scholarly journals ADAMTS13: a new link between thrombosis and inflammation

2008 ◽  
Vol 205 (9) ◽  
pp. 2065-2074 ◽  
Author(s):  
Anil K. Chauhan ◽  
Janka Kisucka ◽  
Alexander Brill ◽  
Meghan T. Walsh ◽  
Friedrich Scheiflinger ◽  
...  
Keyword(s):  
Leukocyte Adhesion ◽  
Leukocyte Rolling ◽  
Type I ◽  
Wild Type ◽  
Inflammatory Models ◽  
Adamts13 Deficiency ◽  
Acute And Chronic Inflammation ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Body Secretion

von Willebrand factor (VWF) levels are elevated and a disintegrin-like and metalloprotease with thrombospondin type I repeats–13 (ADAMTS13) activity is decreased in both acute and chronic inflammation. We hypothesized that by cleaving hyperactive ultralarge VWF (ULVWF) multimers, ADAMTS13 down-regulates both thrombosis and inflammation. Using intravital microscopy, we show that ADAMTS13 deficiency results in increased leukocyte rolling on unstimulated veins and increased leukocyte adhesion in inflamed veins. Both processes were dependent on the presence of VWF. Depletion of platelets in Adamts13−/− mice reduced leukocyte rolling, suggesting that platelet interaction with ULVWF contributes to this process. Increased levels of endothelial P-selectin and plasma VWF in Adamts13−/− compared with wild-type (WT) mice indicated an elevated release of Weibel-Palade bodies. ULVWF multimers released upon stimulation with histamine, a secretagogue of Weibel-Palade bodies, slowed down leukocyte rolling in Adamts13−/− but not in WT mice. Furthermore, in inflammatory models, ADAMTS13 deficiency resulted in enhanced extravasation of neutrophils, and this process was also dependent on VWF. Our findings reveal an important role for ADAMTS13 in preventing excessive spontaneous Weibel-Palade body secretion, and in the regulation of leukocyte adhesion and extravasation during inflammation.

Deficiency of the VWF-Cleaving Protease ADAMTS13 Results in Increased Leukocyte Rolling and Adhesion in Mice.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 290-290 ◽  
Author(s):  
Anil K. Chauhan ◽  
Janka Kiucka ◽  
Alexander Brill ◽  
Meghan T. Walsh ◽  
Denisa D. Wagner
Keyword(s):  
Endothelial Cells ◽  
Von Willebrand Factor ◽  
Leukocyte Adhesion ◽  
Surface Expression ◽  
Leukocyte Rolling ◽  
Type I ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Adherent Leukocytes

Abstract von Willebrand factor (VWF) is synthesized in megakaryocytes and endothelial cells and stored in a-granules and Weibel-Palade bodies, respectively. VWF levels are elevated in both chronic and acute inflammation. ADAMTS13 (A D isintegrin-like A nd M etalloprotease with T hrombo s pondin type I repeats-13) is a metalloprotease that cleaves ultra large von Willebrand factor (ULVWF) multimers quickly after its release from endothelium. Recent studies have found that VWF promotes leukocyte adhesion in vitro and that ADAMTS13 activity is reduced in inflammation and sepsis. We hypothesized that by cleaving ULVWF multimers, ADAMTS13 not only inhibits thrombosis, but also attenuates leukocyte rolling and adhesion. Using intravital microscopy, we found more leukocyte rolling/min on the unstimulated veins in Adamts13-/- mice (Mean ± SE: 98 ± 16) compared to WT (Mean ± SE: 35 ± 6, P<0.001), n=18–20 from 10–11 mice per group. This process was dependent on VWF because the number of leukocytes rolling in Adamts13-/-/Vwf-/- veins was similar to that in Vwf-/-. Significantly increased soluble P-selectin and VWF concentrations were found in the plasma of Adamts13-/- compared to WT mice as quantitated by ELISA. In addition, endothelial P-selectin surface expression was increased in Adamts13-/- mice compared to WT. These results suggest elevated release of Weibel-Palade bodies in Adamts13-/- mice. Notably, circulating platelets were not activated in the absence of ADAMTS13. Upon stimulation of the mesentery with histamine, leukocyte rolling was slower in Adamts13-/- veins compared to WT. Furthermore, upon stimulation with the inflammatory cytokine TNF-alpha (i.v) 3.5 h prior to surgery, the number of leukocytes adhering/250 um was significantly increased in microvenules (diameter of 25–30 um) of Adamts13-/- mice (Mean ± SD: 21 ± 6) compared to WT (Mean ± SD: 12 ± 5, P<0.001), n=10–11 mice per group. This firm adhesion was also dependent on VWF because the number of adherent leukocytes in veins of Adamts13-/-/Vwf-/- was similar to Vwf-/-. Our studies indicate a crucial role for ADAMTS13 in preventing excessive spontaneous Weibel-Palade secretion and in attenuating leukocyte rolling and adhesion to ultra large VWF presented by endothelial cells during inflammation.

