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)

By using a "slow" fluorogenic thrombin substrate and continuous comparison to a simultaneously run calibrator, thrombin generation can be monitored automatically, on line, in clotting PPP or PRP at a throughput of up to 100 samples per hour. The resulting "Thrombogram" in PPP measures hypocoagulability (haemophilias, oral anticoagulants, heparins (-likes), direct inhibitors) and hypercoagulabilities (AT deficiency, prothrombin hyperexpression, prot. C and S deficiency, factor V Leiden, oral contraceptives). In PRP it is diminished in thrombopathies, in von Willebrand disease, by antibodies blocking GPIIb-IIIa or GPIb, or by antiplatelet drugs like aspirin and clopidogrel. Lupus anticoagulant both retards and increases thrombin generation. The thrombogram thus appears to be a broad function test of the haemostatic-thrombotic mechanism of the blood.
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PMID:The calibrated automated thrombogram (CAT): a universal routine test for hyper- and hypocoagulability. 1367 51

Bites by the Indian cobra (Naja naja naja) are common in India and Sri Lanka because of its close association with humans. Cobra venoms are complex and contain several toxic components, including neurotoxins that cause post-synaptic neuromuscular blockade with respiratory paralysis and even death. Bites may also cause extensive local necrosis by mechanisms not fully elucidated. Although no significant coagulopathy has been reported, N.n. naja venom can form blood clots in vitro by activating prothrombin as demonstrated by thrombin-specific chromogenic substrate. Scanning electron microscopy demonstrates that the clots formed by venom lack the thin fibrin strands of normal blood clots formed by thromboplastin or glass contact. Rheometry shows that clots formed by venom have abnormally low elasticity over an extended period and then, as the platelets contract, a retarded and more feeble increase in elasticity. Purified N.n. naja venom PLA2 inhibits platelet aggregation in PRP and explains the decreased clot retraction and retarded and compromised elasticity build up. The present study shows that the PLA2 and the prothrombin activator from N.n. naja venom have effects on haemostasis and blood clotting, although such effects are not observed systemically in envenomed humans.
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PMID:In vitro procoagulant and anticoagulant properties of Naja naja naja venom. 1455 74

Defective prothrombin consumption has been reported in the proband case of Bernard-Soulier syndrome (BSS). There is no consensus, however, on whether the formation of platelet procoagulant activity (PPA) is impaired in BSS and, if so, whether this is due to the lack of GPIb-V-IX-dependent binding of thrombin or of von Willebrand factor (VWF). We show thrombin generation (TG) in platelet-rich plasma of BSS (BSS-PRP) to be defective provided that fibrin remains present in the reaction mixture and that the giant platelets are not damaged by frequent subsampling. In BSS-PRP addition of (thrombin-free) fibrin did not increase TG as in normal PRP, supporting our previous hypothesis that the interaction of fibrin, VWF and GPIb triggers PPA development. Fibrin formed during the lag phase of TG by a snake venom enzyme which only removed fibrinopeptide A induced an immediate burst of TG, that was inhibited by a monoclonal antibody against GPIb (6D1) that abolishes ristocetin-induced binding of VWF to platelets. Inversely, inhibition of polymerization decreased TG and the residual activity was insensitive to 6D1. We conclude that polymerizing fibrin interacts with VWF so as to activate GPIb.
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PMID:Fibrin polymerization is crucial for thrombin generation in platelet-rich plasma in a VWF-GPIb-dependent process, defective in Bernard-Soulier syndrome. 1471 81

The serine protease, thrombin, plays a crucial role in both coagulation and platelet activation. Anhydrothrombin (AhT) is a catalytically inactive derivative of thrombin in which dehydroalanine replaces the active-site serine. AhT retains affinity for natural substrates of thrombin and may be a competitive inhibitor of thrombin-mediated coagulation and platelet reactions. In the present study, thrombelastography showed that AhT not only delayed the onset and the progress of the coagulation process but impaired clot strength, indicating that AhT may have both anticoagulant and antiplatelet activity. In addition, AhT prolonged the activated partial thromboplastin time dose-dependently, but had little effect on the prothrombin time, suggesting that its principal activity was mediated in the intrinsic coagulation pathway. AhT inhibited thrombin-induced aggregation of platelet-rich plasma. Complete inhibition of aggregation was evident at a concentration of 1.85 microM AhT. Furthermore, 3.7 microM of AhT almost completely abolished shear-induced platelet aggregation in PRP. Interpretation of this in vitro study requires confirmation in vivo, but the findings suggest that thrombin plays a critical role in shear related platelet mechanisms. AhT may be a useful tool for investigating platelet-based coagulation reactions and may provide the basis for a novel class of antithrombotic agents.
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PMID:A critical role for thrombin in platelet aggregation under high shear stress. 1518 43

