Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.69 (APC)
 16,337 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Activated protein C (APC) is an antithrombotic serine proteinase having anticoagulant, profibrinolytic and anti-inflammatory activities. Despite its potential clinical utility, relatively little is known about its clearance mechanisms. In the present study we have characterized the interaction of APC and its active site blocked forms with human umbilical vein endothelial cells (HUVEC). At 4 degrees C 125I-APC bound to HUVEC in a specific, time dependent, saturable and reversible manner. Scatchard analysis of the binding isotherm demonstrated a Kd value of 6.8 nM and total number of binding sites per cell of 359,000. Similar binding isotherms were obtained using radiolabeled protein C (PC) zymogen as well as D-phe-pro-arg-chloromethylketone (PPACK) inhibited APC indicating that a functional active site was not required. Competition studies showed that the binding of APC, PPACK-APC and PC were mutually exclusive suggesting that they bound to the same site(s). Proteolytic removal of the N-terminal gamma-carboxyglutamic acid (gla) domain of PC abolished its ability to compete indicating that the gla-domain was essential for cell binding. Surprisingly, APC binding to these cells appeared to be independent of protein S, a cofactor of APC generally thought to be required for its high affinity binding to cell surfaces. The identity of the cell binding site(s), for the most part, appeared to be distinct from other known APC ligands which are associated with cell membranes or extracellular matrix including phospholipid, thrombomodulin, factor V, plasminogen activator inhibitor type 1 (PAI-1) and heparin. Pretreatment of HUVEC with antifactor VIII antibody caused partial inhibition of 125I-APC binding indicating that factor VIII or a homolog accounted for approximately 30% of APC binding. Studies of the properties of surface bound 125I-APC or 125I-PC and their fate at 4 degrees C compared to 37 degrees C were consistent with association of approximately 25% of the initially bound radioligand with an endocytic receptor. However, most of the radioligand appeared not to be bound to an endocytic receptor and dissociated rapidly at 37 degrees C in an intact and functional state. These data indicate the presence of specific, high affinity binding sites for APC and PC on the surface of HUVEC. While a minor proportion of binding sites may be involved in endocytosis, the identity and function of the major proportion is presently unknown. It is speculated that this putative receptor may be a further mechanisms of localizing the PC antithrombitic system to the vascular endothelium.
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PMID:High affinity binding sites for activated protein C and protein C on cultured human umbilical vein endothelial cells. Independent of protein S and distinct from known ligands. 785 99

The vascular endothelium, by virtue of its position at the interface between blood and the vessel wall, is known to play a critical role in the control of thrombosis and fibrinolysis. Thrombomodulin (TM) is a surface receptor that binds thrombin and is a potent activator of the protein C anticoagulant pathway. Although TM expression is known to be regulated by various cytokines, little is known about its response to ever-present biomechanical stimuli. We have explored the role of fluid shear stress, imparted on the luminal surface of the endothelial cell as a result of blood flow, on the expression of TM mRNA and protein in both bovine aortic endothelial (BAE) and bovine smooth muscle (BSM) cells in an in vitro system. We report in the present study that TM expression is regulated by flow. Subjecting BAE cells to fluid shear stress in the physiological range of magnitude of 15 (moderate shear stress) and 36 (elevated shear stress) dynes/cm2 resulted in a mild transient increase followed by a significant decrease in TM mRNA to 37% and 16% of its resting level, respectively, by 9 hours after the onset of flow. In contrast, shear stress at the low magnitude of 4 dynes/cm2 did not affect TM mRNA levels. The sensitivity of TM mRNA expression by flow was found to be specific to endothelium, since it was not observed in BSM cells exposed to steady laminar shear stress of 15 dynes/cm2. Furthermore, unlike BAE cells, BSM cells did not exhibit altered cell shape nor align in the direction of flow after 24 hours of shear stress at 15 dynes/cm2.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Endothelial expression of thrombomodulin is reversibly regulated by fluid shear stress. 815 32

