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)

Ditazole (4,5-diphenyl-2-bis-(2-hydroxyethyl)-aminoxazol) has been shown to be a strong in vitro inhibitor of human platelet aggregation brought about by release reaction inducers; in contrast, it did not significantly affect primary ADP-induced aggregation. Ditazole strongly inhibited the release of platelet-bound 14C-serotonin under the influence of Thrombofax, whereas it did not interfere with the transport and storage of serotonin in nonstimulated platelets. The effect of ditazole was not potentiated by acetylsalicylic acid. Ditazole also inhibited ADP-reptilase clot retraction and modified thrombin-induced clot formation. The inhibition of platelet aggregation exerted by ditazole in plasma could be removed following gel filtration of platelets on Sepharose 2-B gel. This would indicate that ditazole does not act on platelets by a 'hit and run' mechanism.
 ...
PMID:Ditazole and platelets. I. Effect of ditazole on human platelet function in vitro. 1 12

A haemostatic material suitable for embolization was prepared by the adsorption of haemostatics--ethamsylate and aminocaproic acid in the spherical particles of porous poly(2-hydroxyethyl methacrylate) (p(HEMA)). The degree of purification of ethamsylate-treated particles was tested by an analysis of donor blood in contact with the material. An evaluation of the haemostatic properties of these materials was obtained by the determination of the indicators of blood clotting: activated partial thromboplastin time, thrombin time, and prothrombin time. Ethamsylate or aminocaproic acid-containing p(HEMA) has a distinct haemostatic effect on pathological blood of patients suffering from focal alterations of the liver. These haemostatic emboli materials show promise for the immediate control of various haemorrhages; when introduced into a zone with increased haemorrhage, they may help to correct disturbed haemostasis.
 ...
PMID:Haemostatic activity of ethamsylate and aminocaproic acid adsorbed poly(2-hydroxyethyl methacrylate) particles. 163 25

Recombinant-derived human Factor VIII was labeled intrinsically with [35S]methionine, and its binding to washed human platelets was studied. Binding measurements were performed by incubating Factor VIII and platelets for 15 min at room temperature in Tyrode's solution supplemented with Ca2+ (5.0 mM), 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (5.0 mM), 0.50% bovine serum albumin, and the Factor Xa and thrombin inhibitors 5-dimethylaminonaphthalene-1-sulfonylglutamylglycinylarginyl chloromethyl ketone and 5-dimethylaminonaphthalene-1-sulfonyl-arginine-N-(3-ethyl-1, 5-pentanediyl)amide. Separation of free from bound Factor VIII was accomplished by centrifugation through oil, and nonspecific binding was determined with excess unlabeled Factor VIII. Binding was saturable, reversible, and stimulated 20-fold after platelet activation with thrombin. Furthermore, binding was specific in that bound labeled Factor VIII could be displaced by excess unlabeled Factor VIII, but not by Factor V. Scatchard analysis indicated a single class of binding sites with Kd = 2.9 nM and 450 sites/activated platelet. The time course of displacement indicated a t1/2 of bound Factor VIII of approximately 5 min. When platelets were incubated in Ca2+, both the heavy and light chains of Factor VIII were bound, whereas exposure to EDTA resulted in the binding of the light chain only. These results demonstrate the specific reversible binding of Factor VIII to human platelets, likely mediated through the light chain.
 ...
PMID:The binding of 35S-labeled recombinant factor VIII to activated and unactivated human platelets. 314 7

