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Anetta Undas - One of the best experts on this subject based on the ideXlab platform.
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plasma fibrin clots of pulmonary embolism patients present increased amounts of factor xiii and alpha2 Antiplasmin at 3 months anticoagulation since the acute phase
Journal of Physiology and Pharmacology, 2020Co-Authors: Michal Zabczyk, Agata Hanna Bryk, Joanna Natorska, Zsuzsa Bagoly, Barbara Barath, Éva Katona, Ferenc Sarkady, Katharina Zettl, J R Wisniewski, Anetta UndasAbstract:Fibrin cross-linking by coagulation factor (F)XIII leads to clot stabilization. Reduced plasma FXIII levels have been reported in acute pulmonary embolism (PE) patients. We investigated the impact of anticoagulant therapy on clot-bound amounts of FXIII and α2-Antiplasmin and their associations with fibrin clot properties in patients with PE. Clots generated from plasma of 18 acute symptomatic patients on admission and after a 3-month treatment with rivaroxaban were assessed off anticoagulation using mass spectrometry. Plasma FXIII and α2-Antiplasmin activity were determined at the 2 time points along with thrombin generation markers, plasma fibrin clot permeability (Ks), and clot lysis time (CLT). Following anticoagulant therapy, clot-bound FXIII increased from 2.97 (interquartile range, 1.98 - 4.08) to 4.66 (3.5 - 6.9) mg/g protein and α2-Antiplasmin from 9.4 (7.2 - 10.6) to 11 (9.5 - 14) mg/g protein (both p < 0.0001). The two parameters showed positive correlation at baseline only (r = 0.63, p = 0.0056). Similarly to clot-bound amounts, plasma FXIII (+25.8%) and α2-Antiplasmin activity (+12%) increased at 3 months. Plasma FXIII activity on admission, but not after 3 months since the index PE, was associated with amounts of clot-bound FXIII (r = 0.35, p = 0.043) and α2-Antiplasmin (r = 0.47, p = 0.048). At baseline, clot-bound FXIII correlated with plasma F1+2 prothrombin fragments levels (r = 0.51, p = 0.03), while clot-bound α2-Antiplasmin correlated with CLT (r = 0.43, p = 0.036). At 3 months associations of clot-bound FXIII and α2-Antiplasmin were abolished. This study assessed for the first time changes in the fibrin clot composition following acute PE, suggesting an increase of clot-bound and plasma FXIII and α2-Antiplasmin levels after 3 months.
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Interaction of glycated and acetylated human α2-Antiplasmin with fibrin clots.
Blood coagulation & fibrinolysis : an international journal in haemostasis and thrombosis, 2020Co-Authors: Agata Hanna Bryk, Dorota Satala, Joanna Natorska, Maria Rąpała-kozik, Anetta UndasAbstract:: In type 2 diabetes mellitus (T2DM), increased α2-Antiplasmin incorporation in fibrin and impaired fibrinolysis have been reported. Acetylsalicylic acid (ASA), used in cardiovascular prevention, modulates fibrinolysis and exerts weaker therapeutic effect in this disease. We investigated how glycation and acetylation of α2-Antiplasmin affects its interaction with fibrin. Using surface plasmon resonance, we analyzed fibrin binding by α2-Antiplasmin incubated with no β-D-glucose or ASA (control); incubated with β-D-glucose (5, 10, 50 mmol/l); (3) incubated with 1.6 mmol/l acetylsalicylic acid (ASA) and (4) incubated with 1.6 mmol/l ASA and 50 mmol/l β-D-glucose. Incubation with glucose decreased affinity of α2-Antiplasmin for fibrin compared with control α2-Antiplasmin in a glucose concentration-depending manner. α2-Antiplasmin incubation with ASA did not affect its affinity to fibrin. α2-Antiplasmin incubation with ASA and glucose resulted in 4.2-fold increased affinity to fibrin compared with α2-Antiplasmin incubated with 50 mmol/l glucose (P
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Identification of glycated and acetylated lysine residues in human α2-Antiplasmin.
