Correction of a murine model of von Willebrand disease by gene transfer

Robert G. Pergolizzi, Guangchun Jin, Diane Chan, Lorraine Pierre, James Bussel, Barbara Ferris, Philip L. Leopold, Ronald Crystal

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

von Willebrand disease (VWD), the most common inherited bleeding disorder in the U.S. population, is caused by defects in the expression and processing of von Willebrand factor (VWF), a blood glycoprotein required for normal hemostasis that mediates the adhesion of platelets to sites of vascular damage by binding to specific platelet glycoproteins and to constituents of exposed connective tissue. To assess whether VWF deficiency can be corrected by gene transfer, a plasmid expressing the intact 8.4-kb murine VWF coding sequence, directed by the cytomegalovirus immediate/early promoter/ enhancer, was delivered through hydrodynamic tail vein injection into VWF knockout mice (VWF-/-) that exhibit defects in hemostasis, including highly prolonged bleeding time and spontaneous bleeding events, closely mimicking severe human VWD. VWF antigen levels in plasma from animals receiving VWF cDNA, but not control animals, revealed normalized levels of circulating VWF that persisted for at least 1 week after injection. Western blot analysis of plasma from animals receiving VWF cDNA, but not control animals, revealed high molecular-weight multimers with patterns similar to those observed in wild-type mice. Reverse transcription-polymerase chain reaction (RT-PCR) on RNA isolated from the livers of animals receiving VWF cDNA, but not control animals, demonstrated that VWF was expressed in the liver, and immunohistochemical analysis of the livers of treated VWF-/- mice revealed VWF-specific staining throughout the liver parenchyma but not in endothelial cells. Plasma from treated VWF-/- mice, but not control VWF-/- mice, supported the hypothesis that murine platelets aggregate in the presence of botrocetin. Although levels of circulating factor VIII in untreated VWF -/- mice were less than 10% those in wild-type mice, levels of factor VIII in VWF-/- animals treated with VWF cDNA, but not in control animals, were normalized to values in wildtype mice, indicating the restoration of factor VIII carrier function for VWF in treated mice that persisted for at least 1 week at higher doses of VWF cDNA. Most important, bleeding time was normalized by 48 hours after the delivery of VWF cDNA, but not by the control plasmid. These data suggest that with the use of gene transfer of VWF cDNA, VWF protein can be expressed, processed, and secreted in a physiologically active form; thus, it may be possible to correct VWD using gene transfer.

Original languageEnglish
Pages (from-to)862-869
Number of pages8
JournalBlood
Volume108
Issue number3
DOIs
Publication statusPublished - 1 Aug 2006
Externally publishedYes

Fingerprint

Gene transfer
von Willebrand Diseases
von Willebrand Factor
Genes
Animals
Complementary DNA
Liver
Factor VIII
Bleeding Time
Hemostasis
Platelets
Plasmas
Plasmids
Blood Platelets

ASJC Scopus subject areas

  • Hematology

Cite this

Pergolizzi, R. G., Jin, G., Chan, D., Pierre, L., Bussel, J., Ferris, B., ... Crystal, R. (2006). Correction of a murine model of von Willebrand disease by gene transfer. Blood, 108(3), 862-869. https://doi.org/10.1182/blood-2005-06-2330

Correction of a murine model of von Willebrand disease by gene transfer. / Pergolizzi, Robert G.; Jin, Guangchun; Chan, Diane; Pierre, Lorraine; Bussel, James; Ferris, Barbara; Leopold, Philip L.; Crystal, Ronald.

In: Blood, Vol. 108, No. 3, 01.08.2006, p. 862-869.

