Fibrin Lysis Research Articles

Enzymotherapy of toxic anterior segment syndrome after phacoemulsification

Purpose. To investigate the efficacy of recombinant prourokinase (RPU) treatment in patients with toxic anterior segment syndrome after phacoemulsification.
 Material and methods. We observed 123 patients (123 eyes) with toxic anterior segment syndrome after phacoemulsification; patients of the group I (n = 30) received only antiinflammatory treatment; in treatment of patients of the group II (n = 31), instillations of the RPU solution were used, in the group III (n = 31), RPU solution was injected subconjunctivally, in the group IV (n = 31) RPU solution electrophoresis was used. Treatment result analysis was carried out within 30 days.
 Results. Initial mean visual acuity in groups was 0.09 0.04; 0.1 0.04; 0.09 0.04; 0.08 0.04, and was virtually the same (p 0.05). In 24 hours after treatment initiation, mean visual acuity in the group III was higher, than in the others. In three days and up to the end of observation period, the lowest mean visual acuity was noted in the group I (p 0.05). Anterior chamber assessment showed that beginning from the first 24 hours after treatment initiation, in groups III and IV, fibrin lysis in the anterior chamber was more pronounced, than in groups I and II (p 0.05); by the end of the observation period, worst indices of anterior chamber state were found in the group I (p 0.05), in other groups, they were almost identical (p 0.05). When using RPU, no allergic reaction was noted.
 Conclusions. RPU use in combined toxic anterior segment syndrome therapy after phacoemulsification allows increasing visual acuity, reducing convalescence time, and reducing the number of laser dissections. It was established that all methods of RPU administration are effective. RPU may be administered as eye drops on an outpatient basis, receiving efficacy similar to other administration methods.

The clinical relevance of altered fibrinogen packaging in the presence of 17β-estradiol and progesterone

BackgroundThe effect of endogenous hormone concentrations, specifically 17β-estradiol and progesterone, on fibrin network formation has not been established. ObjectivesIt is essential to understand natural hormone mechanisms since these hormones are still present in circulation while hormonal contraceptives, which are associated with increased risk of venous thromboembolism, are used. MethodsDue to the fact that these hormones are known to increase hypercoagulability and the prothrombotic state scanning electron microscopy (SEM), atomic force microscopy (AFM), thromboelastography (TEG) and turbidimetry were employed to investigate the morphology, surface roughness, viscoelastic properties and formation and lysis of fibrin. Results17β-estradiol and progesterone showed hypercoagulable viscoelastic properties and decreased the diameter and surface roughness of fibrin while increasing dense matted deposit occurrence. Our results suggest that the additional burden of hormonal load, together with the presence of endogenous estrogen and progesterone, may result in a prothrombotic and hypercoagulable state in females with an inflammatory predisposition. ConclusionOur results are of clinical importance when considering hormones as either pathological agent or therapeutic intervention as will be assessed in future investigation.

Plasminogen fragments K 1-3 and K 5 bind to different sites in fibrin fragment DD.

Specific plasminogen-binding sites of fibrin molecule are located in Аα148-160 regions of C-terminal domains. Plasminogen interaction with these sites initiates the activation process of proenzyme and subsequent fibrin lysis. In this study we investigated the binding of plasminogen fragments K 1-3 and K 5 with fibrin fragment DD and their effect on Glu-plasminogen interaction with DD. It was shown that the level of Glu-plasminogen binding to fibrin fragment DD is decreased by 50-60% in the presence of K 1-3 and K 5. Fragments K 1-3 and K 5 have high affinity to fibrin fragment DD (Kd is 0.02 for K 1-3 and 0.054 μМ for K 5). K 5 interaction is independent and K 1-3 is partly dependent on C-terminal lysine residues. K 1-3 interacts with complex of fragment DD-immobilized K 5 as well as K 5 with complex of fragment DD-immobilized K 1-3. The plasminogen fragments do not displace each other from binding sites located in fibrin fragment DD, but can compete for the interaction. The results indicate that fibrin fragment DD contains different binding sites for plasminogen kringle fragments K 1-3 and K 5, which can be located close to each other. The role of amino acid residues of fibrin molecule Аα148-160 region in interaction with fragments K 1-3 and K 5 is discussed.