Biochemical and biophysical research communications, 2019Co-Authors: Agata Hanna Bryk, Dominik Cysewski, Michal Dadlez, Anetta UndasAbstract:Abstract Background The post-translational protein modification via lysine residues can significantly alter its function. α2-Antiplasmin, a key inhibitor of fibrinolysis, contains 19 lysine residues. Aim We sought to identify sites of glycation and acetylation in human α2-Antiplasmin and test whether the competition might occur on the lysine residues of α2-Antiplasmin. Methods We analyzed human α2-Antiplasmin (1) untreated; (2) incubated with increasing concentrations of β- d -glucose (0, 5, 10, 50 mM); (3) incubated with 1.6 mM acetylsalicylic acid (ASA) and (4) incubated with 1.6 mM ASA and 50 mM β- d -glucose, using the ultraperformance liquid chromatography system coupled to mass spectrometer. Results Eleven glycation sites and 10 acetylation sites were found in α2-Antiplasmin. Incubation with β- d -glucose was associated with glycation of 4 (K-418, K-427, K-434, K-441) out of 6 lysine residues, known to be important for mediating the interaction with plasmin. Glycation and acetylation overlapped at 9 sites in samples incubated with β- d -glucose or ASA. Incubation with concomitant ASA and β- d -glucose was associated with the decreased acetylation at all sites overlapping with glycation sites. At K-182 and K-448, decreased acetylation was associated with increased glycation when compared with α2-Antiplasmin incubated with 50 mM β- d -glucose alone. Although K-24 located in the proximity of the α2-Antiplasmin cleavage site, was found to be only acetylated, incubation with ASA and 50 mM β- d -glucose was associated the absence of acetylation at that site. Conclusion Human α2-Antiplasmin is glycated and acetylated at several sites, with the possible competition between acetylation and glycation at K-182 and K-448. Our finding suggests possibly relevant alterations to α2-Antiplasmin function at high glycemia and during aspirin use.
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Sex-specific alteration to α2-Antiplasmin incorporation in patients with type 2 diabetes.
Thrombosis research, 2019Co-Authors: Agata Hanna Bryk, Jakub Siudut, Elżbieta Broniatowska, Zsuzsa Bagoly, Barbara Barath, Éva Katona, Anetta UndasAbstract:Abstract Background Type 2 diabetes mellitus (T2DM) is associated with hypofibrinolysis and increased factor XIII-mediated α2-Antiplasmin incorporation into the fibrin clot. It is unclear whether there are sex-related differences in α2-Antiplasmin incorporation in relation to impaired clot lysis in T2DM. Aim We investigated α2-Antiplasmin incorporation into fibrin clots as a determinant of clot lysability in patients of both sexes with T2DM. Methods In a group of 113 T2DM patients, 54 (47.8%) of which were women, we investigated α2-Antiplasmin incorporation using an in-house sandwich enzyme-linked immunoassay and plasma clot lysis by turbidimetry, along with fibrinogen and thrombin generation using calibrated automated thrombogram and factor XIII activity. Results Female patients had 15.2% greater α2-Antiplasmin incorporation into the fibrin clot (p = 0.008) and slightly higher plasma α2-Antiplasmin concentration (p = 0.005) along with 8.4% longer time to 50% lysis (Lys50MA, p = 0.012) compared with men. Female patients had enhanced thrombin generation represented by shorter lag phase (p = 0.042), shorter time to peak (p = 0.033), and higher endogenous thrombin potential (p = 0.003) compared with men, while factor XIII activity was comparable between sexes (p = 0.085). On multivariate regression, patient sex and glycated hemoglobin (HbA1c) level were the predictors of α2-Antiplasmin incorporation in the entire patient group, while α2-Antiplasmin incorporation was associated with Lys50MA, as were fibrinogen, male sex and body-mass index. Conclusions This study suggests that a more compromised fibrinolysis in diabetic women when compared with men could be in part mediated by increased α2-Antiplasmin incorporation into the fibrin.
Paul Bernard Coughlin - One of the best experts on this subject based on the ideXlab platform.
