More homologies among the vertebrate plasma proteins

1985 ◽  
Vol 5 (10-11) ◽  
pp. 877-884 ◽  
Author(s):  
R. F. Doolittle
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Unknown Function ◽  
Plasma Proteins ◽  
Common Ancestry ◽  
High Molecular Weight Kininogen ◽  
A Minor ◽  
Von Willebrand ◽  
Willebrand Factor

Many of the proteins of vertebrate blood plasma share common ancestry. As more sequences are reported, the network of relationships continues to expand in unexpected directions. Computer analysis now reveals that a minor plasma protein of unknown function, gamma-trace protein, is related to the kininogen family. Some other possible relationships have been uncovered also, including a resemblance between the histidine-rich hinge regions of high molecular weight kininogen and hemopexin and between Factor VIII and Von Willebrand Factor.

scholarly journals Immunologic studies of native and modified human factor VIII/von Willebrand factor

Blood ◽  
1979 ◽  
Vol 54 (2) ◽  
pp. 310-321 ◽  
Author(s):  
ME Switzer ◽  
PA McKee
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Proteolytic Enzymes ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Procoagulant Activity ◽  
Human Factor Viii ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Factor VIII/von Willebrand factor (FVIII/vWF) is a glycoprotein with a molecular weight greater than one-million daltons. Two activities are associated with this large molecule: FVIII procoagulant activity and vWF activity. Incubation of FVIII/vWF with proteolytic enzymes causes rapid inactivation of the FVIII procoagulant activity but has little effect on the vWF activity or antigenicity. In an attempt to gain insight into the structural features required for these two activities, antisera were raised in rabbits to normal, thrombin-inactivated, and plasmin-inactivated FVIII/vWF. All of these proteolytically modified forms of FVIII/vWF cross-reacted with each of the rabbit antisera; each blocked the ability of a human inhibitor to inactivate native active FVIII/vWF. Each of the antisera was a potent inhibitor of vWF activity and inactivated vWF activity at the same titer. The antisera were much less potent inhibitors of FVIII activity than of vWF activity. Antibodies to thrombin-inactivated FVIII/vWF or normal FVIII/vWF had about the same ability to inactivate FVIII procoagulant activity. Surprisingly, those to plasmin-inactivated FVIII/vWF still retained about 50% of this inhibitory capacity. A comparison of the three types of antigens by polyacrylamide gel electrophoresis in sodium dodecyl sulfate-6 M urea demonstrated that the structure of thrombin- inactivated FVIII/vWF was indistinguishable from that of normal FVIII/vWF, while plasmin-inactivated FVII/vWF was completely cleaved to lower molecular weight fragments. Some of the reported variations in the ability of rabbit antibodies to inhibit procoagulant activity may be due to partial degradation of the starting antigen. The retention by FVIII/vWF protein of its immunologic properties even after extensive proteolytic degradation suggests that under nondenaturing conditions, the conformation of the native and degraded molecules are very similar.

scholarly journals Immunologic studies of native and modified human factor VIII/von Willebrand factor

Blood ◽  
1979 ◽  
Vol 54 (2) ◽  
pp. 310-321
Author(s):  
ME Switzer ◽  
PA McKee
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Proteolytic Enzymes ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Procoagulant Activity ◽  
Human Factor Viii ◽  
Von Willebrand ◽  
Willebrand Factor