scholarly journals Activated platelets induce Weibel-Palade–body secretion and leukocyte rolling in vivo: role of P-selectin

Blood ◽  
2005 ◽  
Vol 106 (7) ◽  
pp. 2334-2339 ◽  
Author(s):  
Vandana S. Dole ◽  
Wolfgang Bergmeier ◽  
Heather A. Mitchell ◽  
Sarah C. Eichenberger ◽  
Denisa D. Wagner
Keyword(s):  
Acute Inflammation ◽  
Endothelial Activation ◽  
Leukocyte Rolling ◽  
Activated Platelets ◽  
Baseline Levels ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Body Secretion

AbstractThe presence of activated platelets and platelet-leukocyte aggregates in the circulation accompanies major surgical procedures and occurs in several chronic diseases. Recent findings that activated platelets contribute to the inflammatory disease atherosclerosis made us address the question whether activated platelets stimulate normal healthy endothelium. Infusion of activated platelets into young mice led to the formation of transient platelet-leukocyte aggregates and resulted in a several-fold systemic increase in leukocyte rolling 2 to 4 hours after infusion. Rolling returned to baseline levels 7 hours after infusion. Infusion of activated P-selectin-/- platelets did not induce leukocyte rolling, indicating that platelet P-selectin was involved in the endothelial activation. The endothelial activation did not require platelet CD40L. Leukocyte rolling was mediated solely by the interaction of endothelial P-selectin and leukocyte P-selectin glycoprotein ligand 1 (PSGL-1). Endothelial P-selectin is stored with von Willebrand factor (VWF) in Weibel-Palade bodies. The release of Weibel-Palade bodies on infusion of activated platelets was indicated by both elevation of plasma VWF levels and by an increase in the in vivo staining of endothelial P-selectin. We conclude that the presence of activated platelets in circulation promotes acute inflammation by stimulating secretion of Weibel-Palade bodies and P-selectin–mediated leukocyte rolling.

scholarly journals Systemic antithrombotic effects of ADAMTS13

2006 ◽  
Vol 203 (3) ◽  
pp. 767-776 ◽  
Author(s):  
Anil K. Chauhan ◽  
David G. Motto ◽  
Colin B. Lamb ◽  
Wolfgang Bergmeier ◽  
Michael Dockal ◽  
...  
Keyword(s):  
Platelet Adhesion ◽  
Thrombus Formation ◽  
Type I ◽  
Inhibitory Antibody ◽  
Antithrombotic Agent ◽  
Antithrombotic Effects ◽  
Life Threatening ◽  
Adamts13 Deficiency ◽  
Von Willebrand ◽  
Willebrand Factor

The metalloprotease ADAMTS13 (a disintegrin-like and metalloprotease with thrombospondin type I repeats 13) cleaves highly adhesive large von Willebrand factor (VWF) multimers after their release from the endothelium. ADAMTS13 deficiency is linked to a life-threatening disorder, thrombotic thrombocytopenic purpura (TTP), characterized by platelet-rich thrombi in the microvasculature. Here, we show spontaneous thrombus formation in activated microvenules of Adamts13−/− mice by intravital microscopy. Strikingly, we found that ADAMTS13 down-regulates both platelet adhesion to exposed subendothelium and thrombus formation in injured arterioles. An inhibitory antibody to ADAMTS13 infused in wild-type mice prolonged adhesion of platelets to endothelium and induced thrombi formation with embolization in the activated microvenules. Absence of ADAMTS13 did not promote thrombi formation in αIIbβ3 integrin-inhibited blood. Recombinant ADAMTS13 reduced platelet adhesion and aggregation in histamine-activated venules and promoted thrombus dissolution in injured arterioles. Our findings reveal that ADAMTS13 has a powerful natural antithrombotic activity and recombinant ADAMTS13 could be used as an antithrombotic agent.