Trypsin, porcine pancreatic kallikrein, and several blood coagulation enzymes, including bovine thrombin, bovine factor Xa, human factor Xa, human plasma factor XIa, human plasma factor XIIa, and human plasma kallikrein, were inactivated by a number of substituted isocoumarins containing basic functional groups (aminoalkoxy, guanidino, and isothiureidoalkoxy). 3-Alkoxy-4-chloro-7-guanidinoisocoumarins were found to be the most potent inhibitors for the coagulation enzymes tested with kobsd/[I] values in the range of 10(3)-10(5) M-1 s-1. 4-Chloro-3-isothiureidoalkoxyisocoumarins show high inhibitory potency toward porcine pancreatic kallikrein, human plasma kallikrein, human factor XIa, human factor XIIa, and trypsin with kobsd/[I] values of the order of 10(4)-10(5) M-1 s-1. The inhibition of these serine proteases by the substituted isocoumarins are time dependent, and the inactivation of trypsin by 3-alkoxy-4-chloro-7-guanidinoisocoumarins and 7-amino-4-chloro-3-(3-isothiureidopropoxy)isocoumarin occured concurrently with the loss of the isocoumarin absorbance. The complex formed from inactivation of trypsin by these two types of inhibitors was very stable and regained less than 4% activity in 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid buffer (pH 7.5) after 1 day at 25 degrees C and regained 8-45% activity upon addition of buffered 0.29 M hydroxylamine. Trypsin inactivated by other inhibitors regained full activity upon standing or addition of hydroxylamine. Thrombin inactivated by 3-alkoxy-4-chloro-7-guanidinoisocoumarins was also quite stable and only regained 9-15% activity under similar conditions. These results are consistent with a proposed mechanism, where serine proteases inactivated by aminoalkoxyisocoumarins or isothiureidoalkoxyisocoumarins form acyl enzymes that will deacylate upon standing or addition of hydroxylamine. However, the acyl enzymes formed from 3-alkoxy-4-chloro-7-guanidinoisocoumarins or 7-amino-4-chloro-3-(3-isothiureidopropoxy)-isocoumarin will decompose further, probably through a quinone imine methide, to give an irreversibly inactivated enzyme by reaction with an active-site nucleophile such as His-57. The quinone imine methide intermediate may also react with a solvent nucleophile to give an acyl enzyme that can be reactivated by hydroxylamine. The inhibitors 4-chloro-7-guanidino-3-methoxyisocoumarin and 4-chloro-3-ethoxy-7-guanidinoisocoumarin have been tested as anticoagulants in human plasma and were effective at prolonging the prothrombin time. However, they are unstable in plasma (t1/2 = 4-8 min), and their in vivo utility may be limited.
 ...
PMID:Mechanism-based isocoumarin inhibitors for trypsin and blood coagulation serine proteases: new anticoagulants. 316 17

Treatment of human platelets with alpha-thrombin leads to the selective hydrolysis of only one membrane protein, glycoprotein V. To determine whether glycoprotein V was directly cleaved by alpha-thrombin and to permit further characterization of this glycoprotein as the potential functional thrombin receptor, glycoprotein V was purified to > 98% homogeneity. Washed platelets were prepared from concentrates within 18 h of venipuncture, since clinically expired platelets (> 72 h from venipuncture) were shown to contain little or no detectable glycoprotein V. Glycoprotein V was eluted from the platelet membrane by equilibrating the platelets (4 X 10(9)/ml) at 37 degrees C for 24 h in 0.01 M 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (Hepes), pH 7.6 (1 mM in EDTA, 0.3 M in NaCl). Purification was achieved by initially performing ammonium sulfate fractionation, followed by chromatography on Sephacryl S-200, hydroxylapatite, and DEAE- and CM-cellulose, and yielded 0.5 to 1.0 mg of purified glycoprotein per 100 units of platelet concentrate (approximately 6 X 10(12) platelets). Purified glycoprotein V (Mr = 82,000) was a thrombin substrate, and on hydrolysis yielded a major fragment, GPVf1 (Mr = 69,500), identical in molecular weight with that observed previously in the supernatant of thrombin-treated, periodate-labeled platelets. Glycoprotein V existed as at least eight distinct isoelectric forms with pI values ranging from 5.85 +/- 0.05 to 6.55 +/- 0.05. The purified glycoprotein contained approximately 48% carbohydrate by weight, consisting of neutral hexose, hexosamine, and sialic acid in a molar ratio of approximately 8:2:1. Rabbit antiglycoprotein V antibody gave a single precipitin line against purified glycoprotein V, which showed a line of complete identity with Triton-solubilized washed platelets. The availability of purified glycoprotein V and antiglycoprotein V antibody will be useful in delineating the role of this glycoprotein in thrombin activation of platelets.
 ...
PMID:Purification and preliminary physicochemical characterization of human platelet membrane glycoprotein V. 677 67