The plasminogen activator systems in the blood, the coagulation system, and the complement pathways are reviewed. The review describes the role of the vascular intima in activation of coagulation and fibrinolysis and the interrelations between the complement system and haemostatic mechanisms. Physiological activation of fibrinolysis may be triggered by and limited to fibrin because of a special affinity of plasminogen and plasminogen activators. The binding of plasminogen to fibrin is regulated by histidine-rich glycoprotein, and the primary physiological inhibitor of generated plasmin is alpha 2-antiplasmin and especially the plasminogen-binding form of this immediate plasmin inhibitor. Plasminogen activator inhibitors in the blood, that is, notably plasminogen activator inhibitor type 1 (PAI-1), bind circulating tissue-type plasminogen activator (t-PA). However, local fibrinolysis in vivo mediated by t-PA may be independent of complex formation between plasminogen activator inhibitors and t-PA in the fluid phase. Circulating plasminogen activator inhibitors might regulate fibrinolysis by increasing the clearance of t-PA from the blood. The urokinase-type and factor XII-dependent fibrinolytic proactivator system can be activated following t-PA-mediated generation of plasmin, and could thus serve as an amplification system of t-PA-induced fibrinolysis. It is claimed that the as yet uncharacterized proactivator is essential for optimal generation of plasminogen activator activity by the factor XII-dependent fibrinolytic system. The normal antithrombotic condition of the vascular intima probably results from lack of tissue factor activity and the presence of significant antithrombotic components comprising, among others, antithrombin III and the protein C-protein S system. A number of pathophysiologic stimuli, notably mediators of the acute phase response such as the cytokines interleukin-1 and tumour necrosis factor-alpha (cachectin), have the potential to induce the vascular endothelium to express procoagulant activity. Vascular endothelium promoting coagulant activity releases increased amounts of t-PA antigen and PAI-1 antigen into the circulation, and elevated levels in the blood of both may be regarded as a marker of a generalized procoagulant condition involving the vascular endothelium. In a prospective study in patients with unstable angina pectoris, patients in whom disease progresses and acute myocardial infarction develops, have increased amounts of t-PA antigen and PAI-1 antigen in the blood. This suggests that the procoagulant potential and atherosclerotic process of the vascular intima is more pronounced in the risk group.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Fibrinolysis in patients with acute ischaemic heart disease. With particular reference to systemic effects of tissue-type plasminogen activator treatment on fibrinolysis, coagulation and complement pathways. 822 63

Adhesion molecules of leucocytes Leu-CAMs, also called beta-2 integrins, are heterodimeric glycoproteins which play a crucial role in interactions of leucocytes between themselves or with fundamental intercellular substances. They comprise 3 elements: LFA-1 (CD11a/CD18) expressed at the surface of leucocytes, and in particular lymphocytes, M01 or MAC-1 (CD11b/CD18), and p 150.95 (CD11c/CD18) expressed only on granulocytes, monocytes and macrophages. Their structure, function and regulation are studied. These 3 elements differ in the ligand(s) to which they adhere. LFA-1 intervenes in the adhesion of lymphocytes T and B and of cells presenting with the APC antigen; it therefore plays an important role in lymphoblast proliferation, T-cell cytotoxicity and immunoglobulin synthesis. M01 and p 150.95 intervene in the adhesion of particles and germs opsonized by serum complement, thereby playing a fundamental role in the body's defence against bacterial and fungal infections. They also intervene in the adhesion of leucocytes onto the vascular endothelium and in their migration through this vascular wall towards the inflammatory focus. The pathologies related to a congenital deficiency of these adhesion molecules or to the alterations they acquired are dealt with. The use of monoclonal antibodies directed against leucocyte adhesion in animal experiments opens new therapeutic vistas.
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PMID:[Adhesion molecules of Leu-CAMs leukocytes]. 824 68

