Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.5 (thrombin)
33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

ATP and ADP are simultaneously released from activated platelets in equimolar concentrations. Micromolar concentrations of ATP inhibit platelet aggregation by both competitive and non-competitive mechanisms. The current studies addressed the question of how platelets respond to agonists in the presence of nanomolar and micromolar concentrations of ATP and ADP alone or in combination. This is a significant issue since the concentration of ATP +/- ADP may vary widely within a microenvironment depending upon the source and cause for the release of the nucleotides. ATP (1-10 nM) was found to significantly enhance the thromboxane A2 analog, U44619-, collagen- and thrombin-induced platelet aggregations. Conversely, ATP at 1-100 microM inhibited these same reactions. ADP, in general, behaved exactly opposite to ATP. When equal amounts of ATP and ADP were added together the ADP response appeared to predominate. The observed ATP-induced response was not due to a hydrolytic product as evidenced by an unaltered response to ATP in the presence of adenosine deaminase or the ATP generating system, creatine phosphate plus creatine phosphokinase. Adenosine (1-10 nM), like ADP, inhibited agonist-induced platelet aggregation. The stimulation of agonist-induced platelet aggregation by 1-10 nM extracellular ATP appears to depend upon the phosphorylation of platelet membrane ecto proteins. The ATP analog, beta gamma-methylene ATP, that is incapable of serving as a phosphate donor for protein kinases, inhibited rather than stimulated agonist-induced platelet aggregation. The dual response of platelets to low and high concentrations of extracellular ATP +/- ADP may play a physiological role in hemostasis and thrombosis under normal and pathological conditions.
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PMID:A possible dual physiological role of extracellular ATP in the modulation of platelet aggregation. 904 33

von Willebrand factor (vWF) is stored and released from endothelial secretory granules called Weibel-Palade (WP) bodies. Acute release can be induced by thrombin, histamine, and other mediators of thrombosis or inflammation. Their effect is thought to be mediated by an increase in intracellular free calcium ([Ca2+]i). Purine nucleotides such as adenosine triphosphate (ATP) and adenosine diphosphate (ADP) are released from platelet dense granules and from ischemic tissues and are important regulators of platelet function and vascular tone. In the present study, we investigated whether they could also induce exocytosis from cultured endothelial cells. ATP (1 to 100 micromol/L) induced a dose-related increase in vWF release, with a 2.3-fold maximal increase after 30 minutes. Similar responses were observed with ADP. ATP induced calcium mobilization from intracellular stores, an effect mimicked by 2-methylthio-ATP, a selective agonist for P2y receptors. However, 2-methylthio-ATP-induced vWF release was only 43% of the ATP response. ATP-induced vWF release was also associated with a twofold increase in cellular cyclic adenosine monophosphate (cAMP) content, and was potentiated by 3-isobutyl-1-methylxanthine ([IBMX] added to increase cAMP levels by blocking cellular phosphodiesterases) and 8-bromo-cAMP and inhibited by more than 50% by Rp-8-CPT-cAMPS, a competitive protein kinase A inhibitor. Adenosine but not 2-methylthio-ATP mimicked the ATP-induced increase in cAMP. ATP-induced vWF release was partly inhibited by adenosine deaminase, which degrades adenosine generated from ATP in the incubation medium. Adenosine (1 to 100 micromol/L) failed to induce vWF release, but potentiated the secretory response to 2-methylthio-ATP and thrombin without modifying the calcium response to these agents. Our results suggest that ATP/ADP can induce vWF release from endothelial cells via dual activation of P2y and adenosine A2 receptors. ATP/ADP-induced exocytosis could be involved in the regulation of thrombus formation and ischemia-reperfusion injuries. Further, we provide evidence that a receptor-mediated increase in cellular cAMP can potentiate the secretory response to calcium-mobilizing agents.
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PMID:Purine nucleotides induce regulated secretion of von Willebrand factor: involvement of cytosolic Ca2+ and cyclic adenosine monophosphate-dependent signaling in endothelial exocytosis. 941 75

