Effect of Heparin on the Activation of Factor XII and the Contact System in Plasma

1991 ◽  
Vol 66 (05) ◽  
pp. 540-547 ◽  
Author(s):  
Robin A Pixley ◽  
Anita Cassello ◽  
Raul A De La Cadena ◽  
Nathan Kaufman ◽  
Robert W Colman
Keyword(s):  
Molecular Weight ◽  
Human Plasma ◽  
High Molecular Weight ◽  
Anticoagulant Therapy ◽  
Human Factor ◽  
Dextran Sulfate ◽  
Contact System ◽  
Factor Xii ◽  
High Molecular Weight Kininogen

SummaryWe examined in purified systems and in human plasma whether heparin serves as a contact system activating compound. Purified human factor XII zymogen was not activated by heparin through an autoactivation mechanism, but was activated in the presence of purified prekallikrein. Zn2+ (12 εM) did not support autoactivation by heparin. The activation of factor XII and the contact system by heparin in plasma anticoagulated with citrate or with hirudin (not chelating ions) was examined by the cleavage of 125I-labeled factor XII and high molecular weight kininogen (HK). Heparin at 1.6 and 16 USP U/ml was not able to produce activation, in contrast to dextran sulfate (20 εg/ml) which supported activation of both factor XII and HK. This study indicates that heparinized plasma does not support activation of the contact system mediated through activation of factor XII. It is not expected that heparin anticoagulant therapy will contribute to activation of the contact system.

Analysis of Intrinsic Fibrinolysis in Human Plasma Induced by Dextran Sulfate

1992 ◽  
Vol 67 (04) ◽  
pp. 440-444 ◽  
Author(s):  
Hiroko Tsuda ◽  
Toshiyuki Miyata ◽  
Sadaaki Iwanaga ◽  
Tetsuro Yamamoto
Keyword(s):  
Molecular Weight ◽  
Human Plasma ◽  
Dextran Sulfate ◽  
Tissue Type ◽  
Factor Xii ◽  
Normal Human Plasma ◽  
High Molecular Weight Kininogen ◽  
Single Chain ◽  
Major Pathway ◽  
Normal Human

SummaryThe analysis of normal human plasma by fibrin autography revealed four species of plasminogen activator (PA) activity related to tissue-type PA, factor XII, prekallikrein and urokinase-type PA (u-PA). The u-PA activity increased significantly by incubating plasma with dextran sulfate. This increase was coincident with both the cleavage of factor XII and the complex formation of activated factor XII with its plasma inhibitors, which were determined by immunoblotting procedure. The dextran sulfate-dependent activation of u-PA required both factor XII and prekallikrein, but did not require either plasminogen or factor XI. High molecular weight kininogen was required only at a low concentration of dextran sulfate. Thus the results indicate that the factor XII and prekallikrein-mediated activation of single chain u-PA (scu-PA) operates as a major pathway of scu-PA activation in whole plasma in contact with dextran sulfate.

Functional Characterization of a Variant Prekallikrein (PK Zürich)

1993 ◽  
Vol 70 (03) ◽  
pp. 427-432 ◽  
Author(s):  
W A Wuillemin ◽  
M Furlan ◽  
A von Felten ◽  
B Lämmle
Keyword(s):  
Molecular Weight ◽  
Enzymatic Activity ◽  
High Molecular Weight ◽  
Dextran Sulfate ◽  
Functional Characterization ◽  
Factor Xii ◽  
High Molecular Weight Kininogen ◽  
Single Chain ◽  
Normal Plasma ◽  
Sulfate Activation

SummaryThe plasma of a 68-year-old man with cross reacting material (CRM)-positive prekallikrein (PK) deficiency was studied. PK clotting activity was <0.01 U/ml, and PK antigen was 0.1 U/ml. No circulating anticoagulant against PK was detectable. The abnormal PK molecule, denoted as prekallikrein Zürich, was partially characterized by immunological and functional studies on the propositus’ plasma. Immunobiotting analysis showed the abnormal PK being a single chain molecule of the same M r (80 kDa) as normal PK. Dextran sulfate activation of the propositus’ plasma did not lead to proteolytic cleavage of the variant PK molecule, in contrast to dextran sulfate activation of a mixture of 1 volume normal plasma and 9 volumes CRM-negative PK deficient plasma. Agarose gel electrophoresis followed by immunoblotting demonstrated that PK Zürich was complexed with high molecular weight kininogen similarly to PK in normal plasma. Incubation of the propositus’ plasma with purified β-FXIIa resulted in impaired cleavage of PK Zürich when compared with PK hydrolysis in a mixture of 10% normal plasma and 90% CRM-negative PK deficient plasma. Moreover, proteolytically cleaved PK Zürich showed no enzymatic activity against factor XII and high molecular weight kininogen.These studies show that the functional defect of prekallikrein Zürich is due to an impaired cleavage by activated factor XII and probably the lack of enzymatic activity of the cleaved variant molecule.