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contribution of conserved lysine residues in the α2 Antiplasmin c terminus to plasmin binding and inhibition
Journal of Biological Chemistry, 2011Co-Authors: Trifina Sofian, Paul Bernard Coughlin, Ruby Hp Law, Anita J HorvathAbstract:α2-Antiplasmin is the physiological inhibitor of plasmin and is unique in the serpin family due to N- and C-terminal extensions beyond its core domain. The C-terminal extension comprises 55 amino acids from Asn-410 to Lys-464, and the lysine residues (Lys-418, Lys-427, Lys-434, Lys-441, Lys-448, and Lys-464) within this region are important in mediating the initial interaction with kringle domains of plasmin. To understand the role of lysine residues within the C terminus of α2-Antiplasmin, we systematically and sequentially mutated the C-terminal lysines, studied the effects on the rate of plasmin inhibition, and measured the binding affinity for plasmin via surface plasmon resonance. We determined that the C-terminal lysine (Lys-464) is individually most important in initiating binding to plasmin. Using two independent methods, we also showed that the conserved internal lysine residues play a major role mediating binding of the C terminus of α2-Antiplasmin to kringle domains of plasmin and in accelerating the rate of interaction between α2-Antiplasmin and plasmin. When the C terminus of α2-Antiplasmin was removed, the binding affinity for active site-blocked plasmin remained high, suggesting additional exosite interactions between the serpin core and plasmin.
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X-ray crystal structure of the fibrinolysis inhibitor α2-Antiplasmin
Blood, 2007Co-Authors: Ruby Hp Law, Trifina Sofian, Anita J Horvath, Wan-ting Kan, Corinne R Hitchen, Christopher G. Langendorf, Ashley M. Buckle, James C. Whisstock, Paul Bernard CoughlinAbstract:The serpin alpha(2)-Antiplasmin (SERPINF2) is the principal inhibitor of plasmin and inhibits fibrinolysis. Accordingly, alpha(2)-Antiplasmin deficiency in humans results in uncontrolled fibrinolysis and a bleeding disorder. alpha(2)-Antiplasmin is an unusual serpin, in that it contains extensive N- and C-terminal sequences flanking the serpin domain. The N-terminal sequence is crosslinked to fibrin by factor XIIIa, whereas the C-terminal region mediates the initial interaction with plasmin. To understand how this may happen, we have determined the 2.65A X-ray crystal structure of an N-terminal truncated murine alpha(2)-Antiplasmin. The structure reveals that part of the C-terminal sequence is tightly associated with the body of the serpin. This would be anticipated to position the flexible plasmin-binding portion of the C-terminus in close proximity to the serpin Reactive Center Loop where it may act as a template to accelerate serpin/protease interactions.
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Antiplasmin: the forgotten serpin?
The FEBS journal, 2005Co-Authors: Paul Bernard CoughlinAbstract:Much of the basic biochemistry of Antiplasmin was described more than 20 years ago and yet it remains an enigmatic member of the serine protease inhibitor (serpin) family. It possesses all of the characteristics of other inhibitory serpins but in addition it has unique N- and C-terminal extensions which significantly modify its activities. The N-terminus serves as a substrate for Factor XIIIa leading to crosslinking and incorporation of Antiplasmin into a clot as it is formed. Although free Antiplasmin is an excellent inhibitor of plasmin, the fibrin bound form of the serpin appears to be the major regulator of clot lysis. The C-terminal portion of Antiplasmin is highly conserved between species and contains several charged amino acids including four lysines with one of these at the C-terminus. This portion of the molecule mediates the initial interaction with plasmin and is a key component of Antiplasmin's rapid and efficient inhibitory mechanism. Studies of mice with targeted deletion of Antiplasmin have confirmed its importance as a major regulator of fibrinolysis and re-emphasized its value as a potential therapeutic target.
Colin Longstaff - One of the best experts on this subject based on the ideXlab platform.
