scholarly journals the Diagnostic Value of Plasma Level, Activity, Ratios between von Willebrand Factor and ADAMTS13 in Patients with Cerebral Infarction

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5003-5003
Author(s):  
Le Qu ◽  
Changgeng Ruan ◽  
Yiming Zhao
Keyword(s):  
Cerebral Infarction ◽  
Von Willebrand Factor ◽  
Plasma Levels ◽  
Conflicts Of Interest ◽  
Large Population ◽  
Medical Science ◽  
Diagnostic Value ◽  
Adamts13 Activity ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Introduction: Large population studies have revealed that higher levels of von Willebrand Factor (VWF) and lower levels of ADAMTS13 activity (ADAMTS13) are associated with a risk of thrombosis. In this study, we aimed to investigate the relationship among the plasma levels of VWF, ADAMTS13 and the occurrence of cerebral infarction in patients. Methods: In a case-control study of 94 cerebral infarction patients (CIP) and 103 age-matched healthy controls (Control), the plasma levels of VWF antigen (VWF:Ag ), VWF ristocetin cofactor activity (VWF:Rcof), and VWF collagen binding activity (VWF:CB) were measured using ELISA. ADAMTS13 activity (ADAMTS13) was measured with FREST-VWF73. The study was approved by the institutional Ethics Committee at the First Affiliated Hospital of Soochow University, China. All participants were given write informed consents. Results: The levels of VWF:Ag and VWF:Rcof in cerebral infraction patients (mean±SEM, 223±15% and 256±20%) were significantly higher compared with controls (117±8% and 128±9%). Levels of ADAMTS13 and ratios of VWF:CB/VWF:Ag, ADAMTS13/VWF:Ag and ADAMTS13/VWF:Rcof in patients (107±4%, 0.73±0.08, 0.76±0.07 and 0.86±0.10, respectively) were significantly lower compared with controls (125±3%, 1.37±0.08, 1.69±0.14 and 1.45±0.10, P<0.001, respectively). However, there was no difference between cerebral infarction patients and controls in the levels of VWF:CB, VWF:Rcof/VWF:Ag and ADAMTS13/VWF:CB (P>0.05). Unconditional logistic regression analysis demonstrated the highest quartile of VWF:Ag (odds ratio [OR] = 5.11, 95% confidence interval [CI], 1.49-17.50), VWF:Rcof (OR = 5.04, 95% CI, 1.62-15.66), and the lowest quartile of VWF:CB/VWF:Ag (OR = 5.91, 95% CI, 1.95-17.93), ADAMTS13/VWF:Ag (OR = 9.11, 95% CI, 2.49-33.33), ADAMTS13/VWF:Rcof (OR = 3.73, 95% CI, 1.39-10.03) were associated with cerebral infarction (P<0.01). The receiver operating characteristic (ROC) curves analysis demonstrated the cutoff values of VWF:Ag (137.88%), VWF:Rcof (135.88%) and VWF:CB/VWF:Ag (0.538), ADAMTS13/VWF:Ag (0.974), ADAMTS13/VWF:Rcof (0.946) showed a better diagnostic value of cerebral infarction (AUC > 0.7). Conclusions: High levels of VWF:Ag, VWF:Rcof and lower levels of VWF:CB/VWF:Ag, ADAMTS13/VWF:Ag, ADAMTS13/VWF:Rcof are relative risk factors. The threshold of VWF:Ag ( > 137.88%), VWF:Rcof ( > 135.88%), VWF:CB/VWF:Ag ( ≤ 0.538), ADAMTS13/VWF:Ag ( ≤ 0.974), ADAMTS13/VWF:Rcof ( ≤ 0.946) can be diagnostic indicators of cerebral infarction. Research funding: This work was supported by grants from National Scientific Foundation of China (NSFC) (81270593), Jiangsu Provincial Special Program of Medical Science (BL2012005) and Jiangsu Province’s Key Medical Center (ZX201102), China Disclosures No relevant conflicts of interest to declare.