Complete deficiency in ADAMTS13 is prothrombotic, but it alone is not sufficient to cause thrombotic thrombocytopenic purpura

Blood ◽  
2006 ◽  
Vol 107 (8) ◽  
pp. 3161-3166 ◽  
Author(s):  
Fumiaki Banno ◽  
Koichi Kokame ◽  
Tomohiko Okuda ◽  
Shigenori Honda ◽  
Shigeki Miyata ◽  
...  
Keyword(s):  
Thrombotic Thrombocytopenic Purpura ◽  
Hemolytic Anemia ◽  
Thrombus Formation ◽  
Wild Type ◽  
Thrombocytopenic Purpura ◽  
Phenotypic Differences ◽  
System Functioning ◽  
Adamts13 Deficiency ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract ADAMTS13 is a plasma metalloproteinase that regulates platelet adhesion and aggregation through cleavage of von Willebrand factor (VWF) multimers. In humans, genetic or acquired deficiency in ADAMTS13 causes thrombotic thrombocytopenic purpura (TTP), a condition characterized by thrombocytopenia and hemolytic anemia with microvascular platelet thrombi. In this study, we report characterization of mice bearing a targeted disruption of the Adamts13 gene. ADAMTS13-deficient mice were born in the expected mendelian distribution; homozygous mice were viable and fertile. Hematologic and histologic analyses failed to detect any evidence of thrombocytopenia, hemolytic anemia, or microvascular thrombosis. However, unusually large VWF multimers were observed in plasma of homozygotes. Thrombus formation on immobilized collagen under flow was significantly elevated in homozygotes in comparison with wild-type mice. Thrombocytopenia was more severely induced in homozygotes than in wild-type mice after intravenous injection of a mixture of collagen and epinephrine. Thus, a complete lack of ADAMTS13 in mice was a prothrombotic state, but it alone was not sufficient to cause TTP-like symptoms. The phenotypic differences of ADAMTS13 deficiencies between humans and mice may reflect differences in hemostatic system functioning in these species. Alternatively, factors in addition to ADAMTS13 deficiency may be necessary for development of TTP.

