologic inhibition depends on the C1-inhibitor (C1-Inh.). Septic patients exhibit a relative deficiency of biologically active C1-Inh. Substitution with concentrations of C1-Inh has been safely performed and preliminary results are consistent with a possible beneficial effect on hypotension and vasopressor requirement in septic shock. The extrinsic pathway is the main initial coagulation process involved in sepsis-induced DIC. Endothelial and monocyte generation of tissue factor (TF) is activated by bacterial products and endotoxin. Activation of TF is counteracted by a specific tissue factor pathway inhibitor (TFPI). The potential for TFPI substitution to inhibit the activation of the coagulation cascade in sepsis requires further study. Thrombin generation is inhibited by antithrombin III (AT III) and the protein C-protein S system. During sepsis, AT III is consumed and degraded by elastase. Animal studies have shown that DIC and death were prevented by high doses of AT III concentrates. Although a significant reduction in the duration of biological symptoms of DIC has been reported in most human studies, the usefulness of AT III substitution in human sepsis is still debated. None of the studies was able to document a statistically significant reduction in mortality. Protein C is activated by thrombomodulin and, with its cofactor protein S, inhibits factors Va and VIIIa. The free level of protein S depends on the level of the C4b binding protein (C4bBP), an acute-phase complement regulatory protein. During sepsis, protein C activity is significantly reduced, either by acute consumption or by thrombomodulin down-regulation, and increased levels of plasma C4bBP inhibit protein S. Infusion of activated protein C and protein S substitution both protect animals from the lethal effects of bacteria. Combining these different coagulation inhibitors should be carefully studied before its use in septic patients is recommended.

Topics: Animals; Antithrombin III; Blood Coagulation; Complement C1 Inactivator Proteins; Complement Pathway, Classical; Disseminated Intravascular Coagulation; Humans; Protein C; Protein S; Sepsis; Thromboplastin

Extrinsic pathway inhibitor (EPI) is a Kunitz type serine protease inhibitor. EPI is a potent inhibitor of the factor VIIa/thromboplastin (TP) complex in the presence of factor Xa and is also a direct inhibitor of factor Xa. The inhibitory mechanism is complex and is currently thought to involve, in a first step, the formation of a EPI-factor Xa complex, and, in a second step, the formation a quaternary EPI-factor Xa-factor VIIa-TP complex. In the blood vessels, EPI is confined to three different pools. A major pool of EPI is bound to the endothelial surface, and this fraction may be released by heparin. Plasma contains a second, but smaller pool of EPI (approximately 10-50% of the endothelial surface pool) at a concentration of 50-100 ng/ml. This pool consists mostly of EPI-lipoprotein complexes and only less than 10% is carrier-free EPI. A third pool of EPI is confined to platelets (less than 10% of the plasma pool). The biological role of these pools has not yet been clarified, but some evidence suggest that the carrier-free EPI is biologically most active. In patients, disseminated intravascular coagulation may continue despite normal or even elevated EPI levels. However, evidence has now been provided to indicate that EPI can inhibit factor VIIa/TP complexes formed in vivo to prevent the effect of limited amounts of TP. Taken together, the present knowledge of EPI indicates that EPI functions as a key inhibitor to feedback control of blood coagulation initiated by TP.

Topics: Blood Coagulation; Blood Platelets; Disseminated Intravascular Coagulation; Endothelium, Vascular; Factor VII; Factor VIIa; Factor Xa Inhibitors; Feedback; Heparin; Humans; Lipoproteins; Liver Diseases; Neoplasms; Recombinant Proteins; Reference Values; Sepsis; Thromboplastin

Complement appears to be an important system for cell activation in circulating blood. It is established that granulocytes may be stimulated through complement activation. In the present paper we propose also a central role of complement in the induction of thromboplastin synthesis in blood monocytes and endothelial cells.

Topics: Antigen-Antibody Reactions; Complement Activation; Complement System Proteins; Disseminated Intravascular Coagulation; Dose-Response Relationship, Drug; Endothelium; Endotoxins; Humans; Monocytes; Sepsis; Thromboplastin

Tissue factor is an ubiquitous phospholipid-protein complex, which triggers blood coagulation through the so-called extrinsic pathway. Reactions initiated by tissue factor bypass many of the early stages of coagulation (contact phase) and involve factors VII, X, V, II and fibrinogen but also factor IX (and VIII) as it was recently demonstrated. So, it appears that tissue factor has a key-role in the haemostasic process as it has been suggested by the mildness or the absence of haemorrhagic syndrome in contact factors deficiencies. Tissue factor activity has been found in many types of cells, especially in white bloods cells. Experimental studies have demonstrated the presence of tissue factor activity in polymorphonuclears, lymphocytes, monocytes (or macrophages). This activity is enhanced by gram-negative endotoxin stimulation, inflammation, cell mediated immunologic phenomena or malignancy. These data are in good agreement with a wild range of features observed in human pathology: fibrin deposits in inflammatory lesions, disseminated intravascular coagulation (DIC) during the course of gram-negative septicemias or acute promyelocytic leukemias, local thrombi at the early phase of graft rejection. The protective effect of a phospholipase C against DIC induced in rats by tissue factor infusion suggests in the future, a specific therapy would be possible in man that, in the frequent clinical conditions involving clotting activation through tissue factor pathway.

Topics: Animals; Blood Coagulation; Blood Coagulation Factors; Disseminated Intravascular Coagulation; Fibrin; Graft Rejection; Humans; Inflammation; Leukemia; Leukocytes; Rabbits; Sepsis; Thromboplastin

To investigate the tissue factor (TF) expression on different subsets of monocyte and tissue factor secretion of peripheral blood and evaluate their association with the prognosis of sepsis in the Chinese older adult population.. Monocyte subsets and TF expression on different subsets of monocyte were measured using flow cytometry in 80 older adult sepsis patients and 40 age and sex matched healthy controls. Plasma level of TF was measured using ELISA (enzyme-linked immunosorbent assay) method.. TF expression on CD14++CD16- (MO1) monocyte was lower in death (28-day non-survivor) group compared with survival (28-day survivor) groups [1.01 % (0.58 %, 1.62 %) vs. 3.66 % (1.32 %, 6.93 %), p = 0.001]. The plasma level of TF was increased in death group compared to survival group according to the 28-day mortality [109.2 (67.3, 154.2) vs. 62.1 (44.7, 115.5) pg/mL, p = 0.031]. Logistic regression analysis showed TF expression on MO1 monocyte (β = -0.776, OR = 0.460, CI: 0.251, 0.843, p = 0.012) was independently associated with the 28-day mortality. The ROC (receiver operating characteristic) curve showed that the AUC (area under the curve) of the TF expression on MO1 monocyte for predicting 28-day mortality was 0.846 (p < 0.001).. The TF expression on CD14++CD16- monocyte is a new marker for the prognosis of older adult sepsis.

Topics: Aged; East Asian People; Flow Cytometry; Humans; Lipopolysaccharide Receptors; Monocytes; Prospective Studies; Receptors, IgG; Sepsis; Thromboplastin

Tissue factor expression on monocytes is implicated in the pathophysiology of sepsis-induced coagulopathy. How tissue factor is expressed by monocyte subsets (classical, intermediate and non-classical) is unknown.. Monocytic tissue factor surface expression was investigated during three conditions. Primary human monocytes and microvascular endothelial cell co-cultures were used for in vitro studies. Volunteers received a bolus of lipopolysaccharide (2 ng/kg) to induce endotoxemia. Patients with sepsis, or controls with critical illness unrelated to sepsis, were recruited from four intensive care units.. Contact with endothelium and stimulation with lipopolysaccharide reduced the proportion of intermediate monocytes. Lipopolysaccharide increased tissue factor surface expression on classical and non-classical monocytes. Endotoxemia induced profound, transient monocytopenia, along with activation of coagulation pathways. In the remaining circulating monocytes, tissue factor was up-regulated in intermediate monocytes, though approximately 60 % of individuals (responders) up-regulated tissue factor across all monocyte subsets. In critically ill patients, tissue factor expression on intermediate and non-classical monocytes was significantly higher in patients with established sepsis than among non-septic patients. Upon recovery of sepsis, expression of tissue factor increased significantly in classical monocytes.. Tissue factor expression in monocyte subsets varies significantly during health, endotoxemia and sepsis.

Topics: Endotoxemia; Humans; Lipopolysaccharides; Monocytes; Sepsis; Thromboinflammation; Thromboplastin

Systemic inflammatory response, as observed in sepsis and severe COVID-19, may lead to endothelial damage. Therefore, we aim to compare the extent of endothelial injury and its relationship to inflammation in both diseases. We included patients diagnosed with sepsis (SEPSIS group, n = 21), mild COVID-19 (MILD group, n = 31), and severe COVID-19 (SEVERE group, n = 24). Clinical and routine laboratory data were obtained, circulating cytokines (INF-γ, TNF-α, and IL-10) and endothelial injury markers (E-Selectin, Tissue Factor (TF) and von Willebrand factor (vWF)) were measured. Compared to the SEPSIS group, patients with severe COVID-19 present similar clinical and laboratory data, except for lower circulating IL-10 and E-Selectin levels. Compared to the MILD group, patients in the SEVERE group showed higher levels of TNF-α, IL-10, and TF. There was no clear relationship between cytokines and endothelial injury markers among the three studied groups; however, in SEVERE COVID-19 patients, there is a positive relationship between INF-γ with TF and a negative relationship between IL-10 and vWF. In conclusion, COVID-19 and septic patients have a similar pattern of cytokines and endothelial dysfunction markers. These findings highlight the importance of endothelium dysfunction in COVID-19 and suggest that endothelium should be better evaluated as a therapeutic target for the disease.

Topics: Aged; Aged, 80 and over; Biomarkers; Blood Cell Count; C-Reactive Protein; COVID-19; E-Selectin; Endothelium, Vascular; Female; Humans; Interferon-gamma; Interleukin-10; Male; Middle Aged; Retrospective Studies; SARS-CoV-2; Sepsis; Severity of Illness Index; Systemic Inflammatory Response Syndrome; Thromboplastin; Tumor Necrosis Factor-alpha; von Willebrand Factor

Inflammation and thrombosis are closely intertwined in numerous disorders, including ischemic events and sepsis, as well as coronavirus disease 2019 (COVID-19). Thrombotic complications are markers of disease severity in both sepsis and COVID-19 and are associated with multiorgan failure and increased mortality. Immunothrombosis is driven by the complement/tissue factor/neutrophil axis, as well as by activated platelets, which can trigger the release of neutrophil extracellular traps (NETs) and release further effectors of immunothrombosis, including platelet factor 4 (PF4/CXCL4) and high-mobility box 1 protein (HMGB1). Many of the central effectors of deregulated immunothrombosis, including activated platelets and platelet-derived extracellular vesicles (pEVs) expressing PF4, soluble PF4, HMGB1, histones, as well as histone-decorated NETs, are positively charged and thus bind to heparin. Here, we provide evidence that adsorbents functionalized with endpoint-attached heparin efficiently deplete activated platelets, pEVs, PF4, HMGB1 and histones/nucleosomes. We propose that this elimination of central effectors of immunothrombosis, rather than direct binding of pathogens, could be of clinical relevance for mitigating thrombotic complications in sepsis or COVID-19 using heparin-functionalized adsorbents.

Topics: Blood Coagulation; Blood Platelets; Blood Proteins; COVID-19; Extracellular Traps; Heparin; Histones; HMGB Proteins; HMGB1 Protein; Humans; Neutrophils; Platelet Activation; Platelet Factor 4; SARS-CoV-2; Sepsis; Thromboinflammation; Thromboplastin; Thrombosis

Acquired and isolated deficiencies in FVII are exceptional. They have mainly been reported during states of severe sepsis by the presence of proteases destroying the factor or neoplastic pathologies by the presence of an inhibitor. Consequently, very few cases have been published.. We report two cases of isolated and acquired deficiency of factor VII due to the presence of inhibitors which were related to bacterial sepsis in the first patient and to squamous cell carcinoma in the second patient, diagnosed in the Hematology Laboratory of the CHU Ibn Rochd.. Factor VII deficiency is a rare and poorly described deficiency that can be acquired or constitutional. The search for anti-factor VII antibodies by diluted thromboplastin time should be requested depending on the clinical context.

Topics: Antibodies; Factor VII; Factor VII Deficiency; Humans; Sepsis; Thromboplastin

Topics: Acute Lung Injury; Animals; Gene Expression Regulation; Lidocaine; Lipopolysaccharides; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Sepsis; Thromboplastin; Voltage-Gated Sodium Channel Blockers

Sepsis-induced disseminated intravascular coagulation (DIC) is a common life-threatening terminal-stage disease with high mortality. This study aimed to identify effective miRNAs as therapeutic targets for DIC. Bioinformatics and luciferase reporter gene analyses were performed to predict miR-19a-3p and validate that it targets tissue factor (TF). Quantitative real-time PCR was used to detect the expression of miR-19a-3p and TF, and TF procoagulant activity was determined using the chromogenic substrate method. Western blotting was used to detect the protein levels of TF, AKT serine/threonine kinase (AKT), extracellular regulated protein kinases (ERK), nuclear factor kappa B (NF-κB) P65, NFKB inhibitor alpha (IκB-a) and their phosphorylated counterparts in cell experiments. Furthermore, a rat model was established to explore the potential of miR-19a-3p in DIC treatment. As a result, a human clinical study revealed that miR-19a-3p was downregulated and that TF was upregulated in neonates with sepsis-induced DIC compared with those in the control group. The luciferase reporter assay showed that TF was a direct target of miR-19a-3p. Cell experiments verified that the mRNA and protein levels of TF, and the p-AKT/AKT, p-Erk/Erk, p-P65/P65, p-IκB-a/IκB-a ratios, and TF procoagulant activity were significantly decreased in lipopolysaccharide (LPS) -induced human peripheral blood mononuclear cells (PBMCs) and human umbilical vein endothelial cells (HUVECs) inhibited by overexpression of miR-19a-3p, and that miR-19a-3p regulating TF was dependent on the NF-kB and AKT pathways. In vivo, miR-19a-3p injection into DIC rats suppressed the mRNA expression of TF; more importantly, significant improvements in coagulation function indicators and in histopathologies of lung and kidney were observed. In conclusion, miR-19a-3p may suppress DIC by targeting TF and might be a potential therapeutic target in treating sepsis-induced DIC.

Topics: Animals; Cells, Cultured; Disseminated Intravascular Coagulation; Down-Regulation; Female; Human Umbilical Vein Endothelial Cells; Humans; Infant, Newborn; Lipopolysaccharides; Male; MicroRNAs; Rats; Rats, Sprague-Dawley; Sepsis; Thromboplastin

Patients with sepsis may progress to acute respiratory distress syndrome (ARDS). Evidence of neutrophil extracellular traps (NETs) in sepsis-induced lung injury has been reported. However, the role of circulating NETs in the progression and thrombotic tendency of sepsis-induced lung injury remains elusive. The aim of this study was to investigate the role of tissue factor-enriched NETs in the progression and immunothrombosis of sepsis-induced lung injury.. Human blood samples and an animal model of sepsis-induced lung injury were used to detect and evaluate NET formation in ARDS patients. Immunofluorescence imaging, ELISA, Western blotting, and qPCR were performed to evaluate. Significantly increased levels of TF-enriched NETs were observed in ARDS patients and mice. Blockade of NETs in ARDS mice alleviated disease progression, indicating a reduced lung wet/dry ratio and PaO2 level.. The interaction of thrombin-activated platelets with PMNs in ARDS patients results in local NET formation and delivery of active TF. The notion that NETs represent a mechanism by which PMNs release thrombogenic signals during thrombosis may offer novel therapeutic targets.

