scholarly journals CpaA Is a Glycan-Specific Adamalysin-like Protease Secreted byAcinetobacter baumanniiThat Inactivates Coagulation Factor XII

mBio ◽  
2018 ◽  
Vol 9 (6) ◽  
Author(s):  
Ursula Waack ◽  
Mark Warnock ◽  
Andrew Yee ◽  
Zachary Huttinger ◽  
Sara Smith ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Defense Mechanisms ◽  
Thrombus Formation ◽  
Coagulation Factor ◽  
Factor Xii ◽  
Contact Activation ◽  
Extrinsic Pathway ◽  
Content Type

ABSTRACTAntibiotic-resistantAcinetobacter baumanniiis increasingly recognized as a cause of difficult-to-treat nosocomial infections, including pneumonia, wound infections, and bacteremia. Previous studies have demonstrated that the metalloprotease CpaA contributes to virulence and prolongs clotting time when added to human plasma as measured by the activated partial thromboplastin time (aPTT) assay. Here, we show that CpaA interferes with the intrinsic coagulation pathway, also called the contact activation system, in human as well as murine plasma, but has no discernible effect on the extrinsic pathway. By utilizing a modified aPTT assay, we demonstrate that coagulation factor XII (fXII) is a target of CpaA. In addition, we map the cleavage by CpaA to two positions, 279-280 and 308-309, within the highly glycosylated proline-rich region of human fXII, and show that cleavage at the 308-309 site is responsible for inactivation of fXII. At both sites, cleavage occurs between proline and an O-linked glycosylated threonine, and deglycosylation of fXII prevents cleavage by CpaA. Consistent with this, mutant fXII (fXII-Thr309Lys) from patients with hereditary angioedema type III (HAEIII) is protected from CpaA inactivation. This raises the possibility that individuals with HAEIII who harbor this mutation may be partially protected fromA. baumanniiinfection if CpaA contributes to human disease. By inactivating fXII, CpaA may attenuate important antimicrobial defense mechanisms such as intravascular thrombus formation, thus allowingA. baumanniito disseminate.IMPORTANCEVentilator-associated pneumonia and catheter-related bacteremia are the most common and severe infections caused byAcinetobacter baumannii. Besides the capsule, lipopolysaccharides, and the outer membrane porin OmpA, little is known about the contribution of secreted proteins toA. baumanniisurvivalin vivo. Here we focus on CpaA, a potentially recently acquired virulence factor that inhibits blood coagulationin vitro. We identify coagulation factor XII as a target of CpaA, map the cleavage sites, and show that glycosylation is a prerequisite for CpaA-mediated inactivation of factor XII. We propose adding CpaA to a small, but growing list of bacterial proteases that are specific for highly glycosylated components of the host defense system.

scholarly journals In vivo roles of factor XII

Blood ◽  
2012 ◽  
Vol 120 (22) ◽  
pp. 4296-4303 ◽  
Author(s):  
Thomas Renné ◽  
Alvin H. Schmaier ◽  
Katrin F. Nickel ◽  
Margareta Blombäck ◽  
Coen Maas
Keyword(s):  
Thrombus Formation ◽  
Coagulation Factor ◽  
Special Focus ◽  
Factor Xii ◽  
Excessive Bleeding ◽  
Charged Surfaces ◽  
Factor Xiia ◽  
Knockout Model

Abstract Coagulation factor XII (FXII, Hageman factor, EC = 3.4.21.38) is the zymogen of the serine protease, factor XIIa (FXIIa). FXII is converted to FXIIa through autoactivation induced by “contact” to charged surfaces. FXIIa is of crucial importance for fibrin formation in vitro, but deficiency in the protease is not associated with excessive bleeding. For decades, FXII was considered to have no function for coagulation in vivo. Our laboratory developed the first murine knockout model of FXII. Consistent with their human counterparts, FXII−/− mice have a normal hemostatic capacity. However, thrombus formation in FXII−/− mice is largely defective, and the animals are protected from experimental cerebral ischemia and pulmonary embolism. This murine model has created new interest in FXII because it raises the possibility for safe anticoagulation, which targets thrombosis without influence on hemostasis. We recently have identified platelet polyphosphate (an inorganic polymer) and mast cell heparin as in vivo FXII activators with implications on the initiation of thrombosis and edema during hypersensitivity reactions. Independent of its protease activity, FXII exerts mitogenic activity with implications for angiogenesis. The goal of this review is to summarize the in vivo functions of FXII, with special focus to its functions in thrombosis and vascular biology.

scholarly journals Contradictory functions of sulfatide in the blood coagulation system as coagulant and anticoagulant.

