scholarly journals Role of Shear Stress and tPA Concentration in the Fibrinolytic Potential of Thrombi

2021 ◽  
Vol 22 (4) ◽  
pp. 2115
Author(s):  
Claire S. Whyte ◽  
Hadj Ahmed. Mostefai ◽  
Kim M. Baeten ◽  
Andrew J. Lucking ◽  
David E. Newby ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Ex Vivo ◽  
Active Form ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
High Shear ◽  
Plasminogen Activator Inhibitor 1 ◽  
Fibrinolytic Potential ◽  
Pai 1 ◽  
Low Shear

The resolution of arterial thrombi is critically dependent on the endogenous fibrinolytic system. Using well-established and complementary whole blood models, we investigated the endogenous fibrinolytic potential of the tissue-type plasminogen activator (tPA) and the intra-thrombus distribution of fibrinolytic proteins, formed ex vivo under shear. tPA was present at physiologically relevant concentrations and fibrinolysis was monitored using an FITC-labelled fibrinogen tracer. Thrombi were formed from anticoagulated blood using a Chandler Loop and from non-anticoagulated blood perfused over specially-prepared porcine aorta strips under low (212 s−1) and high shear (1690 s−1) conditions in a Badimon Chamber. Plasminogen, tPA and plasminogen activator inhibitor-1 (PAI-1) concentrations were measured by ELISA. The tPA–PAI-1 complex was abundant in Chandler model thrombi serum. In contrast, free tPA was evident in the head of thrombi and correlated with fibrinolytic activity. Badimon thrombi formed under high shear conditions were more resistant to fibrinolysis than those formed at low shear. Plasminogen and tPA concentrations were elevated in thrombi formed at low shear, while PAI-1 concentrations were augmented at high shear rates. In conclusion, tPA primarily localises to the thrombus head in a free and active form. Thrombi formed at high shear incorporate less tPA and plasminogen and increased PAI-1, thereby enhancing resistance to degradation.

scholarly journals Plasma tissue plasminogen activator and plasminogen activator inhibitor-1 in hospitalized COVID-19 patients

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yu Zuo ◽  
Mark Warnock ◽  
Alyssa Harbaugh ◽  
Srilakshmi Yalavarthi ◽  
Kelsey Gockman ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Ex Vivo ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Bleeding Complications ◽  
Clot Lysis ◽  
Thrombosis Prophylaxis ◽  
Plasminogen Activator Inhibitor 1 ◽  
Activator Inhibitor ◽  
Pai 1

AbstractPatients with coronavirus disease-19 (COVID-19) are at high risk for thrombotic arterial and venous occlusions. However, bleeding complications have also been observed in some patients. Understanding the balance between coagulation and fibrinolysis will help inform optimal approaches to thrombosis prophylaxis and potential utility of fibrinolytic-targeted therapies. 118 hospitalized COVID-19 patients and 30 healthy controls were included in the study. We measured plasma antigen levels of tissue-type plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1) and performed spontaneous clot-lysis assays. We found markedly elevated tPA and PAI-1 levels in patients hospitalized with COVID-19. Both factors demonstrated strong correlations with neutrophil counts and markers of neutrophil activation. High levels of tPA and PAI-1 were associated with worse respiratory status. High levels of tPA, in particular, were strongly correlated with mortality and a significant enhancement in spontaneous ex vivo clot-lysis. While both tPA and PAI-1 are elevated among COVID-19 patients, extremely high levels of tPA enhance spontaneous fibrinolysis and are significantly associated with mortality in some patients. These data indicate that fibrinolytic homeostasis in COVID-19 is complex with a subset of patients expressing a balance of factors that may favor fibrinolysis. Further study of tPA as a biomarker is warranted.

The Amount of Plasminogen, Tissue-Type Plasminogen Activator and Plasminogen Activator Inhibitor Type 1 in Human Thrombi and the Relation to IVVO-TNO Lysibility

1992 ◽  
Vol 67 (01) ◽  
pp. 101-105 ◽  
Author(s):  
B J Potter van Loon ◽  
D C Rijken ◽  
E J P Brommer ◽  
A P C van der Maas
Keyword(s):  
Plasminogen Activator ◽  
Ex Vivo ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Plasminogen Activator Inhibitor 1 ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator ◽  
Activator Inhibitor ◽  
Pai 1