Topics: Animals; Disease Progression; Extracellular Traps; Humans; Lung Injury; Mice; Neutrophils; Sepsis; Thromboplastin

Sepsis is a systemic inflammatory disease causing life-threatening multi-organ dysfunction. Accumulating evidences suggest that two forms of programmed necrosis, necroptosis and pyroptosis triggered by the pathogen component lipopolysaccharide (LPS) and inflammatory cytokines, play important roles in the development of bacterial sepsis-induced shock and tissue injury. Sepsis-induced shock and tissue injury required receptor-interacting protein kinase-3 (RIPK3) and mixed lineage kinase domain-like protein (MLKL) phosphorylation, caspase11 activation and gasdermin D (GSDMD) cleavage. However, the synergistic effect of necroptosis and pyroptosis in the pathological progress of sepsis remains elusive. In this study, we found that blockage of both necroptosis and pyroptosis (double deletion of Ripk3/Gsdmd or Mlkl/Gsdmd) resulted in accumulative protection against septic shock, systemic blood clotting and multi-organ injury in mice. Bone marrow transplantation confirmed that necroptosis and pyroptosis in both myeloid and nonmyeloid cells are indispensable in the progression of sepsis-induced multi-organ injury. Both RIPK3 and GSDMD signaling collaborated to amplify necroinflammation and tissue factor release in macrophages and endothelial cells, which led to tissue injury. Furthermore, cell death induced by inflammatory cytokines and high-mobility group box 1 could be prevented by double ablation of Ripk3/Gsdmd or Mlkl/Gsdmd, suggesting that a positive feedback loop interconnecting RIPK3/MLKL and GSDMD machinery and inflammation facilitated sepsis progression. Collectively, our findings demonstrated that RIPK3-mediated necroptosis and GSDMD-mediated pyroptosis collaborated to amply inflammatory signaling and enhance tissue injury in the process of sepsis, which may shed new light on two potential targets of combined therapeutic interventions for this highly lethal disorder.

Topics: Acute Lung Injury; Animals; Blood Coagulation; Cecum; Cell Movement; Endothelial Cells; Gene Deletion; Inflammation; Interleukin-1beta; Intestines; Intracellular Signaling Peptides and Proteins; Ligation; Mice, Inbred C57BL; Mice, Knockout; Monocytes; Myeloid Cells; Necroptosis; Neutrophils; Organ Specificity; Phosphate-Binding Proteins; Punctures; Pyroptosis; Receptor-Interacting Protein Serine-Threonine Kinases; Sepsis; Signal Transduction; Thromboplastin

Sepsis is an infection-induced aggressive and life-threatening organ dysfunction with high morbidity and mortality worldwide. Infection-associated inflammation and coagulation promote the progression of adverse outcomes in sepsis. Here, we report that phospho-Tyr705 of STAT3 (pY-STAT3), not total STAT3, contributes to systemic inflammation and coagulopathy in sepsis.. Cecal ligation and puncture (CLP)-induced septic mice were treated with BP-1-102, Napabucasin, or vehicle control respectively and then assessed for systemic inflammation, coagulation response, lung function and survival. Human pulmonary microvascular endothelial cells (HPMECs) and Raw264.7 cells were exposed to lipopolysaccharide (LPS) with pharmacological or genetic inhibition of pY-STAT3. Cells were assessed for inflammatory and coagulant factor expression, cell function and signaling.. Pharmacological inhibition of pY-STAT3 expression by BP-1-102 reduced the proinflammatory factors, suppressed coagulation activation, attenuated lung injury, alleviated vascular leakage and improved the survival rate in septic mice. Pharmacological or genetic inhibition of pY-STAT3 diminished LPS-induced cytokine production in macrophages and protected pulmonary endothelial cells via the IL-6/JAK2/STAT3, NF-κB and MAPK signaling pathways. Moreover, the increase in procoagulant indicators induced by sepsis such as tissue factor (TF), the thrombin-antithrombin complex (TAT) and D-Dimer were down-regulated by pY-STAT3 inhibition.. Our results revealed a therapeutic role of pY-STAT3 in modulating the inflammatory response and defective coagulation during sepsis. Video Abstract.

Topics: Aminosalicylic Acids; Animals; Benzofurans; Blood Coagulation; Cecum; Cell Membrane Permeability; Endothelial Cells; Humans; Inflammation; Inflammation Mediators; Ligation; Lipopolysaccharides; Macrophages; Male; Mice; Mice, Inbred C57BL; Models, Biological; Molecular Targeted Therapy; Naphthoquinones; Phosphotyrosine; Punctures; RAW 264.7 Cells; Sepsis; STAT3 Transcription Factor; Sulfonamides; Suppression, Genetic; Survival Analysis; Thromboplastin; Toll-Like Receptor 4

Adult mesenchymal stem cells (MSCs) improve the host response during experimental sepsis in animals. MSCs from various sources express a procoagulant activity that has been linked to the expression of tissue factor. This study sought to determine the role of tissue factor associated with adipose-derived MSCs (ASCs) in their procoagulant and antibacterial effects during pneumonia-derived sepsis.. Mice were infused intravenously with ASCs or vehicle after infection with the common human pathogen Klebsiella pneumoniae via the airways.. Infusion of freshly cultured or cryopreserved ASCs induced the expression of many genes associated with tissue factor signaling and coagulation activation in the lungs. Freshly cultured and cryopreserved ASCs, as well as ASC lysates, exerted procoagulant activity in vitro as determined by a fibrin generation assay, which was almost completely inhibited by an anti-tissue factor antibody. Infusion of cryopreserved ASCs was associated with a rise in plasma thrombin-antithrombin complexes (indicative of coagulation activation) and formation of multiple thrombi in the lungs 4 h post-infusion. Preincubation of ASCs with anti-tissue factor antibody prior to infusion prevented the rise in plasma thrombin-antithrombin complex concentrations but did not influence thrombus formation in the lungs. ASCs reduced bacterial loads in the lungs and liver at 48 h after infection, which was not influenced by preincubation with anti-tissue factor antibody. At this late time point, microthrombi in the lungs were not detected anymore.. These data indicate that ASC-associated tissue factor is responsible for systemic activation of coagulation after infusion of ASCs but not for the formation of microthrombi in the lungs or antibacterial effects.

Topics: Adipose Tissue; Animals; Blood Coagulation; Cells, Cultured; Female; Humans; Klebsiella Infections; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Mice, Inbred C57BL; Sepsis; Thromboplastin

Severe sepsis is critical to health and can result in acute renal failure (ARF). Tissue factor (TF) and thrombomodulin (TM) play key roles in vascular endothelial functions by helping maintain microcirculation in the kidney. Budding uninhibited by benzimidazole-1 (Bub1) plays a role in Akt and JNK signaling, which control TF and TM, respectively. We hypothesized that Bub1 could control vascular endothelial function in sepsis. The aim of this study was to determine the role of Bub1 in septic ARF. We used Mouse cecum ligation and puncture (CLP) using low Bub1 expressing (Bub1) and wild-type (Bub1) mice in vivo and lipopolysaccharide (LPS) stimulation of human aortic endothelial cell (HAEC) in vitro. Bub1 mice had a higher survival rate after CLP than Bub1. Bub1 mice had more severe ARF after CLP than Bub1 with blood biochemical and pathological analyses. TF expression in Bub1 mice and control HAEC (control) significantly increased in the septic model compared with Bub1 and Bub1 silenced HAEC (siBub1). TM expression in the control significantly decreased after LPS stimulation compared with siBub1. Akt and JNK phosphorylation of siBub1 were attenuated after LPS stimulation. Associations of Bub1 with Akt or JNK after LPS stimulation of HAEC were detected using immunoprecipitation, suggesting that Bub1 is involved in the phosphorylation of Akt and JNK after LPS stimulation. Bub1 insufficiency attenuates TF expression and reduces TM suppression by blocking Akt and JNK phosphorylation, respectively, thus leading to the prevention of ARF and death caused by sepsis.

Topics: Acute Kidney Injury; Animals; Disease Models, Animal; Endothelial Cells; Humans; Mice; Mice, Mutant Strains; Protein Serine-Threonine Kinases; Sepsis; Thrombomodulin; Thromboplastin

There is a close cross-talk between complement, Toll-like receptors (TLRs) and coagulation. The role of the central complement component 5 (C5) in physiological and pathophysiological hemostasis has not, however, been fully elucidated. This study examined the effects of C5 in normal hemostasis and in Escherichia coli-induced coagulation and tissue factor (TF) up-regulation. Fresh whole blood obtained from six healthy donors and one C5-deficient individual (C5D) was anti-coagulated with the thrombin inhibitor lepirudin. Blood was incubated with or without E. coli in the presence of the C5 inhibitor eculizumab, a blocking anti-CD14 monoclonal antibody (anti-CD14) or the TLR-4 inhibitor eritoran. C5D blood was reconstituted with purified human C5. TF mRNA was measured by quantitative polymerase chain reaction (qPCR) and monocyte TF and CD11b surface expression by flow cytometry. Prothrombin fragment 1+2 (PTF1·2) in plasma and microparticles exposing TF (TF-MP) was measured by enzyme-linked immunosorbent assay (ELISA). Coagulation kinetics were analyzed by rotational thromboelastometry and platelet function by PFA-200. Normal blood with eculizumab as well as C5D blood with or without reconstitution with C5 displayed completely normal biochemical hemostatic patterns. In contrast, E. coli-induced TF mRNA and TF-MP were significantly reduced by C5 inhibition. C5 inhibition combined with anti-CD14 or eritoran completely inhibited the E. coli-induced monocyte TF, TF-MP and plasma PTF1·2. Addition of C5a alone did not induce TF expression on monocytes. In conclusion, C5 showed no impact on physiological hemostasis, but substantially contributed to E. coli-induced procoagulant events, which were abolished by the combined inhibition of C5 and CD14 or TLR-4.

Topics: Antibodies, Blocking; Antibodies, Monoclonal, Humanized; Blood Cells; Blood Coagulation; Cells, Cultured; Complement C5; Disaccharides; Escherichia coli; Escherichia coli Infections; Female; Hemostasis; Hirudins; Humans; Lipopolysaccharide Receptors; Male; Platelet Function Tests; Receptor Cross-Talk; Recombinant Proteins; Sepsis; Sugar Phosphates; Thrombelastography; Thromboplastin; Toll-Like Receptor 4

To investigate the expression changes of high mobility group box-1 (HMGB-1) and tissue factor (TF) and their correlation in the serum of sepsis rat models.. 30 rats were divided into the sham-operated group, 15 rats were in the control group. The cecal ligation and puncture method was used to make the animal model with abdominal infection induced by sepsis. There were 15 rats in the sepsis group among which they were divided into 3 subgroups at different time points after modeling (after 6 hours, 12 hours, 24 hours). Cardiac function indicators of the rats in each subgroup were monitored, including heart rate (HR), left ventricular end-diastolic pressure (LVEDP) and left ventricular developed pressure (LVDP), and enzyme-linked immunosorbent assay (ELISA) was used to test the changes of the expression levels of HMGB-1 and TF in the serum of the rats after 6 hours, 12 hours, 24 hours. Pearson correlation analysis was used to analyze the correlation between HMGB-1 and TF.. HR and LVEDP of the rats in the sepsis group were significantly higher than those of the rats in the control group. The differences were statistically significant (p<0.050). LVDP of the rats in the sepsis group was markedly lower than that of the rats in the control group. The differences were statistically significant (p<0.050). The expressions of HMGB-1 and TF of the rats in the subgroups of the sepsis group were higher than those of the rats in the control group after 6 hours, 12 hours, 24 hours; the expression levels of HMGB-1 and TF of the rats with sepsis increased with time. The differences were statistically significant (p<0.050). When the expressions of HMGB-1 and TF of the rats in the sepsis group were compared with each other within the group the differences were significantly different (p<0.050). The expressions of HMGB-1 and TF in the subgroups at the 24th hour were significantly higher than those at the 6th hour. The differences were statistically significant (p<0.050). The differences of the expression of TF of the rats in the control group were not statistically significant (p>0.050). There was a significant positive correlation between HMGB-1 and TF of the rats in the sepsis group (r=0.772, p=0.002).. The expression levels of HMGB-1 and TF of the rats with sepsis gradually increased with time, and the level of HMGB-1 was positively correlated with the level of TF.

Topics: Animals; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; HMGB1 Protein; Rats; Rats, Wistar; Sepsis; Thromboplastin

Activated immune cells such as monocytes are key factors in systemic inflammatory response syndrome (SIRS) following trauma and sepsis. Activated monocytes induce almost all tissue factor (TF) expression contributing to inflammation and coagulation. TF and CD13 double-positive microparticles (TF/CD13MPs) are predominantly released from these activated monocytes. This study aimed to evaluate TF/CD13MPs and assess their usefulness as a biomarker of pathogenesis in early SIRS following trauma and sepsis. This prospective study comprising 24 trauma patients, 25 severe sepsis patients, and 23 healthy controls was conducted from November 2012 to February 2015. Blood samples were collected from patients within 24 h after injury and diagnosis of severe sepsis and from healthy controls. Numbers of TF/CD13MPs were measured by flow cytometry immediately thereafter. Injury Severity Score (ISS) and Acute Physiology and Chronic Health Evaluation (APACHE) II and Sequential Organ Failure Assessment (SOFA) scores were calculated at patient enrollment. APACHE II and SOFA scores and International Society of Thrombosis and Haemostasis (ISTH) overt disseminated intravascular coagulation (DIC) diagnostic criteria algorithm were calculated at the time of enrollment of severe sepsis patients. Numbers of TF/CD13MPs were significantly increased in both trauma and severe sepsis patients versus controls and correlated significantly with ISS and APACHE II score in trauma patients and with APACHE II and ISTH DIC scores in severe sepsis patients. Increased numbers of TF/CD13MPs correlated significantly with severities in the acute phase in trauma and severe sepsis patients, suggesting that TF/CD13MPs are important in the pathogenesis of early SIRS following trauma and sepsis.

Topics: Adult; Aged; APACHE; Biomarkers; CD13 Antigens; Cell-Derived Microparticles; Female; Humans; Injury Severity Score; Male; Middle Aged; Prospective Studies; Sepsis; Systemic Inflammatory Response Syndrome; Thromboplastin; Wounds and Injuries

Neisseria meningitidis (N. meningitidis) may cause sepsis and meningitis. N. meningitidis with a mutated lpxL1 gene has five, instead of six, acyl chains in the lipid A moiety. Compared with patients infected with the wild type (wt) meningococcus, patients infected with the lpxL1 mutant have a mild meningococcal disease with less systemic inflammation and less coagulopathy. Circulating tissue factor (TF), the main initiator of coagulation, has a central role in the development of coagulation disturbances during sepsis. To study how TF was influenced by the lpxL1 mutant, human primary monocytes and whole blood were incubated with the lpxL1 mutant or the wt meningococcus (H44/76). Monocyte and microvesicle (MV)-associated TF expression and TF-dependent thrombin generation were measured. In both purified monocytes and whole blood, our data show that the lpxL1 mutant is a weaker inducer of monocyte and MV-associated TF compared with the wt. Our data indicate that low levels of circulating TF may contribute to the reduced coagulopathy reported in patients infected with lpxL1 mutants.

Topics: Acyltransferases; Bacterial Proteins; Blood Coagulation; Cell-Derived Microparticles; Cells, Cultured; Disease Progression; Humans; Inflammation; Meningitis; Monocytes; Mutation; Neisseria meningitidis; Primary Cell Culture; Sepsis; Thromboplastin

To examine whether sodium tanshinone II A sulfonate (STS), the main effective component of Salvia miltiorrhiza is effective in relieving the microcirculatory disturbance of small intestine by suppressing the production of reactive oxygen species (ROS) in rats with sepsis.. A rat model of sepsis was induced by cecal ligation and puncture (CLP). Rats (n =40) were randomly divided into 4 groups: sham-operated group (sham, n =10), sepsis group (CLP, n =10), STS treatment group (STS, n =10) and ROS scavenger dimethylthiourea (DMTU, n =10) group. Animals in the STS group were injected with STS (1 mg/kg) for 10 min through the right external jugular vein after the CLP operation, and animals in the CLP group were given the same volume of normal saline after the CLP operation. Animals in the DMTU group were intraperitoneally injected with 5 mL/kg of 20% DMTU 1 h before CLP. The histopathologic changes in the intestinal tissues and changes of mesenteric microcirculation were observed. The levels of ROS in intestinal tissues from each group were qualitatively evaluated using a fluorescent microscope. The expressions of apoptosis signal-regulating kinase (ASK1), phosphorylated ASK1 (phospho-ASK1), p38 mitogen-activated protein kinases (p38 MAPK), phosphorylated p38 MAPK (phospho-p38 MAPK) and tissue factor (TF) were determined by Western blotting.. It was shown that there were obvious microcirculatory disturbance (P <0.05) and tissue injuries in intestinal tissues after CLP operation. The levels of ROS production, phospho-ASK1, phospho-p38 MAPK and TF were increased. Both STS and DMTU suppressed ROS, phospho-ASK1, phospho-p38 MAPK and TF production, and ameliorated the microcirculatory disturbance and tissues injury (P <0.01).. STS can ameliorate the microcirculatory disturbance of the small intestine by attenuating the production of ROS in rats with sepsis.