Factor VIII/von Willebrand factor (FVIII/vWF) is a glycoprotein with a molecular weight greater than one-million daltons. Two activities are associated with this large molecule: FVIII procoagulant activity and vWF activity. Incubation of FVIII/vWF with proteolytic enzymes causes rapid inactivation of the FVIII procoagulant activity but has little effect on the vWF activity or antigenicity. In an attempt to gain insight into the structural features required for these two activities, antisera were raised in rabbits to normal, thrombin-inactivated, and plasmin-inactivated FVIII/vWF. All of these proteolytically modified forms of FVIII/vWF cross-reacted with each of the rabbit antisera; each blocked the ability of a human inhibitor to inactivate native active FVIII/vWF. Each of the antisera was a potent inhibitor of vWF activity and inactivated vWF activity at the same titer. The antisera were much less potent inhibitors of FVIII activity than of vWF activity. Antibodies to thrombin-inactivated FVIII/vWF or normal FVIII/vWF had about the same ability to inactivate FVIII procoagulant activity. Surprisingly, those to plasmin-inactivated FVIII/vWF still retained about 50% of this inhibitory capacity. A comparison of the three types of antigens by polyacrylamide gel electrophoresis in sodium dodecyl sulfate-6 M urea demonstrated that the structure of thrombin- inactivated FVIII/vWF was indistinguishable from that of normal FVIII/vWF, while plasmin-inactivated FVII/vWF was completely cleaved to lower molecular weight fragments. Some of the reported variations in the ability of rabbit antibodies to inhibit procoagulant activity may be due to partial degradation of the starting antigen. The retention by FVIII/vWF protein of its immunologic properties even after extensive proteolytic degradation suggests that under nondenaturing conditions, the conformation of the native and degraded molecules are very similar.

scholarly journals Heterogeneity of the Factor VIII/Von Willebrand Factor Protein in Von Willebrand’s Disease

1977 ◽  
Author(s):  
H. R. Gralnick ◽  
Y. Sultan ◽  
B. S. Coller
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Apparent Molecular Weight ◽  
Total Carbohydrate ◽  
Von Willebrand's Disease ◽  
Von Willebrand’S Disease ◽  
Parallel Reduction ◽  
Von Willebrand ◽  
Willebrand Factor

The recent advent of techniques to purify the factor VIII/von Willebrand factor (f.VIII/vWf) protein from plasma to quantitate the f.VIII/vWf protein and to measure the vWf (plasma ristocetin cofactor) have greatly added to our understanding of von Willebrand’s disease (vWd). The initial studies of antigen, procoagulant and vWf levels revealed a parallel reduction in all three activities in vWd suggesting a quantitative deficiency of the f. VIII/vWf protein and its biologic activities.Recent studies, however, have suggested three major forms of vWd. The first group of patients have a quantitative defect with a parallel reduction of the f.VIII/vWf protein and of the antigen, vWf and procoagulant activities. The second group of patients appear to have a qualitative defect of the f.VIII/vWf protein. These individuals have normal levels of the antigen and procoagulant activity; however, the vWf activity is reduced or absent. The third group of patients have a combination of a qualitative and quantitative deficiency. These variants resemble both previous groups in that there are reduced levels of the antigen, procoagulant and vWf activities but usually greater reduction of the vWf activity.Three major defects of the f.VIII/vWf protein have been recognized in vWd: 1) decreased plasma concentration of the f. VIII/vWf protein, 2) the apparent molecular weight of the protein is reduced and/or the largest molecular weight polymers are absent, and 3) there is a partial or total carbohydrate deficiency. For the f.VIII/vWf protein to express vWf activity, it must be of a minimal molecular size and have a specific carbohydrate content and/or sequence.

scholarly journals Adhesion of platelets to human artery subendothelium: effect of factor VIII-von Willebrand factor of various multimeric composition

Blood ◽  
1984 ◽  
Vol 63 (1) ◽  
pp. 128-139 ◽  
Author(s):  
JJ Sixma ◽  
KS Sakariassen ◽  
NH Beeser-Visser ◽  
M Ottenhof-Rovers ◽  
PA Bolhuis
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Platelet Adhesion ◽  
Bleeding Time ◽  
Low Molecular Weight ◽  
Perfusion Chamber ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Adhesion Factor