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increased urokinase and consumption of α2 Antiplasmin as an explanation for the loss of benefit of tranexamic acid after treatment delay
Journal of Thrombosis and Haemostasis, 2019Co-Authors: Colin Longstaff, Matthew LockeAbstract:Essentials Delayed treatment with tranexamic acid results in loss of efficacy and poor outcomes. Increasing urokinase activity may account for adverse effects of late tranexamic acid treatment. Urokinase + tranexamic acid produces plasmin in plasma or blood and disrupts clotting. α2 -Antiplasmin consumption with ongoing fibrinolysis increases plasmin-induced coagulopathy. SUMMARY: Background Tranexamic acid (TXA) is an effective antifibrinolytic agent with a proven safety record. However, large clinical trials show TXA becomes ineffective or harmful if treatment is delayed beyond 3 h. The mechanism is unknown but urokinase plasminogen activator (uPA) has been implicated. Methods Inhibitory mechanisms of TXA were explored in a variety of clot lysis systems using plasma and whole blood. Lysis by tissue plasminogen activator (tPA), uPA and plasmin were investigated. Coagulopathy was investigated using ROTEM and activated partial thromboplastin time (APTT). Results IC50 values for antifibrinolytic activity of TXA varied from 1000 μmol L-1 depending on the system, but good fibrin protection was observed in the presence of tPA, uPA and plasmin. However, in plasma or blood, active plasmin was generated by TXA + uPA (but not tPA) and coagulopathy developed leading to no or poor clot formation. The extent of coagulopathy was sensitive to available α2 -Antiplasmin. No clot formed with plasma containing 40% normal α2 -Antiplasmin after short incubation with TXA + uPA. Adding purified α2 -Antiplasmin progressively restored clotting. Plasmin could be inhibited by aprotinin, IC50 = 530 nmol L-1 , in plasma. Conclusions Tranexamic acid protects fibrin but stimulates uPA activity and slows inhibition of plasmin by α2 -Antiplasmin. Plasmin proteolytic activity digests fibrinogen and disrupts coagulation, exacerbated when α2 -Antiplasmin is consumed by ongoing fibrinolysis. Additional direct inhibition of plasmin by aprotinin may prevent development of coagulopathy and extend the useful time window of TXA treatment.
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Differences between neonates and adults in carbohydrate sequences and reaction kinetics of plasmin and α2-Antiplasmin
Thrombosis research, 2002Co-Authors: Martin Ries, Richard L. Easton, Colin Longstaff, Martin Zenker, Howard R. Morris, Anne Dell, Patrick J. GaffneyAbstract:Abstract This study investigates reaction kinetics by slow-binding kinetics methods of both adult and fetal plasmin (Types 1 and 2) with adult and fetal α2-Antiplasmin. In addition, carbohydrate sequences of Fetal and Adult Plasminogen Types 1 and 2, as well as fetal and adult α2-Antiplasmin, were determined by mass spectrometric analysis. All curves of plasmin–α2-Antiplasmin interaction followed the same pattern, indicating reversible slow-binding inhibition with an initial loose complex and a following tight complex. Differences between fetal and adult plasmin reactions with α2-Antiplasmin were predominantly due to the initial loose complex. Values for Ki initial in the reaction with adult α2-Antiplasmin were 1.5 and 1.6 nM for Fetal Plasmin Types 1 and 2, respectively; compared to 0.3 and 0.7 nM for the corresponding adult types. Increasing concentrations of tranexamic acid resulted in a continuous increase of Ki initial until a plateau was reached which was similar for all plasmin types. Almost identical values could be obtained when fetal α2-Antiplasmin was used instead of adult α2-Antiplasmin. Mass spectrometric analyses of the glycans present on plasminogen revealed a higher level of truncated N-glycans on the fetal material compared to the adult. The O-glycans of fetal and adult plasminogen were closely similar and only minor differences were observed between N-glycans of fetal and adult α2-Antiplasmin. In conclusion, both fetal plasmin isoforms are less inhibited by α2-Antiplasmin compared to the adult plasmin variants. These findings are important for the understanding of the physiology of the fibrinolytic system in neonates and provide further evidence that differences in glycosylation could be associated with marked effects on protein function.
