Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.5 (thrombin)
 33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Interaction of von Willebrand Factor with glycoprotein Ib-IX-V induces platelet activation through a still poorly defined mechanism. Previous studies have suggested a possible role for the low affinity receptor for immunoglobulin, Fc gamma RIIA, in GPIb-IX-V signaling. Here we show that binding of vWF to platelets induces the tyrosine phosphorylation of Fc gamma RIIA by a Src kinase. Treatment of platelets with the anti-Fc gamma RIIA monoclonal antibody IV.3 specifically inhibits vWF-induced but not thrombin-induced pleckstrin phosphorylation and serotonin secretion. Moreover, vWF fails to induce pleckstrin phosphorylation in mouse platelets, lacking Fc gamma RIIA, and serotonin secretion is impaired. Pleckstrin phosphorylation and serotonin secretion in human platelets stimulated with vWF are blocked by the cyclooxygenase inhibitor acetylsalicylic acid. However, release of arachidonic acid and synthesis of TxA(2) induced by vWF are not affected by the anti-Fc gamma RIIA monoclonal antibody IV.3. Similarly, vWF-induced tyrosine phosphorylation of Fc gamma RIIA, as well as of Syk and PLC gamma 2, occurs normally in aspirinized platelets. Inhibition of the tyrosine kinase Syk by piceatannol does not affect vWF-induced tyrosine phosphorylation of Fc gamma RIIA but prevents phosphorylation of PLC gamma 2. Pleckstrin phosphorylation and platelet secretion induced by vWF, but not by thrombin, are also inhibited by piceatannol. Pleckstrin phosphorylation is also sensitive to the phosphatidylinositol 3-kinase inhibitor wortmannin. These results indicate that PLC gamma 2 plays a central role in platelet activation by vWF and that the stimulation of this enzyme requires coordinated signals through endogenous TxA(2) and Fc gamma RIIA.
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PMID:Platelet activation by von Willebrand factor requires coordinated signaling through thromboxane A2 and Fc gamma IIA receptor. 1134 69

1. The pharmacological characteristics of solid-phase von Willebrand factor (svWF), a novel platelet agonist, were studied. 2. Washed platelet suspensions were obtained from human blood and the effects of svWF on platelets were measured using aggregometry, phase-contrast microscopy, flow cytometry and zymography. 3. Incubation of platelets with svWF (0.2 - 1.2 microg ml(-1)) resulted in their adhesion to the ligand, while co-incubations of svWF with subthreshold concentrations of ADP, collagen and thrombin resulted in aggregation. 4. 6B4 inhibitory anti-glycoprotein (GP)Ib antibodies abolished platelet adhesion stimulated by svWF, while aggregation was reduced in the presence of 6B4 and N-Acetyl-Pen-Arg-Gly-Asp-Cys, an antagonist of GPIIb/IIIa. 5. Platelet adhesion stimulated with svWF was associated with a concentration-dependent increase in expression of GPIb, but not of GPIIb/IIIa. 6. In contrast, collagen (0.5 - 10.0 microg ml(-1)) caused down-regulation of GPIb and up-regulation of GPIIb/IIIa in platelets. 7. Solid-phase vWF (1.2 microg ml(-1)) resulted in the release of MMP-2 from platelets. 8. Inhibition of MMP-2 with phenanthroline (10 microM), but not with aspirin or apyrase, inhibited platelet adhesion stimulated with svWF. 9. In contrast, human recombinant MMP-2 potentiated both the effects of svWF on adhesion and up-regulation of GPIb. 10. Platelet adhesion and aggregation stimulated with svWF were reduced by S-nitroso-n-acetyl-penicillamine, an NO donor, and prostacyclin. 11. Thus, stimulation of human platelets with svWF leads to adhesion and aggregation that are mediated via activation of GPIb and GPIIb/IIIa, respectively. 12. Mechanisms of activation of GPIb by svWF involve the release of MMP-2, and are regulated by NO and prostacyclin.
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PMID:Pharmacological characteristics of solid-phase von Willebrand factor in human platelets. 1168 49