SummaryThrombolytic therapy successfully reopens obstructed blood vessels in the majority of cases. However, it is not known why a substantial amount of thrombi are resistant to lysis by a fibrinolytic agent. In vitro studies have demonstrated that tissue-type plasminogen activator (t-PA) and plasminogen incorporated in the clot (during formation) increase lysibility. To test whether lysibility of in vivo formed human thrombi is related to their composition, we studied 25 venous thrombi obtained at autopsy and 21 arterial thrombi obtained during embolectomy.Plasminogen activator inhibitor-1 (PAI-1) antigen was measured in a phosphate-buffered saline (PBS) extract of each thrombus; t-PA antigen and plasminogen antigen were determined in a 6 M urea extract of the thrombus, representing bound proteins. Lysibility was measured as weight reduction during 8 h of incubation in PBS containing streptokinase (SK) 100 U/ml, corrected for spontaneous lysis, reflected by weight loss in PBS without SK. In addition, lysibility in SK was compared with lysibility in urokinase (UK) 100 U/ml and in t-PA 200 U/ml.Spontaneous lysis amounted to 29 ± 5% (mean ± SEM) and 33 ± 5% in venous and arterial thrombi, respectively, and inversely correlated with the PAI-1 content of thrombi (r = —0.43, p <0.01). Lysibility amounted to 76 ± 6% in venous and 90 ± 4% in arterial thrombi (venous vs. arterial: p = 0.051). PAI-1-, plasminogen- and t-PA-content of venous thrombi were 902 ± 129 ng, 34.3 ± 4.8 pg and 26.7 ± 3.0 ng per gram of wet thrombus respectively; for arterial thrombi these values were 2,031 ± 401 ng/g (p = 0.011), 64.1 ± 11.4 pg/g (p = 0.088) and 62.2 ± 8.3 ng/g (p = 0.0001), respectively. A correlation was found between t-PA and plasminogen (r = 0.74, p <0.001). Lysibility by SK related to plasminogen content in both venous (r = 0.60, p <0.002) and arterial (r = 0.44, p <0.05) thrombi; PAI-1 and t-PA did not correlate with lysibility. Lysibility in the chosen concentrations of SK, UK and t-PA were similar.We conclude that spontaneous lysis of thrombi in saline is dependent on PAI-1 content and that susceptibility of thrombibi to lysis by SK ex vivo is dependent on the plasminogen content

scholarly journals Plasma tissue plasminogen activator and plasminogen activator inhibitor-1 in hospitalized COVID-19 patients

2020 ◽  
Author(s):  
Yu Zuo ◽  
Mark Warnock ◽  
Alyssa Harbaugh ◽  
Srilakshmi Yalavarthi ◽  
Kelsey Gockman ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Ex Vivo ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Bleeding Complications ◽  
Clot Lysis ◽  
Thrombosis Prophylaxis ◽  
Plasminogen Activator Inhibitor 1 ◽  
Activator Inhibitor ◽  
Pai 1

ABSTRACTPatients with coronavirus disease-19 (COVID-19) are at high risk for thrombotic arterial and venous occlusions. However, bleeding complications have also been observed in some patients. Understanding the balance between coagulation and fibrinolysis will help inform optimal approaches to thrombosis prophylaxis and potential utility of fibrinolytic-targeted therapies. 118 hospitalized COVID-19 patients and 30 healthy controls were included in the study. We measured plasma antigen levels of tissue-type plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1) and performed spontaneous clot-lysis assays. We found markedly elevated tPA and PAI-1 levels in patients hospitalized with COVID-19. Both factors demonstrated strong correlations with neutrophil counts and markers of neutrophil activation. High levels of tPA and PAI-1 were associated with worse respiratory status. High levels of tPA, in particular, were strongly correlated with mortality and a significant enhancement in spontaneous ex vivo clot-lysis. While both tPA and PAI-1 are elevated among COVID-19 patients, extremely high levels of tPA enhance spontaneous fibrinolysis and are significantly associated with mortality in some patients. These data indicate that fibrinolytic homeostasis in COVID-19 is complex with a subset of patients expressing a balance of factors that may favor fibrinolysis. Further study of tPA as a biomarker is warranted.