1998 ◽  
Vol 45 (2) ◽  
pp. 493-499 ◽  
Author(s):  
M Kyogashima ◽  
J Onaya ◽  
A Hara ◽  
T Taketomi
Keyword(s):  
Blood Coagulation ◽  
Thrombus Formation ◽  
Coagulation Factor ◽  
Coagulation System ◽  
Factor Xii ◽  
Blood Coagulation System ◽  
Coagulant Activity ◽  
J 13

Sulfatide (galactosylceramide I3 -sulfate) has been reported to activate blood coagulation factor XII (Hageman factor), which suggests that it exhibits coagulant activity (Fujikama et al., 1980 Biochemistry 19, 1322-1330) However, sulfatide administered into animals as a bolus shot without subsequent thrombus formation, prolonged conventional clotting times and bleeding time (Hara et al., 1996 Glycoconjugate J. 13, 187-194). These findings suggest that it may exhibit anticoagulant rather than coagulant activity. Following this suggestion we found in vitro that binding of sulfatide to fibrinogen resulted in disturbance of fibrin formation. To examine a possible pharmacological effect of sulfatide on blood coagulation in vivo we continuously infused sulfatide into rats through plastic cannulae and found formation of giant thrombi around the tips of the cannulae. These data suggest that sulfatide may exhibit contradictory functions in the blood coagulation system.

scholarly journals Coagulation factor XII in thrombosis and inflammation

Blood ◽  
2018 ◽  
Vol 131 (17) ◽  
pp. 1903-1909 ◽  
Author(s):  
Coen Maas ◽  
Thomas Renné
Keyword(s):  
Thrombus Formation ◽  
Coagulation Factor ◽  
Contact System ◽  
Inflammatory Disorder ◽  
Factor Xii ◽  
Kinin System ◽  
Disease States ◽  
Exciting Opportunity

Abstract Combinations of proinflammatory and procoagulant reactions are the unifying principle for a variety of disorders affecting the cardiovascular system. The factor XII–driven contact system starts coagulation and inflammatory mechanisms via the intrinsic pathway of coagulation and the bradykinin-producing kallikrein-kinin system, respectively. The biochemistry of the contact system in vitro is well understood; however, its in vivo functions are just beginning to emerge. Challenging the concept of the coagulation balance, targeting factor XII or its activator polyphosphate, provides protection from thromboembolic diseases without interfering with hemostasis. This suggests that the polyphosphate/factor XII axis contributes to thrombus formation while being dispensable for hemostatic processes. In contrast to deficiency in factor XII providing safe thromboprotection, excessive FXII activity is associated with the life-threatening inflammatory disorder hereditary angioedema. The current review summarizes recent findings of the polyphosphate/factor XII–driven contact system at the intersection of procoagulant and proinflammatory disease states. Elucidating the contact system offers the exciting opportunity to develop strategies for safe interference with both thrombotic and inflammatory disorders.

scholarly journals Heparin is more effective than apixaban in inhibiting in vitro contact activated coagulation

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
M.M Engelen ◽  
C Van Laer ◽  
M Jacquemin ◽  
C Vandenbriele ◽  
K Peerlinck ◽  
...  
Keyword(s):  
Thrombin Generation ◽  
Heart Valves ◽  
Thrombus Formation ◽  
Oral Anticoagulants ◽  
Direct Oral Anticoagulants ◽  
Factor Xa ◽  
Coagulation Factor ◽  
Mechanical Heart Valves ◽  
Contact Activation