ADAMTS13 In 4 Different VWF/VIII Concentrates and Its Impact on Therapy.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3677-3677
Author(s):  
Mirjeta Qorraj ◽  
Tanja Falter ◽  
Sarah Steinemann ◽  
Thomas Vigh ◽  
Inge Scharrer
Keyword(s):  
Von Willebrand Factor ◽  
Gel Electrophoresis ◽  
Conflicts Of Interest ◽  
Von Willebrand Disease ◽  
Relative Reduction ◽  
Hemorrhagic Diathesis ◽  
Adamts13 Activity ◽  
Kinetic Measurements ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Abstract 3677 Introduction: The hemostatic activity of von Willebrand Factor (VWF) is mainly controlled by the plasma metalloprotease ADAMTS13, which cleaves ultralarge VWF multimers. A qualitative or quantitative deficiency of VWF induces the most common hemorrhagic diathesis, the von Willebrand Disease (VWD). The current classification graduates the VWD in three major types. Depending on severity and the type of VWD the treatment with VWF/FVIII concentrates may by necessary. The commercially available VWF/FVIII concentrates differ in their multimer structure and furthermore also in their pharmacokinetics. We investigated commercial VWF concentrates with respect to their ADAMTS 13 activity and antigen levels with the newest available methods. Moreover, to detect a possible correlation, we analysed the VWF multimer structure of the concentrates. Methods: We analysed 4 human derived VWF/VIII-concentrates (over all 7charges) after reconstitution according to the manufacturer's instructions in different dilutions. Following methods were used: BCS Method according to Böhm detects the capacity of the concentrates for autoproteolysis. The VWF solutions were diluted with 5mol/l urea and then incubated for 14–16h at 37°C in low ionic TRIS buffer containing BaCl2 and different plasma samples: pool plasma; plasma from patients with TTP with neutralizing ADAMTS13 auto-antibodies; plasma from patients with TTP without auto-antibodies. The residual VWF:Ristocetin Cofactor (VWF:RCo) activity was subsequently measured using the BC von Willebrand Reagent from Dade Behring. ELISA Technozym®ADAMTS13 and Actifluor TM ADAMTS13 are based on the kinetic measurements of the activity with fluorescence resonance energy transfer (FRET). ADAMTS13 antigen was measured by use of the Technozym ELISA kit. SDS-Gel electrophoresis in 1% Agarose Gel was used to investigate the structure of VWF multimers. Results: The BCS Method according to Böhm is an indirect measurement for endogenous ADAMTS13 activity in the investigated concentrate. Important is the loss of the residual VWF:RCo in the concentrates in presence of TTP-plasma without antibodies and pool plasma compared to the residual VWF:RCo in presence of TTP-plasma with antibodies. All concentrates show some ADAMTS13 activity, however product 1 contains more ADAMTS13 than the other concentrates. The results of the two FRETS-assays correspond very well to the BCS-method results; in addition the assays detect directly the ADAMTS13 activity also in very low measurement range. In a dilution of 16U VWF per ml concentrate the ADAMTS13 activity in product 1 with 4.3% was the highest compared to product 2: 3.2%, product 3: 2.6% and product 4: 2%. The great variability of the test results in higher concentrations may be caused by interferences between some constituents of the concentrates and the analysis. In the same sample set and dilution the ADAMTS13 antigen values correlate very well with ADAMTS13 activity values. The SDS gel electrophoresis reveals the different VWF structure of product1; it has less large and ultralarge multimers. There could be a correlation to the relatively higher ADAMTS13 activity and antigen level. Conclusion: All the investigated VWF/VIII concentrates contain some ADAMTS13 activity and antigen. This was found especially by FRETs assay due to the high sensitivity. Because of the correlation between ADAMTS13 activity and modified VWF multimer structure we like to conclude that ADAMTS13 has influence on stability and therefore also on quality of the concentrates. This might have a therapeutic consequence especially for VWD type 2A. Type 2A is characterized by a relative reduction of intermediate and large VWF multimer. The multimeric abnormalities are commonly the result of in vivo proteolytic degradation of the von Willebrand factor caused by ADAMTS13. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Active and Ultra-Large Von Willebrand Factor Is Significantly Elevated in Patients with Immune Thrombotic Thrombocytopenic Purpura

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 523-523
Author(s):  
Wenjing Cao ◽  
Alicia Veninga ◽  
Elizabeth M. Staley ◽  
Adam Miszta ◽  
Nicole Kocher ◽  
...  