Topics: Animals; Intestine, Small; Male; MAP Kinase Kinase Kinase 5; Microcirculation; p38 Mitogen-Activated Protein Kinases; Phenanthrenes; Phosphorylation; Rats, Wistar; Reactive Oxygen Species; Sepsis; Thromboplastin

Sepsis-induced acute lung injury (ALI) is characterized by fibrin deposition, which indicates the local activation of coagulation. Tissue factor (TF), expressed in the pulmonary microvasculature, acts as a critical initiator of blood coagulation and ALI in sepsis. The molecular mechanism of lipopolysaccharide (LPS)-induced TF expression in endothelial cells (ECs), however, has not been determined. In this study, we implicate the Rho-associated protein kinase (ROCK)/Yes associated protein (YAP)/early growth response (Egr-1) signaling pathway in LPS-induced TF expression in vitro and in sepsis-induced ALI in vivo.. Human umbilical vein ECs incubated with LPS were pretreated with or without the ROCK inhibitor Y-27632, a YAP small, interfering RNA (siRNA) and an Egr-1 siRNA. ROCK, YAP and Egr-1 signaling-induced protein expression was investigated by Western blot. The LPS-induced activation of YAP was analyzed by an immunofluorescent assay. Furthermore, we intratracheally injected YAP siRNA to assess septic ALI in mice by hematoxylin and eosin staining.. LPS rapidly induced ROCK activation and increased TF expression in ECs. LPS caused YAP shuttling into the nuclei of ECs and combined with Egr-1 via the activation of ROCK. Furthermore, the LPS-mediated TF expression increase was prevented by ROCK inactivation, YAP knockdown and Egr-1 depletion, suggesting that LPS-induced TF expression is closely associated with the ROCK/YAP/Egr-1 signaling pathway in ECs. Finally, an intratracheal injection of YAP siRNA relieved lung injury in septic mice.. This study not only suggests that ROCK/YAP/Egr-1 signaling regulates TF expression after stimulation with LPS in ECs, but it also indicates that LPS-induced activation of YAP signaling plays an important role in septic ALI in mice. Our findings provide a new insight into the pathogenic mechanism of TF expression, which is closely linked to septic ALI, and YAP signaling is considered to be a novel target for therapeutic intervention under septic conditions.

Topics: Acute Lung Injury; Adaptor Proteins, Signal Transducing; Animals; Biomarkers; Blotting, Western; Cell Cycle Proteins; Early Growth Response Protein 1; Human Umbilical Vein Endothelial Cells; Humans; Lipopolysaccharides; Male; Mice; Mice, Inbred C57BL; Phosphoproteins; rho-Associated Kinases; Sepsis; Signal Transduction; Thromboplastin; Transcription Factors; YAP-Signaling Proteins

Sepsis is characterized by systemic activation of inflammatory and coagulation pathways in response to infection. Recently, it was demonstrated that histones released into the circulation by dying/activated cells may contribute to sepsis pathology. Although the ability of extracellular histones to modulate the procoagulant activities of several cell types has been investigated, the influence of histones on the hemostatic functions of circulating monocytes is unknown. To address this, we investigated the ability of histones to modulate the procoagulant potential of THP-1 cells and peripheral blood monocytes, and examined the effects of plasmas obtained from septic patients to induce a procoagulant phenotype on monocytic cells.. Tissue factor (TF) activity assays were performed on histone-treated THP-1 cells and blood monocytes. Exposure of monocytic cells to histones resulted in increases in TF activity, TF antigen, and phosphatidylserine exposure. Histones modulate the procoagulant activity via engagement of Toll-like receptors 2 and 4, and this effect was abrogated with inhibitory antibodies. Increased TF activity of histone-treated cells corresponded to enhanced thrombin generation in plasma determined by calibrated automated thrombography. Finally, TF activity was increased on monocytes exposed to plasma from septic patients, an effect that was attenuated in plasma from patients receiving unfractionated heparin (UFH).. Our studies suggest that increased levels of extracellular histones found in sepsis contribute to dysregulated coagulation by increasing TF activity of monocytes. These procoagulant effects can be partially ameliorated in sepsis patients receiving UFH, thereby identifying extracellular histones as a potential therapeutic target for sepsis treatment.

Topics: Adult; Blood Coagulation; Cell Survival; Cells, Cultured; Histones; Humans; Monocytes; Phosphatidylserines; Plasma; Sepsis; THP-1 Cells; Thrombin; Thromboplastin

Sepsis-induced disseminated intravascular coagulation (DIC) is a major cause of death in patients admitted to intensive care units. Endothelial injury with microparticle production is reported in the pathogenesis of sepsis. Endothelial microparticles (EMPs) present several cell-specific surface antigens with different bioactivities, for example, tissue factor (TF), thrombomodulin (TM), and endothelial protein C receptor (EPCR). We investigated associations between these three different surface antigen-positive EMPs and sepsis-induced DIC. This cross-sectional study composed of 24 patients with sepsis and 23 healthy controls was conducted from November 2012 to September 2013. Blood samples were collected from patients within 24 h of diagnosis of severe sepsis and from healthy controls. Numbers of TF-positive EMPs (TF EMPs), TM-positive EMPs (TM EMPs), and EPCR-positive EMPs (EPCR EMPs) were measured by flow cytometry immediately thereafter. Acute Physiology and Chronic Health Evaluation II and Sequential Organ Failure Assessment scores were assessed in the severe sepsis patients at enrollment. We assessed DIC with the International Society of Thrombosis and Haemostasis (ISTH) overt DIC diagnostic criteria algorithm. Numbers of antigen-positive EMPs were increased significantly in both severe sepsis patients and controls and with the increase in ISTH DIC score. Numbers of TF EMPs and EPCR EMPs correlated significantly with Sequential Organ Failure Assessment score, and numbers of EPCR EMPs correlated significantly with Acute Physiology and Chronic Health Evaluation II score. Numbers of the three antigen-positive EMPs were increased significantly in severe sepsis patients versus those in healthy controls and with the increase of ISTH DIC score, suggesting that the specific bioactivity of each antigen-positive EMP may play a role in the progression of sepsis-induced DIC.

Topics: Adult; Aged; Algorithms; Antigens, CD; Antigens, Surface; Case-Control Studies; Cell-Derived Microparticles; Cross-Sectional Studies; Disease Progression; Disseminated Intravascular Coagulation; Endothelial Protein C Receptor; Endothelium; Female; Fibrin Fibrinogen Degradation Products; Fibrinogen; Flow Cytometry; Gene Expression Regulation; Humans; Intensive Care Units; Interleukin-6; Male; Middle Aged; Receptors, Cell Surface; Sepsis; Surface Properties; Thrombomodulin; Thromboplastin; Time Factors

Tissue factor (TF) and tissue factor pathway inhibitor (TFPI) play a central role in the endothelial permeability regulation and dysfunction, which is associated with the development of sepsis and acute lung injury/acute respiratory distress syndrome (ALI/ARDS). The aim of this study is to assess the diagnostic and prognostic values of TF and TFPI in patients with sepsis and sepsis-induced ARDS.. A total of 62 patients with sepsis, 167 patients with severe sepsis and 32 healthy volunteers were enrolled in this prospective observational study. TF and TFPI levels were measured by enzyme-linked immunosorbent assay (ELISA).. Patients with sepsis-induced ARDS showed significantly higher median levels of TF compared with patients without ARDS (1425.5 (1019.9 to 2595.2) pg/ml vs 916.2 (724.1 to 1618.2) pg/ml, P < 0.001), and compared with sepsis patients (943.5 (786.4 to 992.4) pg/ml, P < 0.001) on the day of admission. However, there was no significant difference between sepsis patients and healthy subjects, or between septic shock and non-septic shock patients (P > 0.05). The AUC of TF for the diagnosis of sepsis-induced ARDS was 0.749 (95% confidence interval (CI) 0.675-0.822). Plasma TF levels in the non-survivors of severe sepsis were significantly higher than those of survivors (1618.6 (1017.1 to 2900.8) pg/ml vs. 979.9 (757.2 to 1645.5) pg/ml, P < 0.001), and multivariate logistic regression showed the plasma value of TF was the independent predictor for 30-day mortality in patients with severe sepsis (P = 0.0022, odds ratio (OR) = 1.41, 95% CI 1.24-1.69). The AUC of TF for predicting 30-day mortality in severe sepsis patients was 0.718 (95% CI 0.641-0.794). However, there was no significant difference in the plasma TFPI values among the healthy control, sepsis and severe sepsis groups (P > 0.05).. Our data showed that tissue factor is a valuable diagnostic biomarker for the diagnosis of sepsis-induced ARDS. Moreover, tissue factor is a strong prognostic marker for short-term mortality in severe sepsis and sepsis-induced ARDS patients.

Topics: Acute Lung Injury; Aged; Case-Control Studies; Female; Humans; Male; Middle Aged; Prognosis; Respiratory Distress Syndrome; ROC Curve; Sepsis; Survival Analysis; Thromboplastin; Treatment Outcome

To explore the effects of rosiglitazone, a synthetic ligand of proliferator-activated receptor-γ (PPAR-γ) on vascular endothelial injuries in septic rats.. A total of 40 male Sprague-Dawley rats were randomly divided into 4 groups of vehicle control, lipopolysaccharide (LPS), pretreatment of rosiglitazone (ROSI) and pretreatment of PPAR-γ antagonist 2-chloro-5-nitroaniline (GW9662) (n=10 each). At 4 hours post-intervention, blood samples were collected to detect the expression of PPAR-γ by immunocytochemistry and image analysis. And the following parameters of vascular endothelial injury were measured: Vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), angiopoietin-2 (Ang-2), thrombomodulin (TM), anti-thrombin III (AT-III), tissue factor (TF), von Willebrand factor (vWF) and circulating endothelial cell (CEC).. ① In ROSI group, the expression of PPAR-γ was significantly higher than that in LPS group (P<0.01). In GW9662 group, the expression of PPAR-γ had no significant difference compared to vehicle control group (P>0.05). ② The serum concentrations of VCAM-1, ICAM-1, Ang-2, TM, AT-III, TF and vWF were significantly higher in LPS group than those in vehicle control group (P<0.01). The concentrations of these parameters in ROSI group were significantly lower than those in LPS group (P<0.01). In GW9662 group, the concentrations of these parameters had no significant difference compared with LPS group (P>0.05). ③ The numbers of CEC were significantly higher in LPS group than those in vehicle control group (P<0.01). And the numbers of CEC were significantly lower in ROSI group than those in LPS group (P<0.01). In GW9662 group, the numbers of CEC had no significant difference compared with LPS group (P>0.05).. Proliferator-activated receptor-γ agonist improves sepsis-induced vascular endothelial injury. And its mechanism may be through stabilizing vascular endothelial cell for improving serious inflammatory reaction and blood coagulation dysfunction.

Topics: Angiopoietin-2; Anilides; Animals; Endothelium, Vascular; Intercellular Adhesion Molecule-1; Lipopolysaccharides; Male; PPAR gamma; Rats; Rats, Sprague-Dawley; Rosiglitazone; Sepsis; Thiazolidinediones; Thromboplastin; Vascular Cell Adhesion Molecule-1

The complement system and the Toll-like (TLR) co-receptor CD14 play important roles in innate immunity and sepsis. Tissue factor (TF) is a key initiating component in intravascular coagulation in sepsis, and long pentraxin 3 (PTX3) enhances the lipopolysaccharide (LPS)-induced transcription of TF. The aim of this study was to study the mechanism by which complement and CD14 affects LPS- and Escherichia coli (E. coli)-induced coagulation in human blood. Fresh whole blood was anti-coagulated with lepirudin, and incubated with ultra-purified LPS (100 ng/ml) or with E. coli (1 × 10(7) /ml). Inhibitors and controls included the C3 blocking peptide compstatin, an anti-CD14 F(ab')2 antibody and a control F(ab')2 . TF mRNA was measured using quantitative polymerase chain reaction (qPCR) and monocyte TF surface expression by flow cytometry. TF functional activity in plasma microparticles was measured using an amidolytic assay. Prothrombin fragment F 1+2 (PTF1.2) and PTX3 were measured by enzyme-linked immunosorbent assay (ELISA). The effect of TF was examined using an anti-TF blocking antibody. E. coli increased plasma PTF1.2 and PTX3 levels markedly. This increase was reduced by 84->99% with compstatin, 55-97% with anti-CD14 and > 99% with combined inhibition (P < 0·05 for all). The combined inhibition was significantly (P < 0·05) more efficient than compstatin and anti-CD14 alone. The LPS- and E. coli-induced TF mRNA levels, monocyte TF surface expression and TF functional activity were reduced by > 99% (P < 0·05) with combined C3 and CD14 inhibition. LPS- and E. coli-induced PTF1.2 was reduced by 76-81% (P < 0·05) with anti-TF antibody. LPS and E. coli activated the coagulation system by a complement- and CD14-dependent up-regulation of TF, leading subsequently to prothrombin activation.

Topics: Antithrombins; Blood Coagulation; C-Reactive Protein; Complement C3; Escherichia coli; Hirudins; Humans; Lipopolysaccharide Receptors; Lipopolysaccharides; Peptide Fragments; Peptides, Cyclic; Prothrombin; Recombinant Proteins; RNA, Messenger; Sepsis; Serum Amyloid P-Component; Thromboplastin; Up-Regulation

The key effector molecule of the natural protein C pathway, activated protein C (aPC), exerts pleiotropic effects on coagulation, fibrinolysis, and inflammation. Coagulation-independent cell signaling by aPC appears to be the predominant mechanism underlying its highly reproducible therapeutic efficacy in most animal models of injury and infection. In this study, using a mouse model of Staphylococcus aureus sepsis, we demonstrate marked disease stage-specific effects of the anticoagulant and cell signaling functions of aPC. aPC resistance of factor (f)V due to the R506Q Leiden mutation protected against detrimental anticoagulant effects of aPC therapy but also abrogated the anti-inflammatory and mortality-reducing effects of the signaling-selective 5A-aPC variant that has minimal anticoagulant function. We found that procofactor V (cleaved by aPC at R506) and protein S were necessary cofactors for the aPC-mediated inhibition of inflammatory tissue-factor signaling. The anti-inflammatory cofactor function of fV involved the same structural features that govern its cofactor function for the anticoagulant effects of aPC, yet its anti-inflammatory activities did not involve proteolysis of activated coagulation factors Va and VIIIa. These findings reveal a novel biological function and mechanism of the protein C pathway in which protein S and the aPC-cleaved form of fV are cofactors for anti-inflammatory cell signaling by aPC in the context of endotoxemia and infection.