THE CHARACTERISTIC OF CHANGES OF THE FIBRINOLYTIC AND PROTEOLYTIC ACTIVITY OF THE BLOOD PLASMA AND TISSUES OF THE INTERNAL ORGANS IN THE BLINDED HYPOTHYROID RATS

In experiments on the blinded (enucleated) male albino rats with hypothyroidism increasing of indicators of fibrinolytic activity, reducing of the lysis of low and high molecular weight proteins and activation of сolagenolysis in blood plasma were found. Significant intensification of process of lysis of fibrin in the heart and liver tissues occurred against the backdrop of the decreasing the background of non-enzymatic and enzymatic fibrinolysis in lung tissue.

Ex vivo recapitulation of trauma-induced coagulopathy and preliminary assessment of trauma patient platelet function under flow using microfluidic technology.

Relevant to trauma-induced coagulopathy diagnostics, microfluidic assays allow controlled hemodynamics for testing of platelet and coagulation function using whole blood. Hemodilution or hyperfibrinolysis was studied under flow with modified healthy whole blood. Furthermore, platelet function was also measured using whole blood from trauma patients admitted to a Level I trauma center. Platelet deposition was measured with PPACK-inhibited blood perfused over collagen surfaces at a wall shear rate of 200 s, whereas platelet/fibrin deposition was measured with corn trypsin inhibitor-treated blood perfused over tissue factor (TF)/collagen. In hemodilution studies, PPACK-treated blood displayed almost no platelet deposition when diluted to 10% hematocrit with saline, platelet-poor plasma, or platelet-rich plasma. Using similar dilutions, platelet/fibrin deposition was essentially absent for corn trypsin inhibitor-treated blood perfused over TF/collagen. To mimic hyperfibrinolysis during trauma, exogenous tissue plasminogen activator (50 nM) was added to blood before perfusion over TF/collagen. At both venous and arterial flows, the generation and subsequent lysis of fibrin were detectable within 6 minutes, with lysis blocked by addition of the plasmin inhibitor, ε-aminocaproic acid. Microfluidic assay of PPACK-inhibited whole blood from trauma patients revealed striking defects in collagen response and secondary platelet aggregation in 14 of 21 patients, whereas platelet hyperfunction was detected in three of 20 patients. Rapid microfluidic detection of (1) hemodilution-dependent impairment of clotting, (2) clot instability because of lysis, (3) blockade of fibrinolysis, or (4) platelet dysfunction during trauma may provide novel diagnostic opportunities to predict trauma-induced coagulopathy risk.

Dynamics of changes in the activation of blood coagulation tests at different variants of thromboprophylaxis

Dynamics of changes in the activation of blood coagulation tests at different variants of thromboprophylaxis

Peroxynitrite and fibrinolytic system—The effects of peroxynitrite on t-PA-induced plasmin activity

The aim of the present study was the investigation of peroxynitrite (ONOO−) effects on fibrinolysis in vitro and in silico. The exposure of human plasminogen to ONOO− (10–1000μM) resulted in a decrease of t-PA-induced amidolytic activity of plasmin; the inhibitory effect was associated with the increasing level of 3-nitrotyrosine in plasminogen/plasmin molecule. Furthermore, ONOO− displayed both the ability to impair the t-PA-induced activation of plasminogen to plasmin, and to reduce the rate of fibrin lysis by plasmin. The susceptibility of plasminogen in blood plasma to nitrative action of ONOO− was revealed by the immunoprecipitation technique. To confirm the hypothesis that 3-nitrotyrosine generation is crucial for the impairment of plasmin activity, (−)-epicatechin, a polyphenolic antioxidant that selectively prevents tyrosine nitration, was used both for in vitro experiments as well as for in silico studies on ONOO−, ONOOH and (−)-epicatechin binding and plasminogen nitration. (−)-Epicatechin effectively protected plasminogen against ONOO−-induced inactivation and significantly reduced the level of 3-nitrotyrosine. The obtained results revealed tyrosine nitration as the most likely mechanism of the inhibitory effect of ONOO− on plasmin(ogen) functions. The possible role of tyrosine modifications was additionally confirmed by bioinformatics calculations with indication of nitration susceptible tyrosine residues.

