scholarly journals A bleeding disorder due to deficiency of alpha 2-antiplasmin

Blood ◽  
1982 ◽  
Vol 59 (6) ◽  
pp. 1246-1251 ◽  
Author(s):  
LA Miles ◽  
EF Plow ◽  
KJ Donnelly ◽  
C Hougie ◽  
JH Griffin
Keyword(s):  
System Analysis ◽  
Bleeding Disorder ◽  
Fibrinolytic System ◽  
In Vitro Assays ◽  
Clot Lysis ◽  
Plasma Clot ◽  
Normal Limits ◽  
Normal Plasma ◽  
Bleeding Episodes

Abstract A deficiency of alpha 2-antiplasmin has been identified in a female patient with severe and frequent bleeding episodes. Routine coagulation and platelet assays of the patient's plasma were within normal limits. However, abnormally rapid whole blood or dilute plasma clot lysis times and an abnormal FXIII test in which clots were lysed in the presence of urea or saline suggested an abnormal fibrinolytic system. Analysis of alpha 2-antiplasmin levels by radioimmunoassay revealed less than 1.0 microgram/ml alpha 2-antiplasmin. Functional assays indicated an alpha 2-antiplasmin level less than or equal to 10% of normal. Addition of purified alpha 2-antiplasmin to the patient's plasma restored its ability to inhibit plasmin in in vitro assays, and mixtures of patient plasma with normal plasma did not interfere with the antiplasmin activity of the normal plasma. Whereas normal platelets contain 68 ng alpha 2-antiplasmin/10(9) platelets, platelets from the patient contained 30% of the normal level of antigen. Analysis of alpha 2- antiplasmin functional and antigenic levels in the plasma of both parents and four siblings of the propositus provided evidence consistent with an autosomal mechanism of inheritance of alpha 2- antiplasmin deficiency. One sibling appeared to be homozygous and three siblings and the parents were heterozygous for the deficiency. Two heterozygotes had positive bleeding histories. The association of a bleeding disorder with a deficiency of alpha 2-antiplasmin emphasizes that lack of regulation of the fibrinolytic system can result in a hemostatic dysfunction.

scholarly journals A bleeding disorder due to deficiency of alpha 2-antiplasmin

Blood ◽  
1982 ◽  
Vol 59 (6) ◽  
pp. 1246-1251 ◽  
Author(s):  
LA Miles ◽  
EF Plow ◽  
KJ Donnelly ◽  
C Hougie ◽  
JH Griffin
Keyword(s):  
System Analysis ◽  
Bleeding Disorder ◽  
Fibrinolytic System ◽  
In Vitro Assays ◽  
Clot Lysis ◽  
Plasma Clot ◽  
Normal Limits ◽  
Normal Plasma ◽  
Bleeding Episodes

A deficiency of alpha 2-antiplasmin has been identified in a female patient with severe and frequent bleeding episodes. Routine coagulation and platelet assays of the patient's plasma were within normal limits. However, abnormally rapid whole blood or dilute plasma clot lysis times and an abnormal FXIII test in which clots were lysed in the presence of urea or saline suggested an abnormal fibrinolytic system. Analysis of alpha 2-antiplasmin levels by radioimmunoassay revealed less than 1.0 microgram/ml alpha 2-antiplasmin. Functional assays indicated an alpha 2-antiplasmin level less than or equal to 10% of normal. Addition of purified alpha 2-antiplasmin to the patient's plasma restored its ability to inhibit plasmin in in vitro assays, and mixtures of patient plasma with normal plasma did not interfere with the antiplasmin activity of the normal plasma. Whereas normal platelets contain 68 ng alpha 2-antiplasmin/10(9) platelets, platelets from the patient contained 30% of the normal level of antigen. Analysis of alpha 2- antiplasmin functional and antigenic levels in the plasma of both parents and four siblings of the propositus provided evidence consistent with an autosomal mechanism of inheritance of alpha 2- antiplasmin deficiency. One sibling appeared to be homozygous and three siblings and the parents were heterozygous for the deficiency. Two heterozygotes had positive bleeding histories. The association of a bleeding disorder with a deficiency of alpha 2-antiplasmin emphasizes that lack of regulation of the fibrinolytic system can result in a hemostatic dysfunction.