Topics: Animals; Factor V; Mice; Mice, Transgenic; Protein C; Protein S; Sepsis; Signal Transduction; Staphylococcal Infections; Staphylococcus aureus; Thromboplastin

Topics: Animals; Factor V; Protein C; Sepsis; Signal Transduction; Staphylococcal Infections; Staphylococcus aureus; Thromboplastin

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

Inflammatory responses can induce microvascular and endothelial dysfunction, which is associated with the development of sepsis. This study is aimed at examining the concentrations of plasma tissue factor (TF), von Willebrand factor (vWF), and tumor necrosis factor-α (TNF-α) in patients with sepsis and at determining how septic plasma (SP) regulates TF and vWF expression and p38 mitogen activated protein kinase (p38 MAPK)/nuclear factor-κB (NF-κB) pathways in human endothelial cells. The concentrations of plasma TF, vWF, and TNF-α in 22 septic patients and eight healthy controls (HCs) were examined by enzyme-linked immunosorbent assay, and their potential association with disease severity was analyzed. Human umbilical vein endothelial cells (HUVECs) were treated with SP from patients or normal plasma (NP) from the HCs, and the levels of TF and vWF were measured. The SP-induced ERK, p38 MAPK, and NF-κB activation was characterized by Western blot and immunofluorescent assays. The SP-induced HUVEC apoptosis was detected by flow cytometry. The concentrations of plasma TF, vWF, and TNF-α in the patients were significantly higher than that in the HCs and were positively correlated with the Acute Physiology and Chronic Health Evaluation II scores in the patients. Furthermore, treatment with SP, but not NP, induced TF and vWF production in HUVECs in a dose- and time-dependent manner, which was associated with sequential activation of the p38 MAPK and NF-κB pathways. Septic plasma induced HUVEC apoptosis, which was inhibited by activating the NF-κB pathway. The sepsis-related inflammatory factors promoted endothelial cell activation, dysfunction, and apoptosis through activation of the p38 MAPK pathway that was regulated by NF-κB signaling.

Topics: Aged; Apoptosis; Cells, Cultured; Endothelial Cells; Endothelium, Vascular; Female; Humans; Male; MAP Kinase Signaling System; Middle Aged; NF-kappa B; p38 Mitogen-Activated Protein Kinases; Sepsis; Thromboplastin; Tumor Necrosis Factor-alpha; von Willebrand Factor

Outer membrane vesicles (OMVs) were previously shown to be capable of initiating the inflammatory response seen in the transition of an infection to sepsis. However, another tenet of sepsis is the development of a hypercoagulable state and the role of OMVs in the development of this hypercoagulability has not been evaluated. The objective of this study was to evaluate the ability of OMVs to elicit endothelial mediators of coagulation and inflammation and induce platelet activation.. Human umbilical vein endothelial cells (HUVECs) were incubated with OMVs and were analyzed for the expression of tissue factor (TF), thrombomodulin, and the adhesion molecules P-selectin and E-selectin. Supernatants of OMV-treated HUVECs were mixed with whole blood and assessed for prothrombotic monocyte-platelet aggregates (MPA).. OMVs induce significantly increased expression of TF, E-selectin, and P-selectin, whereas, the expression of thrombomodulin by HUVECs is significantly decreased (P < 0.05). The lipopolysaccharide inhibitor clearly inhibited the expression of E-selectin following incubation with OMVs, although its impact on TF and thrombomodulin expression was nominal. Incubation of whole blood with supernatant from HUVECs exposed to OVMs resulted in increased MPAs.. This study demonstrates that, at the cellular level, OMVs from pathogenic bacteria play a complex role in endothelial activation. Although OMV-bound lipopolysaccharide modulates inflammatory proteins, including E-selectin, it has a negligible effect on the tested coagulation mediators. Additionally, endothelial activation by OMVs facilitates platelet activation as indicated by increased MPAs. By influencing the inflammatory and coagulation cascades, OMVs may contribute to the hypercoagulable response seen in sepsis.

Topics: Bacterial Outer Membrane Proteins; Blood Coagulation; Cell-Derived Microparticles; Cytoplasmic Vesicles; E-Selectin; Escherichia coli; Human Umbilical Vein Endothelial Cells; Humans; Lipopolysaccharides; Monocytes; P-Selectin; Platelet Activation; Sepsis; Thrombomodulin; Thrombophilia; Thromboplastin

Activation of inflammation and coagulation was closely related and mutually interdependent in sepsis. Tissue factor (TF) and its endogenous inhibitor, tissue factor pathway inhibitor (TFPI) was the main regulators of the initiation of coagulation process. Altered plasma levels of TF and TFPI have been related to worse outcome in sepsis. The objective of this study was to investigate whether single nucleotide polymorphisms (SNPs) in the TF and TFPI genes were associated with risk and outcome for patients with severe sepsis.. Seventeen SNPs in TF and TFPI were genotyped in samples of sepsis (n =577) and severe sepsis patients (n =476), and tested for association in this case-control collection. We then investigated correlation between the associated SNPs and the mRNA expression, and protein level of the corresponding gene. The mRNA levels of TF were determined using real-time quantitative reverse transcription-polymerase chain reaction and the soluble plasma levels of TF were measured using enzyme linked immunosorbent assay (ELISA) method.. Association analysis revealed that three TF SNPs in perfect linkage disequilibrium, rs1361600, rs3917615 and rs958587, were significantly associated with outcome of severe sepsis. G allele frequency of rs1361600 in survivor patients was significantly higher than that in nonsurvivor severe sepsis patients (P =4.91 × 10(-5), odds ratio (OR) =0.48, 95% confidence interval (CI) 0.33 to 0.69). The association remained significant after adjustment for covariates in multiple logistic regression analysis and for multiple comparisons. Lipopolysaccharide-induced TF-mRNA expression levels in peripheral blood mononuclear cells from subjects carrying rs1361600 AG and GG genotypes, were significantly lower than those subjects carrying AA genotype (P =0.0012). Moreover, severe sepsis patients of GG and GA genotypes showed lower serum levels of TF than patients with AA genotype (P adj =0.02). The plasma levels of TF were also associated with outcome of severe sepsis patients (P adj =0.01). However, genotype and allele analyses did not show any significant difference between sepsis and severe sepsis patients.. Our findings indicate that common genetic variation in TF was significantly associated with outcome of severe sepsis in Chinese Han population.

Topics: Aged; Aged, 80 and over; Asian People; Case-Control Studies; Female; Genetic Variation; Humans; Male; Middle Aged; Polymorphism, Single Nucleotide; Sepsis; Thromboplastin; Treatment Outcome

The complement pathway and CD14 play essential roles in inflammation, but little is known about the relative roles of complement and CD14 in E. coli-induced tissue factor (TF) mRNA upregulation, expression by monocytes, and functional activity in human whole blood.. Whole E. coli bacteria were incubated for up to 4 h in human whole blood containing the anticoagulant lepirudin, which does not affect complement activation. TF mRNA levels were analyzed using reverse transcription, quantitative real-time PCR (RT-qPCR), and the expression of TF on the cell surface was analyzed using flow cytometry. Complement was selectively inhibited using the C3 convertase inhibitor compstatin or a C5a receptor antagonist (C5aRa), while CD14 was blocked by an anti-CD14 F(ab')2 monoclonal antibody.. The E. coli-induced TF mRNA upregulation was reduced to virtually background levels by compstatin, whereas anti-CD14 had no effect. Monocyte TF expression and TF activity in plasma microparticles were significantly reduced by C5aRa. Anti-CD14 alone only slightly reduced E. coli-induced monocyte TF expression but showed a modest additive effect when combined with the complement inhibitors. Inhibiting complement and CD14 efficiently reduced the expression of the E. coli-induced cytokines IL-1beta, IL-6, IL-8, and platelet-derived growth factor bb.. Our results indicate that E. coli-induced TF mRNA upregulation is mainly dependent on complement activation, while CDI4 plays a modest role in monocyte TF expression and the plasma TF activity in human whole blood.

Topics: Adult; Anticoagulants; Complement Inactivating Agents; Complement System Proteins; Cytokines; Dose-Response Relationship, Drug; Enzyme-Linked Immunosorbent Assay; Escherichia coli Infections; Flow Cytometry; Hirudins; Humans; Lipopolysaccharide Receptors; Monocytes; Real-Time Polymerase Chain Reaction; Recombinant Proteins; RNA, Messenger; Sepsis; Thromboplastin; Up-Regulation

During severe sepsis, microvesicles that are positive for tissue factor (TF) are at increased levels within blood and in pulmonary lavage. These microvesicles potentially disperse TF, the major initiator of the coagulation cascade, throughout multiple organ systems, initiating fibrin deposition and resultant ischemia. The source of these microvesicles has remained incompletely defined. Although TF(+) microvesicles are shed from cells that express nascent TF transcript in response to injury, recent findings revealed that circulating, full-length TF protein is detectable prior to these nascent transcripts. This finding suggested that the protein is released from constitutive sources as an acute response. We examined whether Staphylococcus aureus, the Gram-positive bacteria that is emerging as one of the most common etiologic agents in sepsis, is capable of stimulating the release of TF(+) microvesicles from a pulmonary cell line that constitutively expresses TF protein. We found that host cell invasion stimulated an acute release of TF(+) microvesicles and that these microvesicles mediated the transfer of the protein to TF-negative endothelial cells. We also found that transfer was inhibited by cholesterol-lowering simvastatin. Taken together, our findings reveal that S. aureus pathogenesis extends to the acute release of TF(+) microvesicles and that inhibiting dispersal by this mechanism may provide a therapeutic target.

Topics: Cell Line; Cytoplasmic Vesicles; Host-Pathogen Interactions; Humans; Lung; Protein Transport; Sepsis; Simvastatin; Staphylococcal Infections; Staphylococcus aureus; Thromboplastin

Both the complement system and tissue factor (TF), a key initiating component of coagulation, are activated in sepsis, and cross-talk occurs between the complement and coagulation systems. C1-inhibitor (C1-INH) can act as a regulator in both systems. Our aim in this study was to examine this cross-talk by investigating the effects of C1-INH on Escherichia coli-induced haemostasis and inflammation. Fresh human whole blood collected in lepirudin was incubated with E. coli or ultrapurified E. coli lipopolysaccharide (LPS) in the absence or presence of C1-INH or protease-inactivated C1-INH. C3 activation was blocked by compstatin, a specific C3 convertase inhibitor. TF mRNA was measured using reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and TF surface expression was measured by flow cytometry. In plasma, the terminal complement complex, prothrombin F1·2 (PTF1·2) and long pentraxin 3 (PTX3) were measured by enzyme-linked immunosorbent assay (ELISA). Cytokines were analysed using a multiplex kit. C1-INH (1·25-5 mg/ml) reduced both LPS- and E. coli-induced coagulation, measured as a reduction of PTF1·2 in plasma, efficiently and dose-dependently (P < 0·05). Both LPS and E. coli induced marked up-regulation of TF mRNA levels and surface expression on whole blood monocytes. This up-regulation was reduced efficiently by treatment with C1-INH (P < 0·05). C1-INH reduced the release of PTX3 (P < 0·05) and virtually all cytokines measured (P < 0·05). Complement activation was inhibited more efficiently with compstatin than with C1-INH. C1-INH inhibited most of the other readouts more efficiently, consistent with additional non-complement-dependent effects. These results indicate that complement plays a role in activating coagulation during sepsis and that C1-INH is a broad-spectrum attenuator of the inflammatory and haemostatic responses.

Topics: Blood Coagulation; C-Reactive Protein; Cells, Cultured; Coculture Techniques; Complement C1 Inactivator Proteins; Complement C1 Inhibitor Protein; Escherichia coli; Female; Gene Expression Regulation; Humans; Lipopolysaccharides; Male; Monocytes; Peptide Fragments; Prothrombin; RNA, Messenger; Sepsis; Serum Amyloid P-Component; Thromboplastin

Sepsis, a leading cause of death worldwide, involves widespread activation of inflammation, massive activation of coagulation, and lymphocyte apoptosis. Calpains, calcium-activated cysteine proteases, have been shown to increase inflammatory reactions and lymphocyte apoptosis. Moreover, calpain plays an essential role in microparticle release.. We investigated the contribution of calpain in eliciting tissue damage during sepsis.. To test our hypothesis, we induced polymicrobial sepsis by cecal ligation and puncture in wild-type (WT) mice and transgenic mice expressing high levels of calpastatin, a calpain-specific inhibitor.. In WT mice, calpain activity increased transiently peaking at 6 hours after cecal ligation and puncture surgery. Calpastatin overexpression improved survival, organ dysfunction (including lung, kidney, and liver damage), and lymphocyte apoptosis. It decreased the sepsis-induced systemic proinflammatory response and disseminated intravascular coagulation, by reducing the number of procoagulant circulating microparticles and therefore delaying thrombin generation. The deleterious effect of microparticles in this model was confirmed by transferring microparticles from septic WT to septic transgenic mice, worsening their survival and coagulopathy.. These results demonstrate an important role of the calpain/calpastatin system in coagulation/inflammation pathways during sepsis, because calpain inhibition is associated with less severe disseminated intravascular coagulation and better overall outcomes in sepsis.

Topics: Animals; Apoptosis; Calcium-Binding Proteins; Calpain; Cell-Derived Microparticles; Cytokines; Disease Models, Animal; Disseminated Intravascular Coagulation; Lymphocytes; Mice; Mice, Inbred C57BL; Mice, Transgenic; Multiple Organ Failure; NF-kappa B; Sepsis; Thromboplastin

Sepsis is associated with systemic inflammatory responses and induction of coagulation system. Neutrophil extracellular traps (NETs) constitute an antimicrobial mechanism, recently implicated in thrombosis via platelet entrapment and aggregation.. In this study, we demonstrate for the first time the localization of thrombogenic tissue factor (TF) in NETs released by neutrophils derived from patients with gram-negative sepsis and normal neutrophils treated with either serum from septic patients or inflammatory mediators involved in the pathogenesis of sepsis. Localization of TF in acidified autophagosomes was observed during this process, as indicated by positive LC3B and LysoTracker staining. Moreover, phosphatidylinositol 3-kinase inhibition with 3-MA or inhibition of endosomal acidification with bafilomycin A1 hindered the release of TF-bearing NETs. TF present in NETs induced thrombin generation in culture supernatants, which further resulted in protease activated receptor-1 signaling.. This study demonstrates the involvement of autophagic machinery in the extracellular delivery of TF in NETs and the subsequent activation of coagulation cascade, providing evidence for the implication of this process in coagulopathy and inflammatory response in sepsis.

Topics: Autophagy; Cells, Cultured; Humans; Macrolides; Neutrophils; Phosphatidylinositol 3-Kinase; Phosphoinositide-3 Kinase Inhibitors; Protein Transport; Sepsis; Thromboplastin

The endoplasmic reticulum (ER) is responsible for the synthesis and folding of secretory, transmembrane and ER-resident proteins. Conditions that impair protein folding or overwhelm its protein folding capacity disrupt ER homeostasis, thereby causing ER stress. ER stress-induced apoptosis and inflammation are involved in the pathogenesis of inflammatory diseases. Activated protein C (APC) inhibits inflammation and apoptosis in monocytes, and this may partly explain the protective effects of APC treatment in severe sepsis. However, the precise molecular pathways by which APC modulates these effects remain unknown.. To investigate whether APC modulates the ER stress response in human monocytes.. We treated monocytes with ER stress-inducing agents in the presence or absence of APC to determine the effect on this response. Protein and mRNA levels were determined by immunoblotting and real-time PCR, respectively. Enzyme assays and flow cytometry were used to determine the role of APC in this model.. In thapsigargin (Tg)-treated cells, APC dampened unfolded protein response activation, as indicated by reduced levels of the 78-kDa glucose-regulated protein (GRP78), in an endothelial protein C receptor-independent and protease-activated receptor-1-independent manner. Consistent with this, APC decreased phosphorylated eukaryotic translational initiation factor 2α and C/EBP homologous protein levels induced by Tg. APC inhibited Tg-induced ER Ca(2+) flux and reactive oxygen species generation. Functionally, APC diminished Tg-induced caspase-3 activity and degradation of the nuclear factor kappaB inhibitor IκBα. Furthermore, APC dampened the induction of tissue factor procoagulant activity facilitated by Tg.. These studies suggest that APC modulates the adverse effects of ER Ca(2+) depletion in human monocytes.

