Genomische Diagnostik thrombophiler Gerinnungsstörungen bei Frauen

2007 ◽  
Vol 27 (01) ◽  
pp. 22-31 ◽  
Author(s):  
E. Lindhoff-Last ◽  
B. Luxembourg
Keyword(s):  
Protein C ◽  
Protein S ◽  
Genetische Diagnostik ◽  
Klinische Relevanz ◽  
Mthfr Mutation ◽  
Pai 1

ZusammenfassungDie Aufklärung der DNA-Sequenzen sowohl der Gerinnungsfaktoren als auch der Gerinnungsinhibitoren hat die Erforschung genetischer Ursachen einer venösen Thromboseneigung ermöglicht. Da die Entstehung venöser Thrombosen ein multifaktorielles Geschehen ist, weisen Frauen auf Grund frauenspezifischer Risikosituationen (z. B. hormonale Kontrazeption, Schwangerschaft, Wochenbett) in bestimmten Lebensphasen ein zusätzliches expositionelles Thromboserisiko auf. Von wesentlicher Bedeutung ist es, die thrombophilen Neigungen zu definieren, bei denen eine genetische Diagnostik von besonderer klinischer Relevanz ist, zumal auch eine habituelle Abortneigung, schwangerschaftsinduzierte hypertensive Erkrankungen und frustrane In-vitro-Fertilisationen in Zusammenhang mit einer Thrombophilie gebracht werden. Trotz der vielfältigen Möglichkeiten einer genomischen Diagnostik ist eine DNA-Analyse nur in wenigen Fällen klinisch notwendig: Sinnvoll ist eine Mutationsanalytik bezüglich einer Faktor- V-Leiden-Mutation, da die Risiken zwischen homo- und heterozygoter Ausprägung differieren und von klinischer Relevanz in der Schwangerschaft und bei Schwangerschaftskomplikationen sind. Ebenso bewiesen ist dies für die Prothrombinmutation G20210A, wobei die Datenlage für deren homozygote im Vergleich zur heterozygoten Form auf Grund der Seltenheit unzureichend ist.Die Analytik der homozygoten Form der MTHFR-Mutation C677T ist nicht zu empfehlen, da die klinische Relevanz für die meisten Indikationen nicht ausreichend belegt ist. Für angeborene Mangelzustände von Antithrombin, Protein C und Protein S ist die Datenlage auf Grund ihrer geringen Inzidenzen selbst in venösen Thrombosekollektiven sehr gering. Die Mutationsanalytik kommt hier nur in Frage, wenn die Aktivitätsuntersuchungen der Proteine trotz wiederholter Messungen keine eindeutige Beurteilung zulassen. Alle übrigen Mutationsanalysen (z. B. PAI-1-Polymorphismus, Faktor-XIII-Val34Leu-Polymorphismus) sind bei Frauen zurzeit ohne klinische Relevanz.

Activated Protein C Increases Fibrin Clot Lysis by Neutralization of Plasminogen Activator Inhibitor No Evidence for a Cofactor Role of Protein S

1988 ◽  
Vol 60 (02) ◽  
pp. 328-333 ◽  
Author(s):  
N J de Fouw ◽  
Y F de Jong ◽  
F Haverkate ◽  
R M Bertina
Keyword(s):  
Plasminogen Activator ◽  
Protein C ◽  
Blood Platelets ◽  
Plasminogen Activator Inhibitor ◽  
Protein S ◽  
Tissue Type ◽  
Clot Lysis ◽  
Activator Inhibitor ◽  
Pai 1

summaryThe effect of purified human activated protein G (APC) on fibrinolysis was studied using a clot iysis system consisting of purified glu-plasminogen, tissue-type plasminogen activator, plasminogen activator inhibitor (released from endothelial cells or blood platelets), fibrinogen, 125T-fibrinogen and thrombin. All proteins were of human origin.In this system APC could increase fibrinolysis in a dose dependent way, without affecting fibrin formation or fibrin crosslinking. However, this profibrinolytic effect of APC could only be observed when plasminogen activator inhibitor (PAI-l) was present. The effect of APC was completely quenched by pretreatment of APC with anti-protein C IgG or di-isopropylfluorophosphate. Addition of the cofactors of APC:protein S, Ca2+-ions and phospholipid-alone or in combination did not enhance the profibrinolytic effect of APC. These observations indicate that human APC can accelerate in vitro clot lysis by the inactivation of PAI-1 activity. However, the neutralization of PAI-1 by APC is independent of the presence or absence of protein S, phospholipid and Ca2+-ions.