Mutations in the von Willebrand Factor A1 Domain That Increase the Affinity for Collagen Have Different Effects on the Binding to Glycoprotein Ib.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3661-3661
Author(s):  
Miguel A. Cruz ◽  
Liza D. Morales
Keyword(s):  
Binding Site ◽  
Platelet Adhesion ◽  
Collagen Type ◽  
Type I ◽  
Fibrillar Collagen ◽  
Wild Type ◽  
Collagen Types ◽  
A1 Domain ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract The interaction of plasma von Willebrand factor (VWF) with collagen at the site of vascular injury plays a critical role in the initiation of thrombus formation under high shear stress. It does this by forming a bridge between the fibrils of collagen in the subendothelium and the platelet glycoprotein Ib-IX-V complex (GPIb). The A1 domain of VWF is the binding site for GPIb whereas the collagen-function of VWF is controlled by both A1 and A3 domains. The VWF-A3 domain is important to support binding to fibrils of collagen Types I and III while the A1 domain is involved in the binding to microfibrillar collagen Type VI. It is assumed that the interaction of VWF with fibrillar collagen (via the A3 domain) may regulate the expression of the GPIb-binding site in the A1 domain. However, there is no a definite data to substantiate that hypothesis. Our goal was to demonstrate that a direct interaction between the A1 domain and fibrillar collagen Types I or III exposes the GPIb binding site. Thus, we postulated that platelet GPIb is able to interact with isolated A1 domain that is bound to collagen. We have demonstrated that the VWF-A1 protein binds specifically to human placenta collagen Types I and III with a KD ~ 200 nM by using surface plasmon resonance (SPR). Using plasma-free blood, we have provided strong evidence that isolated VWF-A1 domain bound to either collagen Type I or III is able to support platelet adhesion under high flow conditions. This platelet interaction was effectively blocked with antibodies against either GPIb or A1 domain. These results clearly show the ability of the A1 domain to concurrently interact with both GPIb and collagen fibrils and they also suggest that the collagen-A1 binding may regulate the expression of the GPIb-binding site in the A1 domain. To test this hypothesis, we analyzed three residues that in a previous mutagenesis study they increased the binding of VWF to GPIb, reasoning that they may have an effect on the collagen binding activity as well. The three residues are located in the a7 helix (rear face) of the folded A1 domain and mutagenesis studies of other I(A)-domains have demonstrated that this helix plays a role in regulating the affinity of the ligand-binding. We introduced point mutations into the 3 residues and the recombinant mutant proteins were expressed in bacteria. The three mutants (R687E, D688R, and E689R) were purified as wild type and their structural integrity was confirmed with three conformation-specific antibodies. All the mutants bound to both collagens Type I or III with an affinity much higher than the wild type (WT) (KD~ 9 -1 nM). The mutants were assessed by their ability to mediate platelet adhesion to collagen, and their ability to inhibit both ristocetin-induced platelet agglutination and shear-induced platelet aggregation. Interestingly, in the three assays the R687E mutant had an activity higher than WT while the D688R had a markedly decrease activity. The mutant E689R had an activity similar to that of WT for the three assays. Together our data indicate that a direct association between the VWF-A1 domain and collagen fibrils influences the expression of GPIb binding function in VWF. Further, these data indicate that residue R687 located in the a7 helix plays a novel and important role in modulating the collagen/A1/GPIb binding.

Platelet Activation and Aggregation Induced by Recombinant von Willebrand Factors Reproducing Four Type 2B von Willebrand Disease Missense Mutations

1998 ◽  
Vol 79 (01) ◽  
pp. 211-216 ◽  
Author(s):  
Lysiane Hilbert ◽  
Claudine Mazurier ◽  
Christophe de Romeuf
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Von Willebrand Disease ◽  
Platelet Glycoprotein ◽  
Missense Mutations ◽  
Inhibit Platelet Aggregation ◽  
Wild Type ◽  
A1 Domain ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryType 2B of von Willebrand disease (vWD) refers to qualitative variants with increased affinity of von Willebrand factor (vWF) for platelet glycoprotein Ib (GPIb). All the mutations responsible for type 2B vWD have been located in the A1 domain of vWF. In this study, various recombinant von Willebrand factors (rvWF) reproducing four type 2B vWD missense mutations were compared to wild-type rvWF (WT-rvWF) for their spontaneous binding to platelets and their capacity to induce platelet activation and aggregation. Our data show that the multimeric pattern of each mutated rvWF is similar to that of WT-rvWF but the extent of spontaneous binding and the capacity to induce platelet activation and aggregation are more important for the R543Q and V553M mutations than for the L697V and A698V mutations. Both the binding of mutated rvWFs to platelets and platelet aggregation induced by type 2B rvWFs are inhibited by monoclonal anti-GPIb and anti-vWF antibodies, inhibitors of vWF binding to platelets in the presence of ristocetin, as well as by aurin tricarboxylic acid. On the other hand, EDTA and a monoclonal antibody directed against GPIIb/IIIa only inhibit platelet aggregation. Furthermore, the incubation of type 2B rvWFs with platelets, under stirring conditions, results in the decrease in high molecular weight vWF multimers in solution, the extent of which appears correlated with that of plasma vWF from type 2B vWD patients harboring the corresponding missense mutation. This study supports that the binding of different mutated type 2B vWFs onto platelet GPIb induces various degrees of platelet activation and aggregation and thus suggests that the phenotypic heterogeneity of type 2B vWD may be related to the nature and/or location of the causative point mutation.