A Specific Immunologic Assay for Functional Plasminogen Activator Inhibitor 1 in Plasma – Standardized Measurements of the Inhibitor and Related Parameters in Patients with Venous Thromboembolic Disease

1992 ◽  
Vol 68 (05) ◽  
pp. 486-494 ◽  
Author(s):  
Malou Philips ◽  
Anne-Grethe Juul ◽  
Johan Selmer ◽  
Bent Lind ◽  
Sixtus Thorsen
Keyword(s):  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Normal Subjects ◽  
Tissue Type ◽  
Blood Collection ◽  
Plasminogen Activator Inhibitor 1 ◽  
Type Plasminogen Activator ◽  
Significant Difference ◽  
Activator Inhibitor ◽  
Pai 1

SummaryA new assay for functional plasminogen activator inhibitor 1 (PAI-1) in plasma was developed. The assay is based on the quantitative conversion of PAI-1 to urokinase-type plasminogen activator (u-PA)-PAI-l complex the concentration of which is then determined by an ELISA employing monoclonal anti-PAI-1 as catching antibody and monoclonal anti-u-PA as detecting antibody. The assay exhibits high sensitivity, specificity, accuracy, and precision. The level of functional PAI-1, tissue-type plasminogen activator (t-PA) activity and t-PA-PAI-1 complex was measured in normal subjects and in patients with venous thromboembolism in a silent phase. Blood collection procedures and calibration of the respective assays were rigorously standardized. It was found that the patients had a decreased fibrinolytic capacity. This could be ascribed to high plasma levels of PAI-1. The release of t-PA during venous occlusion of an arm for 10 min expressed as the increase in t-PA + t-PA-PAI-1 complex exhibited great variation and no significant difference could be demonstrated between the patients with a thrombotic tendency and the normal subjects.

Abstract T P75: Temporal Profiles of Plasminogen Activator Inhibitor-1 Concentration and Activity Changes in Circulation after Focal Stroke and Tissue Type Plasminogen Activator Administration in Rats

Stroke ◽  
2014 ◽  
Vol 45 (suppl_1) ◽  
Author(s):  
Qi Liu ◽  
Xiang Fan ◽  
Helen Brogren ◽  
Ming-Ming Ning ◽  
Eng H Lo ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Plasminogen Activator Inhibitor 1 ◽  
Type Plasminogen Activator ◽  
Temporal Profiles ◽  
Activator Inhibitor ◽  
Pai 1

Aims: Plasminogen activator inhibitor-1 (PAI-1) is the main and potent endogenous tissue-type plasminogen activator (tPA) inhibitor, but an important question on whether PAI-1 in blood stream responds and interferes with the exogenously administered tPA remains unexplored. We for the first time investigated temporal profiles of PAI-1 concentration and activity in circulation after stroke and tPA administration in rats. Methods: Permanent MCAO focal stroke of rats were treated with saline or 10mg/kg tPA at 3 hours after stroke (n=10 per group). Plasma (platelet free) PAI-1 antigen and activity levels were measured by ELISA at before stroke, 3, 4.5 (1.5 hours after saline or tPA treatments) and 24 hours after stroke. Since vascular endothelial cells and platelets are two major cellular sources for PAI-1 in circulation, we measured releases of PAI-1 from cultured endothelial cells and isolated platelets after direct tPA (4 μg/ml) exposures for 60 min in vitro by ELISA (n=4 per group). Results: At 3 hours after stroke, both plasma PAI-1 antigen and activity were significantly increased (3.09±0.67, and 3.42±0.57 fold of before stroke baseline, respectively, all data are expressed as mean±SE). At 4.5 hours after stroke, intravenous tPA administration significantly further elevated PAI-1 antigen levels (5.26±1.24), while as expected that tPA neutralized most elevated PAI-1 activity (0.33±0.05). At 24 hours after stroke, PAI-1 antigen levels returned to the before baseline level, however, there was a significantly higher PAI-1 activity (2.51±0.53) in tPA treated rats. In vitro tPA exposures significantly increased PAI-1 releases into culture medium in cultured endothelial cells (1.65±0.08) and platelets (2.02±0.17). Conclution: Our experimental results suggest that tPA administration may further elevate stroke-increased blood PAI-1 concentration, but also increase PAI-1 activity at late 24 hours after stroke. The increased PAI-1 releases after tPA exposures in vitro suggest tPA may directly stimulate PAI-1 secretions from vascular walls and circulation platelets, which partially contributes to the PAI-1 elevation observed in focal stroke rats. The underlying regulation mechanisms and pathological consequence need further investigation.