Sudden physical exertion is associated with an increased risk of acute myocardial infarction (MI) and sudden cardiac death. In addition, activation of the coagulation cascade and/or reduced fibrinolytic capacity after physical exercise has been reported in patients with cardiovascular disease. We investigated the haemostatic responses to an acute submaximal physical exercise in middle-aged women with a history of MI compared with healthy, age-matched controls. Resting plasma von Willebrand factor antigen (vWF Ag) and tissue plasminogen activator (tPA) antigen concentrations and plasminogen activator inhibitor-1 (PAI-1) activity were higher in the patients compared with control subjects. After 30 min of submaximal exercise on a bicycle ergometer, small, but still significant, increases in fibrinogen and vWF Ag concentrations were found in both groups. However, exercise did not induce thrombin generation and fibrin formation, as assessed by thrombin-antithrombin complex and fibrin D-dimer, in either group. Both tPA antigen concentration and activity increased and PAI-1 activity decreased significantly with exercise in both groups. Interestingly, the magnitude of changes in these latter variables did not differ between the groups (P=.99, P=.88 and P=.24, respectively). The present study demonstrates that some middle-aged women with previous MI have no signs of coagulation activation and retained fibrinolytic response after submaximal exercise. The clinical implication of these results might be that women with stable coronary heart disease can participate in rehabilitative exercise training without exhibiting a procoagulative state.
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PMID:Retained fibrinolytic response and no coagulation activation after acute physical exercise in middle-aged women with previous myocardial infarction. 1209 Oct 46

In an earlier study, a site directed mutant rFVIII (rFVIII(m), Arg(336) --> Gln(336)) expressed in baculovirus-insect cell (Sf9) system was found to sustain high level activity during incubation at 37 degrees celsius for 24 h while the cofactor activity of normal plasma was declined steadily. In this study, a mutant B-domain deleted rFVIII(m), Arg(336) --> Gln(336) expressed in baculovirus-insect cell (Sf9) system was characterized for its enzymatic and chemical properties. The expressed rFVIII(m) and plasma FVIII (pFVIII) were purified by immunoaffinity column chromatography and identified by Western blot analysis. The partially purified rFVIII(m) exhibited cofactor specific activity of 2.01 x 10(3)units/mg protein. The molecular weight of rFVIII(m) ranged between 40 to 150 kDa with a major band at 150 kDa. Treatment of both rFVIII(m) and pFVIII with thrombin increased their cofactor activity in a similar pattern. Treatment of both the activated rFVIII(m) and native FVIII with APC decreased their cofactor activities, however, the former exhibited a slower decrease than the latter, although no significant difference was present. rFVIII(m) formed a complex with vWF, resulting in a stabilized form, and the lag period of thrombin-mediated activating was extended by vWF association. These results implicated that rFVIII(m) expressed in baculovirus-insect cell system had a comparable capacity as FVIII cofactor activity and might be a good candidate for the FVIII replacement therapy for hemophilia A patients.
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PMID:Expression and characterization of a mutant recombinant blood coagulation factor VIII (rFVIII (m)). 1221 15

Von Willebrand factor (vWF) is an adhesive protein involved in primary haemostasis virtually absent in the thoracic aorta of swine, an animal model widely used in thrombosis and atherosclerosis. By RT-PCR analysis we show that porcine aortic endothelial cells (PAEC) express the vWF gene, although vWF mRNA levels were 8+/-0.8-fold (p<0.05) or 290+/-8.9-fold (p<0.0001) lower than those in porcine pulmonary artery EC (PPEC) or human aortic EC (HAEC), respectively. Although vWF was rare in the thoracic aorta of swine, vWF propeptide (vWFpp) was present in the endothelium of this artery and in both primary and passaged PAEC. In addition, vWFpp but not vWF was detected in PAEC by Western blot. In PAEC neither vWFpp nor P-selectin immunostaining depicted Weibel-Palade bodies (WPB)-like structures, and acute stimuli (alpha-thrombin or the calcium ionophore A23187) did not increase vWF secretion. vWFpp co-localized with a Golgi marker, that cycles between the stacked Golgi (SG fraction) and earlier compartments of the secretory pathway. Our results confirm that PAEC express very low levels of vWF mRNA and indicate that in these cells, that do not have WPB, vWF and vWFpp have divergent intracellular trafficking pathways.
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PMID:Differential intracellular trafficking of von Willebrand factor (vWF) and vWF propeptide in porcine endothelial cells lacking Weibel-Palade bodies and in human endothelial cells. 1261 68

