Zebrafish von Willebrand factor

Maira Carrillo, Seongcheol Kim, Surendra Kumar Rajpurohit, Vrinda Kulkarni, Pudur Jagadeeswaran

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

von Willebrand factor (vWF) is a large protein involved in primary hemostasis. A dysfunction in this protein or an insufficient production of the protein leads to improper platelet adhesion/aggregation, resulting in a bleeding phenotype known as von Willebrand disease (vWD). To gain a better understanding of vWF interactions in vivo, the use of zebrafish as a model is ideal because of the transparency of the embryos and larvae. In this article, we examined the presence and function of vWF in hemostasis of zebrafish utilizing a variety of molecular methods. Using RT-PCR and antibody staining, we have shown that vWF mRNA is present in thrombocytes. Through antibody staining, we demonstrated vWF is synthesized in blood vessels. The role of zebrafish vWF in hemostasis was established through knockdown methods using vWF morpholino (vWF MO) antisense oligonucleotides. Embryos injected with vWF MO at the one to four cell stages resulted in a bleeding phenotype. Injection of embryos with vWF MO also caused an increase in time to occlusion within arteries in larvae upon laser induced injury. We then used vWF-specific Vivo-morpholinos (VMO) to induce vWF knockdown in adult zebrafish by targeting the exon homologous to the human exon 28 of the vWF gene. The reduced ristocetin-mediated agglutination of thrombocytes in a plate tilting assay, using blood from adult zebrafish injected with VMO, provided evidence that vWF is involved in the hemostatic process. We also administered desmopressin acetate to larvae and adults which resulted in enhanced aggregation/agglutination of thrombocytes. Zebrafish genome database analysis revealed the presence of GPIbβ gene. It also revealed the exon of zebrafish vWF gene corresponding to exon 28 of human vWF gene is highly similar to the exon 28 of human vWF gene, except that it has an insertion that leads to a translated peptide sequence that separates the two A domains coded by this exon. This exon is also conserved in other fishes. In summary, we established that zebrafish vWF has a role similar to that of vWF found in humans, thus, making zebrafish a useful model for studying the cell biology of vWF in vivo.

Original languageEnglish (US)
Pages (from-to)326-33
Number of pages8
JournalBlood Cells, Molecules, and Diseases
Volume45
Issue number4
DOIs
StatePublished - Dec 15 2010

Fingerprint

von Willebrand Factor
Zebrafish
Exons
Morpholinos
Hemostasis
Larva
Blood Platelets
Embryonic Structures
Genes
Agglutination
Ristocetin
Staining and Labeling
Hemorrhage
Phenotype
von Willebrand Diseases
Deamino Arginine Vasopressin
Proteins
Antibodies
Antisense Oligonucleotides
Hemostatics

Keywords

  • Animals
  • Embryo, Nonmammalian
  • Exons
  • Hemostasis
  • Humans
  • Models, Animal
  • Thrombosis
  • Zebrafish
  • von Willebrand Factor

Cite this

Zebrafish von Willebrand factor. / Carrillo, Maira; Kim, Seongcheol; Rajpurohit, Surendra Kumar; Kulkarni, Vrinda; Jagadeeswaran, Pudur.

In: Blood Cells, Molecules, and Diseases, Vol. 45, No. 4, 15.12.2010, p. 326-33.

Research output: Contribution to journalArticle

Carrillo, M, Kim, S, Rajpurohit, SK, Kulkarni, V & Jagadeeswaran, P 2010, 'Zebrafish von Willebrand factor', Blood Cells, Molecules, and Diseases, vol. 45, no. 4, pp. 326-33. https://doi.org/10.1016/j.bcmd.2010.10.002
Carrillo, Maira ; Kim, Seongcheol ; Rajpurohit, Surendra Kumar ; Kulkarni, Vrinda ; Jagadeeswaran, Pudur. / Zebrafish von Willebrand factor. In: Blood Cells, Molecules, and Diseases. 2010 ; Vol. 45, No. 4. pp. 326-33.
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AU - Rajpurohit, Surendra Kumar

AU - Kulkarni, Vrinda

AU - Jagadeeswaran, Pudur

N1 - Copyright © 2010 Elsevier Inc. All rights reserved.

