Substrate Composition and the Effect of ε-Aminocaproic Acid on Tissue Plasminogen Activator and Urokinase-induced Fibrinolysis

1974 ◽  
Vol 32 (02/03) ◽  
pp. 306-324 ◽  
Author(s):  
Sixtus Thorsen ◽  
Tage Astrup
Keyword(s):  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Aminocaproic Acid ◽  
Fibrin Clot ◽  
Clot Lysis ◽  
Substrate Composition ◽  
Lysis Time ◽  
Biphasic Effect ◽  
Tissue Plasminogen ◽  
Inhibition Curve

SummaryThe influence of variations in substrate composition on the biphasic pattern of inhibition by e-aminocaproic acid (EACA) of urokinase-induced fibrinolysis, first observed on bovine plasminogen-rich fibrin, was studied using a fibrin clot lysis time assay. Different species of fibrinogen and plasminogen or plasmin were used. Preparations of different degrees of purification were compared at varying concentrations. The behavior of urokinase was compared with that of a porcine tissue plasminogen activator. Variations in the conditions of the assay greatly influenced the results. The concentration of fibrinogen had a particularly marked influence. At increased fibrinogen concentrations the biphasic response of urokinase was enhanced and a weak biphasic effect was also produced by the tissue activator which usually yields a uniformly increasing inhibition curve. The phase of fibrinolysis enhancement produced by urokinase with genuine plasminogen substrates occurs at substrate concentrations and EACA levels present in patients treated with EACA. The observed variations may explain discordant findings reported by various investigators.

PHYSICAL ACTIVITY RELEASES TISSUE PLASMINOGEN ACTIVATOR FROM EXERCISING PARTS OF BODY

1987 ◽  
Author(s):  
I Keber ◽  
K Potisk ◽  
D Keber ◽  
M Stegnar ◽  
N Vene
Keyword(s):  
Physical Activity ◽  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Fibrinolytic Activity ◽  
Venous Occlusion ◽  
Clot Lysis ◽  
Lysis Time ◽  
Tissue Plasminogen ◽  
Euglobulin Clot Lysis Time ◽  
The Difference

To determine the origin of tissue plasminogen activator (t-PA) release during physical activity, we studied the separate and combined effects of venous occlusion and acute physical activity on t-PA release in arm and leg. In 15 healthy volunteers 20 min venous occlusions of arm and leg were performed simultaneously before physical activity ( maximal stress testing on treadmill)(occlusion I), immediately after physical activity and 45 min later (occlusion II). Blood samples were drawn from unoccluded arm before occlusion and after physical activity, and from occluded arm and leg after occlusion. Fibrinolytic activity was measured by euglobulin clot lysis time (ECLT) and t-PA activity assay. The amount of released t-PA during different stimuli (fibrinolytic potential) was calculated as the difference between post- and prestimulation fibrinolytic activity. Before physical activity there was a great increase in fibrinolytic activity due to t-PA in the occluded arm but no increase in the occluded leg. Physical activity itself caused a similar increase of systemic fibrinolytic activity as arm occlusion locally. After physical activity arm occlusion evoked equally good response than before it. Fibrinolytic activity during leg occlusion behaved differently: there was an increase in t-PA activity in the occluded leg which persisted one hour after physical activity, when systemic fibrinolytic activity already fell to initial level.These results demonstrated that walking and running triggered t-PA release from the leg vessels. Since leg occlusion was not a stimulus for t-PA release, it served only as a method to demonstrate the effect of physical activity.

