摘要:

Our specific aim was to compare three plasminogen activator-inhibitor type 1 (PAI-1) antigen ELISA kit assays (the Biopool AB, Ltd, TintElize™ PAI-1 Strip-Well Format; the American Diagnostica, Inc., Imubind™ 822/1; and the second generation Imubind™ 822/1S). Within-run coefficients of variation (n= 6) for the TintElize, Imubind 822/1 and Imubind 822/1S methods were 5.5%, 5.9% and 6.8%, respectively. Between-run coefficients of variation for six aliquots per run were 2.9% for TintElize, 3.8% for Imubind 822/1, and 3.5% for Imubind 822/1S. Comparison of the average of duplicate aliquots from hyperlipidaemic patients demonstrated intrclass correlations of 0.75, 0.79 and 0.95 for TintElizevsImubind 822/1 (n= 39), TintElizevsImubind 822/1S (n= 39), and Imubind 822/1vs822/1S (n= 84), respectively. Lower 95% confidence interval limits of the intraclass correlation were 0.55, 0.48 and 0.93, respectively. Mean PAI-1 antigen values (n= 39) were 12.1, 15.8, 15.8 and 16.0 ng/ml, respectively, for TintElize, TintElize without using the quenching well, Imubind 822/1, and Imubind 822/1S. All three methods were easily performed and exhibited high correlation and reproducibility. A significant systematic bias (p<0.006) existed between TintElize and TintElize without using the quenching well, Imubind 822/1, and Imubind 822/1S. However, there was no significant bias when TintElize without using the quenching well is compared with Imubind 822/1 (p>0.8) and to 822/1S (p>0.8) nor is there significant systematic bias between Imubind 822/1 and 822/1S (p>0.3). By convention, interchangeability between assay methods suggests that the lower limit of the 95% intraclass correlation confidence interval be greater than 0.75. Thus, the three assay methods for measuring PAI-1 antigen are very similar and directly interchangeable when quenching well correction is not employed.

ISSN:0957-5235    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

Homocystinuria due to cystathionine-β-synthase deficiency (CBS-def-HOCY) initially often presents with thromboembolic events. In most cases in which coagulation factors have been analysed, a deficiency of AT-IIIc and factor VIIc has been reported, the cause of which has not been elucidated. Activation of coagulation with consumption of coagulation factors has been postulated as the mechanism. This paper reports a longitudinal study of two patients: patient 1 with thromboembolic disease and his asymptomatic sister, patient 2. Before start of therapy in patient 1, a reduction of FVIIc, other coagulation factors, and AT-IIIc was found. Markers of activation of coagulation (F1 + 2, TAT, FM, d-dimers) were elevated only in patient 1, and only at the time of thrombotic complications. In patient 2 reduced levels of FVIIc and other coagulation proteins, and a low borderline AT-IIIc level was found. Thus, in the two patients, sustained activation of coagulation can be reasonably excluded to be the cause of low levels of coagulation proteins. Vitamin therapy with 15 mg folate and 600 mg pyridoxine per day led to almost complete normalization of amino acids in urine and plasma. Thrombosis has not recurred to date. FVIIc and the other coagulation proteins and AT-IIIc increased in parallel with the biochemical remission. Direct inhibition of the activity of AT-III and coagulation factor VII and other factors by homocysteine was attemptedin vitrobut could not be shown at HC concentrations known to occur in the plasma of HOCY patients. Therefore, in these patients, deficient synthesis of coagulation factors and AT-III due to a disturbance of amino acid metabolism is still the most probable explanation for the observed low levels.

ISSN:0957-5235    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

Clot-bound thrombin remains active and is less accessible to heparin-antithrombin III than fluid-phase thrombin. To determine whether clot-bound human thrombin is more susceptible to inactivation by direct thrombin inhibitors, the activity of a novel synthetic competitive thrombin inhibitor Ro 46-6240, recombinant hirudin and unfractionated heparin were compared with fluid-phase thrombin and clot-bound thrombin. Fibrinopeptide A generated in human plasma was used as an index of thrombin activity. Hirudin was the most potent inhibitor of fluid-phase and clot-bound thrombin. However, Ro 46-6240 inhibited clot-bound thrombin three times more potently than fluid-phase thrombin (IC5019vs56 ng/ml) while hirudin was two times (IC508vs3 ng/ml) and heparin six times (IC501205vs200 ng/ml) less active against clot-bound thrombin compared with fluid-phase thrombin. The relative selectivity for clot-bound thrombin is not a unique property of Ro 46-6240 since two other synthetic thrombin inhibitors tested inhibited clot-bound thrombin more effectively than fluid-phase thrombin and a third was equally active against both forms of thrombin. In contrast, the affinities of two chromogenic substrates were similar for both forms of thrombin. This study shows that direct thrombin inhibitors inhibit clot-bound thrombin more potently than heparin and suggests that an apparent selectivity for clot-bound thrombin can be achieved with some synthetic thrombin inhibitors. Further studies have to show whether the high potency of these direct thrombin inhibitors translates into antithrombotic efficacy in clinical situations with pre-existing clots.

