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)

Removal of N-acetylneuraminic acid from the platelet surface causes rapid removal of platelets from the circulation but causes little change in other platelet functions. We have now investigated the effects of sodium periodate which is thought to oxidize the sialic acid of glycoproteins on cell surfaces and has been shown to affect the functions of other cells. NaIO4 (1 to 10 mm) caused aggregation of stirred suspensions of washed platelets from rabbits. Calcium was required in the suspending medium for NaIO4-induced aggregation. Aggregation was not accompanied by the release of amine storage granule contents nor by cell lysis. Aggregation induced by NaIO4 was not inhibited by creatine phosphate-creatine phosphokinase, by platelet inhibitors that raise platelet cyclic AMP levels such as prostaglandin E1 or methylxanthines, by agents that modify platelet surface--SH groups (N-ethylmaleimide, p-chloromercuribenzene sulfonate), nor by cytochalasin B and/or colchicine which interfere with platelet contractile processes. Drugs such as acetylsalicyclic acid, penicillin G, or cephalothin had no effect on NaIO4-induced aggregation. NaIO4-induced aggregation was practically independent of platelet metabolism since it was not affected by low temperatures and was only slightly inhibited by a combination of antimycin and iodoacetate. Periodate treatment enhanced CO2 production by platelets. When rabbit platelets were pretreated, without stirring, with NaIO4 (0.01 to 1 mm), they did not aggregate. They retained their disc shape and granule contents. However, this pretreatment with NaIO4 inhibited aggregation induced by ADP and inhibited both aggregation and release induced by collagen, thrombin, arachidonic acid, and the ionophore A23,187. The extent of inhibition corresponded to the concentration of NaIO4 used to pretreat the platelets. In contrast, concanavalin A-induced aggregation was unchanged by NaIO4 pretreatment. When NaIO4 oxidation was followed by sodium borohydride (NaBH4) reduction, the effects caused by NaIO4 pretreatment on ADP-induced aggregation and collagen- or thrombin-induced aggregation and release were partially reversed. Pretreatment with NaIO4 also diminished the rate of serotonin uptake and decreased the ability of platelets to adhere to collagen-coated surfaces or to the subendothelial structures of the rabbit aorta. Platelets which had been treated with NaIO4 and then reinfused into rabbits did not survive, and in this way were similar to platelets from which surface sialic acid had been removed by neuraminidase treatment. Since NaIO4 has been shown to oxidize sialic acid on red cell membranes, it seems probably that alteration of surface sialic acid resulted in recognition of the periodate-treated platelets as "foreign" by the reticuloendothelial system. When NaIO4 oxidation was followed by NaBH4 reduction, platelet survival returned toward normal values.
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PMID:Effects of sodium periodate on platelet functions. 17 61

We have observed that treatment of rabbit synovial fibroblasts with proteolytic enzymes can induce secretion of collagenase (EC 3.4.24.7) and plasminogen activator (EC 3.4.21.-). Cells treated for 2-24 hr with plasmin, trypsin, chymotrypsin, pancreatic elastase, papain, bromelain, thermolysin, or alpha-protease but not with thrombin or neuraminidase secreted detectable amounts of collagenase within 16-48 hr. Treatment of fibroblasts with trypsin also induced secretion of plasminogen activator. Proteases initiated secretion of collagenase (up to 20 units per 10(6) cells per 24 hr) only when treatment produced decreased cell adhesion. Collagenase production did not depend on continued presence of proteolytic activity or on subsequent cell adhesion, spreading, or proliferation. Routine subculturing with crude trypsin also induced collagenase secretion by cells. Secretion of collagenase was prevented and normal spreading was obtained if the trypsinized cells were placed into medium containing fetal calf serum. Soybean trypsin inhibitor, alpha(1)-antitrypsin, bovine serum albumin, collagen, and fibronectin did not inhibit collagenase production. Although proteases that induced collagenase secretion also removed surface glycoprotein, the kinetics of induction of cell protease secretion were different from those for removal of fibronectin. Physiological inducers of secretion of collagenase and plasminogen activator by cells have not been identified. These results suggest that extracellular proteases in conjunction with plasma proteins may govern protease secretion by cells.
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PMID:Proteases induce secretion of collagenase and plasminogen activator by fibroblasts. 20 72