Effect of von Willebrand factor on clot structure and lysis.

Von Willebrand Factor (vWF) is constitutively secreted by the endothelium and incorporated in the fibrin clots under slow clotting conditions. The aim of the present work was to study the effect of vWF on clot structure and lysis. Purified fibrinogen was mixed with vWF or Tris-buffered saline and clotted with thrombin - activated factor XIII. Fibrin polymerization was followed by turbidity at 350 nm during 2.5 h. After this time, plasmin was added on the top of the clots, and the optical density (OD) was read until baseline values. vWF effect on network[Combining Acute Accent]s porosity was evaluated by permeation using the same clotting conditions as for fibrin polymerization. Clot structure was visualized and analyzed by laser scanning confocal microscopy (LSCM). The rate of fibrin polymerization was 1.47 mOD/s in the presence of vWF and 0.5 mOD/s when vWF was not added (P < 0.05). The fibrin lysis rate was approximately four times faster when vWF was added to fibrinogen. The fibrin network porosity was (20.4 ± 1.6) × 10 cm with vWF and (8.3 ± 1.2) × 10 cm without external vWF (P < 0.05). The analysis of LSCM images showed that vWF increased fibrin fibers diameter and the networks[Combining Acute Accent] pores size. In conclusion, vWF covalently crosslinked to fibrin modify its structure (increases fibrin diameter and the pores filling space of the meshwork) that accelerates the fibrin lysis rate.

Isolation and identification of a novel fibrinolytic Bacillus tequilensis CWD-67 from dumping soils enriched with poultry wastes.

A newly isolated strain, CWD-67, which exhibited high fibrinolytic activity, was screened from dumping soils enriched with poultry wastes. The strain was identified as Bacillus tequilensis (KF897935) by 16Sr RNA gene sequence analysis and biochemical characterization. A fibrinolytic enzyme was purified to homogeneity from the culture supernatant using ammonium sulfate precipitation, membrane concentration, dialysis, ion-exchange, and gel filtration chromatography. SDS-PAGE analysis showed that the purified enzyme was a monomeric protein with an apparent molecular weight of 22 kDa, which is the lowest among Bacillus fibrinolytic enzymes reported to date. The purified enzyme was confirmed to have fibrinolytic activity by a fibrin zymogram. The optimal pH and temperature values of the enzyme were 8.0 and 45 °C, respectively. The enzyme was completely inhibited by PMSF and significantly inhibited by EDTA, TPCK, Co(2+), Zn(2+), and Cu(2+), suggesting a chymotrypsin-like serine metalloprotease. In vitro assays revealed that the purified enzyme could catalyze fibrin lysis effectively, indicating that this enzyme could be a useful fibrinolytic agent.

Abstract 16668: The Effects of Dabigatran on Fibrin Network Structure and Lysis: A Simple Technique to Monitor Efficacy of Treatment?

Background: Thrombus formation represent the final step in the atherothrombotic process and occurs secondary to a complex interaction between platelets and coagulation factors. Increased platelet reactivity and enhanced clot formation can both determine predisposition to vascular events. Direct platelet inhibition by modern antiplatelet therapy is now effective at controlling the cellular component of coagulation but the fibrin network is not generally targeted in arterial disease. Our aim was to investigate the effects of dabigatran on fibrin network characteristics in individuals dual antiplatelet therapy (DAPT). Methods: This was a prospective, randomized, double-blind study conducted in 30 patients on maintenance (at least 30 days) DAPT with aspirin (81mg/day) and clopidogrel (75mg/day). Patients were randomized to receive either dabigatran 150mg bid or matching placebo for 7 days. Fibrin clot properties were studied using a validated turbidimetric assay and the following parameters were recorded: lag phase and time to full clot formation, both of which measure clotting potential; clot maximum absorbance, representing fibrin network density and fibre thickness; clot lysis time to assess fibrinolytic efficiency. Results: Dabigatran treatment was associated with an increase in lag phase, compared with baseline, of 1013±167 sec , whereas no difference was detected in placebo treated patients (-17±14 sec, p&lt;0.001 for dabigatran vs placebo). Similar results were obtained for clot formation time with an increase of 240±37 sec in dabigatran treated patients with no change following placebo treatment (-24±18 sec; p&lt;0.001 comparing the two groups). The change in clot maximum absorbance after dabigatran and placebo treatment showed no differences (-0.02±0.01 and 0.00±0.01 au, respectively; p=0.2) and clot lysis time was similar. Conclusions: In patients on DAPT with aspirin and clopidogrel, additional treatment with dabigatran delays fibrin clot formation without affecting fibrinolysis. Therefore, triple therapy with dabigatran may be one option to reduce thrombosis potential in high risk individuals. Moreover, our assay, which can be conducted on stored samples, offers the opportunity to monitor response to dabigatran therapy.