Abstract The relationship between the multimeric size of factor VIII-von Willebrand factor (FVIII-vWF) and the support of platelet adhesion to subendothelium was studied in an annular perfusion chamber, employing human renal and umbilical arteries. Commercial factor VIII concentrates containing multimers of low molecular weight that had been shown not to correct the bleeding time upon infusion into patients with von Willebrand's disease did not support platelet adhesion in the perfusion chamber. Cryoprecipitate and two experimental FVIII-vWF concentrates containing multimers of high molecular weight supported platelet adhesion. Factor VIII-vWF purified from cryoprecipitate was subdivided into three fractions of different molecular weights (6.0–14.0, 4.0–9.0, and 3.0–7.5 X 10(6) dalton). These fractions appeared to bind equally well and to be equally effective in supporting platelet adhesion. Factor VIII-vWF with multimers of low molecular weight (0.5–1.5 X 10(6) dalton) were prepared by partial reduction. Binding of FVIII-vWF to subendothelium was not impaired, and the support of platelet adhesion appeared to be more resistant to the effect of reduction than the ristocetin cofactor activity. At high shear rate (2,500 sec-1), increased platelet adhesion was observed with partially reduced FVIII- vWF. These data indicate that the ability of FVIII-vWF preparations to correct the bleeding time is reflected in enhanced platelet adhesion to subendothelium in a perfusion chamber. These data also emphasize that multimeric size is not the only factor determining whether FVIII-vWF will support platelet adhesion.

Current Knowledge of the AHF-Like Antigen

1975 ◽  
Author(s):  
Dominique Meyer
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Hemophilia A ◽  
Factor Activity ◽  
Factor Viii Activity ◽  
Von Willebrand's Disease ◽  
Von Willebrand’S Disease ◽  
Von Willebrand ◽  
Willebrand Factor

Factor VIII and von Willebrand Factor activities are associated with a high molecular weight protein which can be isolated from plasma and may be studied by immunological methods. Homologous antibodies to Factor VIII are directed towards the active site of the Factor VIII molecule; they do not neutralize Willebrand Factor activity and do not precipitate with normal plasma. The use of such antibodies has allowed the distinction between Hemophilia A+ and A-. Specific precipitation of Factor VIII antibodies using polyethylene glycol will be reported, allowing typing of heavy and light chains of purified antibodies. Heterologous antisera prepared in rabbits against purified human Factor VIII complex neutralize Factor VIII and von Willebrand Factor activities and precipitate with AHF-like antigen. Estimation of this antigen in plasma has allowed (1) the differenciation of the molecular abnormalities in Hemophilia A and classical von Willebrand’s disease; (2) the comparison between normal and Hemophilic AHF-like antigen; (3) the detection of carriers of Hemophilia A; (4) the study of variants of von Willebrand’s disease; (5) the demonstration of this antigen in platelets and in endothelial cells. Factor VIII activity and AHF-like antigen are probably separate entities, circulating as a complex in normal plasma, as suggested by the following experiments: transfusion studies in von Willebrand’s disease; immuno-adsorption studies; comparison of Factor VIII complex in cryoprecipitate and supernatant; and dissociation in high salt buffer, demonstrating that Factor VIII includes two biologically linked but distinct fragments, of high (HMW) and low (LMW) molecular weight. The non functional HMW subunit, controlled by an autosomal locus, is identified by the presence of AHF-like antigen and Willebrand Factor activity. The LMW subunit, product of an X-chromosome locus, does not contain AHF-like antigen, but it carries Factor VIII activity, as demonstrated by the following facts: inactivation by both human and rabbit antibodies to Factor VIII; transient activation by thrombin; obtention of antisera which specifically inactivate Factor VIII.

scholarly journals Comparison of inhibitory and binding characteristics of an antibody causing acquired von Willebrand syndrome: an assay for von Willebrand factor binding by antibody

Blood ◽  
1985 ◽  
Vol 66 (3) ◽  
pp. 562-569 ◽  
Author(s):  
WA Fricke ◽  
KM Brinkhous ◽  
JB Garris ◽  
HR Roberts
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
High Molecular Weight ◽  
Von Willebrand Factor ◽  
Plasma Levels ◽  
Binding Capacity ◽  
Anamnestic Response ◽  
Von Willebrand's Disease ◽  
Von Willebrand ◽  
Willebrand Factor