The permeability of bovine pulmonary artery endothelial (CPAE) monolayers to Evans blue-labelled albumin (Evans blue-albumin) has been measured in vitro. Thrombin caused a concentration-dependent increase in Evans blue-albumin clearance across endothelial monolayers. Isoprenaline inhibited thrombin-induced Evans blue-albumin clearance in a concentration-dependent manner (EC50 21 nM). This effect was mimicked by the selective beta 2-adrenoceptor agonists salbutamol (EC50 64 nM) and salmeterol (EC50 2.7 nM), but not by the selective beta 1-adrenoceptor agonist, RO-363 ((1-[3',4'-dihydroxyphenoxy]-2-hydroxy-[3",4"- dimethoxyphenethylamino]-propane)oxalate), nor by the selective beta 3-adrenoceptor agonist, CL-316,243 (disodium (R,R)-5-[2-[[2-(3-chlorophenyl)-2-hydroxyethyl]-amino]propyl]-1,3- benzodioxole-2,2-dicarboxylate). Isoprenaline, salbutamol and salmeterol, but not RO-363 or CL-316,243 produced small, but significant reductions in Evans blue-albumin clearance across unstimulated endothelial monolayers. Inhibition of the response to thrombin by isoprenaline was antagonised by the selective beta 2-adrenoceptor antagonist, ICI-118,551 ((erythro-DL-1(7-methylindan-4- yloxy)3-isopropylaminobutan-2-ol), pKB 8.4). Salmeterol also inhibited hydrogen peroxide-stimulated Evans blue-albumin clearance. Hence, the widely used beta 2-adrenoceptor agonists, salbutamol and salmeterol, are able to reduce endothelial permeability at nanomolar concentrations.
 ...
PMID:Beta 2-adrenoceptors mediate a reduction in endothelial permeability in vitro. 776 83

Two new polymeric materials (polymers A and B) containing covalently bound iodine were prepared. These polymers were evaluated with respect to their possible use as radiopaque implant biomaterials--that is, materials that are visible in a noninvasive manner using routine X-ray absorption imaging techniques. Polymer A is a copolymer of methyl methacrylate (MMA) and 1 (80 and 20 mol%, respectively). Polymer B was prepared from MMA, 1, and 2-hydroxyethyl methacrylate (HEMA) (mol ratio 65:20:15, respectively). Compound 1 was synthesized from 4-iodophenol and methacryloyl chloride. The resulting polymers were characterized with GPC, DSC, NMR, and by measuring both the advancing and receding contact angles. Thrombogenicity of the polymers was determined by an in vitro thrombin generation test procedure. The maximum concentration of free thrombin was 76 +/- 1 nM for polymer A, and 64 +/- 3 nM for polymer B. The lag times (i.e., time onset of thrombin generation) were 392 seconds for polymer A and 553 seconds for polymer B. For PVC-T, which is known as a passive material, a lag time of 583 seconds was found. This indicates that polymer B is comparable to PVC-T, and more passive than polymer A. Polymer A exhibited minor activation of platelets. Polymer B did not induce platelet activation at all. The polymers exhibited, even as fibers with a diameter of ca. 0.3 mm, good radiopacity with routine imaging X-ray techniques in the clinic.(ABSTRACT TRUNCATED AT 250 WORDS)
 ...
PMID:Studies on two new radiopaque polymeric biomaterials. 782 55