Residual perfusion defects in patients with pulmonary embolism are related to impaired fibrinolytic capacity

BackgroundFew studies investigated the relationship between fibrinolysis abnormalities and residual pulmonary perfusion defects after acute pulmonary embolism (PE). ObjectiveTo assess the fibrinolytic profile in patients with prior PE in relation to the extent of scintigraphically detectable residual perfusion abnormalities. Patients and methodsWe studied 71 consecutive patients with a prior episode of PE, who were examined after one year of the incident embolic event, and at least one month after anticoagulation withdrawal. They underwent lung scintigraphy to assess the recovery of pulmonary perfusion, echocardiography and chest radiography to look for signs of pulmonary hypertension. Clot formation and lysis were evaluated by two turbidimetric methods: Clot and Lysis Assay and Clot Lysis Time. We also measured the in vitro plasmin-mediated lysis of fibrin from purified fibrinogen, and the circulating levels of fibrinolytic inhibitors. The sample was split in two categories based on the extent of residual perfusion defects: <10% (n=53), ≥10% (n=18). ResultsPatients with perfusion defects >10% had significantly longer lysis time (p<0.05), and higher levels of plasminogen activator inhibitor-1 (p<0.01) than those with perfusion defects <10%. The time interval between symptoms onset and PE diagnosis (time-to-diagnosis) was significantly longer in patients with perfusion defects >10% than in the others (p=0.005). In multivariate logistic regression, both lysis time and time-to-diagnosis were independently associated with perfusion defects >10% (p<0.001). None of the sampled patients had echocardiographic or radiologic signs of pulmonary hypertension. ConclusionProlonged time-to-diagnosis and fibrinolysis imbalance are independent predictors of incomplete perfusion recovery after acute PE.

The possible role of hydrogen sulfide as a modulator of hemostatic parameters of plasma

Hydrogen sulfide (H2S) is a well known toxic gas at high levels. However, at physiological levels, H2S may play a role in the pathogenesis of various cardiovascular diseases. The objective was to study the effects of exogenous H2S on the hemostatic parameters (coagulation and fibrinolytic activity) of human plasma. Human plasma was incubated (5, 15 and 30min) with NaHS as a H2S donor at the final concentration of 0.01–100μM. Hemostatic factors, such as maximum velocity of clot formation, fibrin lysis half-time, the activated partial thromboplastin time (APTT), prothrombin time (PT), and thrombin time (TT) were estimated. Moreover, the aim of our study was to establish the influence of NaHS (10μM; 5, 15 and 30min) on the clot formation using the purified fibrinogen. We demonstrated that coagulation/fibrinolytic properties of human plasma incubated with NaHS were changed. APPT, PT and TT of plasma treated with NaHS at tested concentrations – 0.01–100μM were prolonged. We observed that NaHS (0.01–100μM) reduced fibrin polymerization in whole plasma and 10μM NaHS also reduced polymerization of purified fibrinogen. In the presence of NaHS (at the low tested concentration – 1μM) the decrease was about 18% (in plasma, p<0.05). Our experiments also showed that NaHS (0.01–100μM) stimulated the fibrin lysis in whole plasma. However, the time-dependent (5, 15 and 30min) reduction of fibrin/fibrinogen polymerization and stimulation of fibrin lysis by NaHS (10μM) was not observed. In conclusion, the present study demonstrates the anticoagulant properties of exogenous H2S in vitro.