Desire Collen - One of the best experts on this subject based on the ideXlab platform.
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mechanisms of activation of mammalian plasma fibrinolytic systems with streptokinase and with recombinant staphylokinase
FEBS Journal, 1993Co-Authors: Desire Collen, Berthe Van Hoef, Bernhard Schlott, Manfred Hartmann, Karlheinz Guhrs, Roger H LijnenAbstract:The molecular basis of the marked interspecies variability in the response of plasma fibrinolytic systems to activation by streptokinase (SK) or recombinant staphylokinase (STAR) was studied using highly purified plasminogens and a,-Antiplasmins from five representative species (man, baboon, rabbit, dog and cow). Human plasminogen reacted rapidly and stoichiometrically with both SK and STAR to yield potent plasminogen activators (catalytic efficiencies, k,,,/K,, of 1 .O pM-' . s-' and 0.3 pM-' . sK', respectively). The complex with SK was insensitive to a,-Antiplasmin, which, however, rapidly inhibited the complex with STAR (second-order rate constant, k,.,,, of 8 X 10" M-' . s-'). In a system composed of a 0.06-ml 'z51-fibrin-labeled plasma clot submerged in 0.30 ml plasma, both SK and STAR had potent fibrinolytic properties, causing 50% clot lysis in 2 h (EC,,), with 120 nM and 13 nM, respectively. Clot lysis with SK was non-fibrin specific (residual fibrinogen < lo%), whereas lysis with STAR was highly fibrin specific (residual fibrinogen 76%). Canine plasminogen reacted avidly with SK, but SK was rapidly degraded: it reacted rapidly and quantitatively with STAR to form a potent plasminogen-activating complex (kcd,/Km of 0.4 pM-' SK') which was sensitive to neutralization by a,-Antiplasmin (k,,a,, of 6 X 10' M-' . s-'). In a canine plasma milieu, SK was relatively potent (EC,, 200 nM) and fibrin specific, whereas STAR was very potent (EC,, 1.3 nM) but poorly fibrin specific. Baboon and rabbit plasminogen did not form stable stoichiometric complexes with SK, but reacted stoichiometrically and quantitatively with STAR. The complexes with STAR, however, had low catalytic efficiencies for the activation of their autologous plasminogens (k,,JK,, 0.02 pM-' . s-') and reacted more slowly with a,-Antiplasmin (k,,,,, 5-10 X 10' M-' . s-'). Bovine plasminogen was virtually unreactive towards both SK and STAR as well as to their complexes with human plasminogen, as monitored by measurement of the initial activation rates. The resistance to fibrinogen degradation with STAR observed in the human system could be transferred to the canine system by reconstituting canine plasma, depleted of plasminogen and a,-Antiplasmin, with the human proteins. Conversely, the sensitivity to fibrinogen degradation of the canine system could be transferred to the human system by reconstituting depleted plasma with canine plasminogen and a,-Antiplasmin. It is concluded that the variability in the response of mammalian plasma fibrinolytic systems to activation with SK or STAR is determined mainly by the extent of complex formation of these compounds with plasminogen, by the catalytic efficiencies of the complexes for the activation of autologous plasminogen and by the rate of inhibition of these complexes by a,-Antiplasmin.