We labeled surface glycoproteins of human platelets by the neuraminidase-galactose oxidase/borotritiide and the periodate/borotritiide methods. When labeled platelets were treated with 1-nM thrombin, a minor glycoprotein weighing 68,000-85,000-d was lost from the surface, and a soluble glycoprotein weighing 57,000-68,000-d was found in the supernatant. Treatment of platelets with ADP, collagen, or the calcium ionophore A23187 did not cause loss of the 68,000-85,000-d glycoprotein from platelet surfaces or appearance of the 57,000-68,000-d glycoprotein in the supernatant. However, trace amounts of the intact 68,000-85,000-d glycoprotein were found in the supernatants of platelets that were not treated with thrombin. The numerous effects of thrombin on platelets could be initiated by cleavage and release the thrombin-sensitive glycoprotein.
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PMID:Action of thrombin on surface glycoproteins of human platelets. 21 39

The minimal concentration of the platelet aggregation principle (Platelet Aggregoserpentin, PAS) necessary to induce platelet aggregation was 10 ng/ml, about one-hundredth of that of the crude venom. PAS induced the release of platelet factors 3 and 4 from platelets, but the released platelet factor 3 was easily inactivated by the anti-phospholipid effect of PAS. Pretreatment of platelets with neuraminidase potentiated PAS-induced platelet aggregation. PAS-induced platelet aggregation was independent on released ADP; it could occur in the ADP-removing systems, such as apyrase or a combination of phosphoenolpyruvate and pyruvate kinase. However, PAS-induced platelet aggregation could be inhibited by adenine nucleotides and adenosine. PAS-induced platelet aggregation was inhibited by some anti-inflammatory agents, antimalarial drugs, local anesthetics, antihistamine and smooth muscle relaxants. After deaggregation of PAS-treated platelets, thrombin and sodium arachidonate could further induce platelet aggregation, but ADP and second dose of PAS could not. It is concluded that PAS-induced platelet aggregation is due to prostaglandin synthesis. Recent literatures on the mechanism of platelet aggregation were surveyed and the actions of PAS were discussed.
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PMID:The action mechanism of the purified platelet aggregation principle of Trimeresurus mucrosquamatus venom. 46 15

Human antithrombin III was found to contain covalently linked N-acetylglucosamine, mannose, galactose, and sialic acid in a molar ratio of approximately 1:1:0.6:1. Sialic acid was released upon treatment with neuraminidase. The modified glycoprotein retained the capability to inhibit thrombin and to bind with heparin. Antithrombin III isolated by different procedures was also found to contain glucose in an approximately equimolar ratio with N-acetylglucosamine. Th" glucose-containing component was extractable with lipid solvents and shown to be beta-glucosylceramide. This glycolipid is tightly complexed with antithrombin III and could not be separated by fractional precipitations or ion exchange gels. Although it remains to be established whether the inhibitory actions of antithrombin III are affected by glucosylceramide, the relative amounts which are bound suggest that antithrombin III may be a significant carrier of the glycolipid.
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PMID:Human antithrombin III. Carbohydrate components and associated glycolipid. 61 63

Treatment of human platelets with purified bovine Factor VIII caused three types of aggregation: (a) primary agglutination; (b) secondary aggregation involving the platelet release reaction; and (c) super-aggregation, in which the platelets were gathered into only a few large clumps. Removal of calcium ions or treatment with p-hydroxymercuiriphenyl sulfonate blocked the release reaction, but not primary agglutination or super-aggregation. Platelets treated with formalin were not aggregated by ADP, thrombin, or collagen, but were agglutinated by bovine Factor VIII, although they did not show super-aggregation. For malin-treated platelets were agglutinated by phytohemagglutinin P less extensively and less rapidly than by bovine Factor VIII. Treatment of platelets and Factor VIII with neuraminidase released 60 and 53%, respectively, of the sialic acid residues without affecting the agglutination reaction or the procoagulant activity of the Factor VIII. Agglutination was inhibited by high salt concentrations, dextran sulfate, and heparin. During agglutination, both the procoagulant and platelet-agglutinating activities of Factor VIII became bound to the platelet surface.
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PMID:The interaction of bovine factor VIII with human platelets. 80