Oxidative modification of fibrinogen is associated with altered function and structure in the subacute phase of myocardial infarction.

Among plasma proteins, fibrinogen represents a major target of oxidative modifications. In patients with post-acute myocardial infarction (6 months after the acute event), fibrinogen oxidation-induced carbonyls and fibrinogen function were estimated using in vitro and ex vivo approaches. Fibrinogen structural features and clot architecture were also explored. In 39 patients with post-acute myocardial infarction and 28 age-, sex-, and risk factor-matched controls, oxidative stress markers (in plasma and in purified fibrinogen fractions), thrombin-catalyzed fibrin polymerization, and plasmin-induced fibrin lysis were estimated. Circular dichroism spectra of purified fibrinogen extracts, electron microscopy, and differential interference contrast microscopy analyses of fibrin clots were also performed. Marked signs of oxidative stress in plasma (P<0.01 versus controls) and, correspondingly, an increased extent of fibrinogen carbonylation (3.5-fold over control values; P<0.01 versus controls) were observed in patients. Furthermore, fibrinogen fractions purified from patients exhibited significantly reduced clotting ability and decreased susceptibility to plasmin-induced lysis (P<0.01 versus controls). Alterations in fibrinogen secondary structure, as suggested by circular dichroism spectroscopy, and in fibrin clot architecture, as analyzed by electron and differential interference contrast microscopy, were also identified. Here, we report for the first time that patients with post-acute myocardial infarction present with an overall imbalance in redox status and marked fibrinogen carbonylation associated with altered fibrinogen function, thus suggesting a role for carbonylation as a direct mechanism of fibrinogen function. The observed features occur along with modifications in protein structure and in clot architecture.

Plasma Fibrin Degradation Products (Fdp ) In Patients With Pre Eclamptic Toxaemia (Pre-eclampsia)

Plasma fibrin degradation products (F.D.P.) assay measures the amount of fibrin split products in the blood. The high levels of F.D.P. in pre-eclampsia suggests that abnormal amount of degradation products are most likely due to localized lysis of fibrin in the uterine vascular compartment. The present case control study was designed to determine the relationship between plasma fibrin degradation products and pre-eclampsia. The study was carried out in the department of Biochemistry, Chittagong Medical College during the period of September 2011 to August 2012. The samples were collected from the department of Obstetrics and Gynecology, Chittagong Medical College Hospital, age from 18- 40 years. The data were collected by a structured questionnaire including age, socioeconomic condition, gravida, para, edema, blood pressure, proteinuria, history of hypertension, family history of pre-eclampsia and diabetes mellitus. Considering the cost of experiment and length of time, total 60 patients were included in this study. Among them 40 were considered as case (Blood Pressure &gt; or = 140/90 mm of Hg and proteinuria &gt; 0.3 gram/day) and 20 were considered as control (Blood Pressure &lt; 140/90 mm of Hg and no proteinuria). Plasma fibrin degradation products were measured in all samples. Study showed that plasma FDP were more increased in pre-eclamptic patients than that of normal pregnancy (21.52 + 16.47 ugm/ml Vs 10.63 + 7.12 ugm/ml), P = &lt;0.01. Result showed that percentage of raised plasma FDP is more in preeclamptic patients (100%) than that of normal pregnancy (75%), P= &lt;0.01. Pearson's Correlation Coefficient (r) showed that there were positive correlation between blood pressure and plasma FDP (systolic blood plessure and plasma FDP, r=0.221,P=&gt;.0.05 and diastolic blood pressure and plasma FDP, r=0.285,P=&lt;0.05). Significantly raised level of plasma F.D.P. in pre-eclampsia help to formulate a management plan and thereby reduce the complications of this disease. JCMCTA 2013; 24 (1):9-13

Fibrinolytic Mechanisms of tPA, prouPA, Mutant prouPA and Their Implications for Therapeutic Thrombolysis