Biological and Thrombolytic Properties of Proenzyme and Active Forms of Human Urokinase – I. Fibrinolytic and Fibrinogenolytic Properties in Human Plasma In Vitro of Urokinases Obtained from Human Urine or by Recombinant DNA Technology

1984 ◽  
Vol 52 (01) ◽  
pp. 019-023 ◽  
Author(s):  
C Zamarron ◽  
H R Lijnen ◽  
B Van Hoef ◽  
D Collen
Keyword(s):  
Human Plasma ◽  
Recombinant Dna ◽  
Fibrinolytic System ◽  
Tissue Type ◽  
Clot Lysis ◽  
Recombinant Dna Technology ◽  
Plasma Clot ◽  
Type Plasminogen Activator ◽  
Intermediate Response

SummaryThe fibrinolytic and fibrinogenolytic properties of recombinant pro-urokinase (Rec-pro-UK) and recombinant urokinase (Rec- UK) obtained by expression of the human urokinase cDNA in E. coli, were compared with those of natural urokinase (Nat-UK) of urinary origin and of tissue-type plasminogen activator (t-PA) in a system, composed of a radioactive human plasma clot immersed in citrated human plasma.The specific fibrinolytic effects of Nat-UK, Rec-pro-UK and Rec-UK were very similar, causing significant clot lysis at concentrations of 100 IU/ml plasma or more. t-PA caused equivalent degrees of clot lysis at 10-fold lower concentrations.Activation of the fibrinolytic system in the plasma (fib- rinogenolysis), was not observed with t-PA in concentrations which induced complete clot lysis within 5 hr (20-30 IU/ml plasma). With Nat-UK and Rec-UK, all concentrations which caused significant clot lysis (100-200 IU/ml plasma) also caused extensive activation of the plasma fibrinolytic system. With Rec- pro-UK an intermediate response was obtained. The highest amounts required for complete clot lysis in 5 hr (200 IU/ml plasma) also caused significant fibrinogenolysis. At intermediate concentrations (50-100 IU/ml), however, significant clot lysis (40-80%) was observed without systemic fibrinolytic activation.

Absence of Synergism Between Tissue-Type Plasminogen Activator (t-PA), Single-Chain UrokinaseType Plasminogen Activator (scu-PA) and Urokinase on Clot Lysis in a Plasma Milieu In Vitro

1986 ◽  
Vol 56 (01) ◽  
pp. 035-039 ◽  
Author(s):  
D Collen ◽  
F De Cock ◽  
E Demarsin ◽  
H R Lijnen ◽  
D C Stump
Keyword(s):  
Human Plasma ◽  
Plasminogen Activator ◽  
Dose Response ◽  
Fibrinolytic System ◽  
Tissue Type ◽  
Clot Lysis ◽  
Single Chain ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator

SummaryA potential synergic effect of tissue-type plasminogen activator (t-PA), single-chain urokinase-type plasminogen activator (scuPA) or urokinase on clot lysis was investigated in a whole human plasma system in vitro. The system consisted of a human plasma clot labeled with 125I-fibrinogen, immersed in titrated whole human plasma, to which the thrombolytic agents were added. Clot lysis was quantitated by measurement of released 125I, and activation of the fibrinolytic system in the surrounding plasma by measurements of fibrinogen and α2-antiplasmin.t-PA, scu-PA and urokinase induced a dose-dependent and time-dependent clot lysis; 50 percent lysis after 2 h was obtained with 5 nM t-PA, 20 nM scu-PA and 12 nM urokinase. At these concentrations no significant activation of the fibrinolytic system in the plasma was observed with t-PA and scu-PA, whereas urokinase caused significant α2-antiplasmin consumption and concomitant fibrinogen degradation. The shape of the dose-response curves was different; t-PA and urokinase showed a log linear dose-response whereas that of scu-PA was sigmoidal.