The role of sialic acid in the functional and metabolic properties of purified human fibrinogen was investigated. Fibrinogen treated with Vibrio cholerae neuraminidase released 90 percent of its sialic acid without evidence of proteolysis, as indicated by the presence of intace A alpha, B beta, and gamma chains on sodium dodecylsulfate (SDS)-polyacrylamide gels of the reduced asialoprotein. The thrombin and Reptilase clotting times of human asialofibrinogen were shortened compared to those of normal fibrinogen. Fibrinopeptide release was normal in rate and amount, but asialofibrin monomer aggregation was increased at both low and high ionic strength. Similarly, the asialo-derivative of fibrinogen Philadelphia (functionally characterized by impairment of fibrin monomer aggregation) demonstrated shortening of its thrombin and Reptilase times and improvement in its monomer aggregation especially at high ionic strength. Asialofibrin showed a normal capacity to form cross-linked fibrin as demonstrated by normal gamma-chain dimerization and alpha-chain polymerization. Simultaneous metabolic studies of human normal fibrinogen and asialofibrinogen in rabbits revealed only a modest decrease in the half-life of the asialoprotein compared to the intact protein, with no preferential uptake of the asialo-derivative by the liver. Control studies with rabbit normal fibrinogen and asialofibrinogen in rabbits revealed the same modest difference in half-life. Thus, asialofibrinogen clots faster due to enhancement of its monomer aggregation, has a normal capacity to form cross-linked fibrin, and does not differ significantly in its metabolic properties from normal fibrinogen. The possible influence of sialic acid in the functional abnormality of some congenital dysfibrinogenemias is discussed.
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PMID:Functional and metabolic properties of human asialofibrinogen. 83 73

Lactoperoxidase-catalyzed 125I iodination and sodium dodecyl sulphate-polyacrylamide gel electrophoresis have been performed on whole, washed platelets as well as on isolated platelet membranes and granules. Electrophoresis of the whole platelets demonstrated two major radioactive peaks, corresponding to glycopolypeptides of estimated molecular weights of 120 000 and 100 000. A small, but consistent amount of radioactivity was also associated with a 147 000 dalton glycopolypeptide. The membranes showed the same pattern of radioactivity as the whole platelets, whereas only negligible amounts of labeled material was found in the soluble and granule fractions. Practically all the polypeptides were labeled in membranes iodinated after their isolation. A glycopolypeptide of 147 000 molecular weight was observed also in the soluble and the granule fractions, but no radioactivity was associated with these substances. In unreduced form, the granule glycopolypeptide penetrated only slightly into the polyacrylamide gel. Thrombin induced the relase of this granule-located substance from whole platelets, as observed by gel electrophoresis of the supernatant after release reaction (secretion). The granule glycoproteins were only partly exposed on the granule membrane since about 50% of the acid-hydrolyzable sialic acid could be liberated by neuraminidase treatment of isolated granules. In whole, iodinated granules the bulk of the radioactivity was associated with a polypeptide of estimated molecular weight 46 000 (possibly actin). This polypeptide was not seen in the supernatant after removal of the thrombin-degranulated platelets by centrifugation, which indicates that the granule membrane is retained with the platelets during the secretion process.
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PMID:Studies on subcellular fractions of human platelets by the lactoperoxidase-iodination technique. 99 Mar 26