Topics: Anti-Inflammatory Agents; Apoptosis; Base Sequence; Calcium; Calcium Signaling; Caspase 3; Endoplasmic Reticulum; Endoplasmic Reticulum Chaperone BiP; Gene Expression; Heat-Shock Proteins; Humans; In Vitro Techniques; Monocytes; NF-kappa B; Protein C; Reactive Oxygen Species; Recombinant Proteins; RNA, Messenger; Sepsis; Stress, Physiological; Thapsigargin; Thromboplastin

Sepsis is a systemic host response to invasive infection by bacteria. Despite treatment with antibiotics, current mortality rates are in the range of 20%-25%, which makes sepsis the most important cause of death in intensive care. Gram-negative bacteria are a prominent cause of sepsis. Lipopolysaccharide (LPS), one of the major constituents of the outer membrane of Gram-negative bacteria, plays a major role in activating the host's immune response by binding to monocytes and other cells. Several proteins are involved in neutralization and clearance of LPS from the bloodstream. Here, we provide evidence that β₂-glycoprotein I (β₂GPI) is a scavenger of LPS. In vitro, β₂GPI inhibited LPS-induced expression of tissue factor and IL-6 from monocytes and endothelial cells. Binding of β₂GPI to LPS caused a conformational change in β₂GPI that led to binding of the β₂GPI-LPS complex to monocytes and ultimately clearance of this complex. Furthermore, plasma levels of β₂GPI were inversely correlated with temperature rise and the response of inflammatory markers after a bolus injection of LPS in healthy individuals. Together, these observations provide evidence that β₂GPI is involved in the neutralization and clearance of LPS and identify β₂GPI as a component of innate immunity.

Topics: beta 2-Glycoprotein I; Cell Line; Cells, Cultured; Endothelial Cells; Escherichia coli; Humans; Immunity, Innate; Interleukin-6; Lipopolysaccharides; Monocytes; Sepsis; Thromboplastin

Septic infections dysregulate hemostatic pathways, prompting coagulopathy. Nevertheless, anticoagulant therapies typically fail to protect humans from septic pathology. The data reported in this work may help to explain this discrepancy by demonstrating critical protective roles for coagulation leading to fibrin deposition during host defense against the Gram-negative bacterium Yersinia enterocolitica. After i.p. inoculation with Y. enterocolitica, fibrinogen-deficient mice display impaired cytokine and chemokine production in the peritoneal cavity and suppressed neutrophil recruitment. Moreover, both gene-targeted fibrinogen-deficient mice and wild-type mice treated with the anticoagulant coumadin display increased hepatic bacterial burden and mortality following either i.p. or i.v. inoculation with Y. enterocolitica. Mice with low tissue factor activity succumb to yersiniosis with a phenotype similar to fibrin(ogen)-deficient mice, whereas factor XI-deficient mice show wild-type levels of resistance. Mice deficient in plasminogen activator inhibitor-1 or thrombin-activatable fibrinolysis inhibitor display modest phenotypes, but mice deficient in both plasminogen activator inhibitor-1 and thrombin-activatable fibrinolysis inhibitor succumb to yersiniosis with a phenotype resembling fibrin(ogen)-deficient mice. These findings demonstrate critical protective roles for the tissue factor-dependent extrinsic coagulation pathway during host defense against bacteria and caution that therapeutics targeting major thrombin-generating or antifibrinolytic pathways may disrupt fibrin-mediated host defense during Gram-negative sepsis.

Topics: Animals; Carboxypeptidase B2; Factor XI; Fibrin; Humans; Liver; Mice; Mice, Knockout; Sepsis; Serpin E2; Thromboplastin; Yersinia enterocolitica; Yersinia Infections

A high mortality occurs in dogs with idiopathic immune-mediated haemolytic anaemia (IMHA) during the first 2 weeks after the diagnosis. The aim of this study was to investigate the inflammatory response and coagulation abnormalities in dogs with IMHA in relation to the prognosis and to establish the contribution of whole blood tissue factor (TF) and IL-8 gene expressions. Gene expressions in dogs with IMHA were compared to healthy dogs, dogs with DIC, dogs with sepsis, and in two groups of dogs that underwent intensive care treatment but had no evidence for either DIC or sepsis. The whole blood TF and IL-8 expressions were up regulated in all non-IMHA groups. Similarly, the TF expression in IMHA dogs was high, but the intravascular IL-8 expression was not increased. The dogs with IMHA had a pronounced inflammatory response that included a high WBC, left shift and monocytosis in comparison to the other disease groups. Coagulation factor activities in IMHA dogs were decreased fitting consumptive coagulopathy and the acute phase proteins FVIII and fibrinogen were increased. The platelet parameters suggested platelet activation and high platelet turnover in IMHA dogs. The model that best explained mortality contained monocytosis, increased activated partial thromboplastin time and elevated creatinine. Whole blood TF gene expression is up regulated and may contribute to consumptive coagulopathy in dogs with IMHA. Increased TF expression by activated platelets is an alternative explanation and should be investigated.

Topics: Anemia, Hemolytic, Autoimmune; Animals; Blood Coagulation; Disseminated Intravascular Coagulation; Dog Diseases; Dogs; Female; Inflammation; Interleukin-8; Male; Prognosis; Real-Time Polymerase Chain Reaction; Sepsis; Thromboplastin

To evaluate citrated recalcified thromboelastography (TEG) in healthy newborn foals, and to determine intra-assay, inter-individual and intra-individual (at 12 h, 24 h and 7 days after birth) variations. Additionally, to compare TEG variables, haematological values and conventional coagulation profiles from healthy, sick non-septic, and septic foals.. Prospective study.. The study group comprised 18 healthy, 15 sick non-septic and 17 septic foals. Two citrated (3.2%; 1 : 9 anticoagulant : blood ratio) blood samples were submitted for haemostatic evaluation using a TEG analyser and conventional coagulation profile. TEG values (R time (R), K time (K), angle (α), maximum amplitude (MA) and G value (G)), complete blood count (CBC) and conventional coagulation profile (prothrombin time (PT), activated partial thromboplastin time (aPTT), fibrinogen concentration (Fib) and antithrombin (AT)) were evaluated. Signalment, presenting complaint, sepsis scores, blood culture results and outcome were taken from the medical records of the sick foals.. Mean values ± SD for TEG variables in healthy neonatal foals were: R = 11.82 ± 5.35 min, K = 3.06 ± 1.34 min, α= 51.19 ± 12.66 degrees, MA = 55.06 ± 6.67 mm and G = 6361 ± 1700 dyn/cm(2) . Mean coefficients of variation for intra-assay/inter-individual/intra-individual in healthy foals were: R = 3.5/45.2/43.1%; K = 5.3/58.7/28.7%; α= 1.5/24.7/11.9%; MA = 0.3/12.1/6.1%; G = 1.6/26.7/14.7%. Septic foals had significantly greater α, MA and G values than sick non-septic foals, and significantly greater MA and G than healthy foals, changes that are consistent with hypercoagulability. Weak correlations were detected between TEG variables and haematological or haemostatic values.. TEG could be used to provide additional information about the haemostatic system in equine neonates.

Topics: Animals; Animals, Newborn; Blood Coagulation; Citrates; Female; Hemostasis; Horse Diseases; Horses; Male; Prospective Studies; Reproducibility of Results; Sensitivity and Specificity; Sepsis; Thrombelastography; Thromboplastin

Acadesine, an adenosine-regulating agent and activator of AMP-activated protein kinase, has been shown to possess antiinflammatory activity. This study investigated whether and how acadesine inhibits tissue factor (TF) expression and thrombus formation.. Human umbilical vein endothelial cells and human peripheral blood monocytes were stimulated with lipopolysaccharide to induce TF expression. Pretreatment with acadesine dramatically suppressed the clotting activity and expression of TF (protein and mRNA). These inhibitory effects of acadesine were unchanged for endothelial cells treated with ZM241385 (a specific adenosine A(2A) receptor antagonist) or AMP-activated protein kinase inhibitor compound C, and in macrophages lacking adenosine A(2A) receptor or alpha1-AMP-activated protein kinase. In endothelial cells and macrophages, acadesine activated the phosphoinositide 3-kinase/Akt signaling pathway, reduced the activity of mitogen-activated protein kinases, and consequently suppressed TF expression by inhibiting the activator protein-1 and NF-kappaB pathways. In mice, acadesine suppressed lipopolysaccharide-mediated increases in blood coagulation, decreased TF expression in atherosclerotic lesions, and reduced deep vein thrombus formation.. Acadesine inhibits TF expression and thrombus formation by activating the phosphoinositide 3-kinase/Akt pathway. This novel finding implicates acadesine as a potentially useful treatment for many disorders associated with thrombotic pathology, such as angina pain, deep vein thrombosis, and sepsis.

Topics: Adenosine A2 Receptor Antagonists; Aminoimidazole Carboxamide; AMP-Activated Protein Kinases; Animals; Apolipoproteins E; Atherosclerosis; Blood Coagulation; Cells, Cultured; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelial Cells; Enzyme Activation; Fibrinolytic Agents; Humans; Lipopolysaccharides; Macrophages; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Monocytes; NF-kappa B; Phosphatidylinositol 3-Kinases; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-akt; Pyrazoles; Pyrimidines; Receptor, Adenosine A2A; Ribonucleosides; RNA, Messenger; Sepsis; Signal Transduction; Thromboplastin; Transcription Factor AP-1; Triazines; Triazoles; Up-Regulation; Venous Thrombosis

Sepsis is a generalized inflammatory disease, caused by the hyperinflammatory response of the host, rather than by invading organisms. Endothelial cells play a crucial role in the pathogenesis of sepsis. In this study, we investigated the effects of interleukin-8 (IL-8), a known neutrophil chemoattractant, on lipopolysaccharide (LPS) -induced reactive oxygen species (ROS) production by endothelial cells, and its significance in the pathogenesis of LPS-mediated sepsis. The results revealed that IL-8 directly induced ROS production in human umbilical vein endothelial cells (HUVECs), and also mediated LPS-induced ROS production by HUVECs. Stimulation of HUVECs by LPS strongly enhanced tissue factor expression, a hallmark of severe sepsis, which was suppressed by IL-8 knockdown. We further discovered that NADPH oxidase (Nox) 1 expression in LPS-stimulated HUVECs was markedly repressed by IL-8 knockdown, and Nox1 knockdown reduced tissue factor expression, suggesting that the LPS/IL-8 signalling in endothelial cells was predominantly mediated by Nox1. In conclusion, LPS stimulation of endothelial cells causes activation of the IL-8-Nox1 axis, enhances the production of ROS, and ultimately contributes to the progression of severe sepsis.

Topics: Cell Line; Endothelium, Vascular; Enzyme Activation; Humans; Interleukin-8; Lipopolysaccharides; NADPH Oxidases; Reactive Oxygen Species; RNA, Small Interfering; Sepsis; Signal Transduction; Thromboplastin

Topics: Animals; Disease Models, Animal; Drugs, Chinese Herbal; Male; Random Allocation; Rats; Rats, Wistar; Sepsis; Thromboplastin

To explore the inhibitive effects of exogenous carbon monoxide-releasing molecules 2 (CORM-2) on expression of tissue factor (TF) in sepsis.. Human umbilical vein endothelial cells (HUVEC) were cultured with trypsin digestion method, which were divided into NC group (with normal treatment), LPS group (with culture of 10 microg/mL LPS), LD group (with 10 microg/mL LPS and DMSO in co-culture), LC1 group (with 10 microg/mL LPS and 10 micromol/L CORM-2 in co-culture), LC2 group (with 10 microg/mL LPS and 50 micromol/L CORM-2 in co-culture), LC3 group (with 10 microg/mL LPS and 100 micromol/L CORM-2 in co-culture). After culture for 4 hours, TF activity, TF protein expression, nuclear factor-kappaB (NF-kappaB) activity were examined. Forty-five C57 BL/6 male mice were randomly divided into NC (without treatment, n = 5), CLP (n = 5) and CLP + CORM-2 (with treatment of 8 mg/kg CROM-2 after CLP, n = 5) groups. The serum samples in CLP, CLP + CORM-2 groups were collected at 2, 6, 12 and 24 post operation hour ( POH, 5 mice at each time point) for determination of TF and TFPI levels,which were also examined in NC group.. Compared with those of NC group, TF activity increased (P < 0.01) , TF protein expression and NF-KB activity also increased in LPS group. Compared with those of LPS group, above indices were decreased in LC1, LC2, LC3 groups. The serum level of TF in CLP group at 6 POH was higher than that of NC group (80.0 +/- 11.9 pg/mL vs 58.4 +/- 6.9 pg/mL, P < 0.05), peaked at 12 POH, and still higher than that of NC group at 24 POH, while the serum level of TFPI showed no obvious difference in NC and CLP groups. Compared with that of NC group, TFPI levels obviously increased in CLP + CORM-2 group at 6, 12 POH (23.7 +/-3.5 ng/mL, 24.4 +/- 5.0 ng/mL vs 12.4 +/- 2.8 ng/mL, P < 0.05).. Exogenous CORM can obviously inhibit TF and NF-KB activity,decrease TF protein expression. Meanwhile, it can also decrease serum level of TF, and increase serum level of TFPI, preventing activation of procoagulant system, balancing procoagulant and anticoagulant system in sepsis.

Topics: Animals; Cells, Cultured; Humans; Lipoproteins; Male; Mice; Mice, Inbred C57BL; NF-kappa B; Organometallic Compounds; Sepsis; Thromboplastin

The protective effect of recombinant activated protein C therapy in patients with severe sepsis likely reflects the ability of recombinant activated protein C to modulate multiple pathways implicated in sepsis pathophysiology. In this study, we examined the effects of recombinant activated protein C on the anti-inflammatory cytokine IL-10 and on the procoagulant molecule tissue factor (TF) in LPS-challenged blood monocytes. Treatment of LPS-stimulated monocytes with recombinant activated protein C resulted in an up-regulation of IL-10 protein production and mRNA synthesis. The up-regulation of IL-10 required the serine protease activity of recombinant activated protein C and was dependent on protease-activated receptor-1, but was independent of the endothelial protein C receptor. At the intracellular level, p38 MAPK activation was required for recombinant activated protein C-mediated up-regulation of IL-10. We further observed that incubation of LPS-stimulated monocytes with recombinant activated protein C down-regulated TF Ag and activity levels. This anticoagulant effect of recombinant activated protein C was dependent on IL-10 since neutralization of endogenously produced IL-10 abrogated the effect. In patients with severe sepsis, plasma IL-10 levels were markedly higher in those treated with recombinant activated protein C than in those who did not receive recombinant activated protein C. This study reveals novel regulatory functions of recombinant activated protein C, specifically the up-regulation of IL-10 and the inhibition of TF activity in monocytes. Our data further suggest that these activities of recombinant activated protein C are directly linked: the recombinant activated protein C-mediated up-regulation of IL-10 reduces TF in circulating monocytes.

Topics: Adult; Aged; Aged, 80 and over; Antigens; Antigens, CD; Cells, Cultured; Endothelial Protein C Receptor; Enzyme Activation; Female; Humans; Interleukin-10; Lipopolysaccharides; Male; MAP Kinase Signaling System; Middle Aged; Monocytes; p38 Mitogen-Activated Protein Kinases; Protein C; Receptors, Cell Surface; Recombinant Proteins; RNA, Messenger; Sepsis; Thromboplastin; Up-Regulation

Sepsis is associated with enhanced generation of reactive oxygen species, which leads to multiple organ dysfunctions. Based on the potent antioxidant effects of silymarin, we investigated the putative protective role of silymarin against sepsis-induced oxidative damage in lung and brain tissues.. Sepsis was induced by cecal ligation and perforation (CLP). Sham and CLP groups received either vehicle or silymarin (50 mg/kg, p.o.) or 150 mg/kg i.p. N-acetylcysteine (NAC) for 10 days prior and immediately after the operation. Six hours after the surgery, rats were decapitated and blood was collected for the measurement of proinflammatory cytokines (tumor necrosis factor-alpha, interleukin-1 beta [IL-1 beta], and IL-6) levels, lactate dehydrogenase activity, and total antioxidant capacity. Lung and brain samples were taken for the measurement of malondialdehyde and glutathione levels, myeloperoxidase activity, thromboplastic activity, and also for histological assessment. Formation of reactive oxygen species in tissue samples was monitored by using chemiluminescence technique with luminol and lusigenin probe.. Sepsis increased serum TNF-alpha, IL-1 beta, IL-6 levels, and lactate dehydrogenase activity and decreased total antioxidant capacity. On the other hand, tissue glutathione levels were decreased while malondialdehyde levels and myeloperoxidase activity were increased in both the lung and the brain tissues due to CLP. Furthermore, luminol and lucigenin chemiluminescence were significantly increased in the CLP group, indicating the presence of the oxidative damage. Silymarine and NAC treatment reversed these biochemical parameters and preserved tissue morphology as evidenced by histological evaluation.. Silymarin, like NAC, reduced sepsis-induced remote organ injury, at least in part, through its ability to balance oxidant-antioxidant status, to inhibit neutrophil infiltration, and to regulate the release of inflammatory mediators.