An acquired inhibitor of von Willebrand factor (vWF) activity occurring in a patient with benign gammopathy and von Willebrand syndrome (vWS) has been partially characterized. The inhibitor-induced syndrome resulted in low to undetectable plasma levels of vWF/ristocetin, vWF/botrocetin, FVIIIR:Ag, and FVIII:C with a normal to slightly prolonged bleeding time. Platelet vWF was normal. Intensive and continuous infusion of a heat-treated factor VIII concentrate (Hemofil- T, Hyland, Glendale, Calif) elevated the FVIII:C plasma levels to about 100%, with an increase in FVIIIR:Ag levels to about 340% and vWF/ristocetin levels to about 40%, much lower than expected based on the dose of Hemofil-T and its content of vWF and FVIII:C activities. The inhibitor bound to staphylococcal protein A (SpA) with high affinity, indicating an IgG antibody (Ab). An assay for the vWF-binding capacity was developed on the basis of absorption of the Ab from serially diluted plasma by SpA and removal of vWF and FVIII:C activities from normal plasma by the SpA-Ab complex. The Ab-binding site was on the vWF component of the factor VIII complex. The Ab was unable to bind isolated FVIII:C. The combined use of the new vWF- binding assay and a battery of tests for inhibition of vWF-dependent platelet aggregation with ristocetin (which detects high molecular weight vWF), with botrocetin (which detects high and low molecular weight vWF), and with platelet-aggregating factor (which detects high molecular weight vWF) provided a means of analysis of Ab effect on in vitro vWF function. Using these tests, a comparison was made of the effects of the vWS Ab with those of an Ab inhibitor occurring in homozygous von Willebrand's disease. The Ab of the vWS patient had weak inhibitory action on vWF/ristocetin without having an effect on vWF/botrocetin and platelet-aggregating factor, a high titer vWF- binding capacity, and no anamnestic response following concentrate therapy. These findings contrasted with those of the Ab occurring in inhibitor von Willebrand's disease in which vWF inhibitor and binding values were similar, with a strong anamnestic response. The findings indicate that the vWS Ab binds to an epitope on the molecular vWF in such a way that causes only limited inhibition of vWF/ristocetin function and no inhibition of vWF/botrocetin function, suggesting that these two functional domains are at separate sites.

A Comparative Multi-laboratory Assessment of Three Factor VIII/von Willebrand Factor Concentrates

2002 ◽  
Vol 87 (03) ◽  
pp. 466-476 ◽  
Author(s):  
Melinda Bukuya ◽  
Teresa Martinelli ◽  
Joanne Tzouroutis ◽  
Elizabeth Duncan ◽  
Katie Welldon ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Study Data ◽  
Low Molecular Weight ◽  
Laboratory Assessment ◽  
Factor Concentrate ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Three Factor