Molekularbiologische Grundlagen und Diagnostik der hereditären Defekte von Antithrombin III, Protein C und Protein S

2002 ◽  
Vol 22 (02) ◽  
pp. 57-66
Author(s):  
I. Witt
Keyword(s):  
Protein C ◽  
Antithrombin Iii ◽  
Protein S ◽  
Klinische Relevanz ◽  
At Iii

ZusammenfassungDie enormen Fortschritte in der Molekularbiologie in den letzten Jahren ermöglichten sowohl die Aufklärung der Nukleotidsequenzen der Gene für Antithrombin III (AT III), Protein C (PROC) und Protein S (PROS) als auch die Identifizierung zahlreicher Mutationen bei hereditären Defekten dieser wichtigen Inhibitoren des plasmatischen Gerinnungssystems. Da die Gene für AT III (13,8 kb) und PROC (11,2 kb) nicht groß und relativ leicht zu analysieren sind, gibt es bereits umfangreiche »databases« der Mutationen (50, 73). Für AT III sind 79 und für PROC 160 unterschiedliche Mutationen beschrieben.Sowohl beim AT-III-Mangel als auch beim Protein-C-Mangel hat die Mutationsaufklärung neue Erkenntnisse über die Struktur-Funktions-Beziehung der Proteine gebracht. Beim Protein-C-Mangel steht die klinische Relevanz der DNA-Analyse im Vordergrund, da die Diagnostik des Protein-C-Mangels auf der Proteinebene nicht immer zuverlässig möglich ist.Das Protein-S-Gen ist für die Analytik schwer zugänglich, da es groß ist (80 kb) und außerdem ein Pseudogen existiert. Es sind schon zahlreiche Mutationen bei Patienten mit Protein-S-Mangel identifiziert worden. Eine Database ist bisher nicht publiziert. Die klinische Notwendigkeit zur Mutationsaufklärung besteht ebenso wie beim Protein-C-Mangel. Es ist zu erwarten, dass zukünftig die Identifizierung von Mutationen auch beim Protein-S-Mangel beschleunigt vorangeht.

Platelet-mediated proteolytic down regulation of the anticoagulant activity of protein S in individuals with haematological malignancies

2012 ◽  
Vol 107 (03) ◽  
pp. 468-476 ◽  
Author(s):  
Ilze Dienava-Verdoold ◽  
Marina R. Marchetti ◽  
Liane C. J. te Boome ◽  
Laura Russo ◽  
Anna Falanga ◽  
...  
Keyword(s):  
Protein C ◽  
Normal Values ◽  
Anticoagulant Activity ◽  
Activated Protein C ◽  
Protein S ◽  
Intact Protein ◽  
The Impact ◽  
Independent Protein

SummaryThe natural anticoagulant protein S contains a so-called thrombin-sensitive region (TSR), which is susceptible to proteolytic cleavage. We have previously shown that a platelet-associated protease is able to cleave protein S under physiological plasma conditions in vitro. The aim of the present study was to investigate the relation between platelet-associated protein S cleaving activity and in vivo protein S cleavage, and to evaluate the impact of in vivo protein S cleavage on its anticoagulant activity. Protein S cleavage in healthy subjects and in thrombocytopenic and thrombocythaemic patients was evaluated by immunological techniques. Concentration of cleaved and intact protein S was correlated to levels of activated protein C (APC)-dependent and APC-independent protein S anticoagulant activity. In plasma from healthy volunteers 25% of protein S is cleaved in the TSR. While in plasma there was a clear positive correlation between levels of intact protein S and both APC-dependent and APC-independent protein S anticoagulant activities, these correlations were absent for cleaved protein S. Protein S cleavage was significantly increased in patients with essential thrombocythaemia (ET) and significantly reduced in patients with chemotherapy-induced thrombocytopenia. In ET patients on cytoreductive therapy, both platelet count and protein S cleavage returned to normal values. Accordingly, platelet transfusion restored cleavage of protein S to normal values in patients with chemotherapy-induced thrombocytopenia. In conclusion, proteases from platelets seem to contribute to the presence of cleaved protein S in the circulation and may enhance the coagulation response in vivo by down regulating the anticoagulant activity of protein S.