Topics: Acridines; Animals; Anti-Inflammatory Agents; Antioxidants; Brain; Brain Diseases; Female; Glutathione; L-Lactate Dehydrogenase; Luminescent Agents; Luminol; Male; Peroxidase; Pulmonary Alveoli; Rats; Rats, Wistar; Respiratory Distress Syndrome; Sepsis; Silybum marianum; Silymarin; Survival Rate; Thromboplastin; Tumor Necrosis Factor-alpha

Severe sepsis is associated with an exacerbated procoagulant state with protein C (PC) system impairment. In contrast, the inflammatory and coagulation status of nonseptic patients with organ failure (OF) is less documented.. To compare coagulation activation, focusing on the PC system, and inflammatory status in septic and nonseptic patients with OF.. Thirty patients with severe sepsis and 30 nonseptic patients were recruited at the onset of OF and compared with 30 matched healthy subjects. We performed an extensive analysis of the PC pathway, including plasma protein measurements and quantification of leukocyte expression of PC system receptors. In addition, we analyzed the inflammatory status, based on inflammation-related gene leukocyte expression.. We observed coagulation activation, reflected by a similar increase in tissue factor mRNA expression, in the two patient groups when compared with the healthy subjects. Soluble thrombomodulin levels were higher in septic patients than in healthy control subjects, whereas PC, protein S, and soluble endothelial cell PC receptor levels were lower. Similar results were obtained in nonseptic patients with OF. Monocyte thrombomodulin overexpression, together with increased circulating levels of activated PC, suggests that the capacity for PC activation is at least partly preserved in both settings. No difference in the inflammatory profile was found between septic and nonseptic patients.. The pathogenesis of OF in critical care patients is characterized by an overwhelming systemic inflammatory response and by exacerbated coagulation activation, independently of whether or not infection is the triggering event. Clinical trial registered with www.clinicaltrials.gov (NCT 00361725).

Topics: Adult; Aged; Antigens, CD; Blood Coagulation; Case-Control Studies; Endothelial Protein C Receptor; Female; Humans; Male; Middle Aged; Multiple Organ Failure; Protein C; Protein S; Receptors, Cell Surface; RNA, Messenger; Sepsis; Thrombomodulin; Thromboplastin

Tissue factor (TF) is a transmembrane glycoprotein that binds its zymogen cofactor, Factor VIIa (FVIIa) on the cell surface. Together (TF/FVIIa) they activate Factor X (FX) and Factor IX (FIX) and start the extrinsic pathway of blood coagulation. As such, the TF/FVIIa complex plays an important role in normal physiology as well as in thrombotic diseases such as unstable angina (UA), disseminated intravascular coagulation (DIC), and deep vein thrombosis (DVT). In addition to its function as an initiator of coagulation, TF/FVIIa plays an important role in inflammation. Expression of TF on the cell surface and its appearance as a soluble molecule are characteristic features of acute and chronic inflammation in conditions such as sepsis and atherosclerosis. Here we demonstrate that BCX-3607, a small molecule potent inhibitor of TF/FVIIa, reduces thrombus weight in an animal model of DVT. BCX-3607 also decreases the level of interleukin-6 (IL-6) in a LPS-stimulated mouse model of endotoxemia. Additionally, in vitro studies indicate that BCX-3607 blocks the generation of TF/FVIIa-induced IL-8 mRNA in human keratinocytes and reduces the TF/FVIIa-mediated generation of IL-6 and IL-8 in human umbilical vein endothelial cells (HUVEC). Therefore, BCX-3607 might block the TF/FVIIa-mediated coagulation and inflammation associated with pathological conditions.

Topics: Animals; Anti-Inflammatory Agents; Atherosclerosis; Blotting, Northern; Cells, Cultured; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelium, Vascular; Endotoxemia; Factor VIIa; Fibrinolytic Agents; Humans; Inflammation; Interleukin-6; Interleukin-8; Keratinocytes; Lipopolysaccharides; Male; Mice; Models, Biological; Models, Chemical; Prothrombin Time; Pyridines; Rats; Rats, Sprague-Dawley; RNA, Messenger; Sepsis; Thromboplastin; Time Factors

Topics: Animals; Atherosclerosis; Humans; Neoplasms; Sepsis; Thromboplastin; Wounds and Injuries

Tissue factor (TF) is a transmembrane protein, which is essential for initiation of the coagulation cascade. TF has been reported to play an important role in the progression of endotoxin (lipopolysaccharide, LPS)-mediated endotoxemia, being induced in numerous tissues, such as kidney, spleen and lung. We developed and validated a rabbit anti-murine TF peptide antiserum to localize TF protein in a murine sepsis model. TF protein distribution was compared to localization of TF mRNA and fibrin deposits, the ultimate resultant of procoagulant TF activity. Evident LPS mediated TF mRNA induction was observed in the tubular area at the cortico-medullar junction in the kidney, and TF activity was increased after 6 hours of endotoxemia. In the spleen, however, TF mRNA was induced in the interfollicular region upon LPS injection, corresponding to increased TF protein in the same area. The clusters of TF-protein positive cells in the spleen are predominantly granulocytes, but no TF mRNA expression was observed within these cells. Based on these observations and the presence of TF-protein positive granulocytes after splenectomy, we hypothesize that granulocytes take-up TF for transport to other locations in order to initiate fibrin formation or to induce pro-inflammatory gene expression upon interaction with factor VIIa.

Topics: Animals; Endotoxemia; Female; Fibrin; Gene Expression Regulation; Granulocytes; Immune Sera; Inflammation; Kidney; Lipopolysaccharides; Mice; Protein Transport; Rabbits; RNA, Messenger; Sepsis; Spleen; Thromboplastin; Tissue Distribution

Substantial progress has been made in understanding the contribution of alterations in coagulation and fibrinolysis to the pathogenesis of acute lung injury (ALI). Findings from mouse, rat, baboon, and human studies indicate that alterations in coagulation and fibrinolysis may be of major pathogenetic importance in ALI and other inflammatory conditions in the lung including pneumonia, sepsis, and ventilator-induced lung injury. Therapies targeted at both activation of coagulation through the extrinsic coagulation cascade and modulation of coagulation through the protein C system have the potential to favorably impact clinical ALI/acute respiratory distress syndrome.

Topics: Animals; Blood Coagulation; Disease Models, Animal; Fibrinolysis; Humans; Models, Immunological; Papio; Pneumonia; Protein C; Receptors, Proteinase-Activated; Respiration, Artificial; Respiratory Distress Syndrome; Sepsis; Thrombomodulin; Thromboplastin

Pathologic disseminated intravascular coagulation (PDIC) is a serious complication in sepsis. In an in-vitro system consisting of incubation of fresh citrated blood with lipopolysaccharides (LPS) or glucans and subsequent plasma recalcification plasmatic thrombin was quantified. Five hundred microliters of freshly drawn citrated blood of healthy donors were incubated with up to 800 ng/mL LPS (Escherichia coli) or up to 80 microg/mL Zymosan A (ZyA; Candida albicans) for 30 minutes at room temperature (RT). The samples were centrifuged, and 30 microL plasma were recalcified with 1 volume or less of CaCl(2) (25 micromoles Ca(2+)/mL plasma). After 0 to 12 minutes (37 degrees C), 20 microL 2.5 M arginine, pH 8.6, were added. Thirty microliters 0.9 mM HD-CHG-Ala-Arg-pNA in 2.3 M arginine were added, and the absorbance increase at 405 nm was determined. Fifty microliters plasma were also incubated with 5 microL 250 mM CaCl2 for 5, 10, or 15 minutes (37 degrees C). Fifty microliters 2.5 M arginine stops coagulation, and 50 microL 0.77 mM HD-CHG-Ala-Arg-pNA in 2.3 M arginine starts the thrombin detection. The standard was 1 IU/mL thrombin in 7% human albumin instead of plasma. Arginine was also added in the endotoxin exposure time (EET) or in the plasma coagulation reaction time (CRT). Tissue factor (TF)-antigen and soluble CD14 were determined. LPS at blood concentrations greater than 10 ng/mL or ZyA at greater than 1 microg/mL severalfold enhance thrombin generation, when the respective plasmas are recalcified. After 30 minutes EET at RT, the thrombin activity at 12 minutes CRT generated by the addition of 200 ng/mL LPS or 20 microg/mL ZyA is approximately 200 mIU/mL compared to approximately 20 mIU/mL without addition of endotoxin, or compared to about 7 mIU/mL thrombin at 0 minutes CRT. Arginine added to blood or to plasma inhibits thrombin generation; the inhibitory concentration 50% (IC 50) is approximately 15 mM plasma concentration. Endotoxin incubation of blood increases neither TF nor sCD14. This assay allows the study of the hemostasis alteration in PDIC, particularly in PDIC by sepsis. The thrombin generated by blood plus endotoxin incubation and plasma recalcification suggests that the contact phase of coagulation; e.g., triggered by cell components of (phospholipase-) lysed cells such as monocyte or endothelium DNA or phospholipid-vesicles (microparticles), is of primary pathologic importance in sepsis-PDIC. Arginine at plasma concentrations of 10 to

Topics: Arginine; Blood; Blood Coagulation Tests; Cells, Cultured; Disseminated Intravascular Coagulation; Endotoxins; Humans; Lipopolysaccharide Receptors; Lipopolysaccharides; Models, Cardiovascular; Sepsis; Thrombin; Thromboplastin; Zymosan

Inhibition of blood coagulation appears to be an important therapeutic strategy to improve the outcome in sepsis. However, the beneficial effect of anticoagulant treatment in sepsis is solely based on experimental data using inhibitors of the extrinsic coagulant pathway. The role of the intrinsic pathway of coagulation in the pathogenesis of sepsis has not been explored yet.. In the current study, we contribute to determine the role of factor (F)VIII, the key player of the intrinsic coagulant pathway, on host defense against peritonitis.. Hemizygous FVIII-deficient mice and their wild-type littermates were challenged with 1 x 10(4) bacteria in a septic peritonitis model.. The intraperitoneal injection of Escherichia coli led to growth and dissemination of bacteria and provoked an inflammatory response as evident from elevated cytokine levels, increased cell influx into tissues, liver necrosis, and endothelialitis resulting in mortality. The FVIII-deficient genotype slightly reduced bacterial outgrowth but had no effect on markers of inflammation and/or survival. In addition, FVIII-deficient mice showed profound activation of coagulation, thereby improving the hemophilic phenotype of FVIII-deficient mice.. FVIII deficiency slightly modifies host defense in septic peritonitis in mice, but does not influence the final outcome of peritonitis. Therefore, we question the importance of the intrinsic coagulant pathway during sepsis.

Topics: Animals; Blood Coagulation; Colony Count, Microbial; Escherichia coli; Factor VIII; Hemophilia A; Immunity; Inflammation; Lipoproteins; Mice; Peritonitis; RNA, Messenger; Sepsis; Survival Rate; Thromboplastin

The biphasic waveform that can predict for disseminated intravascular coagulation (DIC) is due to the formation of a calcium-dependent complex between C reactive protein (CRP) and very low density lipoprotein (VLDL). As thrombin generation is pivotal to DIC, this aspect has been specifically investigated and the VLDL component has been found to increase prothrombinase activity via both quantitative and qualitative changes. The specific prothrombinase activity of VLDL from patients manifesting the biphasic waveform was 2.5 times that of normal individuals or critically ill patients without the biphasic waveform. This activity was due to an increase in anionic phospholipid surfaces that could be inhibited with excess annexin V and which was dependent on structurally intact apolipoprotein B. The qualitative change appeared to be due to a deficiency of phosphatidylethanolamine in VLDL from patients with the biphasic waveform. The functional consequence of this enhanced prothrombinase activity was an increased procoagulant effect in plasma. Using a modified activated partial thromboplastin time assay, the mean normal clot time decreased significantly when VLDL from patients with biphasic waveforms was substituted. These results indicate that VLDL derived from patients with the biphasic waveform can enhance thrombin procoagulant activity. As the CRP-VLDL complex exists in vivo, it could have a pathogenic role in disseminating the process of intravascular coagulation.

Topics: Adsorption; Annexin A5; Apolipoproteins B; Blood Coagulation Tests; C-Reactive Protein; Cell Separation; Cerebrosides; Chromatography, Thin Layer; Coagulants; Disseminated Intravascular Coagulation; Dose-Response Relationship, Drug; Flow Cytometry; Humans; Immunosorbent Techniques; Lipoproteins; Lipoproteins, VLDL; Partial Thromboplastin Time; Phosphatidylethanolamines; Protein Binding; Sepsis; Thrombin; Thromboplastin; Time Factors; Triglycerides

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

Blockade of tissue factor before lethal sepsis prevents acute lung injury and renal failure in baboons, indicating that activation of coagulation by tissue factor is an early event in the pathogenesis of acute lung injury and organ dysfunction. We hypothesized that blockade of tissue factor would also attenuate these injuries in established sepsis by prevention of further fibrin deposition and inflammation. Twelve male baboons received heat-killed Escherichia coli intravenously followed 12 hours later by live E. coli infusion. Six animals were treated 2 hours after the live bacteria with site-inactivated Factor VIIa, a competitive tissue factor inhibitor, and six animals were vehicle-treated sepsis control subjects. Animals were ventilated and monitored for 48 hours. Physiologic and hematologic parameters were measured every 6 hours, and pathologic evaluation was performed after 48 hours. Animals treated with site inactivated Factor VIIa had less severe lung injury, with preserved gas exchange, better lung compliance and histology scores, and decreased lung wet/dry weight. In treated animals, urine output was higher, metabolic acidosis was attenuated, and renal tubular architecture was protected. Coagulopathy was attenuated, and plasma interleukin-6, interleukin-8, and soluble tumor necrosis factor receptor-1 levels were significantly lower in the treated animals. These results show that blockade of coagulation attenuates acute lung and renal injury in established Gram-negative sepsis accompanied by antiinflammatory effects of therapy.