Blood platelets become activated and aggregate at the site of vessel injury. Upon activation by thrombin, platelets release storage pools of proteins and growth factors (GFs), including those involved in tissue repair. Our goal was to evaluate the potential beneficial effect of proteins released from platelet-rich clots on tendon healing. PDGF, TGF-beta-1, IGF-I, HGF, VEGF and EGF were measured in human platelet-poor plasma (PPP) and in the releasates collected from either platelet-poor or platelet-rich clots prepared in vitro. We then studied the effects of the releasates on human tendon cells in culture. Releasates from both platelet-rich and platelet-poor clots stimulated tendon cell proliferation, in contrast to un-clotted PPP. The mitogenic activity of the supernatants was not decreased by the thrombin inhibitor, hirudin. Cultured tendon cells synthesise VEGF and HGF in the presence of PPP-clots and PRP-clot releasates, thus the synthesised amount was significantly higher with supernatants from platelet-rich clots than supernatants from a platelet-poor clot (p < 0.05). These results suggest that administering autologous platelet-rich clots may be beneficial to the treatment of tendon injuries by inducing cell proliferation and promoting the synthesis of angiogenic factors during the healing process.
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PMID:Autologous preparations rich in growth factors promote proliferation and induce VEGF and HGF production by human tendon cells in culture. 1577 47

Activated platelets participate in arterial thrombosis by forming aggregates and potentiating the coagulation through exposure of procoagulant phosphatidylserine. The function of the two receptors for ADP, P2Y(1) and P2Y(12), is well-established in aggregation, but is incompletely understood in the platelet procoagulant response. We established that, in PRP from healthy subjects, ADP accelerated and potentiated tissue factor induced thrombin generation exclusively via stimulation of P2Y(12) and not via P2Y(1) receptors. The P2Y(12) receptors also mediated the potentiating effect of PAR-1 stimulation on thrombin generation. Furthermore, ADP enhanced in a P2Y(12)-dependent manner the Ca(2+) response induced by thrombin, which was either added externally or generated in-situ. This ADP effect was in part dependent of phosphoinositide 3-kinase and was paralleled by increased phosphatidylserine exposure. In PRP from (young) patients with either stroke or type-II diabetes, platelet-dependent thrombin generation was similarly enhanced byADP or SFLLRN as in healthy subjects. In PRP from stroke patients of older age, the P2Y(12)-mediated contribution to thrombin generation was variably reduced by two weeks of clopidogrel medication. Remaining P2Y(12) activity after medication correlated with remaining P2Y(12)-dependent P-selectin exposure, i.e. Ca(2+)-dependent secretion, likely due to incomplete antagonism of P2Y(12) receptors. Together, these results indicate that physiological platelet agonists amplify phosphatidylserine exposure and subsequent thrombin generation by release of ADP and P2Y(12)-receptor stimulation. This P2Y(12) response is accomplished by a novel Ca(2+) signalling pathway. It is similarly active in platelets from control subjects and patients at thrombotic risk. Finally, the thrombogram method is useful for measuring incomplete P2Y(12) inhibition with clopidogrel.
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PMID:Platelet P2Y12 receptors enhance signalling towards procoagulant activity and thrombin generation. A study with healthy subjects and patients at thrombotic risk. 1596 99

Nonhealing diabetic foot ulcers are a common cause of amputation. Emerging cellular therapies such as platelet-rich plasma gel provide ulcer management options to avoid loss of limb. The purpose of this prospective, randomized, controlled, blinded, multicenter clinical study was to evaluate the safety and efficacy of autologous platelet-rich plasma gel for the treatment of nonhealing diabetic foot ulcers. One hundred, twenty-nine (129) patients were screened; 72 completed a 7-day screening period and met the study inclusion criteria. Patients were randomized into two groups - the standard care with platelet-rich plasma gel or control (saline gel) dressing group - and evaluated biweekly for 12 weeks or until healing. Healing was confirmed 1 week following closure and monitored for another 11 weeks. An independent audit led to the exclusion of 32 patients from the final per-protocol analysis because of protocol violations and failure to complete treatment. In this group, 13 out of 19 (68.4%) of the platelet-rich plasma gel and nine out of 21 (42.9%) of the control wounds healed. After adjusting for wound size outliers (n = 5), significantly more platelet-rich plasma gel (13 out of 16, 81.3%) than control gel (eight out of 19, 42.1%) treated wounds healed (P = 0.036, Fisher's exact test). Kaplan-Meier time-to-healing also was significantly different between groups (log-rank, P = 0.0177). No treatment-related serious adverse events were reported and bovine thrombin used in the preparation of PRP did not cause Factor V inhibition. When used with good standards of care, the majority of nonhealing diabetic foot ulcers treated with autologous platelet-rich plasma gel can be expected to heal.
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PMID:A prospective, randomized, controlled trial of autologous platelet-rich plasma gel for the treatment of diabetic foot ulcers. 1679 84