Thrombin activatable fibrinolysis inhibitor (TAFI) affects fibrinolysis in a plasminogen activator concentration-dependent manner

2004 ◽  
Vol 91 (03) ◽  
pp. 473-479 ◽  
Author(s):  
Ana Guimarães ◽  
Dingeman Rijken
Keyword(s):  
Plasminogen Activator ◽  
In Vitro Study ◽  
Inhibitory Effect ◽  
Retardation Factor ◽  
Clot Lysis ◽  
Dependent Manner ◽  
Plasma Clot ◽  
Concentration Dependent Manner ◽  
Plasmin Inhibitor

SummaryTAFIa was shown to attenuate fibrinolysis. In our in vitro study, we investigated how the inhibitory effect of TAFIa depended on the type and concentration of the plasminogen activator (PA). We measured PA-mediated lysis times of plasma clots under conditions of maximal TAFI activation by thrombin-thrombomodulin in the absence and presence of potato carboxypeptidase inhibitor. Seven different PAs were compared comprising both tPA-related (tPA, TNK-tPA, DSPA), bacterial PA-related (staphylokinase and APSAC) and urokinase-related (tcu-PA and k2tu-PA) PAs. The lysis times and the retardation factor were plotted against the PA concentration. The retardation factor plots were bell-shaped. At low PA concentrations, the retardation factor was low, probably due to the limited stability of TAFIa. At intermediate PA concentrations the retardation factor was maximal (3-6 depending on the PA), with TNK-tPA, APSAC and DSPA exhibiting the strongest effect. At high PA concentrations, the retardation factor was again low, possibly due to inactivation of TAFIa by plasmin or to a complete conversion of glu-plasminogen into lys-plasminogen. Using individual plasmas with a reduced plasmin inhibitor activity (plasmin inhibitor Enschede) the bell-shaped curve of the retardation factor shifted towards lower tPA and DSPA concentrations, but the height did not decrease. In conclusion, TAFIa delays the lysis of plasma clots mediated by all the plasminogen activators tested. This delay is dependent on the type and concentration of the plasminogen activator, but not on the fibrin specificity of the plasminogen activator. Furthermore, plasmin inhibitor does not play a significant role in the inhibition of plasma clot lysis by TAFI.

A Novel Congenital Bleeding Disorder Related to High Plasma Heparin-Like Anticoagulant Activity.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4074-4074
Author(s):  
Zhaoyue Wang ◽  
Haiyen Yang ◽  
Xia Bai ◽  
Wei Zhang ◽  
Changgeng Ruan
Keyword(s):  
Ex Vivo ◽  
Anticoagulant Activity ◽  
Coagulation Factors ◽  
High Plasma ◽  
Bleeding Disorder ◽  
Normal Plasma ◽  
Normal Ranges ◽  
Coagulation Tests ◽  
Plasma Heparin