Relationships between Euglobulin Clot Lysis Time and the Plasma Levels of Tissue Plasminogen Activator and Plasminogen Activator Inhibitor 1

1990 ◽  
Vol 63 (01) ◽  
pp. 082-086 ◽  
Author(s):  
Tetsumei Urano ◽  
Kenji Sakakibara ◽  
Andrzej Rydzewski ◽  
Shoko Urano ◽  
Yumiko Takada ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Slight Modification ◽  
Clot Lysis ◽  
Plasminogen Activator Inhibitor 1 ◽  
Lysis Time ◽  
Tissue Plasminogen ◽  
Euglobulin Clot Lysis Time ◽  
Pai 1 ◽  
Clot Lysis Time

SummaryThe relationships between tissue plasminogen activator (tPA), its fast acting inhibitor (PAI-1) and euglobulin clot lysis time (ELT) were investigated with healthy volunteers’ plasma. Turbidimetric clot lysis assay by the microtiter plate reader was utilized for ELT with a slight modification. Both tPA and PAI-1 showed the significant correlation with ELT. tPA had a significantly positive, not negative, correlation with ELT (R = 0.387, p <0.001). Higher correlation coefficients (R = 0.580, p <0.001 and R = 0.599, p <0.001) were obtained between ELT and total PAI-1 or free PAI-1 than tPA or tPA-PAI-1 complex (R = 0.427, p <0.001). The positive correlation was also obtained between tPA and PAI-1. These data suggest that PAI-1 is a highly important factor for ELT, especially, the amounts of free PAI-1 being the key factor to determine the ELT, which can represent the potential activity of the fibrinolytic system.

scholarly journals Severe SARS-CoV-2 infection inhibits fibrinolysis leading to changes in viscoelastic properties of blood clot: A descriptive study of fibrinolysis

2021 ◽  
Author(s):  
Stefanie Hammer ◽  
Helene Haeberle ◽  
Christian Schlensak ◽  
Michael Bitzer ◽  
Nisar Malek ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Point Of Care ◽  
Blood Clot ◽  
Clot Lysis ◽  
Plasminogen Activator Inhibitor 1 ◽  
Blood Samples ◽  
Lysis Time ◽  
Tissue Plasminogen ◽  
Pai 1

Background: Accumulating evidence indicates towards an association between SARS-CoV-2 infection and procoagulatory state in blood. Thromboelastographic investigations are useful point-of-care devices to assess coagulation and fibrinolysis. Objectives: We investigated the hypothesis that the procoagulatory state in COVID-19 patients is caused by impaired fibrinolysis system. Methods: COVID-19 patients admitted to normal wards or to the intensive care unit (ICU) were included in the study. Whole blood samples were investigated by thromboelastography. Additionally, global parameters of coagulation and factors of fibrinolysis as plasminogen activator inhibitor-1 (PAI-1), tissue plasminogen activator (tPA), plasminogen activity and alpha 2-antiplasmin (A2AP) were determined. Results and conclusion: A significantly increased lysis-resistance and a significantly longer time of lysis after adding tissue plasminogen activator was observed in blood samples from ICU COVID-19 patients compared to controls (maximal lysis: 3.25% ± 0.56 vs. 6.20% ± 0.89, p=0.0127; lysis time: 365.7s ± 44.6 vs. 193.2s ± 16.3, p=0.0014). PAI-1 activity was significantly higher in plasma samples of ICU COVID-19 patients (PAI-1: 4.92U/ml ± 0.91 vs. 1.28U/ml ± 0.33, p=0.001). Interestingly, a positively correlation between the activity of PAI-1 and the lysis time of the formed clot (r=0.7015, p=0.0006) was observed. Our data suggest that severe SARS-CoV-2 infection is associated with impaired fibrinolytic activity in blood, where fibrinolytic inhibitors are elevated leading to an increased resistance to clot lysis. Future clinical studies should address the contribution of the fibrinolysis system to the procoagulatory state in blood of COVID-19 patients and investigate potential therapeutic targets in this system.

scholarly journals The pathogenesis of accelerated fibrinolysis in liver cirrhosis: a critical role for tissue plasminogen activator inhibitor

Blood ◽  
1987 ◽  
Vol 69 (5) ◽  
pp. 1315-1319 ◽  
Author(s):  
SL Hersch ◽  
T Kunelis ◽  
RB Jr Francis
Keyword(s):  
Liver Cirrhosis ◽  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Fibrinolytic Activity ◽  
Blood Clot ◽  
Critical Role ◽  
Plasminogen Activator Inhibitor ◽  
Clot Lysis ◽  
Lysis Time ◽  
Tissue Plasminogen