Topics: Acute Kidney Injury; Animals; Antigens, CD; Cytokines; Disease Models, Animal; Drug Monitoring; Escherichia coli Infections; Factor VIIa; Hemodynamics; Inflammation; Interleukin-6; Interleukin-8; Lung Compliance; Male; Papio; Pulmonary Gas Exchange; Receptors, Tumor Necrosis Factor; Receptors, Tumor Necrosis Factor, Type I; Respiratory Distress Syndrome; Sepsis; Severity of Illness Index; Thromboplastin

To determine the precise relationship among tissue factor, tissue factor pathway inhibitor (TFPI), and neutrophil elastase in sepsis, as well as to test the hypothesis that low TFPI concentrations are not sufficient to prevent tissue factor-dependent intravascular coagulation, leading to multiple organ dysfunction syndrome and death.. Prospective, cohort study.. General intensive care unit of tertiary care emergency department.. Thirty-one consecutive patients with sepsis, classified as 15 survivors and 16 nonsurvivors. Ten normal, healthy volunteers served as controls.. None.. Tissue factor antigen concentration (tissue factor), TFPI, neutrophil elastase, and global variables of coagulation and fibrinolysis were measured on the day of diagnosis of sepsis, severe sepsis, and septic shock and days on 1-4 after diagnosis. The number of systemic inflammatory response syndrome criteria that patients met and the disseminated intravascular coagulation score were determined simultaneously. The results of these measurements were compared between the survivors and the nonsurvivors. In the nonsurvivors, significantly higher concentrations of tissue factor and neutrophil elastase were found compared with the survivors and control subjects. However, the TFPI values showed no difference between the two groups. No correlation was found between the peak concentrations of tissue factor and TFPI. Disseminated intravascular coagulation scores and numbers of the SIRS criteria met by the survivors significantly decreased from day 0 to day 4, but those of the nonsurvivors did not improve during the study period. The nonsurvivors showed thrombocytopenia and higher numbers of dysfunctioning organs than did the survivors.. We systematically elucidated the relationship between tissue factor and TFPI in patients with sepsis, severe sepsis, and septic shock. Activation of tissue factor-dependent coagulation pathway not adequately balanced by TFPI has important roles in sustaining DIC and systemic inflammatory response syndrome, and it contributes to multiple organ dysfunction syndrome and death. High concentrations of neutrophil elastase released from activated neutrophils may explain, in part, the imbalance of tissue factor and TFPI in sepsis.

Topics: Adult; Anticoagulants; Cohort Studies; Disseminated Intravascular Coagulation; Female; Humans; Incidence; Japan; Leukocyte Elastase; Lipoproteins; Male; Middle Aged; Multiple Organ Failure; Platelet Count; Prognosis; Prospective Studies; Respiratory Distress Syndrome; Sepsis; Serine Proteinase Inhibitors; Survival Analysis; Thromboplastin; Treatment Outcome

Topics: Anticoagulants; Humans; Lipoproteins; Sepsis; Thromboplastin

Soluble beta-1,3-glucan has been demonstrated to protect against infection and shock in rats and mice, and clinical studies suggest that administration of soluble glucans to trauma/surgical patients decreases septic complications and improves survival. However, little is known about the precise mechanisms by which glucans influence the state of activation of blood cells, which are responsible for the fulminant cytokine production and the activation of the coagulation system observed in serious gram-negative infection. We studied therefore the effect of an underivatized, soluble yeast beta-1,3-glucan and lipopolysaccharide (LPS), either alone or in combination, on tumor necrosis factor-alpha (TNFalpha), interleukin-6 (IL-6), IL-8 and IL-10 secretion and monocyte tissue factor (TF) expression in human whole blood. As expected, LPS induced the secretion of substantial amounts of all measured parameters, whereas only minor amounts of TNFalpha, IL-6, and IL-10 were induced by beta-glucan itself. However, beta-glucan itself induced the production of significant amounts of IL-8 and TF. Soluble beta-1,3-glucan had a strong synergistic effect on the LPS-induced secretion of IL-8, IL-10, and on monocyte TF activity, but not on TNFalpha and 1L-6 production. On the other hand, soluble beta-glucan strongly primed LPS stimulation of all parameters, including TNFalpha and IL-6. beta-Glucan also induced detectable neutrophil degranulation within 15 min, whereas a response to LPS was first detected after 90 min. In conclusion, soluble beta-1,3-glucan upregulated leukocyte activity, both on its own and in concert with LPS.

Topics: beta-Glucans; Biomarkers; Blood Coagulation; Cytokines; Glucans; Humans; In Vitro Techniques; Indicators and Reagents; Lipopolysaccharides; Platelet Activation; Sepsis; Thromboplastin

Cytokines increase endothelial tissue factor (TF) and tissue plasminogen activator inhibitor type-1 (PAI-1) expression in vitro. Tissue factor interacts with factor VII to facilitate thrombosis and PAI-1 inhibits fibrinolysis by endogenous plasminogen activators. Because cytokine release is increased in children with sepsis-induced multiple organ failure (MOF), we hypothesized a cytokine associated increase in circulating TF and PAI-1 antigen release, and systemic activity in these patients.. One hundred and seven consecutive children, who met the criteria for sepsis, and 10 critically ill children without sepsis, were enrolled in the study. Plasma TF and PAI-1 antigen and activity levels, Interleukin-6 antigen levels (IL-6), nitrite + nitrate levels (marker of nitric oxide production) and number of organs failing were measured on days 1-3 of sepsis.. Increased TF and PAI-1 antigen, and PAI-1 activity levels were associated with increasing IL-6 and nitrite + nitrate levels (p <0.05), the development of MOF (p <0.05), and mortality (p <0.05). Increased systemic PAI-1 activity was associated with cardiovascular, renal. and hepatic failure (p <0.05). Increased systemic TF activity was associated with the development of coagulopathy (p <0.05) and tended to be associated with mortality (p = 0.06, power .77). A shift to an anti-fibrinolytic endothelium phenotype characterizes children who develop sepsis-induced MOF and mortality. Children with coagulopathy have a shift to a pro-coagulant phenotype. These findings support potential therapeutic roles for PAI-1 and TF pathway inhibitors in reversal of this devastating pathophysiologic process.

Topics: Adolescent; Child; Child, Preschool; Cytokines; Endothelium, Vascular; Female; Fibrinolysis; Humans; Infant; Infant, Newborn; Interleukin-6; Male; Multiple Organ Failure; Nitrates; Nitrites; Pennsylvania; Plasminogen Activator Inhibitor 1; Prospective Studies; Sepsis; Thrombophilia; Thromboplastin

To investigate the effects of antithrombin on lipopolysaccharide (LPS)-induced tissue factor and interleukin-6 (IL-6) production in three different in vitro cellular systems: whole blood, human umbilical vein endothelial cells, and mononuclear cells.. Laboratory in vitro study of the effects of antithrombin on procoagulant activity and cytokine release by LPS-stimulated endothelial and peripheral blood cells.. In vitro whole blood, isolated human umbilical vein endothelial cell, and mononuclear cell cultures.. Addition of antithrombin to LPS-treated whole blood, human umbilical vein endothelial cells, and mononuclear cells.. Citrated whole blood, isolated human umbilical vein endothelial cells, or mononuclear cells were stimulated with LPS for 4-6 hrs in the presence or absence of antithrombin. Tissue factor activity was estimated by a tissue factor-dependent clotting or chromogenic assay and IL-6 was measured by specific ELISA. Antithrombin was found to inhibit tissue factor and IL-6 production in all three systems in a dose-dependent manner (0-40 IU/mL). Flow-through fractions of immunoadsorbed antithrombin concentrate were found to be ineffective. Five different batches of the same antithrombin concentrate were tested and the inhibitory activity was found to be consistent throughout all batches. Up to 40 microM of recombinant hirudin, a specific thrombin inhibitor, did not inhibit the production of tissue factor or IL-6 in either of the three cell systems, suggesting that the observed inhibition by antithrombin was not due solely to its ability to inhibit thrombin.. Apart from the inhibition of thrombin and other activated clotting factors, antithrombin may also down-regulate the cellular expression of proinflammatory cytokines. Consequently, antithrombin concentrates may have value in the treatment of sepsis-induced disseminated intravascular coagulation.

Topics: Antithrombins; Blood; Cells, Cultured; Endothelium, Vascular; Hirudins; Humans; In Vitro Techniques; Interleukin-6; Leukocytes, Mononuclear; Lipopolysaccharides; Sepsis; Thromboplastin; Umbilical Veins

1Alpha,25-dihydroxyvitamin D3 (active form of vitamin D3; vitamin D3) has been reported to induce the upregulation of thrombomodulin and downregulation of tissue factor (TF) on monocytes. The possibility exists that vitamin D3 prevents the development of disseminated intravascular coagulation (DIC). In particular, monocyte TF production plays an important role in the pathophysiology of DIC in septic patients. We have attempted to determine whether vitamin D3 is effective against DIC in a rat model induced by lipopolysaccharides (LPS) (30 mg/kg, 4 h) or TF (3.75 U/kg, 4 h) using selective hemostatic parameters, markers of organ dysfunction and pathological findings (assessment of glomelular fibrin deposition). Vitamin D3 was administered orally each day at a dose of 2.0 mg/kg/day for 3 days, or low molecular weight heparin (LMWH 200 u/kg; i.v.) was given 10 min before the injection of TF or LPS in each treatment group. Vitamin D3 was effective against DIC in the rat model induced by LPS only, whereas LMWH was effective against DIC in both rat models induced by either TF or LPS. The anti-DIC effect of vitamin D3 was equal to (or more potent than) that of LMWH. The results suggested that vitamin D3 was useful for the treatment of LPS-induced DIC, and that the assessment of a drug's efficacy should be done carefully given the markedly different results obtained according to the agents used to induce DIC.

Topics: Administration, Oral; Animals; Anticoagulants; Cholecalciferol; Coagulants; Disease Models, Animal; Disseminated Intravascular Coagulation; Fibrin; Heparin; Kidney Diseases; Kidney Glomerulus; Lipopolysaccharides; Male; Rats; Rats, Wistar; Sepsis; Thromboplastin; Thrombosis

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

Although tissue factor (TF) is involved in hemostasis, thrombogenesis, inflammation, and cellular immune response, its source in sepsis remains controversial. Recently, we found that, in addition to monocytes and endothelial cells, neutrophils may express TF in a rabbit model. The purpose of this study was to determine whether neutrophils could be a source of TF in a monkey model of sepsis.. TF messenger RNA (mRNA) and protein in neutrophils were assayed by in situ hybridization and immunohistochemistry in tissues obtained from monkeys after injection of lipopolysaccharide (LPS) (n = 3) and after injection of saline as a control (n = 2). Coagulation parameters were measured before and at 1.5 and 3 hours after injections.. In LPS-treated monkeys, TF mRNA and protein were induced not only in monocytes and endothelial cells, but also in neutrophils accumulating in the liver 3 hours after LPS injection. Thrombin-antithrombin III complex and fibrin degradation products D-dimer levels were significantly increased at 3 and 1.5 hours after LPS injection compared with controls.. Neutrophils are a source of TF and are implicated in direct activation of the coagulation cascade in the early phases of sepsis in the monkey. These results give important information for the treatment of sepsis.

Topics: Animals; Antithrombin III; Escherichia coli; Fibrin; Fibrin Fibrinogen Degradation Products; Immunohistochemistry; In Situ Hybridization; Lipopolysaccharides; Liver; Lung; Macaca; Macaca mulatta; Male; Neutrophils; Peptide Hydrolases; RNA, Messenger; Sepsis; Thromboplastin

Topics: Acute Disease; Animals; Antithrombin III; Blood Coagulation Disorders; Complement Activation; Cytokines; Disseminated Intravascular Coagulation; Endotoxemia; Fibrin; Gene Expression Regulation; Hemorrhage; Humans; Lipoproteins; Lung Injury; Primates; Protein C; Reactive Oxygen Species; Respiratory Distress Syndrome; Sepsis; Thromboplastin; Transcription, Genetic; Tumor Necrosis Factor-alpha

Topics: Blood Coagulation; Blood Coagulation Disorders; Cell Communication; Cytokines; Humans; Inflammation; Sepsis; Thromboplastin

Extravascular coagulation and fibrin deposition coupled with perturbations of intravascular coagulation occurs in association with acute respiratory distress syndrome (ARDS). To evaluate the pathogenetic role of an extrinsic coagulation pathway in the intravascular coagulation of ARDS patients and to explore the time course of the changes of tissue factor levels, platelet counts, and disseminated intravascular coagulation (DIC), we performed a prospective cohort study.. The study subjects consisted of 113 patients: 27 patients with ARDS, 31 patients at risk for but not developing the syndrome, and 55 patients without ARDS. According to the underlying disease, the patients were further subdivided into two groups: patients with trauma (n = 76) and patients with sepsis (n = 37). Ten normal healthy volunteers served as control subjects. Plasma tissue factor antigen (tissue factor) levels and platelet counts were measured on the day of admission and on days 1 through 4 after admission. Simultaneously, the DIC scores were determined.. The values of tissue factor in the patients with ARDS were significantly more elevated than those measured in the other two groups (p < 0.001) and control subjects (p < 0.001) on the day of admission. The values continued to be markedly high up to day 4 of admission. On the day of admission, the platelet counts in the ARDS patients showed significantly lower values (p < 0.05) than those in the other two groups. The incidence of DIC and the DIC scores in ARDS patients were significantly higher than those in the other two groups. The tissue factor levels (r(s) = 0.428, p < 0.0001) and DIC scores (r(s) = 0.357, p < 0.0002) correlated significantly with Lung Injury Score. When the patients were subdivided into two subgroups, i.e., trauma and sepsis, some differences of the tissue factor levels were noted between the two groups.. We demonstrated that tissue-factor dependent coagulation pathway of plasma is extensively activated in patients with ARDS, followed by intravascular coagulation and platelet consumption. We further provide precise information on the time course of tissue factor levels and DIC in patients with ARDS and those at risk for developing this syndrome.

Topics: Analysis of Variance; APACHE; Blood Coagulation; Case-Control Studies; Disseminated Intravascular Coagulation; Female; Fibrin; Fibrinogen; Humans; Male; Middle Aged; Multiple Trauma; Platelet Count; Prospective Studies; Prothrombin Time; Respiratory Distress Syndrome; Risk Factors; Sepsis; Thromboplastin; Time Factors

To determine the role of plasma tissue factor on disseminated intravascular coagulation (DIC) in trauma and septic patients, and also to investigate the relationships between tissue factor and various thrombin markers, we made a prospective cohort study. Forty trauma patients and 20 patients with sepsis were classified into subgroups according to the complication of DIC. Plasma tissue factor antigen concentration (tissue factor), prothrombin fragment F1+2 (PF1+2), thrombin antithrombin complex (TAT), fibrinopeptide A (FPA), and D-dimer were measured on the day of admission (day 0), and on days 1, 2, 3, and 4 after admission. The levels of plasma tissue factor in the DIC group were more elevated than those of the non-DIC group in both the trauma and the septic patients. In patients with sepsis, tissue factor levels on days 0 through 4 in the non-DIC group showed markedly higher values than those in the control patients (135 +/- 8 pg/ml). Significant correlations between tissue factor and PF1+2, TAT, FPA, and D-dimer were observed in the DIC patients, however, no such correlations were found in the non-DIC patients. These results suggest that elevated plasma tissue factor in patients with trauma and sepsis gives rise to thrombin generation, followed by intravascular coagulation.

Topics: Adult; Aged; Aged, 80 and over; Antithrombin III; APACHE; Biomarkers; Disseminated Intravascular Coagulation; Endothelium, Vascular; Female; Fibrin Fibrinogen Degradation Products; Fibrinopeptide A; Humans; Male; Middle Aged; Monocytes; Peptide Fragments; Peptide Hydrolases; Prospective Studies; Prothrombin; Sepsis; Thrombin; Thromboplastin; Wounds and Injuries

Topics: Adult; Aged; Aged, 80 and over; Cohort Studies; Disseminated Intravascular Coagulation; Female; Flow Cytometry; Gene Expression Regulation; Humans; Macrophage-1 Antigen; Male; Middle Aged; Monocytes; NF-kappa B; Sepsis; Thromboplastin; Time Factors

To obtain systematic information on the extrinsic coagulation pathway, as well as to investigate the time course of the coagulation abnormalities in sepsis.. Prospective observational study.. General intensive care unit.. Nineteen patients with the diagnosis of severe sepsis or septic shock and nine control patients.. None.. Tissue factor antigen concentration (tissue factor antigen), prothrombin fragment F1+2, thrombin antithrombin III complex, fibrinopeptide A, D-dimer, and antithrombin III concentrations were measured on the day of diagnosis of severe sepsis and septic shock, and on days 1, 2, 3, and 4 after diagnosis. The concentrations of tissue factor antigen, prothrombin fragment F1+2, fibrinopeptide A, and D-dimer were significantly increased in patients with severe sepsis and septic shock compared with control subjects. However, the concentrations of thrombin antithrombin III complex showed no statistical differences between the septic patients and the control subjects. Significantly, low antithrombin III concentrations were observed in the septic patient groups compared with control subjects. With the exception of D-dimer, the concentrations of the hemostatic markers were similar between severe sepsis and septic shock patients. Significant correlations were noted between tissue factor antigen and the disseminated intravascular coagulation score (r2=.236, p< .0001) and the number of dysfunctioning organs (r2=.229, p=.035).. We systematically elucidated coagulation disorders in newly defined sepsis. The extrinsic coagulation pathway is activated in patients with severe sepsis and septic shock. In these patients, enhanced thrombin generation and activation, and fibrin formation were demonstrated when compared with the control subjects. Furthermore, the thrombin generated appears not to be fully neutralized by antithrombin III.