Intravenous gamma-immunoglobulin (i.v.Ig) is commonly used in the treatment of autoimmune and inflammatory vascular disorders to prevent thrombotic complications. The mechanism of action of i.v.Ig is, however, not yet elucidated. In view of this, we investigated the ability of i.v.Ig to modulate i) Ca(2+) signals of fura-2 loaded endothelial cells, and ii) the associated release of nitric oxide (NO) and von Willebrand factor (vWf). NO was measured either indirectly by radioimmunoassay of cGMP in unstimulated cells or directly by electrochemistry at the surface of stimulated endothelial cells from human umbilical cord veins (HUVEC). Short-term treatment of unstimulated HUVEC with intact i.v.Ig decreased the basal cytosolic Ca(2+) concentration by 20% while it activated the NO/cGMP synthesis. Following i.v.Ig treatment of HUVEC, the Ca(2+) liberation from internal stores and the vWf secretion induced by ATP, thrombin or histamine were significantly reduced by 38 and 60%, respectively. The effects on Ca(2+) signals were observed with intact i.v.Ig as well as with the F(ab')2 or the Fc fragments indicating that both portions are involved in the mechanism of action. The i.v.Ig treatment of HUVECs had no effect on the NO release induced by thrombin or histamine. By contrast, the i.v.Ig treatment increased the ATP-activated NO release by amplifying the Ser1177-eNOS phosphorylation. The i.v.Ig also activated the NO-dependent cGMP release in resting and collagen-stimulated platelets. Since NO is a potent inhibitor of platelet activation and vWF is a platelet adhesion cofactor, the beneficial effects of therapeutic i.v.Ig may lie in the inhibition of platelet adhesion to damaged endothelium.
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PMID:Therapeutic immunoglobulin reduces Ca2+ mobilization and von Willebrand factor secretion, and increases nitric oxide release in human endothelial cells. 1465 35

At diagnosis, there is evidence of increased thrombin generation in children with acute lymphoblastic leukemia (ALL), the etiology of which is unclear. However, thromboembolism (TE) in children with ALL is most commonly reported after the initiation of antileukemic therapy indicating a possible interaction of the disease and therapy. Antileukemic therapy influences the haemostatic system either by direct effect of the chemotherapeutic agents or indirectly through the effect of supportive care, e.g. central venous line (CVL) or infectious complications secondary to immunosuppression. Asparaginase and steroids are shown to induce hypercoagulable state by suppression of natural anticoagulants, especially AT and plasminogen, and by elevations in F VIII/vWF complex, respectively. In addition, steroid therapy causes hypofibrinolytic state by dose-dependent increase in plasminogen activator inhibitor 1 (PAI-1) levels. Combination of these effects coupled with increased thrombin generation may be responsible for the increased incidence of TE observed with concomitant administration of asparaginase and steroids. Further studies to delineate the mechanism of increased thrombin in generation children with ALL and effects of various chemotherapeutic agents, in isolation and in combination, on haemostatic system are needed.
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PMID:Thrombosis in children with acute lymphoblastic leukemia. Part II. Pathogenesis of thrombosis in children with acute lymphoblastic leukemia: effects of the disease and therapy. 1469 64

Virchow rightly recognised that blood flow plays an important role in thrombosis. The roles of blood flow in haemostasis, and in arterial, intra-cardiac, and venous thrombosis are reviewed. In streamline (laminar) flow, shear stresses are maximal at the vessel wall, and affect endothelial cell morphology and function (e.g. secretion of NO, prostacyclin,t-PA and vWF). Platelets are also concentrated at the vessel wall (due to axial concentration of red cells)where they can be activated by high shear stresses and are well-placed to interact with vWF and subendothelium,resulting in platelet adhesion and the initial stages of haemostasis. On the other hand, increasing wall shear forces increase removal of thrombin and fibrin monomer, hence stasis (induced by internal or external pressure) is required to allow fibrin formation and secondary haemostasis. Atherogenesis occurs in areas of arterial flow separation,which promotes platelet, leucocyte, LDL and fibrinogen adhesion and wall infiltration. Rheological variables (e.g. wall shear stress, viscosity, haematocrit,fibrinogen, LDL) have been correlated with the extent of ultrasonic carotid intima-media thickening. Arterial thrombosis usually follows rupture of atherosclerotic plaques and intra-plaque haemorrhage: high intra-stenotic shear stresses may activate platelets,promoting the initial platelet-rich "white-head" of arterial thrombi, while low post-stenotic shear stresses may promote the subsequent, fibrin--and red cell-rich "red tail". Blood viscosity, platelet microemboli, and activated leucocytes may each reduce post-stenotic microcirculatory blood flow, promoting infarction. Such mechanisms may explain the associations of increased levels of blood and plasma viscosity, haematocrit, white cell count, fibrinogen and vWF with risk and outcome of myocardial, cerebral and limb infarction. Areas of recirculating blood flow under low shear stresses predispose to intracardiac thromboembolism(e.g. atrial fibrillation, in which elevated fibrin D-dimer levels are normalised after cardioversion) and venous thromboembolism (fibrin D-dimer levels are associated with most risk factors). There is good evidence that reduction of venous stasis in the legs reduces the risk of venous thromboembolism. There is increasing evidence that regular exercise and avoidance of immobility reduces the risk of both arterial and venous thrombosis and also has systemic antithrombotic and anti-inflammatory effects. So: "Go with the flow!"
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PMID:Virchow's triad revisited: abnormal flow. 1569 60