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interaction between staphylokinase plasmin ogen and alpha 2 Antiplasmin recycling of staphylokinase after neutralization of the plasmin staphylokinase complex by alpha 2 Antiplasmin
Journal of Biological Chemistry, 1993Co-Authors: Karen Silence, Desire Collen, H. Roger LijnenAbstract:Abstract Although the plasminogen activating equimolar complex of staphylokinase (STA) with human plasmin is very rapidly inhibited by alpha 2-Antiplasmin, STA is a potent fibrinolytic agent in a human plasma milieu which contains 1 microM alpha 2-Antiplasmin. In the present study, it was found that the complex of plasmin with recombinant STA (STAR), after neutralization with alpha 2-Antiplasmin, retained the full plasminogen activating potential of STAR when added to a plasminogen solution (93 +/- 5% residual activity). When added to human plasma containing a 125I-fibrin-labeled plasma clot, equi-effective concentrations (causing 50% lysis in 2 h) were 17 +/- 3.0, 13 +/- 1.0, and 20 +/- 1.0 nM for STAR, equimolar plasmin-STAR mixtures, and plasmin-STAR mixtures neutralized by alpha 2-Antiplasmin, respectively. Gel filtration of mixtures of plasmin(ogen) and STAR revealed elution as plasmin-STAR complex (Mr approximately 100,000), whereas after addition of alpha 2-Antiplasmin, STAR eluted with an apparent Mr of 20,000. When mixtures of plasmin and STAR were adsorbed to lysine-Sepharose, STAR adsorbed quantitatively (96 +/- 1%) to the gel, whereas it was nearly quantitatively recovered in the unbound fraction (92 +/- 4%) after addition of alpha 2-Antiplasmin to the mixture. Scatchard analysis of the binding of STAR to plasmin-Sepharose yielded a dissociation constant of 55 nM, whereas no specific binding of STAR to plasmin-alpha 2-Antiplasmin-Sepharose could be demonstrated. These findings indicate that, both in purified systems and in a human plasma milieu containing a 125I-fibrin-labeled plasma clot, neutralization of the plasmin-STAR complex by alpha 2-Antiplasmin results in dissociation of functionally active STAR from the complex and recycling of STAR to other plasminogen molecules. This dissociation-recycling process may explain the high fibrinolytic potency of STAR in a plasma milieu in the presence of high concentrations of alpha 2-Antiplasmin.
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on the molecular interactions between plasminogen staphylokinase α2 Antiplasmin formula and fibrin
Biochimica et Biophysica Acta, 1992Co-Authors: Roger H Lijnen, Berthe Van Hoef, Osamu Matsuo, Desire CollenAbstract:The molecular interactions between the plasminogen-staphylokinase complex, α 2 -Antiplasmin-formula> and fibrin were studied by measuring the effect of CNBr-digested fibrinogen on the inhibition rate of the plasminogen-staphylokinase complex by α 2 -Antiplasmin-formula>. The second-order rate constant for the inhibition of plasminogen-staphylokinase by α 2 -Antiplasmin-formula> was 2.7±0.3 · 10 6 M −1 s −1 -formula> (mean±S.D.-formula>; n = 7-formula>). Addition of CNBr-digested fibrinogen, but not of fibrinogen, resulted in a concentration-dependent reduction of the apparent inhibition rate constant, with a 50 percent reduction at a concentration of 5 nM CNBr-digested fibrinogen. The second-order rate constant for the inhibition of the low- M r -formula> plasminogen-staphylokinase complex (plasminogen lacking the kringle structures comprising the lysine-binding sites) by α 2 -Antiplasmin-formula> was about 30-fold lower (9.3 ± 0.7 · 10 4 M −1 -formula>, mean±S.D.-formula>; n = 4-formula>) than that of plasminogen-staphylokinase and was not affected by addition of CNBr-digested fibrinogen. Inhibition of the plasminogen-staphylokinase complex by the chloromethylketone d -Val-Phe-Lys-CH 2 Cl is 9-fold less efficient than that of plasmin ( k 2 /K i -formula> of 700 M −1 s −1 versus 6300 M −1 s −1 ). Our results confirm and establish that rapid inhibition of plasminogen-staphylokinase by α 2 -Antiplasmin-formula> requires the availability of the lysine-binding sites in the plasminogen moiety of the complex. Fibrin, but not fibrinogen, reduces the inhibition rate by α 2 -Antiplasmin-formula> by competition for interaction with the lysine-binding site. Protection of the plasminogen-staphylokinase complex bound to fibrin from rapid inhibition by α 2 -Antiplasmin-formula> thus appears to contribute to the fibrin-specificity of clot lysis with staphylokinase in a plasma milieu, by allowing preferential plasminogen activation at the fibrin surface, while the free complex is rapidly inhibited in plasma.