Keyword(s):  
Thrombotic Thrombocytopenic Purpura ◽  
Von Willebrand Factor ◽  
Plasma Levels ◽  
Acute Episode ◽  
Healthy Controls ◽  
Plasma Samples ◽  
Adamts13 Activity ◽  
Thrombocytopenic Purpura ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Background: Immune thrombotic thrombocytopenic purpura (iTTP), a potentially fatal hematological emergency, is primarily caused by acquired deficiency of ADAMTS13 activity due to autoantibodies. Immunoglobulin G (IgG)-type autoantibodies bind ADAMTS13 and inhibit its ability to cleave endothelium-derived ultra large von Willebrand factor (ULVWF). However, it remains poorly understood whether plasma VWF status can be used as a disease marker for diagnosis and monitoring therapy in patients with acute iTTP. Objective: To address this question, we determined plasma levels of VWF antigen (VWF:Ag), collagen-binding activity (VWF:CB), active forms of VWF (VWF:Ac), and VWF multimers in iTTP patients during acute episode and in early remission. Patients and Methods: From the Alabama registry, we identified 69 unique patients with a confirmed diagnosis of iTTP in whom plasma ADAMTS13 activity was <10 U/dL with positive inhibitors and elevated anti-ADAMTS13 IgGs. Of 69 patients, 21 had longitudinal plasma samples collected. Plasma samples from 56 healthy individuals, who did not have a hematological disease, cancer, and infection, were recruited as controls. Plasma levels of VWF:Ag, VWF:CB, and VWF:Ac were determined by an ELISA-based assay. Plasma VWF multimer distribution was assessed by an in-gel Western blotting assay following electrophoresis on a 1% SDS-agarose gel. Results: The mean age for our cohort iTTP patients was 43.9 ± 13.4 years. Twenty-six patients were male and 43 were female with male to female ratio of 1 to 1.7. Fifty-three patients were African American descents, 14 Caucasians, 1 Hispanic, and 1 unknown race. Plasma levels of VWF:Ag in acute iTTP patients were 289.4 ± 17.7%, significantly increased compared with those in the healthy controls (144.9 ± 7.6%) (p<0.0001); plasma levels of VWF:CB in these patients were 241 ± 17.9%, also significantly elevated compared with those in the healthy controls (149.9 ± 12.01%) (p=0.0001); additionally, plasma levels of VWF:Ac (304.6 ± 23.2%), assessed by its ability to bind anti-VWF-A1 nanobody, were more dramatically elevated compared with those in the controls (101.6 ± 5.9%) (p<0.0001). More interestingly, while the ratios of VWF:CB to VWF:Ag in patients with acute iTTP (0.8 ± 0.04) were lower than those in the healthy controls (1.0 ± 0.05) (p=0.0036), the ratios of VWF:Ac to VWF:Ag were significantly higher in patients with acute episode (1.2 ± 0.1) than those in the controls (0.8 ± 0.05) (p=0.0003). Furthermore, there was no statistically significant difference in the patient plasma levels of VWF:Ag (p=0.69) and VWF:CB (p=0.08) during acute episode and during early remission. However, the plasma levels of VWF:Ac in patients with acute disease were significantly higher than those in the early remission (p=0.002). Surprisingly, 90% (36/40) of out iTTP patients during acute episode showed the presence of ULVWF in their plasma using in-gel Western blotting, which allows the ULVWF to be detected without the transfer step to avoid any potential loss of larger VWF multimers during protein transfer. These ULVWF multimers disappeared in 3/4 iTTP patients in remission when ADAMTS13 activity recovered. In 28 healthy control samples, only one showed ULVWF. Conclusion: Our results demonstrate, for the first time in a large cohort, that active forms of VWF and ultra large VWF multimers are present in iTTP patient's plasma during the acute period, which is reduced or disappears during the early remission. Therefore, measuring active forms of VWF and ultra large VWF multimers may aid in diagnosis of iTTP and help monitoring of disease processes following therapy. Our ongoing study is to determine whether these biomarkers can be used to predict responses to treatment and long-term outcome. Disclosures Zheng: Alexion: Research Funding, Speakers Bureau.