ISSN:0957-5235    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

The prevalence of thrombosis in patients with heterozygous type I plasminogen deficiency was studied. Fifteen kindreds described in the literature and a further five additional kindreds from the authors' Department were gathered. The prevalence of thrombotic events in all the patients with plasminogen deficiency was 23.6% (22 out of 93 patients), which decreased to 9.5% (seven of 74 patients) when the propositi were excluded. The 95 unaffected siblings were asymptomatic. The comparison between the prevalence of thrombosis in patients with plasminogen deficiency, with or without inclusion of the index patients, and in unaffected family members was statistically significant in both instances (P<0.0001 andP= 0.002, respectively). Analogous results were obtained from construction of thrombosis-free survival curves, which showed that in plasminogen deficient patients, either with or without inclusion of the propositi, the probability of developing thrombotic manifestations is significantly higher than in unaffected siblings (P= 0.002 andP= 0.043, respectively). It is concluded that congenital heterozygous plasminogen deficiency should be considered a risk factor for thrombosis, even though the probability of having a thrombotic event seems to be lower than in other thrombophilic conditions, such as hereditary defects of clotting factor inhibitors.

ISSN:0957-5235    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

The aim of this study was to examine whether there was a relationship between haemostatic factors and ultrasound-assessed morphology of the common carotid artery and cardiovascular disease in 57- to 77-year-old men at high risk for atherosclerotic disease (hypertension and at least one of the following risk factors: hypercholesterolaemia, smoking, diabetes mellitus). They were divided into one group with (n= 59) and one group without (n= 70) manifest cardiovascular disease. An age-matched reference group with no cardiovascular risk factors was used as a comparison (n= 51). Significant associations, independent of smoking, were found between plasma fibrinogen and both the maximal intima–media thickness and the occurrence of plaque in the high-risk group. High-risk patients with clinical signs of cardiovascular disease had higher levels of plasma fibrinogen and prothrombin 1 + 2 fragment compared with both high-risk patients without concomitant cardiovascular disease and low-risk subjects. Plasminogen activator inhibitor, von Willebrand factor and thrombin/antithrombin complex were increased in the high-risk group with signs of cardiovascular disease in comparison with the low-risk group. In conclusion the results indicate that plasma fibrinogen may be operative in the development of atherosclerosis. Clinical signs of cardiovascular disease were associated with increased plasma levels of fibrinogen, von Willebrand factor, plasminogen activator inhibitor, thrombin/antithrombin complex and prothrombin 1 + 2 fragment.

ISSN:0957-5235    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

The effect of heparin on the inactivation rates of fibrin-bound plasmin, miniplasmin and neutrophil leukocyte elastase (PMN-elastase) by their plasma inhibitors was studied. While plasmin and miniplasmin bound to fibrin are not inactivated by antithrombin, heparin (800 nM) makes these enzymes available for the inhibitor; the second-order rate constant increases from zero to 1.3×103M−1s−1and 3.3×103M−1s−1, respectively. Heparin slightly increases the rate of fibrin-bound enzyme inactivation by plasmin inhibitor.α1-Protease inhibitor, on the other hand, is unable to inactivate plasmin or miniplasmin bound to fibrin and heparin has no facilitating effect. In the case of PMN-elastase, heparin (300 nM) further increases enzyme protection againstα1-protease inhibitor; the rate constant decreases from 41×103M−1s−1; to 23×103M−1s−1.α2-Macroglobulin inhibits fibrin-bound miniplasmin and PMN-elastase with a second-order rate constant of 1.8×104M−1s−1; and heparin (300 nM) increases the rate insignificantly for miniplasmin and by a factor of two for PMN-elastase. It is remarkable that plasmin bound to fibrin is not inhibited byα2-macroglobulin independently of the presence of heparin. On the basts of the reported kinetic data a lifespan of 420 s for plasmin, 66 s for miniplasmin and 4 s for PMN-elastase was calculated, when the enzymes are bound to fibrin in the presence of the four protease inhibitors at physiological plasma concentration. If heparin is present (300 nM) these values decrease to 240 s for plasmin and 42 s for miniplasmin, whereas that of PMN-elastase is unchanged. Thus, the presentin vitrokinetic model suggests an antifibrinolytic effect of heparin in a plasma milieu.