Growing evidence suggests that moderately elevated levels of homocysteine are associated not only with arterial thrombosis and atherosclerosis but also with venous thrombosis as well. We have reviewed recent studies that indicate that homocysteine inhibits several different anticoagulant mechanisms that are mediated by the vascular endothelium. The protein C enzyme system appears to be one of the most important anticoagulant pathways in the blood. Homocysteine inhibits the expression and activity of endothelial cell surface thrombomodulin, the thrombin cofactor responsible for protein C activation. Homocysteine inhibits the antithrombin III binding activity of endothelial heparan sulfate proteoglycan, thereby suppressing the anticoagulant effect of antithrombin III. Homocysteine also inhibits the ecto-ADPase activity of human umbilical vein endothelial cells (HUVECS). Because ADP is a potent platelet aggregatory agent, this action of homocysteine is prothrombotic. Homocysteine also interferes with the fibrinolytic properties of the endothelial surface because it inhibits the binding of tissue plasminogen activator. Homocysteine stimulates HUVEC tissue factor activity. We have found that lipoprotein(a) [Lp(a)] also stimulates HUVEC tissue factor activity. The combination of Lp(a) plus homocysteine induced more tissue factor activity than either agent alone. These disruptions in several different vessel wall-related anticoagulant functions provide plausable mechanisms for the occurrence of thrombosis in hyperhomocysteinemia.
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PMID:Homocysteine and hemostasis: pathogenic mechanisms predisposing to thrombosis. 864 72

Markers of endothelial cell activation were measured in 28 patients presenting with various forms of limited or focal type cutaneous vasculitis. Plasma levels of tissue plasminogen activator antigen (t-PA:Ag), plasminogen activator inhibitor type 1 antigen (PAI-1:Ag) and PAI-1 activity, fibrin plate, von Willebrand factor antigen (vWF:Ag), tissue factor (TF) and soluble thrombomodulin (sTM) were measured. In comparison with the control group (n = 20) there was a significant increase in t-PA:Ag, vWF:Ag and TF (P < 0.05, Mann-Whitney U-test) in the cutaneous vasculitis group. This study confirms that measurable degrees of endothelial activation occur in cutaneous vasculitis. Cutaneous vasculitis includes a diverse group of clinical conditions, which are associated with inflammatory changes in cutaneous blood vessels with local fibrin deposition. The aetiology and pathogenesis of the majority of these entities remain unknown. Causative mediators are thought to include immune complexes, anti-endothelial cell antibodies, cytotoxic lymphocytes and viruses. Histologically, immune complexes and complement are frequently detected on the vessel wall, and serologically anti-endothelial antibodies are often detected in patients with vasculitis and in systemic lupus erythematosus (SLE) which correlate with the severity of cutaneous vasculitis, arthritis and nephritis. Lymphocyte-mediated toxicity to endothelial cells has been reported in a small number of patients with giant cell arteritis and Takayasu's arteritis. The vascular endothelium plays a central part in the control of haemostasis. Under physiological conditions endothelial cells present an anticoagulant surface to blood constituents, partially due to surface expression of heparan sulphate and thrombomodulin (TM). Heparan sulphate binds antithrombin III (ATIII), thereby accelerating inactivation of intrinsic coagulation enzymes. Thrombomodulin is an endothelial cell surface glycoprotein which promotes anticoagulation by forming a complex with thrombin which then activates protein C. Activated protein C together with a cofactor, protein S, inactivates FVa and FVIIIa. von Willebrand factor (vWF) is synthesized by endothelial cells, stored in Weibel-Palade bodies and released into the circulation upon endothelial stimulation. vWF mediates the binding of platelets to the subendothelium and is the carrier molecule for FVIIIC. The endothelium controls fibrinolysis by producing t-PA and its inhibitor PAI-1. Inflammatory cytokines such as interleukin-1 (IL-1) and tumour necrosis factor (TNF) activate endothelial cells, causing a shift from an antithrombotic to prothrombotic state, including expression of tissue factor, increased synthesis of PAI-1 and decreased expression of TM. Fibrin deposition and intravascular thrombosis are seen in cutaneous vasculitis syndromes, suggesting local endothelial cell activation. The aim of this pilot study was to assess whether perturbation of the endothelium in cutaneous vasculitis could be detected in the patients' plasma samples. If so, further studies to assess any correlation in levels of these markers with disease activity might prove useful in the future.
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PMID:Endothelial cell activation in cutaneous vasculitis. 868 65