The activation of platelets and the formation of neutrophil- platelet conjugates may lead to the development of thromboemboli. We studied whether blockade of adenosine receptors during coronary hypoperfusion may cause thromboemboli via P-selectin-dependent mechanisms in 30 open-chest dogs. When coronary blood flow was reduced to 20% of the control, it was stable at low levels with increases in adenosine levels. When 8-p-sulfophenyltheophylline, an adenosine receptor antagonist, was infused during coronary hypoperfusion, coronary blood flow decreased gradually and approached almost zero 20 min after its administration. Histological examination revealed thromboemboli in the small coronary vessels. During hypoperfusion in the presence of 8-p-sulfophenyltheophylline, the mAb against P-selectin attenuated both the reduction in coronary blood flow and the formation of thromboemboli, and improved contractile and metabolic dysfunction of the myocardium. Flow cytometric analysis indicated that the expression of P-selectin on platelet and neutrophil-platelet adhesion were increased during coronary hypoperfusion, and that both were further augmented by 8-p-sulfophenyltheophylline. Immunohistochemical examination showed no staining of P-selectin in the ischemic myocardium. Adenosine inhibited the thrombin-induced expression of P-selectin on platelet and neutrophil- platelet adhesion via adenosine A2 receptors. Adenosine appears to inhibit the formation of thromboemboli during coronary hypoperfusion by suppressing the expression of P-selectin on platelets and neutrophil-platelet adhesion.
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PMID:Endogenous adenosine inhibits P-selectin-dependent formation of coronary thromboemboli during hypoperfusion in dogs. 954 94

Proliferation, differentiation, and survival of erythroid progenitor cells are mainly regulated by stem cell factor (SCF) and erythropoietin (Epo). Using normal human progenitors, we analyzed the role of Ca2+-sensitive protein kinase C (PKC) subtypes and of G-protein-coupled receptor ligands on growth factor-dependent DNA synthesis. We show that stimulation of DNA synthesis by the two growth factors requires activation of PKCalpha. Inhibitors of Ca2+-activated PKC subtypes blocked the growth factor-induced 3H-thymidine incorporation. SCF and Epo caused no significant translocation of PKCalpha into the membrane, but treatment of intact cells with either of the two cytokines resulted in enhanced activity of immunoprecipitated cytosolic PKCalpha. Stimulation of PKC with the phorbol ester PMA mimicked the cytokine effect on DNA synthesis. Epo-, SCF-, and PMA-induced thymidine incorporation was potently inhibited by thrombin (half-maximal inhibition with 0.1 U/mL). This effect was mediated via the G-protein-coupled thrombin receptor and the Rho guanosine triphosphatase. Adenosine diphosphate caused a modest Ca2+-dependent stimulation of DNA synthesis in the absence of cytokines and specifically enhanced the effect of SCF. Cyclic 3', 5'-adenosine monophosphate exerted a selective inhibitory effect on Epo-stimulated thymidine incorporation. Our results define PKCalpha as major intermediate effector of cytokine signaling and suggest a role for thrombin in controlling erythroid progenitor proliferation.
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PMID:Erythropoietin- and stem cell factor-induced DNA synthesis in normal human erythroid progenitor cells requires activation of protein kinase Calpha and is strongly inhibited by thrombin. 1038 4

Sulfatides are glycolipid constituents of human platelet cell membranes and have been shown to interact with platelet-binding proteins involved in hemostasis. Because little is known about the physiological role of sulfatides in platelet function, the effect of sulfatide on platelet adhesion, aggregation, release, and ristocetin-induced platelet agglutination (RIPA) was studied. These processes are inhibited when exogenous sulfatide is present in vitro. Inhibition of aggregation induced by collagen, thrombin, and ristocetin by sulfatide was dose dependent. Adenosine diphosphate-mediated adhesion and aggregation were not significantly affected by sulfatide, nor was serotonin- and epinephrine-mediated aggregation. Collagen mediate release of serotonin was reduced sulfatide. RIPA demonstrated dose-dependent inhibition in response to sulfatide. These results suggest that sulfatide may play a role in modulating platelet activation.
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PMID:Inhibition of human platelet function by sulfatides. 1099 94