Enhancement by Activated Protein C of tPA-induced Lysis in a Cell-free System

1987 ◽  
Author(s):  
J C Fredenburgh ◽  
D Collen ◽  
M E Nesheim
Keyword(s):  
Protein C ◽  
Activated Protein C ◽  
Protein S ◽  
Plasminogen Activation ◽  
Free System ◽  
Cell Free System ◽  
Normal Plasma ◽  
Lysis Time ◽  
A Cell

The profibrinolytic activity of human activated protein C (APC) was studied in a cell-free system using human plasma. Normal and Ba+* citrate adsorbed human plasmas were dialyzed against 150mM NaCl, 20mM Hepes, pH 7.4 and diluted to an A280 of 16. Reactions were initiated by the addition of aliquots of plasma to cuvettes containing human melanoma tPA and human thrombin at final concentrations of 1 and 30nM, respectively. The effects of Ca+* and varying concentrations of APC on clotlysis times were examined by monitoring turbidity at 600nM while maintaining the temperature at 37°C. The lysis time, defined as the midpoint of turbidity change, was 128 min for normal plasma containing 10 mM Ca+* and showed progressive and saturable shortening to about 90 min at > 50nM APC. In the absence of Ca+*, lysis time was 55 min for normal plasma and did not shorten in response to APC. With Ba+* citrate adsorbed plasma, the lysis time was 82 min in the presence of 10mM Ca+*, and shortened to 42 min without Ca+*. APC had no effect on lysis time in Ba+* adsorbed plasma either with or without Ca+*. Both bovine and human APC were equally potent. Electrophoresis in DodSO4 and autoradiography of plasma samples containing 125I-labelled plasminogen indicated enhanced rates of plasminogen activation in the presence of APC. These data indicate that APC decreases lysis time in vitro at the level of plasminogen activation. This effect is dependent on Ca+* and may involve additional vitamin K-dependent protein ( s).

Changes in Clotting and Fibrinolytic Parameters Produced by Venous Stasis

1996 ◽  
Vol 2 (1) ◽  
pp. 64-68
Author(s):  
Raul Altman ◽  
Alejandra Scazziota ◽  
Jorge Rouvier
Keyword(s):  
Sodium Citrate ◽  
Vascular Endothelial Cells ◽  
Protein S ◽  
Venous Occlusion ◽  
Venous Stasis ◽  
Clot Lysis ◽  
Before And After ◽  
Fibrinolytic Parameters ◽  
Pai 1

Vascular endothelial cells have thrombosis- prevention properties through the release of some fibri nolytic factors. This capacity of endothelium was studied using the venous occlusion test (VOT) and measuring proteins released during stasis. Blood samples were col lected before and after 10 and 20 min VOT in tubes con taining sodium citrate at pH 7.0 or 4.3. A significant shortening of the euglobulin clot lysis time (ECLT) was obtained 10 and 20 min after VOT in plasma collected in citrate pH 7.0 or 4.3. No statistical difference was noted between results obtained 10 and 20 minutes after VOT. The shortening was related to the increase of tissue plas minogen activator (t-PA) released during VOT. No differ ence was detected in the concentration of tissue plasmin ogen activator inhibitor 1 (PAI-1), but PAI-1 activity mea sured in plasma from blood collected in citrate pH 7.0 decreased progressively from 10.8 ± 9.2 arbitrary units (AU)/ml to 3.76 ± 4.5 AU/ml and 0.8 ± 1.02 AU/ml (p < 0.001) after 10 and 20 min, respectively, of stasis. When the PAI-1 activity was determined in blood collected in citrate pH 4.3 for preventing in vitro complexing of PAI-1 and t-PA, results were different: PAI-1 activity decreased from a basal value of 29.2 ± 15.3 AU/ml to 24.9 ± 12.2 ( p = NS) and 18.7 ± 11.5; p < 0.02) 10 and 20 min after VOT. As result of fibrinolytic activation, other factors were modified: increase of plasminogen activators, de crease of plasminogen, and increase of D-dimer. Fibrin ogen, fibronectin, protein C, protein S, von Willebrand factor, and tissue factor pathway inhibitor concentration remained unchanged after VOT. According to our find ings, VOT can be performed with a stasis of 10 min. Good responders are related to the endothelial capacity for re leasing t-PA and urokinase-type PA (u-PA) and defined as those with an ECLT of <60 min after 10 min VOT when blood was collected in sodium citrate pH 7.0.