A number of investigators have implicated sialic acid on the surface of platelets in platelet function. In this study we have quantitated the amount of sialic acid removed by purified neuraminidase from the surface of washed platelets of man, rabbit, or pig and examined the effects of this removal. Purified neuraminidase did not induce the release of platelet granule contents. Platelets were pre-labeled with 14C-serotonin for measurement of the release reaction or with 51Cr for determination of adherence to a collagen-coated surface or damaged aortic surface, and for in vivo platelet survival studies. Washed, neuraminidase-treated platelets were resuspended in Tyrode's solution containing 0.35 per cent albumin or in citrated platelet-free plasma from the same species. Both resuspending fluids contained apyrase. Aggregating agents tested were ADP, acid-soluble collagen, thrombin, ristocetin (with human platelets), polylysine, and serotonin (with rabbit platelets). With all of these agents except polylysine, aggregation of neuraminidase-treated human or rabbit platelets was slightly enhanced compared with control platelets; aggregation of pig platelets was unchanged. When release-inducing agents were used, neuraminidase-treated platelets released more of their 14C-serotonin than control platelets. The extent to which rabbit platelets adhered to a collagen-coated surface or to the damaged surface of everted rabbit aorta was unchanged by pretreatment of platelets with neuraminidase. Therefore it seems unlikely that sialic acid is involved in platelet adherence to collagen. When more than 15 per cent of total sialic acid had been removed from rabbit platelets, they were completely cleared from the circulation within 1 hour of their injection into rabbits. When 8 to 10 per cent of total sialic acid had been removed, the platelets were not cleared immediately from the circulation but were cleared more quickly than control platelets. Thus, although removal of up to 65 per cent of platelet sialic acid has only a slightly enhancing effect on platelet aggregation and release in vitro, removal of as little as 8 to 10 per cent results in the recognition of platelets as "foreign" in vivo.
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PMID:Effects on platelet function of removal of platelet sialic acid by neuraminidase. 112 70

Previous studies have shown that thrombin-activated platelets interact through the P-selectin with neutrophils and monocytes. To identify other types of leukocytes capable of such an interaction, eosinophils, basophils, and lymphocytes were isolated from whole blood. Binding of these cells to activated platelets was examined in a double immunofluorescence assay and the results show that activated platelets not only bind to neutrophils and monocytes, but also to eosinophils, basophils, and subpopulations of T lymphocytes. Using monoclonal antibodies (MoAbs) specific for subsets of T cells, we could further demonstrate that the T cells which bind activated platelets are natural killer (NK) cells and an undefined subpopulation of CD4+ and CD8+ cells. All these interactions were dependent on divalent cations and were completely inhibited by an MoAb against P-selectin. Thus, P-selectin mediates the binding of activated platelets to many different types of leukocytes. Studies with leukocytes treated with proteases or neuraminidase have shown that the structures recognized by P-selectin are glycoproteins carrying sialic acid residues. Because the loss of binding of activated platelets to neuraminidase-treated neutrophils was almost complete, but only partial to treated eosinophils, basophils, and monocytes, the latter cell types may have different P-selectin ligands in addition to those present on neutrophils. We found that two previously identified ligands for P-selectin, the oligosaccharides Le(x) and sialyl-Le(x), had little or no inhibitory effect on adhesion of activated platelets to leukocytes and that binding was not inhibited by MoAbs against these oligosaccharides. In addition, there was no correlation between the expression of Le(x) on several cell types and their capacity to bind activated platelets. In contrast, the expression of sialyl-Le(x) on cells was almost perfectly correlated with their ability to bind activated platelets. Thus, while Le(x) cannot be a major ligand for P-selectin, a possible role for sialyl-Le(x) in P-selectin-mediated adhesion processes cannot be dismissed. Finally, activated platelets were found to bind normally to monocytes and neutrophils of patients with paroxysmal nocturnal hemoglobulinuria (PNH) and to neutrophils from which phosphatidyl inositol (PI)-linked proteins had been removed by glycosylphosphatidyl inositol-specific phospholipase C (GPI-PLC) digestion. This suggests that at least part of the P-selectin ligands on these cells are not GPI-anchored.
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PMID:P-selectin mediates Ca(2+)-dependent adhesion of activated platelets to many different types of leukocytes: detection by flow cytometry. 137 47


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