Abstract Introduction Contact of blood with artificial surfaces such as mechanical support devices, catheters, and mechanical heart valves activates the contact activation (CA) pathway of coagulation. Furthermore, recent animal data and clinical studies suggest a more important contribution of CA in pathological thrombus formation in other cardiovascular diseases. Direct oral anticoagulants (DOACs) are recommended as first-line treatment in most patients who require long-term anticoagulation. However, because DOACs directly inhibit a single downstream coagulation factor (thrombin (fXIIa) or factor Xa (fXa)), it has been suggested that their efficacy could be reduced in the presence of strong activation of the CA pathway as compared to anticoagulants that target multiple, more upstream located coagulation factors. Purpose To compare the efficacy of a DOAC (apixaban) and heparin to suppress thrombin generation in the presence of strong CA pathway activation. Methods Pooled platelet-poor plasma was spiked with either apixaban (dissolved in DMSO and PBS) or unfractionated heparin to achieve therapeutic plasma levels. SynthASil, a commercially available mixture of phospholipids and silica, was used to stimulate the CA pathway in two different dilutions (1–80 and 5–80). Downstream coagulation was accessed by Thrombin Generation Test using Thrombinoscope by Stago and associated Thrombin Calibrator (activity 640 nM). The endogenous thrombin potential (area under the thrombin generation curve; ETP), peak thrombin generation (PTG), time to peak (ttPeak) and time to start (ttStart) were accessed. Results With decreasing concentrations of apixaban, stimulation with the lower dose SynthASil reveals an increasing ETP and PTG. As expected, ttPeak and ttStart decreased. Even supratherapeutic levels of apixaban (i.e. 1120 ng/mL) could not inhibit thrombin from being generated, in striking contrast with UFH where no thrombin was formed. Using a five times higher dose of SynthASil showed comparable ETP for all concentrations of apixaban, allocated around the control value. PTG, however, slightly increased with decreasing concentrations of apixaban. ttPeak and ttStart slightly decreased. Except for the subtherapeutic UFH concentration of 0,114 IU/mL, no thrombin was generated with UFH. Conclusion UFH is more effective in inhibiting downstream thrombin generation compared to apixaban as a response to activation of the CA pathway in vitro. These findings could help explain why direct inhibitors were not able to show non-inferiority in patients with mechanical heart valves and support the development of specific CA pathway inhibitors for patients with conditions that activate the CA pathway. Thrombin generation curves Funding Acknowledgement Type of funding source: None

scholarly journals Assessment of Minocycline and Polymyxin B Combination against Acinetobacter baumannii

2015 ◽  
Vol 59 (5) ◽  
pp. 2720-2725 ◽  
Author(s):  
Dana R. Bowers ◽  
Henry Cao ◽  
Jian Zhou ◽  
Kimberly R. Ledesma ◽  
Dongxu Sun ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Clinical Utility ◽  
Efflux Pump ◽  
Polymyxin B ◽  
Intracellular Concentration ◽  
Bacterial Cells ◽  
Efflux Pump Inhibitor ◽  
Content Type

ABSTRACTAntimicrobial resistance amongAcinetobacter baumanniiis increasing worldwide, often necessitating combination therapy. The clinical utility of using minocycline with polymyxin B is not well established. In this study, we investigated the activity of minocycline and polymyxin B against 1 laboratory isolate and 3 clinical isolates ofA. baumannii. Minocycline susceptibility testing was performed with and without an efflux pump inhibitor, phenylalanine-arginine β-naphthylamide (PAβN). The intracellular minocycline concentration was determined with and without polymyxin B (0.5 μg/ml). Time-kill studies were performed over 24 h using approximately 106CFU/ml of each strain with clinically relevant minocycline concentrations (2 μg/ml and 8 μg/ml), with and without polymyxin B (0.5 μg/ml). Thein vivoefficacy of the combination was assessed in a neutropenic murine pneumonia model. Infected animals were administered minocycline (50 mg/kg), polymyxin B (10 mg/kg), or both to achieve clinically equivalent exposures in humans. A reduction in the minocycline MIC (≥4×) was observed in the presence of PAβN. The intracellular concentration andin vitrobactericidal effect of minocycline were both enhanced by polymyxin B. With 2 minocycline-susceptible strains, the bacterial burden in lung tissue at 24 h was considerably reduced by the combination compared to monotherapy with minocycline or polymyxin B. In addition, the combination prolonged survival of animals infected with a minocycline-susceptible strain. Polymyxin B increased the intracellular concentration of minocycline in bacterial cells and enhanced the bactericidal activity of minocycline, presumably due to efflux pump disruption. The clinical utility of this combination should be further investigated.

scholarly journals Targeting coagulation factor XII provides protection from pathological thrombosis in cerebral ischemia without interfering with hemostasis