Abstract Heparin or heparin-like compounds present in human plasma in minute amounts. It has been reported that a very few patients with such diseases as plasma cell neoplasms, acute monoblastic leukemia and acquired immune deficincy syndrome have an increased plasma heparin-like anticoagulant activity. Recently, we found a 10-year-old girl who was physically and developmentally normal, but had recurrent episodes of prolonged bleeding and hematoma starting in her early childhood, which could be stopped by transfusion of fresh frozen plasma or prothrombin complex concentrate. The coagulation tests of her plasma were regularly repeated since she was 2 years old, and always revealed a markedly prolonged APTT (61.8–104 seconds, normal 28–40 seconds) and TT (36–50.1 seconds, normal 14–21 seconds), and a slightly prolonged PT (15.9–25 seconds, normal 11–14.5 seconds). Fibrinogen, prothrombin and other coagulation factors as well as anticoagulant and fibrinolytic systems were all normal. The results of immunologic measurements were either negative or within normal ranges. Treatment of the patient’s plasma in vitro with either protamine or heparinase could completely normalize the coagulation abnormalities, but not with normal plasma. The anticoagulant activity of her plasma corresponded to 0.2 heparin U/mL when measured by a TT assay using normal plasma as substrate and standardized with porcine heparin. Her plasma heparin concentration was 0.22 heparin U/mL when measured using a colometric assay. In ex vivo study, the abnormal coagulation tests could effectively be corrected when the patient was intravenously administed with protamine. Considering these characteristic laboratory features of the patient, we suppose it would probably represent a novel congenital bleeding disorder related to high plasma heparin-like anticoagulant activity which, to our knowledge, had not been described before.

Molecular Conversions of Recombinant Staphylokinase During Plasminogen Activation in Purified Systems and in Human Plasma

1993 ◽  
Vol 70 (03) ◽  
pp. 495-499 ◽  
Author(s):  
S Ueshima ◽  
K Silence ◽  
D Collen ◽  
H R Lijnen
Keyword(s):  
Amino Acid ◽  
Human Plasma ◽  
Catalytic Amount ◽  
Clot Lysis ◽  
Lag Phase ◽  
Plasminogen Activation ◽  
Plasma Clot ◽  
Acid Protein ◽  
Human Plasminogen

SummaryRecombinant staphylokinase (STAR) is produced as a 136 amino acid protein with NH2-terminal sequence Ser-Ser-Ser (mature STAR, HMW-STAR), which may be converted to lower molecular weight forms (LMW-STAR) by removal of the first six residues (yielding STAR-Δ6 with NH2-terminal Gly-Lys-Tyr-) or the first ten residues (yielding STAR-Δ10 with NH2-terminal Lys-Gly-Asp-). In the present study the occurrence and effects of these conversions during plasminogen activation by HMW-STAR were studied in purified systems and in human plasma.In stoichiometric mixtures of HMW-STAR and native human plasminogen (Glu-plasminogen), rapid and quantitative conversion of HMW-STAR to LMW-STAR occurred, concomitant with exposure of the active site in the plasmin-STAR complex. NH2-terminal amino acid sequence analysis revealed the sequence Lys-Gly-Asp- in addition to the known sequences of the Lys-plasmin chains, identifying STAR-Δ10 as the derivative generated from HMW-STAR. In mixtures of catalytic amount of HMW-STAR and human plasminogen, plasmin generation occurred progressively, following an initial lag phase, during which HMW-STAR was converted to LMW-STAR. Plasmin-mediated conversion of HMW-STAR to LMW-STAR obeyed Michaelis-Menten kinetics with K m = 3.6 μM and k 2 = 0.38 s−1. The specific clot lysis activities of HMW-STAR (122,000 ± 8,000 units/mg) and LMW-STAR (129,000 ± 8,000 units/mg) were indistinguishable.In an in vitro system consisting of a 60 μl plasma clot submerged in 250 μl plasma, 80% clot lysis within 1 h was obtained with 70 nM HMW-STAR. This was associated with fibrinogen depletion and conversion of 20% of the HMW-STAR to LMW-STAR. Addition of 100 nM HMW-STAR to human plasma in the absence of a clot did not induce significant fibrinogen breakdown (≥ 90% residual fibrinogen after 2 h), and was not associated with significant coversion to LMW-STAR (≤10% within 2 h). With 400 nM HMW-STAR, fibrinogen depletion in plasma occurred within 1 h, and 80% conversion to LMW-STAR was observed. Thus, at fibrinolytically active concentrations which do not cause fibrinogen breakdown, no significant conversion of HMW-STAR to LMW-STAR occurs in human plasma in the absence of a clot.These findings indicate that the conversion of HMW-STAR to LMW-STAR may occur in association with clot lysis, but is not required to induce it.