PY - 2010/12/15

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N2 - von Willebrand factor (vWF) is a large protein involved in primary hemostasis. A dysfunction in this protein or an insufficient production of the protein leads to improper platelet adhesion/aggregation, resulting in a bleeding phenotype known as von Willebrand disease (vWD). To gain a better understanding of vWF interactions in vivo, the use of zebrafish as a model is ideal because of the transparency of the embryos and larvae. In this article, we examined the presence and function of vWF in hemostasis of zebrafish utilizing a variety of molecular methods. Using RT-PCR and antibody staining, we have shown that vWF mRNA is present in thrombocytes. Through antibody staining, we demonstrated vWF is synthesized in blood vessels. The role of zebrafish vWF in hemostasis was established through knockdown methods using vWF morpholino (vWF MO) antisense oligonucleotides. Embryos injected with vWF MO at the one to four cell stages resulted in a bleeding phenotype. Injection of embryos with vWF MO also caused an increase in time to occlusion within arteries in larvae upon laser induced injury. We then used vWF-specific Vivo-morpholinos (VMO) to induce vWF knockdown in adult zebrafish by targeting the exon homologous to the human exon 28 of the vWF gene. The reduced ristocetin-mediated agglutination of thrombocytes in a plate tilting assay, using blood from adult zebrafish injected with VMO, provided evidence that vWF is involved in the hemostatic process. We also administered desmopressin acetate to larvae and adults which resulted in enhanced aggregation/agglutination of thrombocytes. Zebrafish genome database analysis revealed the presence of GPIbβ gene. It also revealed the exon of zebrafish vWF gene corresponding to exon 28 of human vWF gene is highly similar to the exon 28 of human vWF gene, except that it has an insertion that leads to a translated peptide sequence that separates the two A domains coded by this exon. This exon is also conserved in other fishes. In summary, we established that zebrafish vWF has a role similar to that of vWF found in humans, thus, making zebrafish a useful model for studying the cell biology of vWF in vivo.

AB - von Willebrand factor (vWF) is a large protein involved in primary hemostasis. A dysfunction in this protein or an insufficient production of the protein leads to improper platelet adhesion/aggregation, resulting in a bleeding phenotype known as von Willebrand disease (vWD). To gain a better understanding of vWF interactions in vivo, the use of zebrafish as a model is ideal because of the transparency of the embryos and larvae. In this article, we examined the presence and function of vWF in hemostasis of zebrafish utilizing a variety of molecular methods. Using RT-PCR and antibody staining, we have shown that vWF mRNA is present in thrombocytes. Through antibody staining, we demonstrated vWF is synthesized in blood vessels. The role of zebrafish vWF in hemostasis was established through knockdown methods using vWF morpholino (vWF MO) antisense oligonucleotides. Embryos injected with vWF MO at the one to four cell stages resulted in a bleeding phenotype. Injection of embryos with vWF MO also caused an increase in time to occlusion within arteries in larvae upon laser induced injury. We then used vWF-specific Vivo-morpholinos (VMO) to induce vWF knockdown in adult zebrafish by targeting the exon homologous to the human exon 28 of the vWF gene. The reduced ristocetin-mediated agglutination of thrombocytes in a plate tilting assay, using blood from adult zebrafish injected with VMO, provided evidence that vWF is involved in the hemostatic process. We also administered desmopressin acetate to larvae and adults which resulted in enhanced aggregation/agglutination of thrombocytes. Zebrafish genome database analysis revealed the presence of GPIbβ gene. It also revealed the exon of zebrafish vWF gene corresponding to exon 28 of human vWF gene is highly similar to the exon 28 of human vWF gene, except that it has an insertion that leads to a translated peptide sequence that separates the two A domains coded by this exon. This exon is also conserved in other fishes. In summary, we established that zebrafish vWF has a role similar to that of vWF found in humans, thus, making zebrafish a useful model for studying the cell biology of vWF in vivo.

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KW - Embryo, Nonmammalian

KW - Exons

KW - Hemostasis

KW - Humans

KW - Models, Animal

KW - Thrombosis

KW - Zebrafish

KW - von Willebrand Factor

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JO - Blood Cells, Molecules, and Diseases

JF - Blood Cells, Molecules, and Diseases

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