Experience with thrombolysis for 25 years has been essentially limited to tissue plasminogen activator (tPA). Therefore, tPA has been perceived as synonymous with all thrombolysis. However, tPA has a unique mode of action different from that of urokinase plasminogen activator (uPA), whose native form is prouPA. At the pharmacological concentrations, both physiological activators induced unanticipated effects that were also different. Reperfusion with tPA is almost twice as effective as streptokinase, the former thrombolytic standard. Nevertheless, three mega-trials with a total of 94,740 patients were required to show a significant difference in acute myocardial infarction. Surprisingly, intracranial hemorrhage was also significantly more with tPA. These paradoxical findings can be explained by tPA’s rethrombosis rate undermining its efficacy, and lysis of hemostatic fibrin causing tPA bleeding. By contrast, prouPA has a low rethrombosis rate and it spares hemostatic fibrin by virtue of its different mechanism of fibrin-bound plasminogen activation. Unfortunately, these potential advantages are out of reach in practice since at high therapeutic concentrations in blood, prouPA is vulnerable to non-specific conversion to uPA, a non-specific plasminogen activator that induces a hemorrhagic diathesis. A single-site mutant, M5, was designed to correct this prouPA problem. In vitro and in vivo studies showed that M5 clot lysis was more effective than prouPA or tPA and was accompanied by a sparing of hemostatic fibrin. When M5 administration was preceded by a bolus of plasma C1-inhibitor, it further improved the plasma stability of M5 and permitted a maximum clot lysis rate to be achieved without fibrinogenolysis.

Lytic and mechanical stability of clots composed of fibrin and blood vessel wall components

BackgroundProteases expressed in atherosclerotic plaque lesions generate collagen fragments, release glycosaminoglycans (chondroitin sulfate [CS] and dermatan sulfate [DS]) and expose extracellular matrix (ECM) proteins (e.g. decorin) at sites of fibrin formation.ObjectiveHere we address the effect of these vessel wall components on the lysis of fibrin by the tissue plasminogen activator (tPA)/plasminogen system and on the mechanical stability of clots.Methods and resultsMMP-8-digested collagen fragments, isolated CS, DS, glycosylated decorin and its core protein were used to prepare mixed matrices with fibrin (additives present at a 50-fold lower mass concentration than fibrinogen). Scanning electron microscopy (SEM) showed that the presence of ECM components resulted in a coarse fibrin structure, most pronounced for glycosylated decorin causing an increase in the median fiber diameter from 85 to 187 nm. Rheological measurements indicated that these structural alterations were coupled to decreased shear resistance (1.8-fold lower shear stress needed for gel/fluid transition of the clots containing glycosylated decorin) and rigidity (reduction of the storage modulus from 54.3 to 33.2 Pa). The lytic susceptibility of the modified fibrin structures was increased. The time to 50% lysis by plasmin was reduced approximately 2-fold for all investigated ECM components (apart from the core protein of decorin which produced a moderate reduction of the lysis time by 25%), whereas fibrin-dependent plasminogen activation by tPA was inhibited by up to 30%.ConclusionECM components compromise the chemical and mechanical stability of fibrin as a result of changes in its ultrastructure.

Chronic Spontaneous Urticaria: the Emerging Role of Coagulation

Chronic urticaria (CU) is a cutaneous disease characterized by the recurrent eruption of short-lived wheals for at least six weeks. The wheals, which are surface swelling of the dermis accompanied by itching, may be elicited by different stimuli, or occur spontaneously. The pathogenesis of spontaneous CU has long remained a mystery, with food allergy, intolerance to food additives and chronic infections having been indicated as the main causes of the disease. The demonstration of histamine releasing autoantibodies in the serum of CU patients led to consider it of autoimmune origin in about 45 % of cases, this condition being “idiopathic” in the remaining 55 %. We recently evaluated the possible involvement of blood coagulation in its pathophysiology by measuring in CU patients the plasma levels of prothrombin fragment F1 + 2, a polypeptide released into the circulation during the activation of prothrombin to thrombin, and D-dimer, a marker of fibrin lysis. Interestingly, both F1 + 2 and D-dimer plasma levels were extremely high during the acute phase of disease, but were completely normal after remission, paralleling urticaria severity. It is noteworthy that thrombin causes an increase in vascular permeability, thus amplifying the CU inflammatory network. Moreover, immunohistochemical studies showed expression of tissue factor, the main initiator of coagulation, in CU skin lesions, and co-localization experiments demonstrated that tissue factor is expressed by eosinophils in the inflammatory infiltrate. All these findings support that coagulation activation contributes in the pathophysiology of spontaneous CU, and provide the rationale for using also anticoagulant and antifibrinolytic agents to treat it.