The Levels of Factor XIIa Generated in Human Plasma on an Electronegative Surface Are Insensitive to Wide Variation in the Concentration of FXII, Prekallikrein, High Molecular Weight Kininogen or FXI

1999 ◽  
Vol 82 (09) ◽  
pp. 1033-1040 ◽  
Author(s):  
K. A. Mitropoulos
Keyword(s):  
Molecular Weight ◽  
Human Plasma ◽  
High Molecular Weight ◽  
Bulk Phase ◽  
Factor Xii ◽  
Normal Human Plasma ◽  
High Molecular Weight Kininogen ◽  
Wide Range ◽  
Factor Xiia ◽  
Normal Human

SummaryThe contribution of the various components of the contact system in the generation of factor XIIa (FXIIa) and of kallikrein (KRN) on an electronegative surface and the release of the generated enzymes to the bulk phase was examined in mixtures of normal human plasma and plasmas congenitally deficient in these components. The incubation of normal human plasma in the presence of sulphatide vesicles (40 μM) resulted in a fast generation of amidolytic activities due to FXIIa and to KRN followed by slower first-order inactivation rates of FXIIa (k’FXIIa) and of KRN (k’KRN) due to the presence of esterase inhibitors. Variation of the levels of factor XII (FXII), over a wide range, showed little effect on levels of FXIIa and of KRN but no activities were detected in 100% FXII-deficient plasma. The variation of prekallikrein (PKRN) concentration showed little effect on the generation of FXIIa but the generation of KRN declined linearly with the decrease in the level of PKRN. No activities were detected on treatment of PKRN-deficient plasma. The variation in the concentration of high molecular weight kininogen (HK) showed effects on FXIIa and KRN that were qualitatively similar to those seen on variation of PKRN but 100% HK-deficient plasma generated considerable activities of both FXIIa and KRN. The variation in the concentration of factor XI (FXI) showed no effect on the generation of FXIIa, whereas KRN levels increased linearly with the contribution of FXI-deficient in normal plasma. The present results suggest that the contiguous binding of FXIIa, FXII, PKRN-HK and FXI-HK onto the electronegative surface induces a rapid generation of FXIIa and KRN. The bound PKRN-HK complex prevents the release of generated FXIIa and therefore further binding and activation of FXII from the bulk phase. Consequently, the turnover of FXII is independent of its levels in the bulk phase and is rather related to the concentration of contact surface. The generated KRN is also protected by HK. However, since the enzyme responsible for the activation of PKRN-HK is FXIIa, the levels of generated KRN are positively related to the concentration of substrate.

Bacterial Expression of Biologically Active High Molecular Weight Kininogen Light Chain

1992 ◽  
Vol 67 (04) ◽  
pp. 428-433 ◽  
Author(s):  
Satya P Kunapuli ◽  
Raul A DeLa Cadena ◽  
Robert W Colman
Keyword(s):  
Molecular Weight ◽  
Human Plasma ◽  
High Molecular Weight ◽  
Light Chain ◽  
Specific Activity ◽  
Bacterial Expression ◽  
Contact System ◽  
Biologically Active ◽  
High Molecular Weight Kininogen ◽  
Single Chain

SummaryHuman high molecular weight kininogen (HK), a single chain plasma glycoprotein, serves as a cofactor in the contact system of blood coagulation. After cleavage by human plasma kallikrein, the nonapeptide bradykinin is released. The HK light chain (LC) contains coagulant activity, which requires both the ability to bind the contact system zymogens, prekallikrein and factor XI, and the ability to interact with negatively charged surfaces. Since bacterial expression might not be successful if carbohydrate was required for activity, we evaluated that possibility by incubating plasma HK with endoglycosydase F. Although the procedure removed detectable N-linked carbohydrate, no change in specific activity occurred. We then developed a bacterial expression system to produce recombinant HK LC. The cDNA coding for the HK LC was prepared by polymerase chain reaction (PCR), digested with restriction enzymes EcoRI and PstI, and introduced into the bacterial expression vector pKK223-3. E. coli harboring this recombinant plasmid (pSKl) expressed HK LC upon induction with isopropylthio-galactoside (IPTG). The recombinant protein (27 kDa), when transferred onto a PVDF membrane, was recognized by monospecific polyclonal anti-HK LC-antibodies. The recombinant HK LC was purified by heparin agarose affinity chromatography to homogeneity and found to have a specific activity of 28 coagulant units per mg protein, similar to the specific activity of the LC derived by proteolytic digestion of human plasma HK. We conclude: 1) The HK LC synthesized in bacteria is biologically active, and 2) the 40% carbohydrate content of the HK LC is not required for its cofactor activity.