Research output: Contribution to journalArticle

Pergolizzi, RG, Jin, G, Chan, D, Pierre, L, Bussel, J, Ferris, B, Leopold, PL & Crystal, R 2006, 'Correction of a murine model of von Willebrand disease by gene transfer', Blood, vol. 108, no. 3, pp. 862-869. https://doi.org/10.1182/blood-2005-06-2330
Pergolizzi RG, Jin G, Chan D, Pierre L, Bussel J, Ferris B et al. Correction of a murine model of von Willebrand disease by gene transfer. Blood. 2006 Aug 1;108(3):862-869. https://doi.org/10.1182/blood-2005-06-2330
Pergolizzi, Robert G. ; Jin, Guangchun ; Chan, Diane ; Pierre, Lorraine ; Bussel, James ; Ferris, Barbara ; Leopold, Philip L. ; Crystal, Ronald. / Correction of a murine model of von Willebrand disease by gene transfer. In: Blood. 2006 ; Vol. 108, No. 3. pp. 862-869.
@article{369bf2daaf034c30b4962de53256d2ae,
title = "Correction of a murine model of von Willebrand disease by gene transfer",
abstract = "von Willebrand disease (VWD), the most common inherited bleeding disorder in the U.S. population, is caused by defects in the expression and processing of von Willebrand factor (VWF), a blood glycoprotein required for normal hemostasis that mediates the adhesion of platelets to sites of vascular damage by binding to specific platelet glycoproteins and to constituents of exposed connective tissue. To assess whether VWF deficiency can be corrected by gene transfer, a plasmid expressing the intact 8.4-kb murine VWF coding sequence, directed by the cytomegalovirus immediate/early promoter/ enhancer, was delivered through hydrodynamic tail vein injection into VWF knockout mice (VWF-/-) that exhibit defects in hemostasis, including highly prolonged bleeding time and spontaneous bleeding events, closely mimicking severe human VWD. VWF antigen levels in plasma from animals receiving VWF cDNA, but not control animals, revealed normalized levels of circulating VWF that persisted for at least 1 week after injection. Western blot analysis of plasma from animals receiving VWF cDNA, but not control animals, revealed high molecular-weight multimers with patterns similar to those observed in wild-type mice. Reverse transcription-polymerase chain reaction (RT-PCR) on RNA isolated from the livers of animals receiving VWF cDNA, but not control animals, demonstrated that VWF was expressed in the liver, and immunohistochemical analysis of the livers of treated VWF-/- mice revealed VWF-specific staining throughout the liver parenchyma but not in endothelial cells. Plasma from treated VWF-/- mice, but not control VWF-/- mice, supported the hypothesis that murine platelets aggregate in the presence of botrocetin. Although levels of circulating factor VIII in untreated VWF -/- mice were less than 10{\%} those in wild-type mice, levels of factor VIII in VWF-/- animals treated with VWF cDNA, but not in control animals, were normalized to values in wildtype mice, indicating the restoration of factor VIII carrier function for VWF in treated mice that persisted for at least 1 week at higher doses of VWF cDNA. Most important, bleeding time was normalized by 48 hours after the delivery of VWF cDNA, but not by the control plasmid. These data suggest that with the use of gene transfer of VWF cDNA, VWF protein can be expressed, processed, and secreted in a physiologically active form; thus, it may be possible to correct VWD using gene transfer.",
author = "Pergolizzi, {Robert G.} and Guangchun Jin and Diane Chan and Lorraine Pierre and James Bussel and Barbara Ferris and Leopold, {Philip L.} and Ronald Crystal",
year = "2006",
month = "8",
day = "1",
doi = "10.1182/blood-2005-06-2330",
language = "English",
volume = "108",
pages = "862--869",
journal = "Blood",
issn = "0006-4971",
publisher = "American Society of Hematology",
number = "3",

}

TY - JOUR

T1 - Correction of a murine model of von Willebrand disease by gene transfer

AU - Pergolizzi, Robert G.

AU - Jin, Guangchun

AU - Chan, Diane

AU - Pierre, Lorraine

AU - Bussel, James

AU - Ferris, Barbara

AU - Leopold, Philip L.