Normal Cleavage of Von Willebrand Factor by ADAMTS13 In the Absence of Factor VIII In Patients with Severe Hemophilia A

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2114-2114
Author(s):  
Junmei Chen ◽  
Dominic W. Chung ◽  
Jennie Le ◽  
Barbara A. Konkle ◽  
José A. López
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Hemophilia A ◽  
Conflicts Of Interest ◽  
Ex Vivo ◽  
Adamts13 Activity ◽  
Severe Hemophilia ◽  
Post Infusion ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Abstract 2114 The size of von Willebrand factor (VWF), a carrier protein for factor VIII (FVIII), is regulated by plasma metalloprotease ADAMTS13 proteolytic activity. Recently studies by Cao et al. (PNAS, 2008; 105: 7416–7421) found that under shear stress, exogenous FVIII enhanced ADAMTS13 cleavage of VWF, especially the high molecular weight multimers, in a system using purified proteins. Based on this result, the authors suggested that in the absence of FVIII, such as in patients with severe hemophilia A, VWF will have ultra-large multimers due to defects in ADAMTS13 proteolytic process, which can be corrected by infusion of FVIII. Here, we assessed VWF multimers, antigen, and ADAMTS13 activity in citrated plasma from seven patients with severe hemophilia A. The FVIII levels in six patients were less than 1% and in one was 4%. Plasma from two patients was available both pre- and post-FVIII replacement therapy (recombinant FVIII). All patients displayed VWF multimer patterns similar to those in pooled normal plasma (PNP), and the two patients receiving FVIII infusions displayed no change in VWF multimer size or pattern between their pre- and post-infusion samples. In all patients, the VWF antigen level (0.32–0.76) was below the PNP value (designated as 1), and all had increased ADAMTS13 activity (measured by the ability of plasma to cleave a small A2 peptide substrate) (1.09–1.79, PNP designated as 1), yielding an increased ratio of ADAMTS13 activity to VWF antigen in these patients (1.4–5.2 compared to PNP). We also examined cleavage of endogenous VWF by ADAMTS13 in the plasmas of the two patients studied pre- and post-infusion, yielding different FVIII levels. In this assay, we first diluted plasma 10-fold with a buffer containing 10 mM HEPES, 6.5 mM BaCl2, and 1.5 M urea, incubated at 37°C, and ADAMTS13 cleavage was stopped at different time points with EDTA. VWF multimer patterns were examined on a 1.5% agarose gel. We found that ADAMTS13 cleaved VWF efficiently in patient plasma deficient in FVIII, similar to that in PNP. The extent of cleavage was correlated with the ratio of ADAMTS13 activity to VWF antigen, rather than with the FVIII levels. In conclusion, patients with severe hemophilia A appear to have normal ADAMTS13 processing of VWF multimers in vivo and ex vivo. Further studies of the effect of FVIII and VWF levels on ADAMTS13 cleavage of VWF and clinical correlation are needed. Disclosures: No relevant conflicts of interest to declare.