Three-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors protect the vasculature from inflammation and atherosclerosis by cholesterol dependent and cholesterol independent mechanisms. We hypothesized that HMG-CoA reductase inhibitors decrease exocytosis of Weibel-Palade bodies, endothelial cell granules whose contents promote thrombosis and vascular inflammation. We pretreated human aortic endothelial cells with simvastatin for 24 hours, then stimulated the cells with thrombin, and measured the amount of vWF released into the media. We then measured the effect of simvastatin on myocardial infarction in mice. Simvastatin decreased thrombin-stimulated Weibel-Palade body exocytosis by 89%. Simvastatin inhibited exocytosis in part by increasing synthesis of nitric oxide (NO), which S-nitrosylated N-ethylmaleimide sensitive factor (NSF), a critical regulator of exocytosis. Simvastatin treatment attenuated myocardial infarct size by 58% in wild-type but not eNOS knockout mice. Furthermore, simvastatin decreased endothelial exocytosis and neutrophil infiltration into ischemic-reperfused myocardium, which was mediated in part by P-selectin contained in Weibel-Palade bodies. However, simvastatin did not affect exocytosis and inflammation in myocardial infarcts of eNOS knockout mice. Inhibition of endothelial exocytosis is a novel mechanism by which HMG-CoA reductase inhibitors may reduce vascular inflammation, inhibit thrombosis, and protect the ischemic myocardium. These findings may explain part of the pleiotropic effects of statin therapy for patients with cardiovascular disease.
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PMID:HMG-CoA reductase inhibitors inhibit endothelial exocytosis and decrease myocardial infarct size. 1590 63

Hemostatic serine proteinases-thrombin, Factor VIIa, Factor Xa, play the central role in blood coagulation and thrombosis. Activation of coagulation and generation of active proteinases is initiated by tissue factor (TF) that is expressed by cells of the innate immune system and endothelial cells after tissue damage and cell activation induced by trauma, infection, hypoxia and other cell injury. Coagulation and inflammation are the essential part of the defensive host response. These processes have several connecting points account for the associate and/or the interaction between coagulation and inflammation pathways. The first link between these processes is endothelium, which after damage expresses the adhesive proteins (vWF,P-selectin), inductors and receptors, involved in both coagulation and inflammation. The second link is platelets, which stored in and after activation release proteins with procoagulant and proinflammatory properties. The third link is the serine proteinases, which produced for blood coagulation and activate via its specifical receptors--PARs (proteinase activated receptors) the cells of both coagulation and inflammation system thereby controlling these processes. The generation of these proteinases is initiated by tissue factor (TF) which triggers blood coagulation at sites of tissue injury by selective binding of FVIIa. TF/VIIa complexes with substrate--FX that is activated to FXa. TF/VIIa/Xa can activate both the inflammatory responses of endothelial and other cells and also blood coagulation through stimulation of thrombin generation. This review summarizes the latest data on the blood coagulation activation that include generation of active surface for coagulation, generation of hemostatic serine proteinases and its role as signalling molecules that via PARs and other receptors involved in regulation and control of the interaction of blood coagulation and inflammation and illustrates the potential for therapeutic intervention.
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PMID:Blood coagulation-dependent inflammation. Coagulation-dependent inflammation and inflammation-dependent thrombosis. 1614 14


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