Molekularbiologische Grundlagen und Diagnostik der hereditären Defekte von Antithrombin III, Protein C und Protein S

1994 ◽  
Vol 14 (04) ◽  
pp. 199-208
Author(s):  
Irene Witt
Keyword(s):  
Protein C ◽  
Antithrombin Iii ◽  
Protein S ◽  
Klinische Relevanz ◽  
At Iii

ZusammenfassungDie enormen Fortschritte in der Molekularbiologie in den letzten Jahren ermöglichten sowohl die Aufklärung der Nukleotidsequenzen der Gene für Antithrombin III (AT III), Protein C (PROC) und Protein S (PROS) als auch die Identifizierung zahlreicher Mutationen bei hereditären Defekten dieser wichtigen Inhibitoren des plasmatischen Gerinnungssystems. Da die Gene für AT III (13,8 kb) und PROC (11,2 kb) nicht groß und relativ leicht zu analysieren sind, gibt es bereits umfangreiche »databases« der Mutationen (50, 73). Für AT III sind 79 und für RPOC 160 unterschiedliche Mutationen beschrieben.Sowohl beim AT-Ill-Mangel als auch beim Protein-C-Mangel hat die Mutationsaufklärung neue Erkenntnisse über die Struktur-Funktions-Beziehung der Proteine gebracht. Beim Protein-C-Mangel steht die klinische Relevanz der DNA- Analyse im Vordergrund, da die Diagnostik des Protein-C-Mangels auf der Proteinebene nicht immer zuverlässig möglich ist.Das Protein-S-Gen ist für die Analytik schwer zugänglich, da es groß ist (80 kb) und außerdem ein Pseudogen existiert. Es sind schon zahlreiche Mutationen bei Patienten mit Protein-S-Mangel identifiziert worden. Eine Database ist bisher nicht publiziert. Die klinische Notwendigkeit zur Mutationsaufklärung besteht ebenso wie beim Protein-C-Mangel. Es ist zu erwarten, daß zukünftig die Identifizierung von Mutationen auch beim Protein-S-Mangel beschleunigt vorangeht.

Prevention of thrombosis following deep arterial injury in rats by bovine activated protein C requiring co-administration of bovine protein S

2003 ◽  
Vol 90 (08) ◽  
pp. 227-234 ◽  
Author(s):  
Björn Dahlbäck ◽  
Björn Arnljots ◽  
Karl Malm
Keyword(s):  
Protein C ◽  
Anticoagulant Activity ◽  
Activated Protein C ◽  
Protein S ◽  
Arterial Injury ◽  
Control Group ◽  
Clotting Time ◽  
Antithrombotic Effect

SummaryThe antithrombotic effect of bovine activated protein C (bAPC) given with or without bovine protein S (bPS) was investigated in a rat model of deep arterial injury. A segment of the left common carotid artery was isolated between vascular clamps and opened longitudinally. An endarterectomy was performed and the arteriotomy was closed with a running suture, whereafter the vessel was reperfused by removing the clamps. The antithrombotic effect (vascular patency rates 31 minutes after reperfusion) and the arteriotomy bleeding were measured. Ten treatment groups each containing 10 rats and a control group of 20 animals were in a blind random fashion given intravenous bolus injections of increasing doses of activated protein C, with or without co-administration of protein S. The groups received either bAPC alone (0.8, 0.4, 0.2 or 0.1 mg/kg), bAPC (0.8, 0.4, 0.2, 0.1 or 0.05 mg/kg) combined with bPS (0.6 mg/kg), or bPS alone (0.6 mg/kg) whereas the control group received vehicle only. Administered alone, bAPC or bPS had no antithrombotic effect, regardless of dosage. In contrast, all groups that were treated with bAPC in combination with bPS demonstrated a significant antithrombotic effect, as compared to controls. Neither bAPC, bPS, nor the combination of bAPC and bPS increased the arteriotomy bleeding significantly compared to controls. In vitro clotting assays using bAPC or bPS alone yielded only minor prolongation of clotting time, whereas bAPC combined with bPS prolonged the clotting time considerably, demonstrating the dependence on the APC-cofactor activity of bPS for expression of anticoagulant activity by bAPC. In conclusion, our study shows the in vivo significance of protein S as a cofactor to activated protein C, and that potent anti-thrombotic effect can be achieved by low doses of bAPC combined with bPS, without producing hemorrhagic side effects.