Abstract The pathogenesis of accelerated fibrinolysis in liver cirrhosis was investigated by comparing the results of specific assays for tissue plasminogen activator (tpa) antigen, tpa activity, tpa inhibitor, and alpha-2 plasmin inhibitor (a2PI) in 12 patients with cirrhosis and markedly accelerated fibrinolysis (dilute whole blood clot lysis time (DWBCLT) less than two hours), in nine patients with cirrhosis and moderately accelerated fibrinolysis (DWBCLT two to four hours), and in nine patients with cirrhosis and normal fibrinolysis (DWBCLT greater than four hours). Mean tpa antigen was markedly increased in all three groups, but no correlation was observed between overall fibrinolytic activity as measured by the DWBCLT and the level of tpa antigen. In contrast, there was a significant correlation between overall fibrinolytic activity and tpa activity and an equally significant correlation between fibrinolytic activity and decreased tpa inhibition. Mean a2PI activity was significantly lower than normal in groups 1 and 2 but was normal in group 3. The pathogenesis of accelerated fibrinolysis in liver cirrhosis thus appears to depend critically on the capacity of plasma inhibitors to inhibit increased circulating tpa antigen. Reduced a2PI also appears to play a role.

scholarly journals Physico-Chemical and Biological Properties of Biosimilar and Reference Tissue Plasminogen Activator Products

2019 ◽  
Vol 19 (1) ◽  
pp. 39-49
Author(s):  
V. D. Gusarova ◽  
M. S. Pantyushenko ◽  
V. M. Simonov ◽  
R. R. Shukurov ◽  
R. A. Khamitov ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Specific Activity ◽  
Reference Product ◽  
Fibrin Clot ◽  
Biological Properties ◽  
Clot Lysis ◽  
Physico Chemical ◽  
Tissue Plasminogen

Recombinant tissue plasminogen activator (international nonproprietary name — alteplase) which was developed by «GENERIUM» (Russia) and received a marketing authorisation in Russia is completely analogous to Actilyse® which is used to treat medical conditions accompanied by thrombosis, such as acute myocardial infarction, pulmonary embolism, and ischemic stroke. The aim of the study was to carry out a comprehensive comparison of physico-chemical and biological properties of Revelyse® and the reference product Actilyse® in order to assess their biosimilarity. Materials and Methods: comparative peptide mapping and determination of comparability of chromatographic profiles of tryptic hydrolysates was performed using RP-HPLC and massspectrometry; the molecular weight distribution was determined by mass-spectrometry and polyacrylamide gel electrophoresis (Laemmli method). The purity and homogeneity of products as well as the content of related impurities (oligomers and fragments) were determined using gel filtration; N-glycosylation profile was analysed by hydrophilic HPLC, total sialic acid was quantified by the Svennerholm resorcinol method. Protein binding to fibrin and human fibrinogen was assessed by surface plasmon resonance, and the specific activity was compared by fibrin clot lysis. Results: the research demonstrated a complete overlap of the products’ peptide maps, which indicates the identity of аlteplase amino acid sequences in the two medicines being compared. The authors of the study also determined the molecular weight and the content of the intact single-stranded form of the protein, and quantified post-translational modifications, the content of sialic acids and neutral sugars. The analysis of the N-glycosylation profile revealed insignificant differences in the percentage of multiantenna complex glycans. The specificity of alteplase was evaluated by analysing the formation of protein complexes with natural alteplase ligands – fibrin and plasminogen activator inhibitor-1, but no significant differences were found. The comparison of specific activation of plasminogen fibrinolytic activity was performed based on the results of the assay analysing the fibrin clot lysis rate, and it demonstrated comparability of Revelyse® and Actilyse®. Conclusions: comparative experimental studies have shown no differences in the structure, charge distribution heterogeneity, impurities content, and specific activity of alteplase as a component of Revelyse® and the reference product Actilyse®, which leads to the conclusion that they are similar in terms of physicochemical and biological properties.