The influence of adenosine infusion (40 microg/kg/min for 4 h) on inflammatory and hemostatic parameters was investigated in healthy males without (n = 10) or with (n = 11) intravenous endotoxin injection (4 ng/kg). Without endotoxin, adenosine elevated circulating leukocytes and circulating platelet-leukocyte aggregates. Endotoxin activated platelets and leukocytes in vivo. Platelet activation was seen as slightly increased platelet P-selectin expression, decreased platelet counts, and elevated plasma soluble P-selectin (from 39.6 +/- 3.4 to 68.9 +/- 6.6 ng/ml, P<0.01). Leukocyte activation was evidenced by increased CD1 lb expression (from MFI of 0.54 +/- 0.02 to 2.21 +/- 0.17; P<0.01) and plasma elastase levels (from 25.3 +/- 2.5 to 169.3 +/- 22.5 ng/ml: P <0.01). Endotoxin also enhanced platelet and leukocyte responsiveness to in vitro stimulation. Endotoxin induced von Willebrand factor secretion (from 92 +/- 8 units to 265 +/- 19 units at 4 h; P <0.001) and enhanced thrombin generation in vivo. Endotoxin induced leukocytosis and thus increased circulating platelet-leukocyte, mainly platelet-neutrophil, aggregates. Adenosine caused slight attenuation of platelet reactivity to agonist stimulation, enhanced the endotoxin-induced leukocytosis, and detained more platelet-leukocyte aggregates in circulation, but did not attenuate endotoxin-induced neutrophil elastase secretion, von Willebrand factor secretion, or thrombin generation. Thus, endotoxemia induces multi-cellular activation in vivo. Adenosine inhibits leukocyte adhesion and extravasation, and mildly attenuates platelet responsiveness and soluble P-selectin release. Adenosine has the potential of becoming a therapeutic antiinflammatory drug, but an optimal treatment strategy needs to be developed.
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PMID:Multi-cellular activation in vivo by endotoxin in humans--limited protection by adenosine infusion. 1101 59

Prevention and management of acute coronary syndromes (ACS) are focal points of interest among health care providers. Acute coronary syndromes is an all-encompassing term that refers to unstable angina, non-Q wave myocardial infarction, and Q wave myocardial infarction. These syndromes are usually the result of atherosclerotic plaque rupture leading to thrombus formation in a coronary artery. Heparin and aspirin are traditional antithrombotic treatments. They typically are administered with antiischemic therapies and often with fibrinolytic agents for patients with ST segment elevation associated with acute myocardial infarction. Although aspirin and heparin are important, they have significant limitations that have prompted development of newer antithrombotic approaches. Adenosine diphosphate inhibitors have been evaluated as either alternatives or adjunctive treatment to aspirin. Glycoprotein IIb-IIIa receptor inhibitors, low-molecular-weight heparins, and direct thrombin inhibitors have been studied as concurrent therapy with, or as alternatives to, heparin.
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PMID:Clinical use of new antithrombotic therapies for medical management of acute coronary syndromes. 1125 56

The aim of this study was to examine if acute hyperglycemia (an oral glucose tolerance test) activates platelet function, endothelial cells or thrombin generation in diabetic patients and healthy controls. Eleven males with mild type II diabetes mellitus and 11 healthy male volunteers, matched for age and body mass index, were investigated before and after the glucose load. Soluble P-selectin, von Willebrand factor antigen and markers of thrombin generation in plasma were determined by immunoassays, and platelet P-selectin expression (unstimulated and agonist-stimulated) by flow cytometry in whole blood. Acute hyperglycemia elevated plasma soluble P-selectin from 32.5 to 50.9 ng/ml in the diabetic group (P = 0.05) but not in the controls (from 27.3 to 28.8 ng/ml; P = 0.6). Also, soluble P-selectin levels were higher in patients with diabetes than in healthy controls during hyperglycemia, but not in the fasting state. Adenosine diphosphate- and thrombin-induced platelet P-selectin expression was slightly, but significantly, decreased by the glucose load, whereas platelet P-selectin expression in unstimulated samples was not affected. Plasma levels of von Willebrand factor and thrombin generation were similar in patients and controls, and were not altered by hyperglycemia. In conclusion, we found that acute hyperglycemia elevates soluble P-selectin in plasma in males with mild type II diabetes mellitus. Our observation of unaltered plasma levels of the endothelial marker von Willebrand factor is in agreement with platelets being the main source of P-selectin released into plasma following hyperglycemia. Thus, platelets in individuals with type II diabetes may be more susceptible to hyperglycemia than platelets in non-diabetic individuals.
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PMID:Acute hyperglycemia increases soluble P-selectin in male patients with mild diabetes mellitus. 1130 72