Fibrin Clot Structure and Mechanics Associated with Specific Oxidation of Methionine Residues in Fibrinogen

Using a combination of structural and mechanical characterization, we examine the effect of fibrinogen oxidation on the formation of fibrin clots. We find that treatment with hypochlorous acid preferentially oxidizes specific methionine residues on the α, β, and γ chains of fibrinogen. Oxidation is associated with the formation of a dense network of thin fibers after activation by thrombin. Additionally, both the linear and nonlinear mechanical properties of oxidized fibrin gels are found to be altered with oxidation. Finally, the structural modifications induced by oxidation are associated with delayed fibrin lysis via plasminogen and tissue plasminogen activator. Based on these results, we speculate that methionine oxidation of specific residues may be related to hindered lateral aggregation of protofibrils in fibrin gels.

Coagulation and fibrynolitic parameters in women and the effects of hormone therapy; comparison of transdermal and oral administration

It is established that hormone therapy (HT) is related with significant increased prothrombotic risk factor. The aim of our study was to assess the effects of oral hormone therapy (o-HT) and transdermal hormone therapy (t-HT) on hemostasis parameters: fibrinogen (Fg) concentration, the maximum velocity of polymerization of clot formation, fibrin half-time lysis, plasma level of thrombin inhibitor of fibrinolysis (TAFI) and activity of generated thrombin and plasmin amidolytic activity. We observed that values of initial velocity of polymerization in o-HT group were increased (94.64 mOD/min vs. 131.50 mOD/min, p < 0.001) compared to control group. Fibrin lysis half-time increased in both groups with HT (controls - 18.26 min vs. 32.43 min (o-HT); 23.34 min transdermal hormone therapy (t-HT) p < 0.001) compared to controls. The activity of thrombin was statistically higher in plasma of women after o-HT (72.6 ± 8.5 mOD/min) than in patients with t-HT (53.7 ± 10.1 mOD/min) and controls (51.2 ± 10 mOD/min. Plasmin activity was the highest in controls (84.5 ± 10.2 mOD/min). The highest level of TAFI we observed in patients after oral hormones (80.38 ± 8.23%); women on transdermal HT had 61.58 ± 9.81% and the lowest concentration of TAFI we noted in controls 44.70 ± 10.16). The results of our study show that HT may partly explain the increase in venous thrombosis (VTE) and cardiovascular events reported after the use of it, especially the oral form of treatment.

Clot penetration and retention by plasminogen activators promote fibrinolysis

Tissue-type plasminogen activator (tPA) remains the sole thrombolytic approved by the FDA for the treatment of pulmonary embolism (PE). tPA has not been replaced by third generation plasminogen activators, e.g. Reteplase (Ret) and Tenecteplase (TNK) that circulate with longer life-spans and in theory should have more extended potency in vivo. One reason for this paradox is the inability to assign units of activity to plasminogen activators based on specific biologically relevant standards, which impairs objective comparison. Here, we compare clot permeation, retention and fibrinolytic activities of tPA, TNK and Ret in vitro and clot composition over time with outcome in a mouse model of disseminated pulmonary microembolism (ME). When clots were incubated in the continuous presence of drug, tPA, TNK and Ret lysed fibrin clots identically in the absence of PA inhibitor-1 (e.g. PAI-1). Ret, which has lower fibrin affinity and greater susceptibility to inhibition by PAI-1 than tPA, was less effective in lysing plasma clots, while TNK was less effective when the fibrin content of the clots was enhanced. However, when clots were afforded only brief exposure to drug, as occurs in vivo, Ret showed more extensive clot permeation, greater retention and lysis than tPA or TNK. These results were reproduced in vivo in a mouse model of ME. These studies indicate the need for more relevant tests of plasminogen activator activity in vitro and in vivo and they show that clot permeation and retention are important potential predictors of clinical utility.