In Vitro Studies on the Fibrinolytic, Thrombolytic and Fibrinogenolytic Properties of a Tissue Plasminogen Activator from Guinea Pig Keratocytes

1985 ◽  
Vol 53 (02) ◽  
pp. 200-203 ◽  
Author(s):  
A Electricwala ◽  
R J Ling ◽  
P M Sutton ◽  
B Griffiths ◽  
P A Riley ◽  
...  
Keyword(s):  
Guinea Pig ◽  
Human Plasma ◽  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Fibrin Clot ◽  
Clot Lysis ◽  
Thrombolytic Activity ◽  
Tissue Plasminogen ◽  
In Vitro Systems

SummaryThe fibrinolytic and thrombolytic properties of a tissue plasminogen activator (tPA) purified from the conditioned medium of an established guinea pig keratocyte (GPK) cell line were investigated in in vitro systems and compared with urokinase. Using the fibrin clot lysis assay, GPK activator appears to be similar to human melanoma tPA and not to human urokinase. GPK activator also caused negligible fibrinogen breakdown, when incubated with human plasma at 37° C over 23 hr. Urokinase on the other hand caused significant fibrinogenolysis, under similar conditions. Comparison of the lysis of plasma clots by GPK activator and human urokinase have shown that GPK activator was a much more effective fibrinolytic agent than urokinase, especially at lower concentrations (<50 IU/ml). Studies on the thrombolytic effect of GPK activator on the lysis of aged and cross-linked whole human blood clots and plasma clots hanging in artificially circulating human plasma suggest that GPK activator can lyse both these types of clots equally well. The lysis is dose dependent, attaining complete lysis within 3–6 hr with the concentration of GPK activator in the range of 1–5 μg/ml plasma. It is concluded that GPK activator has a higher fibrinolytic and thrombolytic activity and lower fibrinogenolytic activity than urokinase.

COVALENT Mr ∼92,000 HYBRID PLASMINOGEN ACTIVATOR DERIVED FROM THE PLASMIN FIBRIN-BINDING DOMAIN AND THE TISSUE PLASMINOGEN ACTIVATOR CATALYTIC DOMAIN

1987 ◽  
Author(s):  
K C Robbins ◽  
I Boreisha
Keyword(s):  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Catalytic Domain ◽  
Specific Activity ◽  
Fibrin Clot ◽  
Binding Domain ◽  
Molar Ratio ◽  
Clot Lysis ◽  
Chromatography Method ◽  
Tissue Plasminogen

A covalent hybrid plasminogen activator was prepared from the sulfhydryl forms of the NH2-terminal A chain of human plasmin (Pln^) containing the fibrin-binding domain, and the COOH-terminal B chain of tissue plasminogen activator (t-PAB) containing the catalytic domain. The PlnA (SH)2 and t-PAB(SH) chains were mixed in a 1:1 molar ratio, and hybridization was allowed to proceed at 4 °C for 6 days. The covalent PlnA-t-PAB hybrid activator was isolated from the mixture by a two-step affinity chromatography method, with L-lysine-substituted Sepharose and Zn-chelated agarose. The protein yield of purified hybrid was 10% with a major component (77%) of Mr ∼92,000. The covalent PlnA-t-PAB hybrid activator, contained 1 mol of each chain; after reduction, it gave the two parent chains, PlnA and t-PAA, also shown to be present by double immunodiffusion. The specific plasminogen activator activity, with soluble fibrin, and the specific amidolytic activity, of the purified covalent hybrid activator was determined to be 200,000 IU/mg of protein, about 40% of the specific activity of the parent t-PA. In a fibrin clot lysis assay, the covalent hybrid activator and t-PA have similar specific fibrinolytic activities, 500,000 IU/mg of protein; however, the clot lysis time curves were not parallel. The binding of the covalent PlnA-t-PAB hybrid activator and t-PA to forming fibrin were found to be similar; at physiological fibrinogen concentrations, binding of both activators to forming fibrin was about 90%.

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