Interventional Thermal Injury of the Arterial Wall: Unfolding of von Willebrand Factor and Its Increased Binding to Collagen after 55° C Heating

1996 ◽  
Vol 75 (03) ◽  
pp. 515-519 ◽  
Author(s):  
Mark J Post ◽  
Anke N de Graaf-Bos ◽  
George Posthuma ◽  
Philip G de Groot ◽  
Jan J Sixma ◽  
...  
Keyword(s):  
Von Willebrand Factor ◽  
Conformational Changes ◽  
Arterial Wall ◽  
Platelet Adhesion ◽  
Type I ◽  
Temperature Dependent ◽  
Collagen Types ◽  
Electron Microscopic ◽  
Von Willebrand ◽  
Willebrand Factor

Summary Purpose. Thermal angioplasty alters the thrombogenicity of the arterial wall. In previous studies, platelet adhesion was found to increase after heating human subendothelium to 55° C and decrease after heating to 90° C. In the present electron microscopic study, the mechanism of this temperature-dependent platelet adhesion to the heated arterial wall is elucidated by investigating temperature-dependent conformational changes of von Willebrand factor (vWF) and collagen types I and III and the binding of vWF to heated collagen. Methods. Purified vWF and/or collagen was applied to electron microscopic grids and heated by floating on a salt-solution of 37° C, 55° C or 90° C for 15 s. After incubation with a polyclonal antibody against vWF and incubation with protein A/gold, the grids were examined by electron microscopy. Results. At 37° C, vWF was coiled. At 55° C, vWF unfolded, whereas heating at 90° C caused a reduction in antigenicity. Collagen fibers heated to 37° C were 60.3 ± 3.1 nm wide. Heating to 55° C resulted in the unwinding of the fibers, increasing the width to 87.5 ± 8.2 nm (p < 0.01). Heating to 90° C resulted in denatured fibers with an enlarged width of 85.1 ± 6.1 nm (p < 0.05). Heating of collagen to 55° C resulted in an increased vWF binding as compared to collagen heated to 37° C or to 90° C. Incubation of collagen with vWF, prior to heating, resulted in a vWF binding after heating to 55° C that was similar to the 37° C binding and a decreased binding after 90° C. Conclusions. After 55° C heating, the von Willebrand factor molecule unfolds and collagen types I and III exhibit an increased adhesiveness for von Willebrand factor. Heating to 90° C denatures von Willebrand factor and collagen. The conformation changes of von Willebrand factor and its altered binding to collagen type I and III may explain the increased and decreased platelet adhesion to subendothelium after 55° C and 90° C heating, respectively.

The Genetic Defect of Type I von Willebrand Disease “Vicenza” Is Linked to the von Willebrand Factor Gene

1993 ◽  
Vol 69 (02) ◽  
pp. 173-176 ◽  
Author(s):  
Anna M Randi ◽  
Elisabetta Sacchi ◽  
Gian Carlo Castaman ◽  
Francesco Rodeghiero ◽  
Pier Mannuccio Mannucci
Keyword(s):  
Von Willebrand Factor ◽  
Autosomal Dominant ◽  
Genetic Defect ◽  
Von Willebrand Disease ◽  
Type I ◽  
Bleeding Tendency ◽  
Factor Gene ◽  
Low Levels ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryType I von Willebrand disease (vWD) Vicenza is a rare variant with autosomal dominant transmission, characterized by the presence of supranormal von Willebrand factor (vWF) multimers in plasma, similar to those normally found in endothelial cells and megakaryocytes. The patients have very low levels of plasma vWF contrasting with a mild bleeding tendency. The pathophysiology of this subtype is still unknown. The presence of supranormal multimers in the patients’ plasma could be due to a mutation in the vWF molecule which affects post-translational processing, or to a defect in the cells’ processing machinery, independent of the vWF molecule. In order to determne if type I vWD Vicenza is linked to the vWF gene, we studied six polymorphic systems identified within the vWF gene in two apparently unrelated families with type I vWD Vicenza. The results of this study indicate a linkage between vWF gene and the type I vWD Vicenza trait. This strongly suggests that type I vWD Vicenza is due to a mutation in one of the vWF alleles, which results in an abnormal vWF molecule that is processed to a lesser extent than normal vWF.

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