2006 ◽  
Vol 203 (3) ◽  
pp. 513-518 ◽  
Author(s):  
Christoph Kleinschnitz ◽  
Guido Stoll ◽  
Martin Bendszus ◽  
Kai Schuh ◽  
Hans-Ulrich Pauer ◽  
...  
Keyword(s):  
Thrombus Formation ◽  
Coagulation Factor ◽  
Artery Occlusion ◽  
Factor Xii ◽  
Intrinsic Pathway ◽  
Fibrin Formation ◽  
Vessel Injury ◽  
Normal Hemostasis ◽  
A Site

Formation of fibrin is critical for limiting blood loss at a site of blood vessel injury (hemostasis), but may also contribute to vascular thrombosis. Hereditary deficiency of factor XII (FXII), the protease that triggers the intrinsic pathway of coagulation in vitro, is not associated with spontaneous or excessive injury-related bleeding, indicating FXII is not required for hemostasis. We demonstrate that deficiency or inhibition of FXII protects mice from ischemic brain injury. After transient middle cerebral artery occlusion, the volume of infarcted brain in FXII-deficient and FXII inhibitor–treated mice was substantially less than in wild-type controls, without an increase in infarct-associated hemorrhage. Targeting FXII reduced fibrin formation in ischemic vessels, and reconstitution of FXII-deficient mice with human FXII restored fibrin deposition. Mice deficient in the FXII substrate factor XI were similarly protected from vessel-occluding fibrin formation, suggesting that FXII contributes to pathologic clotting through the intrinsic pathway. These data demonstrate that some processes involved in pathologic thrombus formation are distinct from those required for normal hemostasis. As FXII appears to be instrumental in pathologic fibrin formation but dispensable for hemostasis, FXII inhibition may offer a selective and safe strategy for preventing stroke and other thromboembolic diseases.

scholarly journals A mechanism for hereditary angioedema with normal C1 inhibitor: an inhibitory regulatory role for the factor XII heavy chain

Blood ◽  
2019 ◽  
Vol 133 (10) ◽  
pp. 1152-1163 ◽  
Author(s):  
Ivan Ivanov ◽  
Anton Matafonov ◽  
Mao-fu Sun ◽  
Bassem M. Mohammed ◽  
Qiufang Cheng ◽  
...  
Keyword(s):  
Heavy Chain ◽  
Hereditary Angioedema ◽  
Catalytic Efficiency ◽  
Regulatory Function ◽  
Factor Xii ◽  
C1 Inhibitor ◽  
Contact Activation ◽  
Inhibitory Function

Abstract The plasma proteins factor XII (FXII) and prekallikrein (PK) undergo reciprocal activation to the proteases FXIIa and kallikrein by a process that is enhanced by surfaces (contact activation) and regulated by the serpin C1 inhibitor. Kallikrein cleaves high-molecular-weight kininogen (HK), releasing the vasoactive peptide bradykinin. Patients with hereditary angioedema (HAE) experience episodes of soft tissue swelling as a consequence of unregulated kallikrein activity or increased prekallikrein activation. Although most HAE cases are caused by reduced plasma C1-inhibitor activity, HAE has been linked to lysine/arginine substitutions for Thr309 in FXII (FXII-Lys/Arg309). Here, we show that FXII-Lys/Arg309 is susceptible to cleavage after residue 309 by coagulation proteases (thrombin and FXIa), resulting in generation of a truncated form of FXII (δFXII). The catalytic efficiency of δFXII activation by kallikrein is 15-fold greater than for full-length FXII. The enhanced rate of reciprocal activation of PK and δFXII in human plasma and in mice appears to overwhelm the normal inhibitory function of C1 inhibitor, leading to increased HK cleavage. In mice given human FXII-Lys/Arg309, induction of thrombin generation by infusion of tissue factor results in enhanced HK cleavage as a consequence of δFXII formation. The effects of δFXII in vitro and in vivo are reproduced when wild-type FXII is bound by an antibody to the FXII heavy chain (HC; 15H8). The results contribute to our understanding of the predisposition of patients carrying FXII-Lys/Arg309 to angioedema after trauma, and reveal a regulatory function for the FXII HC that normally limits PK activation in plasma.