Topics: Adult; Aged; Antithrombin III; Biomarkers; Blood Coagulation; Blood Coagulation Disorders; Case-Control Studies; Female; Fibrin Fibrinogen Degradation Products; Fibrinopeptide A; Humans; Male; Middle Aged; Peptide Fragments; Peptide Hydrolases; Prospective Studies; Protein Precursors; Prothrombin; Sepsis; Shock, Septic; Thromboplastin

Binding activity for nuclear factor kappa B (NFkappaB) consensus probes was studied in nuclear extracts from peripheral blood mononuclear cells of 15 septic patients (10 surviving and 5 not surviving). Nonsurvivors could be distinguished from survivors by an increase in NFkappaB binding activity during the observation period (P < 0.001). The increase in NFkappaB binding activity was comparable to the APACHE-II score as a predictor of outcome. Intravenous somatic gene transfer with an expression plasmid coding for IkappaBalpha was used to investigate the role of members of the NFkappaB family in a mouse model of endotoxemia. In this model, increased NFkappaB binding activity was present after injection of LPS. Intravenous somatic gene transfer with IkappaBalpha given before LPS attenuated renal NFkappaB binding activity and increased survival. Endothelial cells and monocytes/macrophages were the major target cells for somatic gene transfer, transfected with an average transfection efficiency of 20-35%. Tissue factor, a gene under regulatory control of NFkappaB, was induced by LPS. Somatic gene transfer with a reporter plasmid containing the functional tissue factor promoter demonstrated NFkappaB-dependent stimulation by LPS. Intravenous somatic gene transfer with IkappaBalpha reduced LPS-induced renal tissue factor expression, activation of the plasmatic coagulation system (decrease of thrombin-antithrombin III complexes) and renal fibrin/fibrinogen deposition. Somatic gene transfer with an expression plasmid with tissue factor cDNA in the antisense direction (in contrast to sense or vector alone) also increased survival. Furthermore, antisense tissue factor decreased renal tissue factor expression and the activation of the plasmatic coagulation system.

Topics: Adult; Aged; Animals; Blood Coagulation; Female; Humans; Lipopolysaccharides; Male; Mice; Mice, Inbred BALB C; Middle Aged; NF-kappa B; Sepsis; Thromboplastin; Tumor Necrosis Factor-alpha

Tissue factor (TF), a transmembrane surface protein, is known to initiate thrombogenesis through plasmatic and cellular activation processes. Besides complexing with factor VII, eventually leading to fibrin generation via the extrinsic pathway, TF can also activate factor IX, resulting in the intrinsic activation of coagulation. Other functions of TF are currently unknown, although various cells are believed to have TF receptors. Many of the post-surgical and post-interventional thrombotic events are due to the release of TF. Increased levels of TF are associated with several pathologic conditions such as cancer, sepsis and inflammation. Cellular necrosis also results in an increase of TF as the cells in the traumatized area lyse and release endogenous cell surface-bound TF. An ELISA method (American Diagnostica, Greenwich, CT) has been developed to assay TF antigen levels in various biological fluids. This ELISA employs a murine monoclonal antibody raised against native human TF for antigen capture. In this study, cerebrospinal fluid, peritoneal fluid, pleural effusion and urine from patients were assayed for their TF content using this ELISA method. Normal individual serum and plasma were also assayed as controls against which the levels of TF in the patients' body fluids could be compared. The amount of TF antigen in normal human plasma and serum was 165 +/- 139 pg/ml and 165 +/- 110 pg/ml, respectively. Concentrations of TF antigen in other fluids were: cerebrospinal fluid 868 +/- 721 pg/ml, peritoneal fluid 124 +/- 247 pg/ml, pleural effusion 385 +/- 569 pg/ml, synovial fluid 97 +/- 23 pg/ml, seminal plasma 11,485 +/- 875 pg/ml and urine 86 +/- 57 pg/ml.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Amino Acid Sequence; Angioplasty; Ascitic Fluid; Body Fluids; Coronary Angiography; Coronary Disease; Enzyme-Linked Immunosorbent Assay; Humans; Molecular Sequence Data; Pleural Effusion; Semen; Sepsis; Synovial Fluid; Thromboplastin

Reciprocal interactions between elements of the acute inflammatory response and the coagulation system play important roles in host defense homeostasis during Gram-negative bacterial sepsis. However, derangements in the regulation of the inflammatory-coagulant axis in this setting may result in progressive tissue damage and disseminated intravascular coagulation. In this article, the integrated responses in the baboon model of Escherichia coli sepsis are analyzed as a basis of understanding these response interactions in the critically ill. In particular, three topics will be reviewed. First, the role of tissue factor in mediating the coagulant response to inflammation and the role of tumor necrosis factor (TNF) in initiating and amplifying this coagulant response into a full-blown consumptive coagulopathy are defined. A second and parallel topic concerns the role played by tissue factor pathway inhibitor and other anticoagulant systems in not only regulating this coagulant response, but also in attenuating the initial inflammatory response. The third topic concerns the use of assays of enzyme inhibitor complexes composed of components of these regulatory anticoagulant systems to help define the hypercoagulable state and possibly to make an early, specific diagnosis of sepsis prior to overt failure of the hemostatic system.

Topics: Animals; Blood Coagulation; Blood Coagulation Factors; Disseminated Intravascular Coagulation; Endothelium, Vascular; Escherichia coli Infections; Fibrin; Inflammation; Lipoproteins; Papio; Sepsis; Thromboplastin; Tumor Necrosis Factor-alpha

Nafamostat mesilate (NM), a synthetic protease inhibitor, is frequently used for the treatment of disseminated intravascular coagulation (DIC) in Japan. NM inhibits several proteases which may be importantly involved in the pathophysiology of DIC. Since tissue factor (TF) plays a critical role in DIC associated with septicemia, inhibition of the extrinsic pathway of coagulation by coagulation inhibitors may be useful for the treatment of DIC. NM inhibited extrinsic pathway activity (TF-F.VIIa mediated-F.Xa generation) in a concentration dependent manner; the IC50 was 1.0 x 10(-7) M. F.Xa was not inhibited by NM at the concentrations used in the experiment, suggesting that NM might inhibit TF-F.VIIa complex activity. When incubated with TF-F.VIIa complex, NM inhibited the complex activity with an IC50 of 1.5 x 10(-7) M, the same value that found for inhibition of extrinsic pathway activity. A Lineweaver-Bulk's plot of the inhibition demonstrated that NM inhibited TF-F.VIIa complex in a competitive fashion, with an inhibition constant (Ki) of 2.0 x 10(-7) M. These findings suggested that NM may be a potent inhibitor of TF-F.VIIa complex and the therapeutic effect of NM in DIC patients could be partly explained by inhibition of the extrinsic pathway of the coagulation system.

Topics: Amino Acid Sequence; Benzamidines; Blood Coagulation; Disseminated Intravascular Coagulation; Factor VIIa; Guanidines; Humans; Molecular Sequence Data; Protease Inhibitors; Sepsis; Thromboplastin

Using a whole blood in vitro model, we have investigated the effect of Escherichia coli (E. coli), Streptococcus agalactiae (group B streptococci, GBS) and tumor necrosis factor alpha (TNF) on the generation of lactoferrin (LF), interleukin-1 beta (IL-1) and tissue thromboplastin (TPL) in healthy newborns at term and their mothers. E. coli (at a final concentration of 10(7)/ml) significantly increased the release of LF in whole blood from newborns after 20 as well as 60 min stimulation, and in samples from their mothers after 60 min stimulation. A significant increase in the release of LF was observed in both newborns and their mothers after 20 and 60 min stimulation with TNF (at a final concentration of 1000 pg/ml). A combination of TNF/E. coli or TNF/GBS never gave any significant additional stimulatory effect. After stimulation with E. coli or GBS (both at a final concentration of 10(7)/ml) for 60 min a significant increase in production of TNF and TPL was observed in newborns. In newborns a significant increase in production of TNF and TPL was observed also after 20 min stimulation with E. coli. TNF (at a final concentration of 1000 pg/ml) significantly increased the generation of TPL after 20 and 60 min stimulation in both groups. There was a tendency for a greater release of LF and generation of TNF and TPL in samples from newborns compared with their mothers, but the differences were not statistically significant. E. coli, GBS and TNF had no significant effect on the production of IL-1.

Topics: Escherichia coli; Female; Humans; In Vitro Techniques; Infant, Newborn; Interleukin-1; Lactoferrin; Sepsis; Streptococcus agalactiae; Thromboplastin; Tumor Necrosis Factor-alpha

Human plasma contains an inhibitor of tissue factor-initiated coagulation known as the lipoprotein-associated coagulation inhibitor (LACI) or also known as the extrinsic pathway inhibitor (EPI). A competitive fluorescent immunoassay was developed to measure the plasma concentration of LACI in samples from normal individuals and patients with a variety of diseases. The LACI concentration in an adult control population varied from 60% to 160% of the mean with a mean value corresponding to 89 ng/mL or 2.25 nmol/L. Plasma LACI levels were not decreased in patients with severe chronic hepatic failure, warfarin therapy, primary pulmonary hypertension, thrombosis, or the lupus anticoagulant. Plasma LACI antigen was decreased in some, but not all patients with gram-negative bacteremia and evidence for disseminated intravascular coagulation. Plasma LACI levels were elevated in women undergoing the early stages of labor (29%), in patients receiving intravenous tissue-type plasminogen activator (45%), and in patients receiving intravenous heparin (375%). A radioligand blot of the pre- and post-heparin plasma samples shows the increase to be in a 40-Kd form of LACI. Very low levels of plasma LACI antigen were found in patients with homozygous abetalipoproteinemia and hypobetalipoproteinemia, diseases associated with low plasma levels of apolipoprotein B containing lipoproteins. Following the injection of heparin into one patient with homozygous abetalipoproteinemia, the plasma LACI antigen level increased to a level comparable with that in normal individuals after heparin treatment.

Topics: Abetalipoproteinemia; Antibodies, Monoclonal; Apolipoproteins E; Cell Line; Factor VII; Female; Fluorescent Antibody Technique; Humans; Hypertension, Pulmonary; Labor, Obstetric; Lipoproteins; Pregnancy; Protease Inhibitors; Reference Values; Sepsis; Tangier Disease; Thromboplastin; Thrombosis

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

Endotoxin and monocyte thromboplastin activity were evaluated in rats with gram-negative septicaemia induced by caecal perforation or intravenous Escherichia coli challenge and treated with antibiotics or placebo. Endotoxin burst was not detected in either form of septicaemia during antibiotic treatment. Thromboplastin synthesis in monocytes is known to be stimulated by endotoxin, but the rats showed no increase of monocyte thromboplastin activity after antibiotic treatment, which constituted further evidence against the concept of massive endotoxin liberation during antibiotic therapy for gram-negative septicaemia.

Topics: Animals; Anti-Bacterial Agents; Cefotaxime; Endotoxins; Escherichia coli Infections; Gentamicins; Male; Monocytes; Rats; Rats, Inbred Strains; Sepsis; Thromboplastin; Tinidazole

Rats were subjected to gram-negative septicemia induced by cecal perforation or were sham-operated. Thromboplastin values increased in blood monocytes (40-fold), peritoneal macrophages (115-fold) pleural macrophages (5-fold), splenic macrophages (3-fold), and lung alveolar macrophages (1.4-fold) in septic animals as compared to controls. In septic animals disseminated intravascular coagulation was evidenced by a significant (p less than 0.05) fall in fibrinogen, factor VII, X and platelets. A simultaneous and significant (p less than 0.05) decrease in thromboplastin content of tissue-specimens from lung and spleen was observed in rats with septicemia, whereas increased thromboplastin values were demonstrated in tissue-samples from cecum - the infectious focus. This might reflect mobilization of mononuclear phagocytes in favour of the site of infection.

Topics: Animals; Blood Coagulation Factors; Cell Movement; Disease Models, Animal; Disseminated Intravascular Coagulation; Gram-Negative Bacteria; Macrophages; Male; Monocytes; Peritoneal Cavity; Pleura; Pulmonary Alveoli; Rats; Rats, Inbred Strains; Sepsis; Spleen; Thromboplastin; Tissue Distribution

The relationship between mononuclear phagocyte thromboplastin activity, microorganisms and levels of endotoxin in peritoneal fluid and splanchnic and systemic circulation was evaluated during experimental endogenous gram-negative peritonitis in the rat. Significant rise in thromboplastin activity of mononuclear phagocytes was demonstrated in all three compartments. This newly synthesized thromboplastin is a trigger for important biologic systems such as the coagulation cascade, and thus may play a major role in the development of disseminated intravascular coagulation so often occurring in gram-negative sepsis. It probably also participates in the formation of fibrous intraabdominal adhesions. Aerobic and anaerobic microorganisms together with endotoxin were detected already 1 1/2 hours after induction of peritonitis, and subsequently were found to increase in parallel fashion. Determination of endotoxin is a rapid and seemingly reliable method for early detection of gram-negative infection and thus may be of clinical value.

Topics: Animals; Ascitic Fluid; Endotoxins; Gram-Negative Bacteria; Male; Phagocytes; Rats; Rats, Inbred Strains; Sepsis; Thromboplastin

Topics: Blood Coagulation; Complement Activation; Humans; Lymphocytes; Macrophages; Monocytes; Neutrophils; Phytohemagglutinins; Plasminogen Activators; Risk; Sepsis; Thromboplastin; Wounds and Injuries

Topics: Afibrinogenemia; Aneurysm; Bacteria; Blood Coagulation; Blood Coagulation Tests; Blood Platelets; Diagnosis, Differential; Disseminated Intravascular Coagulation; Enzyme Activation; Fibrin; Fibrinogen; Fibrinolysis; Hemostasis; Heparin; Models, Biological; Prothrombin; Sepsis; Shwartzman Phenomenon; Thrombin; Thromboplastin

Topics: Blood Coagulation Disorders; Disseminated Intravascular Coagulation; Humans; Oxygen Consumption; Prothrombin Time; Pulmonary Edema; Sepsis; Shock; Thromboplastin

Topics: Animals; Blood Coagulation Disorders; Blood Platelets; Blood Pressure; Cardiac Output; Disseminated Intravascular Coagulation; Escherichia coli Infections; Fibrinolysis; Hematocrit; Hydrogen-Ion Concentration; Male; Oxygen; Papio; Prothrombin Time; Sepsis; Shock, Septic; Thromboplastin; Vascular Resistance

Topics: Blood Cell Count; Blood Coagulation Disorders; Blood Coagulation Factors; Blood Coagulation Tests; Blood Platelets; Blood Pressure; Child; Child, Preschool; Factor V; Fibrinogen; Heparin; Humans; Hypotension; Infant; Infant, Newborn; Prothrombin Time; Rocky Mountain Spotted Fever; Sepsis; Shock, Septic; Thrombocytopenia; 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

Topics: Animals; Blood Coagulation Disorders; Dogs; Endotoxins; Escherichia coli; Fibrinogen; Heparin; Prothrombin Time; Sepsis; Thromboplastin

Topics: Animals; Electrons; Female; Fibrin; Glomerulonephritis; Humans; Kidney Diseases; Kidney Glomerulus; Microscopy; Microscopy, Electron; Pathology; Phagocytosis; Pre-Eclampsia; Pregnancy; Rabbits; Research; Sepsis; Thromboplastin