AU - Crystal, Ronald

PY - 2006/8/1

Y1 - 2006/8/1

N2 - von Willebrand disease (VWD), the most common inherited bleeding disorder in the U.S. population, is caused by defects in the expression and processing of von Willebrand factor (VWF), a blood glycoprotein required for normal hemostasis that mediates the adhesion of platelets to sites of vascular damage by binding to specific platelet glycoproteins and to constituents of exposed connective tissue. To assess whether VWF deficiency can be corrected by gene transfer, a plasmid expressing the intact 8.4-kb murine VWF coding sequence, directed by the cytomegalovirus immediate/early promoter/ enhancer, was delivered through hydrodynamic tail vein injection into VWF knockout mice (VWF-/-) that exhibit defects in hemostasis, including highly prolonged bleeding time and spontaneous bleeding events, closely mimicking severe human VWD. VWF antigen levels in plasma from animals receiving VWF cDNA, but not control animals, revealed normalized levels of circulating VWF that persisted for at least 1 week after injection. Western blot analysis of plasma from animals receiving VWF cDNA, but not control animals, revealed high molecular-weight multimers with patterns similar to those observed in wild-type mice. Reverse transcription-polymerase chain reaction (RT-PCR) on RNA isolated from the livers of animals receiving VWF cDNA, but not control animals, demonstrated that VWF was expressed in the liver, and immunohistochemical analysis of the livers of treated VWF-/- mice revealed VWF-specific staining throughout the liver parenchyma but not in endothelial cells. Plasma from treated VWF-/- mice, but not control VWF-/- mice, supported the hypothesis that murine platelets aggregate in the presence of botrocetin. Although levels of circulating factor VIII in untreated VWF -/- mice were less than 10% those in wild-type mice, levels of factor VIII in VWF-/- animals treated with VWF cDNA, but not in control animals, were normalized to values in wildtype mice, indicating the restoration of factor VIII carrier function for VWF in treated mice that persisted for at least 1 week at higher doses of VWF cDNA. Most important, bleeding time was normalized by 48 hours after the delivery of VWF cDNA, but not by the control plasmid. These data suggest that with the use of gene transfer of VWF cDNA, VWF protein can be expressed, processed, and secreted in a physiologically active form; thus, it may be possible to correct VWD using gene transfer.

AB - von Willebrand disease (VWD), the most common inherited bleeding disorder in the U.S. population, is caused by defects in the expression and processing of von Willebrand factor (VWF), a blood glycoprotein required for normal hemostasis that mediates the adhesion of platelets to sites of vascular damage by binding to specific platelet glycoproteins and to constituents of exposed connective tissue. To assess whether VWF deficiency can be corrected by gene transfer, a plasmid expressing the intact 8.4-kb murine VWF coding sequence, directed by the cytomegalovirus immediate/early promoter/ enhancer, was delivered through hydrodynamic tail vein injection into VWF knockout mice (VWF-/-) that exhibit defects in hemostasis, including highly prolonged bleeding time and spontaneous bleeding events, closely mimicking severe human VWD. VWF antigen levels in plasma from animals receiving VWF cDNA, but not control animals, revealed normalized levels of circulating VWF that persisted for at least 1 week after injection. Western blot analysis of plasma from animals receiving VWF cDNA, but not control animals, revealed high molecular-weight multimers with patterns similar to those observed in wild-type mice. Reverse transcription-polymerase chain reaction (RT-PCR) on RNA isolated from the livers of animals receiving VWF cDNA, but not control animals, demonstrated that VWF was expressed in the liver, and immunohistochemical analysis of the livers of treated VWF-/- mice revealed VWF-specific staining throughout the liver parenchyma but not in endothelial cells. Plasma from treated VWF-/- mice, but not control VWF-/- mice, supported the hypothesis that murine platelets aggregate in the presence of botrocetin. Although levels of circulating factor VIII in untreated VWF -/- mice were less than 10% those in wild-type mice, levels of factor VIII in VWF-/- animals treated with VWF cDNA, but not in control animals, were normalized to values in wildtype mice, indicating the restoration of factor VIII carrier function for VWF in treated mice that persisted for at least 1 week at higher doses of VWF cDNA. Most important, bleeding time was normalized by 48 hours after the delivery of VWF cDNA, but not by the control plasmid. These data suggest that with the use of gene transfer of VWF cDNA, VWF protein can be expressed, processed, and secreted in a physiologically active form; thus, it may be possible to correct VWD using gene transfer.

UR - http://www.scopus.com/inward/record.url?scp=33746661500&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33746661500&partnerID=8YFLogxK

U2 - 10.1182/blood-2005-06-2330

DO - 10.1182/blood-2005-06-2330

M3 - Article

VL - 108

SP - 862

EP - 869

JO - Blood

JF - Blood

SN - 0006-4971

IS - 3

ER -