Plasminogen activator inhibitor-1 rises after hemorrhage in rats

1995 ◽  
Vol 268 (6) ◽  
pp. E1065-E1069 ◽  
Author(s):  
M. Yamashita ◽  
D. N. Darlington ◽  
E. J. Weeks ◽  
R. O. Jones ◽  
D. S. Gann
Keyword(s):  
Plasminogen Activator ◽  
High Performance ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Sprague Dawley ◽  
Plasminogen Activator Inhibitor 1 ◽  
Sprague Dawley Rats ◽  
Type Plasminogen Activator ◽  
Activator Inhibitor ◽  
Pai 1

Large hemorrhage leads to hypercoagulability, a phenomenon that has never been well explained. Because an elevation of plasminogen activator inhibitor (PAI)-1 increases procoagulant activity, we have determined whether plasma PAI activity and tissue PAI-1 mRNA are elevated after hemorrhage. Sprague-Dawley rats were bled (20 or 15 ml/kg) 4 days after cannulation. Plasma PAI activity was determined by the capacity of plasma to inhibit tissue-type plasminogen activator activity. Changes of PAI-1 mRNA in various tissues were detected by high-performance liquid chromatography after reverse transcription and polymerase chain reaction. Hemorrhage (20 ml/kg) significantly elevated plasma PAI activity at 0.5, 1, 2, 4, 6, and 8 h after hemorrhage and PAI-1 mRNA in liver at 1, 2, 4, and 6 h after hemorrhage. The PAI-1 message was also significantly elevated in lung, heart, and kidney at 4 h after hemorrhage. The increases of PAI-1 mRNA after 20 ml/kg hemorrhage were significantly greater than those after 15 ml/kg hemorrhage. These findings indicate that large hemorrhage can induce the increases in PAI activity and PAI-1 message and suggest that induction of PAI-1 may be involved in the thrombogenic responses observed after large hemorrhage.

scholarly journals Spironolactone Abolishes the Relationship between Aldosterone and Plasminogen Activator Inhibitor-1 in Humans

2002 ◽  
Vol 87 (2) ◽  
pp. 448-452 ◽  
Author(s):  
Pairunyar Sawathiparnich ◽  
Sandeep Kumar ◽  
Douglas E. Vaughan ◽  
Nancy J. Brown
Keyword(s):  
Angiotensin Ii ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Plasminogen Activator Inhibitor 1 ◽  
Type Plasminogen Activator ◽  
Serum Aldosterone ◽  
The Relationship ◽  
Activator Inhibitor ◽  
Pai 1

Recent studies have defined a link between the renin-angiotensin-aldosterone system and fibrinolysis. The present study tests the hypothesis that endogenous aldosterone regulates plasminogen activator inhibitor-1 (PAI-1) production in humans. Hemodynamic parameters, PAI-1 and tissue-type plasminogen activator (t-PA) antigen, potassium, PRA, angiotensin II, and aldosterone were measured in nine male hypertensive subjects after a 3-wk washout, after 2 wk of hydrochlorothiazide (HCTZ; 25 mg plus 20 mmol KCl/d), and after 2 wk of spironolactone (100 mg/d plus KCl placebo). Spironolactone (P = 0.04), but not HCTZ (P = 0.57 vs. baseline; P = 0.1 vs. spironolactone), significantly lowered systolic blood pressure. Angiotensin II increased from baseline during both HCTZ (P = 0.02) and spironolactone (P = 0.02 vs. baseline; P = 0.19 vs. HCTZ) treatments. Although both HCTZ (P = 0.004) and spironolactone (P &lt; 0.001 vs. baseline) increased aldosterone, the effect was greater with spironolactone (P &lt; 0.001 vs. HCTZ). HCTZ increased PAI-1 antigen (P = 0.02), but did not alter t-PA antigen. In contrast, there was no effect of spironolactone on PAI-1 antigen (P = 0.28), whereas t-PA antigen was increased (P = 0.01). There was a significant correlation between PAI-1 antigen and serum aldosterone during both baseline and HCTZ study days (r2 = 0.57; P = 0.0003); however, treatment with spironolactone abolished this correlation (r2 = 0.13; P = 0.33). This study provides evidence that endogenous aldosterone influences PAI-1 production in humans.