Blood coagulation and fibrinolytic factors in 105 patients with hemorrhagic syndrome caused by accidental contact with Lonomia obliqua caterpillar in Santa Catarina, Southern Brazil

2003 ◽  
Vol 89 (02) ◽  
pp. 355-364 ◽  
Author(s):  
Marlene Zannin ◽  
Dayse Lourenço ◽  
Guacyara Motta ◽  
Luis Dalla Costa ◽  
Margaret Grando ◽  
...  
Keyword(s):  
Protein C ◽  
Protein S ◽  
Southern Brazil ◽  
Coagulation Tests ◽  
Group A ◽  
Group B ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Pai 1 ◽  
Lonomia Obliqua

SummaryHaemostatic disorders caused by Lonomia obliqua caterpillars has reached epidemic proportions in southern Brazil. Here we evaluated coagulation and fibrinolysis in 105 patients after accidental contact with Lonomia obliqua caterpillars. Global coagulation tests were prolonged in most cases and patients were divided into 3 groups according to fibrinogen (Fg) level: 0.5 g/l (group A); 0.51–1.5 g/l (group B), >1.5 g/l (group C). There was a significant reduction of factors V, XIII, VIII and prekallikrein in group A, with no change in factors X, II and von Willebrand factor. Thrombin-antithrombin and prothrombin F1+2 were elevated in most patients. Antithrombin and protein S were not changed whereas protein C levels were reduced in group A. Plasminogen and alfa2-antiplasmin levels were significantly reduced in group A and D-Dimer levels were extremely high in all groups, showing that fibrinolysis had been activated, possibly secondary to fibrin production. Levels of t-PA were normal and PAI-1 was mildly elevated in group A. The platelet count remained above 150 × 109 platelets/ml in 97% of cases. In summary, our results suggest that Lonomia obliqua envenoming is characterized by a consumption coagulopathy and secondary fibrinolysis.

4g/5g Prevalence in 223 Patients with Thrombosis and in 162 Healthy Controls, from the Centre Region of Portugal.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4057-4057
Author(s):  
Rosa Maia ◽  
Emilia Cortesao ◽  
Catarina Geraldes ◽  
Luis Simoes ◽  
Carla Simoes ◽  
...  
Keyword(s):  
Protein C ◽  
Antithrombin Iii ◽  
Factor V Leiden ◽  
Protein S ◽  
Mthfr C677t ◽  
Insertion Polymorphism ◽  
Factor V ◽  
Centre Region ◽  
History Of ◽  
Pai 1

Abstract A deletion/insertion polymorphism (4G or 5G) in the promoter of the PAI-1 gene has been suggested to be involved in regulation of the synthesis of the inhibitor, the 4G allele being associated with enhanced gene expression, and therefore related with thrombosis. In the present work we studied the prevalence of 4G/5G polymorphism in 223 unrelated patients with history of objectively confirmed thromboembolism, and in 162 healthy unrelated controls, both groups natural from all centre regions of Portugal. In this normal cohort, the prevalence of 4G/4G is 23%, 4G/5G is 38% and 5G/5G is 39%; in the affected population is, respectively, 47%, 21.5% and 30%, which means that 4G/4G is twice more frequent in the patients with thrombosis. When we relate the age of the first thrombosis episodes in the three groups, we find no significative difference, as the respective media is 36.8; 38.6 and 35.5 years in the 4G/4G, 4G/5G and 5G/5G group, respectively. This data suggest that this polymorphism by itself, even in homozygosity, is not associated with earlier thrombosis. In our patients, we studied the presence of Lupus Anticoagulant, Factor V Leiden, Factor IIG20210A, MTHFR C677T, and also Antithrombin III, Protein S and Protein C levels. We analyse the prevalence of the three mutations in patients with DVP, PTE, ischemic and venous CVA and we only find a significative difference in the 4G/4G group: 46.2% patients with DVP and 48.2% patients with PTE (23% in normal cohort). In conclusion, in the centre region of Portugal, the prevalence of 4G/4G is 23%, 4G/5G is 38% and 5G/5G is 39%; in our cohort of unrelated patients the only significative difference is in the 4G/4G group (47%); this variation maintain in the DVP and PTE group. We did not find difference at the age of the first thrombotic episode, in the three groups.