SummaryFive expert laboratories have participated in a cross-laboratory study to co-evaluate and compare three commercial Factor VIII/von Willebrand factor (VWF) concentrates. A total of nine factor concentrate lots were evaluated, comprising AHF (High Purity) (AHF HP; X3), Biostate® (X3) and Humate/Haemate (X3). All laboratories blind tested for FVIII:C, VWF:Ag and VWF:CB, four tested for VWF:RCo, and one performed VWF:Multimers. The study yielded inter-laboratory CVs for VWF:Ag and FVIII:C around 10–15%, and for VWF:CB and VWF:RCo around 20%, significantly lower than those of previous multi-laboratory surveys. All three lots of AHF HP contained in the vicinity of 25 U/ml FVIII:C, around 60–75 U/ml of VWF:Ag, but only 30–45 U/ml of VWF:CB and 40–50 U/ml of VWF:RCo (thus, CB/Ag ratio around 0.5–0.6 and RCo/Ag ratio around 0.6–0.7). Study determined that FVIII:C and VWF:RCo levels were similar to manufacturer assigned levels. Some loss of the high molecular weight (HMW) multimers was observed, together with an intense low molecular weight (LMW) VWF band consistent with some reduction or proteolysis of HMW VWF. All three lots of Humate/Haemate contained in the vicinity of 23–32 U/ml of FVIII:C, 70–105 U/ml of VWF:Ag, 50–90 U/ml of VWF:CB and VWF:RCo (i. e. CB/Ag ratio around 0.6–0.9 and RCo/Ag ratio around 0.6–1.1). Study-determined FVIII:C and VWF:RCo levels were similar to manufacturer-assigned levels. The LMW multimer band seen with AHF HP was also observed with Humate/Haemate. All three lots of Biostate contained in the vicinity of 40–55 U/ml of FVIII:C, 105–170 U/ml of VWF:Ag, 90–150 U/ml of VWF:CB, and 90–135 U/ml of VWF:RCo (i. e. CB/Ag and RCo/Ag ratios around 0.7–1.0). Study-determined FVIII:C levels were similar to manufacturer-assigned levels. The LMW multimer band seen with AHF HP was not observed with Biostate. The defined pattern of increasing CB/ Ag from AHF HP to Humate/Haemate and Biostate was consistently observed in study data from each of the five laboratories. In conclusion, study findings indicate some differences in the retention of functional/ HMW VWF between factor concentrates, and this is expected to have significant implications in terms of clinical efficacy for therapy in VWD.

scholarly journals Comparison of inhibitory and binding characteristics of an antibody causing acquired von Willebrand syndrome: an assay for von Willebrand factor binding by antibody

Blood ◽  
1985 ◽  
Vol 66 (3) ◽  
pp. 562-569 ◽  
Author(s):  
WA Fricke ◽  
KM Brinkhous ◽  
JB Garris ◽  
HR Roberts
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
High Molecular Weight ◽  
Von Willebrand Factor ◽  
Plasma Levels ◽  
Binding Capacity ◽  
Anamnestic Response ◽  
Von Willebrand's Disease ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract An acquired inhibitor of von Willebrand factor (vWF) activity occurring in a patient with benign gammopathy and von Willebrand syndrome (vWS) has been partially characterized. The inhibitor-induced syndrome resulted in low to undetectable plasma levels of vWF/ristocetin, vWF/botrocetin, FVIIIR:Ag, and FVIII:C with a normal to slightly prolonged bleeding time. Platelet vWF was normal. Intensive and continuous infusion of a heat-treated factor VIII concentrate (Hemofil- T, Hyland, Glendale, Calif) elevated the FVIII:C plasma levels to about 100%, with an increase in FVIIIR:Ag levels to about 340% and vWF/ristocetin levels to about 40%, much lower than expected based on the dose of Hemofil-T and its content of vWF and FVIII:C activities. The inhibitor bound to staphylococcal protein A (SpA) with high affinity, indicating an IgG antibody (Ab). An assay for the vWF-binding capacity was developed on the basis of absorption of the Ab from serially diluted plasma by SpA and removal of vWF and FVIII:C activities from normal plasma by the SpA-Ab complex. The Ab-binding site was on the vWF component of the factor VIII complex. The Ab was unable to bind isolated FVIII:C. The combined use of the new vWF- binding assay and a battery of tests for inhibition of vWF-dependent platelet aggregation with ristocetin (which detects high molecular weight vWF), with botrocetin (which detects high and low molecular weight vWF), and with platelet-aggregating factor (which detects high molecular weight vWF) provided a means of analysis of Ab effect on in vitro vWF function. Using these tests, a comparison was made of the effects of the vWS Ab with those of an Ab inhibitor occurring in homozygous von Willebrand's disease. The Ab of the vWS patient had weak inhibitory action on vWF/ristocetin without having an effect on vWF/botrocetin and platelet-aggregating factor, a high titer vWF- binding capacity, and no anamnestic response following concentrate therapy. These findings contrasted with those of the Ab occurring in inhibitor von Willebrand's disease in which vWF inhibitor and binding values were similar, with a strong anamnestic response. The findings indicate that the vWS Ab binds to an epitope on the molecular vWF in such a way that causes only limited inhibition of vWF/ristocetin function and no inhibition of vWF/botrocetin function, suggesting that these two functional domains are at separate sites.