The pathogenesis of depressed platelet activity in uremia is still unknown. The influence of some uremic toxins on platelet aggregation (PLA) and prostaglandin metabolism in 50 uremic patients treated by hemodialysis (HD) or continuous ambulatory peritoneal dialysis (CAPD) was studied. Fifty-seven healthy volunteers (HVs) served for reference values. Adenosine diphosphate (ADP) and thrombin (Thr) were used as agonists of PLA. PLA was determined using the Born method. Malonyldialdehyde (MDA) levels in platelets as an indicator of prostaglandin metabolism, after stimulation with arachidonic acid, were measured according to Stuart. The relationship of PLA and prostaglandin metabolism with plasma concentrations of methylguanidine (MG), guanidinosuccinic acid (GSA), and creatinine (Cr) was assessed. PLA-ADP values in regular HD patients (42 +/- 5 mm) were significantly lower than in CAPD patients (65 +/- 8 mm) and HVs (73 +/- 3 mm). PLA-Thr values in HD patients (25 +/- 4 mm) were significantly lower than in CAPD patients (34.9 mm) and HVs (36 +/- 3 mm). MDA levels in HD patients (7 +/- 1 nmol/L/10(9)) were significantly lower than in CAPD patients (12 +/- 2 nmol/L/10(9)) and HVs (15 +/- 1 nmol/L/10(9)). In HD patients, inverse correlations of PLA-ADP with MG levels (r = -0.92), PLA-Thr with Cr levels (r = -89), and MDA levels with GSA levels (r = -0.86) were found. In CAPD patients, no relationship of PLA and MDA with uremic toxins was observed. Depressed activity of platelets and prostaglandin metabolism was strongly expressed in HD patients.
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PMID:Platelet aggregation and prostaglandin metabolism in uremic patients. 1157 34

In vivo platelet activation results are often confounded by activation induced in vitro during the preparative procedures. We measured ex vivo (basal) and in vitro (thrombin-induced) platelet activation in sodium citrate, ethylenediaminetetraacetic acid (EDTA), and Citrate Theophylline Dipyridamole Adenosine (CTAD) whole blood specimens. Determinations were made by measurements of platelet density (mean platelet component: MPC concentration) on the Advia 120 Hematology System. The MPC has been previously shown to correlate with a fluorescence flow cytometric method, also determined in this study, using the surface expression of CD62P. Moreover, platelet shape and structure changes in EDTA and CTAD anticoagulated whole blood specimens were characterized by transmission electron microscopy (TEM). Observations made using the Advia 120 Hematology System platelet density parameter, MPC, in the absence of thrombin were 25.7 +/- 0.9 g/dl, 27.9 +/- 0.9 g/dl and 24.8 +/- 1.2 g/dl in sodium citrate, EDTA and CTAD whole blood specimens, respectively. Addition of thrombin induced a significant change in platelet MPC for sodium citrate (21.9 +/- 1.9 g/dl; p<0.0001) and EDTA (23.2 +/- 0.9 g/dl; p<0.0001) whole blood specimens. In contrast, thrombin had no effect on MPC measured in whole blood taken into CTAD tubes. In vitro fluorescence flow cytometric platelet activation experiments measuring the percentage of platelets expressing anti-CD62P showed increase in sodium citrate specimens from 9.2 +/- 7.0 to 55.5 +/- 23.1 % (p<0.0001) and in EDTA specimens from 1.9 +/- 1.7 to 64.6 +/- 12.4 % (p<0.0001) after addition of thrombin. However, in blood taken into CTAD tubes, there was no significant change. Studies on platelets isolated from whole blood in CTAD showed activation by thrombin indicating that platelets in CTAD, while protected in its presence remained functional upon its removal. When observed by TEM over time, platelets in EDTA appear more activated and contain fewer granules than platelets in CTAD. We conclude that CTAD demonstrates in vitro platelet activation inhibition and may be useful in stabilizing ex vivo platelet activation. The novel platelet activation parameter, MPC, measured by an automated routine hematology system, using customized proprietary software, may be used in conjunction with CTAD, a stabilizing anticoagulant, to measure the ex vivo platelet activation state in whole blood specimens. TEM studies verify shape modifications and simultaneous retention of intracellular granules at early post-venipuncture time periods in CTAD specimens.
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PMID:Assessment of platelet activation in several different anticoagulants by the Advia 120 Hematology System, fluorescence flow cytometry, and electron microscopy. 1459 91


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