Pregnancies Complicated By Severe Preeclampsia Exhibit Perturbed Von Willebrand Factor (VWF)-Associated Parameters

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3525-3525 ◽  
Author(s):  
Danielle N. Drury-Stewart ◽  
Kara K. Hoppe ◽  
Kerry W. Lannert ◽  
Dominic W. Chung ◽  
Hilary S. Gammill ◽  
...  
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Conflicts Of Interest ◽  
Statistical Significance ◽  
Severe Preeclampsia ◽  
Adamts13 Activity ◽  
Healthy Pregnancy ◽  
Elevated Liver Enzymes ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Background Elevated levels of the critical coagulation glycoproteins von Willebrand Factor (VWF) and Factor VIII (FVIII) are associated with thrombovascular disease. VWF and FVIII levels are elevated in healthy pregnancy compared with the nonpregnant state, and further increases have been reported in pregnancies complicated by preeclampsia, a potentially life-threatening vascular disorder of pregnancy. Aside from the quantitative increases in VWF and FVIII, data is inconsistent or little is known regarding other key VWF-associated parameters in pregnancies complicated by preeclampsia. Hypothesis We hypothesized that pregnancies complicated by severe preeclampsia would exhibit distinct changes in VWF-associated parameters compared to healthy pregnancies. Methods We assayed VWF-associated parameters in subjects newly diagnosed with severe preeclampsia, based on standard blood pressure (BP) and proteinuria criteria. Severe preeclampsia was defined on the basis of one or more of the following: severe hypertension (systolic BP >160 or diastolic BP >110) for at least 6 hours, seizures (eclampsia), hemolysis, elevated liver enzymes, thrombocytopenia, pulmonary edema, renal dysfunction, or fetal growth restriction. We compared results from preeclamptic pregnancies to gestational age-matched healthy pregnant controls. VWF antigen (VWF:Ag), VWF propeptide (VWF:pp), and FVIII antigen (FVIII:Ag) levels were determined by ELISA. Factor VIII activity (FVIII:C) was measured using a one-stage assay and ADAMTS13 activity was determined via cleavage of an enzyme-linked peptide substrate. Results Forty-two patients with severe preeclampsia and a cohort of thirty-nine controls of similar gestational ages met study inclusion criteria (Table 1). The results of the coagulation assays are presented in Table 2. In summary, preeclamptic pregnancies were notable for statistically significant (p<0.001) increases in VWF:Ag (1.4-fold), VWF:pp (1.5-fold), and FVIII:Ag (1.2-fold) compared to controls. FVIII:Ag and FVIII:C were similar, and FVIII:Ag was used in the calculation of the VWF:FVIII ratio. The VWF:FVIII ratio trended higher in preeclamptic pregnancies, but this did not reach statistical significance (p=0.058). The mean ADAMTS13 activity in patients with preeclampsia was also significantly lower (0.76-fold, p<0.001) than healthy pregnancy controls. Conclusions In pregnancies complicated by severe preeclampsia, VWF:Ag and FVIII were significantly elevated, as expected. VWF:FVIII ratios also tended to be higher in preeclamptic pregnancies compared to controls. Interestingly, activity of the VWF-cleaving protease ADAMTS13 was modestly but significantly decreased in preeclampsia pregnancies, a finding for which there is conflicted literature. Three subjects with thrombocytopenia had ADAMTS13 activities >85%, indicating the thrombocytopenia was unrelated to a relative ADAMTS13 deficiency. These findings support the hypothesis that pregnancies complicated by severe preeclampsia exhibit VWF biology that is distinct from healthy pregnancy. We speculate that preeclampsia-associated VWF differences could play a direct role in the vascular pathogenesis and end-organ damage of preeclampsia. These data offer new clues towards better understanding of the mechanisms of severe preeclampsia and represent promising future targets for diagnosis and/or treatment. Disclosures: No relevant conflicts of interest to declare.