Role of α2-Antiplasmin in Fibrin-Specific Clot Lysis with Single-Chain Urokinase-Type Plasminogen Activator in Human Plasma

1991 ◽  
Vol 65 (04) ◽  
pp. 394-398 ◽  
Author(s):  
P J Declerck ◽  
H R Lijnen ◽  
M Verstreken ◽  
D Collen
Keyword(s):  
Human Plasma ◽  
Plasminogen Activator ◽  
Clot Lysis ◽  
Minor Role ◽  
Single Chain ◽  
Plasma Clot ◽  
Normal Plasma ◽  
Type Plasminogen Activator ◽  
Urokinase Type Plasminogen Activator

SummaryThe role of plasma α2-antiplasmin (α2-AP) in the fibrinspecificity of clot lysis by recombinant single-chain urokinase-type plasminogen activator (rscu-PA) and in the conversion of rscu-PA to its two-chain derivative (rtcu-PA, urokinase) was investigated in an in vitro human plasma clot lysis system. Fifty % lysis in 2 h of a 0.1 ml 125l-fibrin labeled human plasma clot immersed in 0.5 ml normal human plasma was obtained with 1.4 ± 0.15 µg/ml rscu-PA (mean ± SD, n = 8). This was associated with degradation of 23 ± 7% of fibrinogen and generation of 0.20 ± 0.09 µg/ml rtcu-PA. In α2-AP-depleted plasrna 50% clot lysis in 2 h required 2-fold less rscu-PA which was associated with 3-fold more extensive fibrinogen degradation and 2-fold more rtcu-PA generation. Fifty % lysis in? h, of a 0.1 ml α2-AP-depleted plasma clot, subriersed in 0.5 ml normal plasma, was obtained with 0.80 ± 0.05 µg/ml rscu-PA (n = 3, p <0.001 vs normal clot) and was associated with 17 ± 6% fibrinogen breakdown (p : 0.22 vs normal clot) and 0.08 ± 0.02 µg/ml rtcu-PA generation (p < 0.05 vs normal clot). In α2-AP-depleted plasma the equipotent rscu-PA concentration was 4-fold lower than in normal plasma and was associated with 3-fold more fibrinogen degradation and a similar extent of rtcu-PA generationThus, α2-AP in plasma contributes significantly to the fibrinspecificity of rscu-PA, primarily via prevention of conversion in plasma of rscu-PA to rtcu-PA. Clot associated α2-AP increases the resistance of the clot to lysis with rscu-PA, but plays an only minor role in the fibrin-specificity of clot lysis in normal plasma.

Effect of Standard Heparin and a Low Molecular Weight Heparin on Thrombolytic and Fibrinolytic Activity of Single-Chain Urokinase Plasminogen Activator In Vitro

1989 ◽  
Vol 62 (03) ◽  
pp. 923-926 ◽  
Author(s):  
D M Lourenço ◽  
A M Dosne ◽  
A Kher ◽  
M Samama
Keyword(s):  
Molecular Weight ◽  
Low Molecular Weight Heparin ◽  
Clot Lysis ◽  
Low Molecular Weight ◽  
Single Chain ◽  
Plasma Clot ◽  
Thrombolytic Activity ◽  
Significant Difference ◽  
Plasma Factor