Changes in platelet cytoplasmic free calcium levels were investigated in contact with cast film surfaces of a block copolymer containing 2-hydroxyethyl methacrylate (HEMA) and styrene (St) (0.5 mole fraction HEMA). These copolymer surfaces demonstrate microdomain alternating lamellae structures composed of hydrophilic HEMA domains (5 nm width) and hydrophobic St domains (20 nm width). The results were compared with those obtained from platelets contacting a random copolymer of HEMA-St (0.5 mole fraction HEMA) and from homopolymers of polystyrene (PSt) and poly(2-hydroxyethyl methacrylate) (PHEMA). Cytoplasmic free calcium levels in platelets contacting the microdomain structured surfaces of the HEMA-St block copolymer remained relatively constant in contrast to the significant increases observed for the radically prepared HEMA-St copolymer, PSt, and PHEMA surfaces. Adhering platelets were stimulated by exogenously introduced thrombin and calcium ionophore A23187 20 min after platelet adherence to the polymer surfaces. Only platelets on the block copolymer surfaces showed active metabolic responses. These results suggest that adhering platelets on the microdomain structured surfaces maintain high sensitivities to external stimulation due to an intrinsic strong inhibition of platelet functional changes induced by surface contact.
 ...
PMID:Prevention of changes in platelet cytoplasmic free calcium levels by interaction with 2-hydroxyethyl methacrylate/styrene block copolymer surfaces. 811 39

Four kinds of monomers carrying a thrombin inhibitor, (2R,4R)-4-methyl-1-[N2-[(3-methyl-1,2,3,4-tetrahydro-8-quinolinyl)sulfon yl]-L-arginyl]-2-piperidinecarboxylic acid (argatroban), were synthesized. These monomers were copolymerized with acrylamide to yield water-soluble polymeric conjugates possessing the argatroban moiety in the side chain. Their antithrombogenic activities were determined from the inhibitory effect on thrombin action and the prolongation effect on blood clotting time. The monomeric conjugates of 2-hydroxyethyl acrylate (HEA), 2-hydroxyethyl methacrylate (HEMA), and 4-hydroxybutyl acrylate (HBA) linked with argatroban through an ester bond were potent inhibitors of thrombin, prolonging the blood-clotting time, whereas a conjugate of amino methyl styrene (AMS) and argatroban through an amide bond was a less potent inhibitor than argatroban. None of the copolymers could prolong blood clotting when assessed just after preparation of their aqueous solutions, but the antithrombogenic activity of the aqueous solutions increased after incubation for 7 days at 37 degrees C for the polymeric conjugates through an ester bond. Free argatroban was detected in the aqueous solutions of polymeric conjugates after incubation, suggesting that argatroban was released by hydrolysis of the ester bond during incubation.
 ...
PMID:Synthesis of monomeric and polymeric conjugates carrying a thrombin inhibitor through an ester bond. 949 24

A bifunctional derivative of the thrombin-binding aptamer with a redox-active Fc moiety and a thiol group at the termini of the aptamer strand was synthesized. The ferrocene-labeled aptamer thiol was self-assembled through S-Au bonding on a polycrystalline gold electrode surface and the surface was blocked with 2-mercaptoethanol to form a mixed monolayer. By use of a fluorescent molecular beacon, the effect of counterions on quadruplex formation was established. The aptamer-modified electrode was characterized electrochemically by cyclic voltammetry (CV), differential pulse voltammetry (DPV), and electrochemical impedance spectroscopy (EIS). The modified electrode showed a voltammetric signal due to a one-step redox reaction of the surface-confined ferrocenyl moiety of the aptamer immobilized on the electrode surface in 10 mM N-(2-hydroxyethyl)piperazine-N'-2-ethanesulfonic acid (HEPES) buffer of pH 8.0. An increase in the DPV current signal was evident after blocking with 2-mercaptoethanol, effectively removing aptamer nonspecifically absorbed rather than bound to electrode surface or due to the formation of the aptamer-thrombin affinity interaction. The impedance measurement, in agreement with the differential pulse voltammetry (DPV), showed decreased Faradaic resistances in the same sequence. The "signal-on" upon thrombin association could be attributed to a change in conformation from random coil-like configuration on the probe-modified film to the quadruplex structure. The DPV of the modified electrode showed a linear response of the Fc oxidation signal to the increase in the thrombin concentration in the range between 5.0 and 35.0 nM with a linear correlation of r = 0.9988 and a detection limit of 0.5 nM. The molecular beacon aptasensor was amenable to full regeneration by simply unfolding the aptamer in 1.0 M HCl, and could be regenerated 25 times with no loss in electrochemical signal upon subsequent thrombin binding.
 ...
PMID:Reagentless, reusable, ultrasensitive electrochemical molecular beacon aptasensor. 1639 Jan 38


1 2 Next >>