ISSN:0957-5235    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

Rabbits were repeatedly immunized with a chimeric plasminogen activator composed of the kringle-2 domain of human tissue-type plasminogen activator (t-PA) attached to the serine protease domain of the human urokinase-type plasminogen activator (u-PA). IgG from these rabbits was purified, biotinylated and subjected to affinity chromatography on K2tu-PA covalently attached to Sepharose. Roughly half the antibodies recovered from the K2tu-PA column could subsequently be bound to a column with similarly immobilized t-PA, whereas the other half bound to a u-PA column. Less than 0.01% of the biotinylated anti-K2tu-PA antibodies did not bind to either t-PA or u-PA and perhaps are directed against neoantigenic determinants on K2tu-PA, not present in the natural plasminogen activators.

ISSN:0957-5235    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

Qualitative ultrastructural autoradiography was used to study the binding of the vascular anticoagulant alpha (annexin V) to normal and atherosclerotic (AS) rabbit aortic intima. Recombinant annexin V was labelled with m125I by the Iodogen method. Rabbits were fed a hypercholesterolemic (HC) diet up to 10 months. After laparotomy and exsanguination, the aortae were perfused with dilutions of125I-annexin V (I-AV) for 10–15 min, either on ice or at 22°C and then perfusion-fixed with aldehydes. Fragments of the labelled aortae were used foren facecontact autoradiography, followed by Sudan Black staining of intimal lipid. Specimens were also included in Epon and sectioned for light- and electron-microscopic autoradiography. The binding of I-AV was increased on the AS aortae as compared with the normal ones, with an apparent preference for the lesioned areas. Microscopically, I-AV was found at the luminal front of aortic intima, on endothelial cells (EC), on macrophage foam cells, and on their disrupted remnants. The presence of the AV binding sites (reportedly known to interact with high affinity with phosphatidylserine) in the rabbit AS aortic intima, together with other known procoagulant conditions, may contribute to the initiation of coagulation events into the lesioned vascular wall, and may offer a rationale for the use of annexin V as an anticoagulant drug.

ISSN:0957-5235    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

Inflammatory agents including bacterial endotoxin (LPS) and low concentrations of phorbol myristate acetate (PMA) stimulate human peripheral blood monocytes to transiently express tissue factor procoagulant activity. Concentrations of PMA that cause the cytosol-to-membrane translocation of protein kinase C (PKC) (10−9–10−7M) induce a rapid decrease in monocyte tissue factor activity. Staurosporine, an inhibitor of protein kinase C, enhances the stimulatory effect of low concentrations of PMA on monocyte expression of tissue factor activity and blocks suppression of tissue factor activity at high PMA concentrations. Furthermore, staurosporine prolongs LPS-induced tissue factor expression in monocytes. These results suggest that protein kinases modulate tissue factor activity in human monocytes by regulating both induction and down-regulation.

ISSN:0957-5235    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

Metal chelate affinity chromatography on copper-charged Chelating Sepharose has been used to purify a factor IX concentrate from 4 000- to 5 000-kg pools of human plasma, with an overall yield of 194 IU/kg. Unwanted proteins and solvent-detergent reagents added to inactivate lipid-enveloped viruses were removed during the chromatographic step. The freeze-dried product was >80% pure factor IX with a mean specific activity of >160 IU/mg protein. The concentrate showed no evidence of clotting factor activation byin vitrotests for potential thrombogenicity or by direct assay for activated factor IX. The concentrate did not exhibit proteolytic activity against a range of synthetic peptide chromogenic substrates. Full functional factor IX activity was retained and there was no evidence of protein degradation. Metal chelate affinity chromatography therefore appears to present less physicochemical challenge to the protein than other factor IX purification methods, while allowing the preparation of a clinical factor IX concentrate at a large scale.

ISSN:0957-5235    

出版商:OVID    

年代:1994

数据来源: OVID