High Altitude: A Hyper Coagulable State: Results of a Prospective Cohort Study.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4040-4040
Author(s):  
Jyoti Kotwal ◽  
G. S. Chopra ◽  
A. Kotwal ◽  
Y. V. Sharma ◽  
J. R. Bhardwaj
Keyword(s):  
Cohort Study ◽  
Platelet Count ◽  
Platelet Activation ◽  
Protein C ◽  
Protein S ◽  
Prolonged Stay ◽  
C Protein ◽  
Apc Resistance ◽  
The Mean ◽  
Pai 1

Abstract Thrombosis has been recognized as a complication of rapid ascent as well as long-term stay at high altitude (HA). Researchers have presented conflicting results and majority of them studied only short-term stay. A prospective cohort study was carried out at a height of 3500 m to study the hematological factors, which may lead to increased propensity to thrombosis on prolonged stay at HA. The subjects were healthy low landers (initial N=38, complete follow up N=32) in age group of 20–40 years, inducted to HA and investigated at induction and subsequently at 3 and 8 months. The comparison cohort was age and sex matched low landers not inducted to HA. Bleeding time, clotting time, hemoglobin (Hb), HCT, platelet count, prothrombin time, APTT, fibrinogen, d-dimers. Protein C, protein S, anti-thrombin III levels, APC resistance, PAI-1, Beta thromboglobulin (BTG) and PF4 were estimated. On induction to HA the mean Hb, platelet count, fibrinogen levels, PAI-1 levels and levels of platelet activation factors were in normal range and not statistically different from those at low altitude (P>0.05). None of the subjects had thrombophilia. There was no statistically significant change in Protein C, Protein S, AT III levels, APC resistance, BT, CT, PT and APTT. The mean values were: Hb (13.9, 15.6, 16.6 gm/dl); platelet count (255, 309, 343 x 103/mm3); fibrinogen (253, 304, 346 mg/dl); BTG (30.3, 38.5, 47.3 IU/ml); PF 4 (3.9, 7.6, 13.7 IU/ml) and; PAI-1 (23.7, 40.1, 49.3 ng/ml), at (induction, 3 months, 8 months) respectively. The P value for all was 0.000 by repeated measure analysis. The high Hb itself is unlikely to be the single cause of the thrombotic tendency at HA as its maximum value was 18.0 gm/dl. Erythrocytosis and hyper viscosity are known to activate platelets, however, platelet activation may be the result of hypoxic stress and injury to platelets or endothelium. An important finding of the study was the rise in PAI-1 levels that correlated with rise in fibrinogen levels and duration of stay at HA (Figure 1 and 2). The endothelial injury and clotting activation followed by increased fibrinolysis maintains homeostasis. We hypothesize that increased PAI-1 level tilts the balance by decreasing fibrinolytic activity and increasing propensity to thrombosis. Hypoxia during surgery is known to cause raised postoperative PAI-1 levels leading to a tendency of postoperative thrombosis. The prolonged hypoxia in a lowlander staying in HA may have a similar effect. By virtue of increased platelet count, hematocrit, platelet activation factors, PAI-1 and fibrinogen, prolonged stay at HA is a pro thrombotic state. Thus, there may be a role for antiplatelet drugs in prevention in view of the platelet hyperactivity and interventional trials with homocysteine lowering vitamins or aspirin also need to be considered.

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