Two novel type 2N von Willebrand disease–causing mutations that result in defective factor VIII binding, multimerization, and secretion of von Willebrand factor

Blood ◽  
2000 ◽  
Vol 95 (6) ◽  
pp. 2000-2007 ◽  
Author(s):  
Simon Allen ◽  
Adel M. Abuzenadah ◽  
Joanna L. Blagg ◽  
Joanna Hinks ◽  
I. Mandy Nesbitt ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
High Molecular Weight ◽  
Von Willebrand Factor ◽  
Amino Acid Position ◽  
Von Willebrand Disease ◽  
Mild Reduction ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
High Molecular Weight Multimers

Abstract Two novel mutations, a T-to-C transition at nucleotide 2612 and a T-to-G transversion at nucleotide 3923 of the von Willebrand factor (vWF) complementary DNA, were detected by analysis of the vWF gene in DNA from members of 2 families with atypical von Willebrand disease. The T2612C transition predicts substitution of cysteine by arginine at amino acid position 788 (C788R), and the T3923G transversion predicts substitution of cysteine by glycine at position 1225 (C1225G) of pre-pro-vWF. The patients homozygous for the C788R and C1225G mutations both had a partial vWF deficiency (0.18 IU/mL and 0.07 IU/mL vWF antigen, respectively); vWF in plasma from patients homozygous for either the C788R or the C1225G mutation failed to bind factor VIII and lacked high molecular weight multimers. Recombinant (r) vWF molecules having the C788R or C1225G mutation were expressed in COS-7 cells. Both rvWF C788R and rvWF C1225G exhibited significantly impaired secretion and failed to bind factor VIII. Recombinant vWF C788R in COS-7 culture medium showed a severe reduction in high molecular weight multimers, whereas rvWF C1225G showed a very mild reduction in high molecular weight multimers when compared with wild-type rvWF.

scholarly journals Adhesion of platelets to human artery subendothelium: effect of factor VIII-von Willebrand factor of various multimeric composition

Blood ◽  
1984 ◽  
Vol 63 (1) ◽  
pp. 128-139
Author(s):  
JJ Sixma ◽  
KS Sakariassen ◽  
NH Beeser-Visser ◽  
M Ottenhof-Rovers ◽  
PA Bolhuis
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Platelet Adhesion ◽  
Bleeding Time ◽  
Low Molecular Weight ◽  
Perfusion Chamber ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Adhesion Factor

The relationship between the multimeric size of factor VIII-von Willebrand factor (FVIII-vWF) and the support of platelet adhesion to subendothelium was studied in an annular perfusion chamber, employing human renal and umbilical arteries. Commercial factor VIII concentrates containing multimers of low molecular weight that had been shown not to correct the bleeding time upon infusion into patients with von Willebrand's disease did not support platelet adhesion in the perfusion chamber. Cryoprecipitate and two experimental FVIII-vWF concentrates containing multimers of high molecular weight supported platelet adhesion. Factor VIII-vWF purified from cryoprecipitate was subdivided into three fractions of different molecular weights (6.0–14.0, 4.0–9.0, and 3.0–7.5 X 10(6) dalton). These fractions appeared to bind equally well and to be equally effective in supporting platelet adhesion. Factor VIII-vWF with multimers of low molecular weight (0.5–1.5 X 10(6) dalton) were prepared by partial reduction. Binding of FVIII-vWF to subendothelium was not impaired, and the support of platelet adhesion appeared to be more resistant to the effect of reduction than the ristocetin cofactor activity. At high shear rate (2,500 sec-1), increased platelet adhesion was observed with partially reduced FVIII- vWF. These data indicate that the ability of FVIII-vWF preparations to correct the bleeding time is reflected in enhanced platelet adhesion to subendothelium in a perfusion chamber. These data also emphasize that multimeric size is not the only factor determining whether FVIII-vWF will support platelet adhesion.

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