Agata Hanna Bryk - One of the best experts on this subject based on the ideXlab platform.
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plasma fibrin clots of pulmonary embolism patients present increased amounts of factor xiii and alpha2 Antiplasmin at 3 months anticoagulation since the acute phase
Journal of Physiology and Pharmacology, 2020Co-Authors: Michal Zabczyk, Agata Hanna Bryk, Joanna Natorska, Zsuzsa Bagoly, Barbara Barath, Éva Katona, Ferenc Sarkady, Katharina Zettl, J R Wisniewski, Anetta UndasAbstract:Fibrin cross-linking by coagulation factor (F)XIII leads to clot stabilization. Reduced plasma FXIII levels have been reported in acute pulmonary embolism (PE) patients. We investigated the impact of anticoagulant therapy on clot-bound amounts of FXIII and α2-Antiplasmin and their associations with fibrin clot properties in patients with PE. Clots generated from plasma of 18 acute symptomatic patients on admission and after a 3-month treatment with rivaroxaban were assessed off anticoagulation using mass spectrometry. Plasma FXIII and α2-Antiplasmin activity were determined at the 2 time points along with thrombin generation markers, plasma fibrin clot permeability (Ks), and clot lysis time (CLT). Following anticoagulant therapy, clot-bound FXIII increased from 2.97 (interquartile range, 1.98 - 4.08) to 4.66 (3.5 - 6.9) mg/g protein and α2-Antiplasmin from 9.4 (7.2 - 10.6) to 11 (9.5 - 14) mg/g protein (both p < 0.0001). The two parameters showed positive correlation at baseline only (r = 0.63, p = 0.0056). Similarly to clot-bound amounts, plasma FXIII (+25.8%) and α2-Antiplasmin activity (+12%) increased at 3 months. Plasma FXIII activity on admission, but not after 3 months since the index PE, was associated with amounts of clot-bound FXIII (r = 0.35, p = 0.043) and α2-Antiplasmin (r = 0.47, p = 0.048). At baseline, clot-bound FXIII correlated with plasma F1+2 prothrombin fragments levels (r = 0.51, p = 0.03), while clot-bound α2-Antiplasmin correlated with CLT (r = 0.43, p = 0.036). At 3 months associations of clot-bound FXIII and α2-Antiplasmin were abolished. This study assessed for the first time changes in the fibrin clot composition following acute PE, suggesting an increase of clot-bound and plasma FXIII and α2-Antiplasmin levels after 3 months.
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Interaction of glycated and acetylated human α2-Antiplasmin with fibrin clots.
Blood coagulation & fibrinolysis : an international journal in haemostasis and thrombosis, 2020Co-Authors: Agata Hanna Bryk, Dorota Satala, Joanna Natorska, Maria Rąpała-kozik, Anetta UndasAbstract:: In type 2 diabetes mellitus (T2DM), increased α2-Antiplasmin incorporation in fibrin and impaired fibrinolysis have been reported. Acetylsalicylic acid (ASA), used in cardiovascular prevention, modulates fibrinolysis and exerts weaker therapeutic effect in this disease. We investigated how glycation and acetylation of α2-Antiplasmin affects its interaction with fibrin. Using surface plasmon resonance, we analyzed fibrin binding by α2-Antiplasmin incubated with no β-D-glucose or ASA (control); incubated with β-D-glucose (5, 10, 50 mmol/l); (3) incubated with 1.6 mmol/l acetylsalicylic acid (ASA) and (4) incubated with 1.6 mmol/l ASA and 50 mmol/l β-D-glucose. Incubation with glucose decreased affinity of α2-Antiplasmin for fibrin compared with control α2-Antiplasmin in a glucose concentration-depending manner. α2-Antiplasmin incubation with ASA did not affect its affinity to fibrin. α2-Antiplasmin incubation with ASA and glucose resulted in 4.2-fold increased affinity to fibrin compared with α2-Antiplasmin incubated with 50 mmol/l glucose (P
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Identification of glycated and acetylated lysine residues in human α2-Antiplasmin.