SECONDARY FIBRINOLYSIS, PROTEIN C ACTIVATION, PLATELET DECREASE, BUT NOT CONTACT ACTIVATION CAN BE CORRELATED TO FREE THROMBIN ACTION IN EXPERIMENTAL DIC

1987 ◽  
Author(s):  
G A Marbet ◽  
P Satiropas ◽  
C Pantaleoni ◽  
F Duckert
Keyword(s):  
Protein C ◽  
Defense Mechanisms ◽  
Dermatan Sulfate ◽  
Correlation Coefficients ◽  
Factor Xii ◽  
Spearman Correlation ◽  
Contact Activation ◽  
Defence Mechanisms ◽  
Tissue Thromboplastin

We have studied the influence of activated coagulation on antithrombotic defence mechanisms in vivo. Conventional coagulation variables, platelets (Tcy), plasminogen (Pig), α2-antiplasmin (AP) , protein C (PC), factor VIII C, factor XII and Cl-inhibitor have been measured before and during reversible tissue thromboplastin-induced DIC in the dog. Free thrombin action as derived from fibrinogen (Fbg) decrease has been expressed as integral of active thrombin concentration over time (φ). Protection by heparin H, pentosan polysulfate PPS or dermatan sulfate DS was studied. DIC had no consistent effect on the behaviour of factor XII and Cl-inhibitor, but led to the consumption (Δ) of the following variables:The Spearman correlation coefficients between and φ in the Δ whole group were all statistically significant and ranged from rs=0.51 (ΔPlg) to rs =0.94 (ΔFbg). The response of major defense mechanisms in vivo quantitatively depends on active thrombin.

scholarly journals In Vivo Fitness Adaptations of Colistin-Resistant Acinetobacter baumannii Isolates to Oxidative Stress

2016 ◽  
Vol 61 (3) ◽  
Author(s):  
Crystal L. Jones ◽  
Shweta S. Singh ◽  
Yonas Alamneh ◽  
Leila G. Casella ◽  
Robert K. Ernst ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Acinetobacter Baumannii ◽  
Catalase Activity ◽  
Content Type ◽  
Increased Susceptibility

ABSTRACT The loss of fitness in colistin-resistant (CR) Acinetobacter baumannii was investigated using longitudinal isolates from the same patient. Early CR isolates were outcompeted by late CR isolates for growth in broth and survival in the lungs of mice. Fitness loss was associated with an increased susceptibility to oxidative stress since early CR strains had reduced in vitro survival in the presence of hydrogen peroxide and decreased catalase activity compared to that of late CR and colistin-susceptible (CS) strains.

scholarly journals Tissue factor–positive neutrophils bind to injured endothelial wall and initiate thrombus formation

Blood ◽  
2012 ◽  
Vol 120 (10) ◽  
pp. 2133-2143 ◽  
Author(s):  
Roxane Darbousset ◽  
Grace M. Thomas ◽  
Soraya Mezouar ◽  
Corinne Frère ◽  
Rénaté Bonier ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Tissue Factor ◽  
Blood Coagulation ◽  
Thrombus Formation ◽  
Coagulation Cascade ◽  
Factor Xii ◽  
Long Time ◽  
Platelet Accumulation

AbstractFor a long time, blood coagulation and innate immunity have been viewed as interrelated responses. Recently, the presence of leukocytes at the sites of vessel injury has been described. Here we analyzed interaction of neutrophils, monocytes, and platelets in thrombus formation after a laser-induced injury in vivo. Neutrophils immediately adhered to injured vessels, preceding platelets, by binding to the activated endothelium via leukocyte function antigen-1–ICAM-1 interactions. Monocytes rolled on a thrombus 3 to 5 minutes postinjury. The kinetics of thrombus formation and fibrin generation were drastically reduced in low tissue factor (TF) mice whereas the absence of factor XII had no effect. In vitro, TF was detected in neutrophils. In vivo, the inhibition of neutrophil binding to the vessel wall reduced the presence of TF and diminished the generation of fibrin and platelet accumulation. Injection of wild-type neutrophils into low TF mice partially restored the activation of the blood coagulation cascade and accumulation of platelets. Our results show that the interaction of neutrophils with endothelial cells is a critical step preceding platelet accumulation for initiating arterial thrombosis in injured vessels. Targeting neutrophils interacting with endothelial cells may constitute an efficient strategy to reduce thrombosis.

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