Inactivation of Plasminogen Activator Inhibitor-1 Accelerates Thrombolysis of a Platelet-rich Thrombus in Rat Mesenteric Arterioles

2001 ◽  
Vol 86 (12) ◽  
pp. 1528-1531 ◽  
Author(s):  
Alain Rupin ◽  
Frédéric Martin ◽  
Marie-Odile Vallez ◽  
Edith Bonhomme ◽  
Tony Verbeuren
Keyword(s):  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Cerebral Stroke ◽  
Plasminogen Activator Inhibitor 1 ◽  
Thrombosis Model ◽  
Rat Mesentery ◽  
Student’S T ◽  
Activator Inhibitor ◽  
Pai 1

SummaryTo investigate the role of active plasminogen activator inhibitor 1 (PAI-1) in the evolution of a microthrombus generated in the arteriolar microcirculation, the monoclonal antibody, 33H1F7, which transforms active PAI-1 to a tissue type plasminogen activator (t-PA) substrate, was evaluated in an arteriolar thrombosis model in the rat mesentery. Arterioles (200-300 μm) were stimulated electrically to create an endothelial lesion; ADP was then perfused for 2 min to induce the formation of a platelet-rich thrombus which lysed spontaneously in 140 ± 24 s. Two successive ADP superfusions produced comparable thrombi which lysed in comparable times. Different doses of 33H1F7 were infused to rats for 30 min and the dose which inactivates rapidly and totally active rat PAI-1 (300 μg/kg/min) was selected to be tested on the thrombosis model. Infusion of 33H1F7 beginning 10 min before the ADP application significantly reduced the lysis time in comparison to the control (123 ± 30 s versus 169 ± 33 s, P < 0.05, paired Student’s t-test) and the cumulative thrombus area during the lysis period was decreased by 56 ± 7%. These results demonstrate that inactivation of PAI-1 is able to accelerate lysis of a platelet-rich clot in a mesenteric arteriole of the rat. Thus active PAI-1 most likely participates to the resistance to thrombolysis in the arteriolar microcirculation and its inactivation may shorten ischemic periods after microvascular obstruction such as e.g. during cerebral stroke.

PURIFICATION AND CHARACTERIZATION OF REACTIVE AND NON-REACTIVE PLASMINOGEN ACTIVATOR INHIBITOR-1 (PAI-1) FROM HUMAN PLACENTA

1987 ◽  
Author(s):  
M Philips ◽  
A G Juul ◽  
S Thorsen ◽  
J Selmer ◽  
L Thim
Keyword(s):  
Monoclonal Antibody ◽  
Plasminogen Activator ◽  
Human Placenta ◽  
Solid Phase ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Single Chain ◽  
Plasminogen Activator Inhibitor 1 ◽  
Sds Page ◽  
Pai 1

Reactive and non-reactive forms of PAI-1 have been identified in various biological materials. The structural differences between these forms remain to be determined.A monoclonal antibody specific for a non-reactive PAI-1 and a monoclonal antibody reacting with both the reactive and nonreactive form of the inhibitor were obtained by immunization with a tissue-type plasminogen activator (t-PA)-PAI-1 complex (Philips et al., Thromb Haemostas 1986; 55:213-7). These antibodies were used for the isolation of reactive and non-reactive PAI-1 by solid-phase immunoadsorption from extracts of human placenta. The inhibitor preparations were further purified by HPLC. Reactive and non-reactive PAI-1 both migrated with a Mr ∼ 52,000 when analyzed by SDS-PAGE. Furthermore, the two inhibitor forms were indistinguishable by N-terminal sequence analysis. Two N-terminal sequences were found in about equal ammounts for both the reactive and non-reactive PAI-1. They were Ser-Ala-Val-His-His-Pro-Pro- and a two residues shorter sequence (Val-His-His-Pro-Pro-). These sequences are in agreement with the published cDNA sequence of PAI-1 and shows that the inhibitor is N-terminally heterogeneously processed. The second order rate constant (ki) for the reaction between reactive PAI-1 and single-chain t-PA was about 6 106 M-1s-1. Treatment with 4 M guanidinium-HCl partially converted the non-reactive PAI-1 to a reactive form exhibiting a similar k1 for inhibition of single-chain t-PA. SDS-PAGE showed that the t-PA-PAI-1 complex could be dissociated by 1,5 M NH4OH/ 39 mM SDS resulting in the release of a PAI-1 with approximately the same Mr as native PAI-1. This indicates either that t-PA does not cleave the inhibitor or that it cleaves a peptide bond close to the C-terminus.In conclusion a non-reactive and a reactive form of PAI-1 can be purified from placenta. The two forms are distinguishable by monoclonal antibodies but they show similar Mr′ls and the same N-terminal sequences.

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