Three commercial systems for whole blood separation were compared to obtain the buffy coat composed of platelet-rich plasma (BC-PRP) and leucocytes . These samples of the buffy coat were used to make a platelet gel (PG), which was used to measure platelet growth factor (PGF) release, to perform a white blood cell (WBC) count and to measure myeloperoxidase (MPO) release from WBCs. Aliquots of whole blood obtained from ten volunteers were distributed either to a blood cell separator (The Electa Cell-Separator, E-CS) or to a tabletop centrifuge (Gravitational Platelet Sequestration System, GPS) to prepare the BC-PRP. The third system combines the BC-PRP production by E-CS with a micro porous filter (Autologous Growth Factor filter, AGF) to enrich for the BC-PRP. Autologous thrombin was used to activate the BC-PRP and to prepare the PG and subsequently to degranulate the platelet concentrate. Platelet-derived growth factor-AB and transforming growth factor-beta1 were present in high levels after thrombin activation of the E-CS or GPS prepared samples. However, the AGF prepared samples released their growth factors before thrombin activation. The WBCs were significantly increased with each of the three systems. Contrary to the AGF, no leucocyte degranulation occurred with the E-CS or GPS prepared samples, based upon the low MPO concentrations in the BC-PRP. The three types of apparatus had different harvesting capacities for collecting the enriched platelets and the release of high concentrations of PGF. When the E-CS and GPS, but not the AGF, were used, low levels of MPO were maintained in the PG, which potentially contributes to antimicrobial properties of platelet gel at the site of application.
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PMID:Differences in platelet growth factor release and leucocyte kinetics during autologous platelet gel formation. 1699 60

Patients with haemophilia requires different amounts of FVIII to prevent and treat bleeds. We hypothesise that this is because FVIII has variable effects on individual patients' global haemostasis. Twelve patients with severe haemophilia A were infused with 50 IU/kg FVIII and thrombin generation in platelet rich (PRP) and platelet poor plasma (PPP) and velocity of changing clot elasticity were measured preinfusion and at nine subsequent time points over 72 h. Despite a close correlation between median FVIII and median initial rate of thrombin generation (R(2) 0.94), endogenous thrombin potential (ETP; R(2) 0.94) and peak thrombin (R(2) 0.91) in PPP, there was wide inter-patient variability at each time point. There was, however, a highly predictable intra-patient relationship between FVIII level and thrombin generation. Inter-patient variability was due to both differences in FVIII levels and the variable effect FVIII had on an individuals' thrombin generation. The utility of PRP was limited because, at low-FVIII levels, only rate of thrombin generation was measurable. At low-FVIII concentrations, the rate of thrombin generation in PPP was the most useful test whilst at higher levels ETP and peak thrombin could also be used.
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PMID:Measurement of global haemostasis in severe haemophilia A following factor VIII infusion. 1767 84

We have developed a whole blood thrombin generation (TG) assay whereby TG is initiated with a low-tissue factor concentration and monitored using a fluorogenic thrombin substrate. Significantly higher values were found in blood samples from 50 patients with a history of venous thromboembolism (VTE) compared with 31 healthy controls (HC), for peak height (P = 0.0034) and endogenous thrombin potential (ETP) (P = 0.0027). Results from 31 VTE patients and the 31 controls in the absence of corn trypsin inhibitor (CTI) showed significantly higher values in the VTE group for peak height (P = 0.0013) and ETP (P = 0.002). In the presence of CTI, significantly higher values were only seen in ETP (P = 0.024). No significant increases in TG were found using platelet poor (PPP) or -rich (PRP) plasma with or without CTI. The whole blood TG assay in the absence or presence of CTI showed a higher proportion (25/50 and 12/31, respectively) of raised peak height and/or ETP values than plasma assays (PPP 9/50 and 5/31 respectively and PRP 13/50 and 6/31, respectively). Our results show the whole blood TG assay is more sensitive for determining the increases in TG in patients with a history of VTE than PPP and PRP TG assays.
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PMID:Thrombin generation: a comparison of assays using platelet-poor and -rich plasma and whole blood samples from healthy controls and patients with a history of venous thromboembolism. 1844 88


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