Fibrinolysis and the Contact System: A Role for Factor XI in the Down-Regulation of Fibrinolysis

1999 ◽  
Vol 82 (08) ◽  
pp. 243-250 ◽  
Author(s):  
Joost Meijers ◽  
Bonno Bouma
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Fibrinolytic System ◽  
Contact System ◽  
Factor Xii ◽  
Charged Surface ◽  
High Molecular Weight Kininogen ◽  
Factor Xi ◽  
Factor Xiia

IntroductionExposure of blood to negatively-charged surfaces, such as collagen, kaolin, or glass, results in the activation of the contact system of the intrinsic pathway of coagulation. Prekallikrein, factor XII, high molecular weight kininogen, and factor XI are the proteins involved in this contact reaction. The assembly of these components on a negatively-charged surface leads to the activation of factor XI, thereby propagating the intrinsic coagulation pathway. Simultaneously, several other reactions occur, such as the activation of factor VII and the initiation of the fibrinolytic system, kinin-forming pathway, and renin-angiotensin pathway.The first step in the contact phase is to bind factor XII to the negatively-charged surface, making it highly susceptible for proteolysis by kallikrein.1-3 Activated factor XII (α-factor XIIa) is formed in a process that may involve autoactivation.4-7 Prekallikrein is bound to high molecular weight kininogen in plasma. High molecular weight kininogen associates with a negatively-charged surface, thereby localizing prekallikrein to the surface. Limited proteolysis by α-factor XIIa converts prekallikrein to kallikrein. Kallikrein can dissociate from the surface and act on surface-bound factor XII at distant sites, thereby propagating the reciprocal cycle.7 Factor XI circulates plasma in a complex with high molcular weight kininogen. High molecular weight kininogen links factor XI to a negatively charged surface where it is activated by surface bound:α-factor XIIa. Although the in vivo, activating, negatively-charged surface is unknown, assembly and activation of the contact system on biological membranes of endothelial cells, platelets, neutrophils, and monocytes can take place, suggesting that these surfaces are the actual activating surfaces in vivo.8 The physiological significance of the contact system in blood coagulation remains unclear, however, because a deficiency of factor XII, prekallikrein, and high molecular weight kininogen does not result in a bleeding disorder. In contrast, patients deficient in factor XI, most common among Ashkenazi Jews, do suffer from variable bleeding abnormalities, especially from tissues with high local fibrinolytic activity (e.g., urinary tract, nose, oral cavity, tonsils).9,10 This suggested that there was an alternative route for the activation of factor XI, and recently, such a route was described.11,12 Thrombin was found to activate factor XI even in the absence of a negatively-charged surface,11-15 and factor XI was shown to play a role in the downregulation of fibrinolysis.16 In this article, the role of the contact system, with an emphasis on factor XI in the regulation of the fibrinolytic system, will be described.

Factor XI and Protection of the Fibrin Clot against Lysis – a Role for the Intrinsic Pathway of Coagulation in Fibrinolysis

1998 ◽  
Vol 80 (07) ◽  
pp. 24-27 ◽  
Author(s):  
Peter von dem Borne ◽  
Joost Meijers ◽  
Bonno Bouma
Keyword(s):  
Molecular Weight ◽  
Blood Coagulation ◽  
High Molecular Weight ◽  
Fibrin Clot ◽  
Contact System ◽  
Activate Factor ◽  
Factor Xii ◽  
High Molecular Weight Kininogen ◽  
Intrinsic Pathway ◽  
Factor Xi