Protein C activation on the surface of the endothelium is critical to the negative regulation of blood coagulation. We now demonstrate that monoclonal antibodies that block protein C binding to the endothelial cell protein C receptor (EPCR) reduce protein C activation rates by the thrombin-thrombomodulin complex on endothelium, but that antibodies that bind to EPCR without blocking protein C binding have no effect. The kinetic result of blocking the EPCR-protein C interaction is an increased apparent Km for the activation without altering the affinity of thrombin for thrombomodulin. Activation rates of the protein C derivative lacking the gamma-carboxyglutamic acid domain, which is required for binding to EPCR, are not altered by the anti-EPCR antibodies. These data indicate that the protein C activation complex involves protein C, thrombin, thrombomodulin, and EPCR. These observations open new questions about the control of coagulation reactions on vascular endothelium.
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PMID:The endothelial cell protein C receptor augments protein C activation by the thrombin-thrombomodulin complex. 881 78

Circulating plasma thrombomodulin (TM) is an endothelial cell marker which may reflect endothelial injury. To find out to what extent diagnostic cardiac catheterization irritates vascular endothelium we conducted a prospective study in 91 children. Soluble TM concentrations, along with thrombin generation, were measured before, at the end of and 24 hours after cardiac catheterization. Compared to starting values, TM concentrations showed a clearly significant increase at the end of cardiac catheterization and returned to pretreatment values 24 hours later. Thrombin generation followed a similar pattern. Five out of the 91 children demonstrated resistance to activated protein C (APCR). With respect to the remaining 86 children, all five APCR cases showed increased thrombomodulin concentrations along with enhanced thrombin generation. Data from this study indicate that increased TM concentrations after cardiac catheterization in children are a sign of short-term endothelial damage. Furthermore, together with enhanced thrombin generation, elevated plasma concentration of soluble TM may reflect this receptor's possible anticoagulant properties.
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PMID:Plasma thrombomodulin concentrations in infants and children undergoing cardiac catheterization. 895 60

Thrombomodulin is a principle thrombin receptor located on the vascular endothelium. Thrombomodulin alters the specificity of thrombin, redirecting its procoagulant function to an anticoagulant function by making it a more efficient activator of protein C. While mutation of the genes of other components of this anticoagulant mechanism, protein C, protein S and factor V, is known to predispose towards venous thromboembolism, there are only a few reports of the investigation of thrombomodulin gene mutation. We present the design and evaluation of a strategy to investigate thrombomodulin gene mutation in arterial and venous thrombosis.
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PMID:Directed search for thrombomodulin gene mutations. 897 28

Thrombomodulin (TM), an endothelial receptor for thrombin, endowed with a powerful anticoagulant activity, plays an important role in the antithrombogenicity of the vascular endothelium. Its presence within the human renal glomerulus is already known but was thought to be only endothelial. We looked for TM expression in human mesangial cells (MC), both in situ, in freshly prepared glomeruli, and in primary culture. Both fresh and cultured MC were strongly reactive for TM by immunocytochemical methods. Total TM antigen measured on MC lysates and surface TM activity on MC were 0.292 +/- 0.075 ng/mg of cellular proteins and 1.20 +/- 0.02 pmole of activated protein C/min/mg of cellular proteins, respectively. As shown by the presence of numerous transcripts detected by in situ hybridization, TM was shown to be synthesized by MC in vivo and in culture. The synthesis of active TM by both endothelial and mesangial cells within the renal glomerulus stresses the importance of its role in maintaining renal hemostatic equilibrium, and sheds some light on the conflicting reports of TM over- and underexpression in glomerulopathies to open a new field for investigation.
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PMID:Thrombomodulin is synthesized by human mesangial cells. 906


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