SummaryThe effect of unfractioned heparin (UH) and low molecular weight heparin (LMWH) (Kabi 2165 - Fragmin®) on in vitro scu-PA thrombolytic and fibrinogenolytic activity was investigated. Thrombolytic activity was evaluated by following lysis of radiolabeled plasma clot immersed in plasma in presence of scu-PA alone or with either form of heparin. A 200 IU/ml scu-PA concentration produced clot lysis within 7 hr. UH or LMWH led to a slightly faster clot lysis which was statistically significant only at the 2nd and 3rd hour. No significant difference could be evidenced between UH and LMWH effect. During clot lysis, plasmin, generated within the clot led to a gradual transformation of scu-PA to tcu-PA, specially after a 4-hr incubation. Appearance of tcu-PA activity in the plasma surrounding the clot was significantly inhibited by either form of heparin. This finding contrasts with results observed in purified systems and suggests the presence of heparin-dependent plasma factor(s) inhibiting tcu-PA formation or its activity. Possible candidates might be anti-thrombin III and PAI-3.No fibrinogen breakdown was observed when plasma was incubated for 7 hr at 37° C in presence of scu-PA alone (200 IU/ ml) or with either form of heparin. However, in presence of a plasma clot, an important fibrinogen breakdown was observed during clot lysis reflecting the action of plasmin and/or tcu-PA generated within the clot, in the surrounding plasma. Fibrinogenolysis was less pronounced in the presence of both heparin preparations possibly as a consequence of the reduction in the tcu-PA level. These results underline the importance of plasma factors in the interaction of heparin with plasminogen activators such as scu-PA.

EVIDENCE FOR SYNERGISM BETWEEN TISSUE-TYPE PLASMINOGEN ACTIVATOR AND SINGLE CHAIN UROKINASE ON IN VITRO PLASMA CLOT LYSIS

1987 ◽  
Author(s):  
R S Rappaport ◽  
M R Blume ◽  
R L Vogel ◽  
M H Levner ◽  
P P Hung
Keyword(s):  
Plasminogen Activator ◽  
Tissue Type ◽  
Clot Lysis ◽  
Single Chain ◽  
Plasma Clot ◽  
Tissue Type Plasminogen Activator ◽  
Molar Ratios ◽  
Type Plasminogen Activator

There is mounting evidence from animal models and the clinic that combination thrombolytic therapy with tissue-type plasminogen activator (tPA) and single chain urokinase (scuPA) is synergistic. Yet, efforts to demonstrate synergism between these two plasminogen activators in vitro have met with discordant results. Collen et al (Thromb. Haemostasis, 56:35, 1986) reported an absence of synergism between these two agents on clot lysis in an in vitro plasma milieu when they were evaluated at molar ratios of 1:4 (tPA:scuPA and vice versa). Gurewich and Pannell (Thromb. Res., 44:217, 1986), however, reported a synergistic effect on fibrin-specific clot lysis in vitro when the agents were combined in concentrations exceeding molar ratios of 1:4 (tPA:scuPA). Here, we present evidence that synergism between tPA and scuPA may be demonstrated in vitro provided that the molar ratio of tPA to scuPA exceeds 1:4 and that the concentration of clot bound or unbound tPA is minimized. In order to achieve this experimental condition, the standard in vitro plasma clot lysis assay was modified. Human plasma clots were incubated first for a short time in plasma containing varying amounts of tPA. After incubation, the clots were washed thoroughly and reimmersed in plasma alone or in plasma containing varying amounts of scuPA or tPA. Under these conditions, lysis proceeded at a greater rate and to a greater extent when tPA clots were immersed in plasma containing an appropriate amount of scuPA than when they were immersed in plasma alone or in plasma containing appropriate amounts of tPA. Lysis of untreated clots or clots exposed first to scuPA and then to plasma containing varying amounts of scuPA proceeded far less efficiently with a characteristic lag. The enhanced lysis produced by tPA and scuPA obeyed the classical definition of synergy: the same biological effect can be obtained with two drugs together at algebraic fractional combinations of less than 1 (Berenbaum, M.C., Clin. Exp. Immunol., 28:1-18, 1977). Thus, conditions that more closely mimic the in vivo situation resulting from a bolus injection of tPA followed by infusion with scuPA, may provide a system for duplication of in vivo synergism in. vi tro and investigation of the mechanism thereof.

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