IntroducationBlood coagulation is an important mechanism that maintains the integrity of the vascular system to prevent blood loss after injury. The conceptions on the working mechanism of coagulation are based on the waterfall or cascade model, which was already proposed more than 30 years ago, independently by Davie and Ratnoff (1) and MacFarlane (2). Blood coagulation was viewed as a series of linked proteolytic reactions in which zymogens are converted into serine proteases, ultimately leading to the formation of thrombin, which converts soluble fibrinogen into insoluble fibrin. Coagulation was thought to proceed via two pathways, an extrinsic and an intrinsic pathway. Activation of the extrinsic pathway of coagulation occurs by the exposition of tissue factor at the site of injury (3) whereas the intrinsic system is activated after exposure of plasma to an activating surface. Although the in vivo activating surface is unknown, the contact system was believed to play a role in the initiation of the intrinsic pathway. This system consists of factor XII, prekallikrein, high molecular weight kininogen and factor XI. The physiological relevance of the contact system is unclear, since a deficiency of factor XII, prekallikrein or high molecular weight kininogen does not result in a bleeding disorder. In contrast, patients deficient in factor XI, most common among Ashkenazi Jews, do suffer from variable bleeding abnormalities especially from tissues with high local fibrinolytic activity (urinary tract, nose, oral cavity, tonsils) (4, 5). This suggested there was an alternative route for the activation of factor XI, and recently such a route was described (6, 7). Thrombin was found to activate factor XI, even in the absence of a negatively charged surface (6-11), and factor XI was shown to play a role in the protection of the fibrin clot against lysis (9). In plasma the possibility cannot be excluded that the activation of factor XI by thrombin takes place via an intermediary component. Recently, it was shown that meizothrombin was capable of activating factor XI (12).

Zinc-dependent contact system activation induces vascular leakage and hypotension in rodents

2013 ◽  
Vol 394 (9) ◽  
pp. 1195-1204 ◽  
Author(s):  
Jenny Björkqvist ◽  
Bernd Lecher ◽  
Coen Maas ◽  
Thomas Renné
Keyword(s):  
Blood Pressure ◽  
Molecular Weight ◽  
High Molecular Weight ◽  
Dextran Sulfate ◽  
Vascular Leakage ◽  
Laser Scanning Microscopy ◽  
Factor Xii ◽  
Dependent Manner ◽  
High Molecular Weight Kininogen

Abstract Contact to polyanions induces autoactivation of the serine protease factor XII that triggers the kallikrei-kinin system. Recent studies indicate that polysaccharide-induced autoactivation of factor XII has a role in allergy-related vascular leakage, and angioedema. Here, we characterize in vivo effects of the synthetic polysaccharide dextran sulfate in human plasma and in rodent models. Minute amounts of high-molecular-weight dextran sulfate-initiated factor XII-autoactivation and triggered formation of the inflammatory mediator bradykinin via plasma kallikrein-mediated cleavage of high-molecular-weight kininogen. High-molecular-weight kininogen fragments, containing the HKH20 sequence in domain D5H, blocked dextran sulfate-initiated bradykinin-generation by depleting plasma Zn2+ ions. Topical application of high molecular weight dextran sulfate increased leakage in murine skin microvessels, in a bradykinin-dependent manner. Intravital laser scanning microscopy showed a greater than two-fold elevated and accelerated fluid extravasation in C1 esterase inhibitor deficient mice that lack the major inhibitor of factor XII, compared to wild-type controls. Intra-arterial infusion of dextran sulfate induced a rapid transient drop in arterial blood pressure in rats and preinjection of kinin B2 receptor antagonists or HKH20 peptide blunted dextran sulfate-triggered hypotensive reactions. The data characterize dextran sulfate as a potent in vivo activator of factor XII with implications for bradykinin-mediated vascular permeability and blood pressure control.

The Contact Phase of Blood Coagulation in Diabetes Mellitus and in Patients with Vasculopathy

1984 ◽  
Vol 52 (03) ◽  
pp. 221-223 ◽  
Author(s):  
M Christe ◽  
P Gattlen ◽  
J Fritschi ◽  
B Lämmle ◽  
W Berger ◽  
...  
Keyword(s):  
Diabetes Mellitus ◽  
Molecular Weight ◽  
Blood Coagulation ◽  
High Molecular Weight ◽  
Negative Correlation ◽  
Factor Xii ◽  
C1 Inhibitor ◽  
High Molecular Weight Kininogen ◽  
Contact Phase ◽  
Α2 Macroglobulin

SummaryThe contact phase has been studied in diabetics and patients with macroangiopathy. Factor XII and high molecular weight kininogen (HMWK) are normal. C1-inhibitor and also α2-macroglobulin are significantly elevated in diabetics with complications, for α1-macroglobulin especially in patients with nephropathy, 137.5% ± 36.0 (p <0.001). C1-inhibitor is also increased in vasculopathy without diabetes 113.2 ± 22.1 (p <0.01).Prekallikrein (PK) is increased in all patients’ groups (Table 2) as compared to normals. PK is particularly high (134% ± 32) in 5 diabetics without macroangiopathy but with sensomotor neuropathy. This difference is remarkable because of the older age of diabetics and the negative correlation of PK with age in normals.

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