Biochemical and biophysical research communications, 2019Co-Authors: Agata Hanna Bryk, Dominik Cysewski, Michal Dadlez, Anetta UndasAbstract:Abstract Background The post-translational protein modification via lysine residues can significantly alter its function. α2-Antiplasmin, a key inhibitor of fibrinolysis, contains 19 lysine residues. Aim We sought to identify sites of glycation and acetylation in human α2-Antiplasmin and test whether the competition might occur on the lysine residues of α2-Antiplasmin. Methods We analyzed human α2-Antiplasmin (1) untreated; (2) incubated with increasing concentrations of β- d -glucose (0, 5, 10, 50 mM); (3) incubated with 1.6 mM acetylsalicylic acid (ASA) and (4) incubated with 1.6 mM ASA and 50 mM β- d -glucose, using the ultraperformance liquid chromatography system coupled to mass spectrometer. Results Eleven glycation sites and 10 acetylation sites were found in α2-Antiplasmin. Incubation with β- d -glucose was associated with glycation of 4 (K-418, K-427, K-434, K-441) out of 6 lysine residues, known to be important for mediating the interaction with plasmin. Glycation and acetylation overlapped at 9 sites in samples incubated with β- d -glucose or ASA. Incubation with concomitant ASA and β- d -glucose was associated with the decreased acetylation at all sites overlapping with glycation sites. At K-182 and K-448, decreased acetylation was associated with increased glycation when compared with α2-Antiplasmin incubated with 50 mM β- d -glucose alone. Although K-24 located in the proximity of the α2-Antiplasmin cleavage site, was found to be only acetylated, incubation with ASA and 50 mM β- d -glucose was associated the absence of acetylation at that site. Conclusion Human α2-Antiplasmin is glycated and acetylated at several sites, with the possible competition between acetylation and glycation at K-182 and K-448. Our finding suggests possibly relevant alterations to α2-Antiplasmin function at high glycemia and during aspirin use.
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Sex-specific alteration to α2-Antiplasmin incorporation in patients with type 2 diabetes.
Thrombosis research, 2019Co-Authors: Agata Hanna Bryk, Jakub Siudut, Elżbieta Broniatowska, Zsuzsa Bagoly, Barbara Barath, Éva Katona, Anetta UndasAbstract:Abstract Background Type 2 diabetes mellitus (T2DM) is associated with hypofibrinolysis and increased factor XIII-mediated α2-Antiplasmin incorporation into the fibrin clot. It is unclear whether there are sex-related differences in α2-Antiplasmin incorporation in relation to impaired clot lysis in T2DM. Aim We investigated α2-Antiplasmin incorporation into fibrin clots as a determinant of clot lysability in patients of both sexes with T2DM. Methods In a group of 113 T2DM patients, 54 (47.8%) of which were women, we investigated α2-Antiplasmin incorporation using an in-house sandwich enzyme-linked immunoassay and plasma clot lysis by turbidimetry, along with fibrinogen and thrombin generation using calibrated automated thrombogram and factor XIII activity. Results Female patients had 15.2% greater α2-Antiplasmin incorporation into the fibrin clot (p = 0.008) and slightly higher plasma α2-Antiplasmin concentration (p = 0.005) along with 8.4% longer time to 50% lysis (Lys50MA, p = 0.012) compared with men. Female patients had enhanced thrombin generation represented by shorter lag phase (p = 0.042), shorter time to peak (p = 0.033), and higher endogenous thrombin potential (p = 0.003) compared with men, while factor XIII activity was comparable between sexes (p = 0.085). On multivariate regression, patient sex and glycated hemoglobin (HbA1c) level were the predictors of α2-Antiplasmin incorporation in the entire patient group, while α2-Antiplasmin incorporation was associated with Lys50MA, as were fibrinogen, male sex and body-mass index. Conclusions This study suggests that a more compromised fibrinolysis in diabetic women when compared with men could be in part mediated by increased α2-Antiplasmin incorporation into the fibrin.