Oxidation of Key Methionine Residues in the N-Terminal Domains of ADAMTS13 Correlates with Loss of Its Proteolytic Activity,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3276-3276
Author(s):  
Yi Wang ◽  
Junmei Chen ◽  
Minhua Ling ◽  
José A. López ◽  
Dominic W Chung ◽  
...  
Keyword(s):  
Proteolytic Activity ◽  
Von Willebrand Factor ◽  
Hypochlorous Acid ◽  
Conflicts Of Interest ◽  
Human Neutrophils ◽  
Adamts13 Activity ◽  
Oxidative Inactivation ◽  
Rich Domain ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Abstract 3276 Deficiency of ADAMTS13 is associated with a life-threatening thrombotic microangiopathic disease—thrombotic thrombocytopenic purpura (TTP), which is characterized by the accumulation of aggregates of hyper-reactive ultra-large von Willebrand factor (VWF) and platelets in the microvasculature. In addition to TTP, systemic inflammatory syndromes such as acute systemic inflammation caused by endotoxin, acute pancreatitis, and severe sepsis demonstrate reduced ADAMTS13 activity. The cause of low ADAMTS13 activity is not known. One possible cause is oxidative inactivation of the enzyme by hypochlorous acid (HOCl), a potent oxidant released from activated neutrophils that is known to damage proteins. In this study, we exposed ADAMTS13 to HOCl produced by a myeloperoxidase (MPO)-H2O2-Cl− system and measured the proteolytic activity of oxidized ADAMTS13 using a small VWF A2 peptide and plasma VWFas substrates. ADAMTS13 activity decreased as a function of oxidant concentration. Treatment with 25 nM MPO plus 50 μM H2O2 reduced ADAMTS13 activity by over 85%. Such concentrations of MPO and H2O2 are routinely found in vivo at sites of inflammation. ADAMTS13 contains a series of structural domains: a metalloprotease domain (M), a disintegrin-like domain (D), a thrombospondin type 1 repeat (TSP1,T), a Cys-rich domain (C), a spacer domain (S), 7 additional TSP1 repeats, and 2 CUB domains. The MDTCS domains are essential for its proteolytic activity, while the C-terminal TSP1 and CUB domains may act as modulators. The MDTCS region contains 10 Met residues, Met being the amino acid residue most vulnerable to oxidation by HOCl. Using mass spectrometry, we identified 7 Met-containing peptides in this region after proteolytic digestion of ADAMTS13. Three of these 7 methionines are highly sensitive to oxidation by HOCl, including M249, M331 and M496. M249 is situated in the ”Met-turn” at the catalytic center of the metalloprotease domain. M331 and M496 are located in the disintegrin-like domain and the Cys-rich domain, respectively. The extent of oxidation of these Met residues was proportional to the HOCl concentration, and strongly correlated with loss of enzymatic activity. The same three Met residues were also oxidized after exposure of ADAMTS13 to activated human neutrophils. These observations suggest an oxidative mechanism for ADAMTS13 inactivation in systemic inflammatory syndromes and that the oxidation-sensitive Mets may serve as biomarkers for this effect. Coupled with our earlier observation that HOCl oxidation of von Willebrand factor enhances its adhesive function and renders it resistant to cleavage by ADAMTS13, these findings indicate that pathological situations during which neutrophils are activated produce extremely prothrombotic conditions, perhaps explaining why many inflammatory syndromes are associated with thrombosis in small and large blood vessels. Disclosures: No relevant conflicts of interest to declare.

Plasma levels of platelet aggregating factor/von Willebrand factor in various species

1977 ◽  
Vol 11 (3) ◽  
pp. 345-355 ◽  
Author(s):  
K.M. Brinkhous ◽  
B.D. Thomas ◽  
S.A. Ibrahim ◽  
M.S. Read
Keyword(s):  
Von Willebrand Factor ◽  
Plasma Levels ◽  
Von Willebrand ◽  
Willebrand Factor
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