Altered bioavailability of platelet-derived factor VIII during thrombocytosis reverses phenotypic efficacy in haemophilic mice

Andrea L. Damon; Lesley E. Scudder; Dmitri V. Gnatenko; Varsha Sitaraman; Patrick Hearing; Jolyon Jesty; Wadie F. Bahou

doi:10.1160/TH08-04-0242

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2008; 100(06): 1111-1122
DOI: 10.1160/TH08-04-0242

Platelets and Blood Cells

Schattauer GmbH

Altered bioavailability of platelet-derived factor VIII during thrombocytosis reverses phenotypic efficacy in haemophilic mice

Andrea L. Damon

¹Department of Medicine, State University of New York, Stony Brook, New York, USA

²Program in Genetics, State University of New York, Stony Brook, New York, USA

,

Lesley E. Scudder

¹Department of Medicine, State University of New York, Stony Brook, New York, USA

,

Dmitri V. Gnatenko

¹Department of Medicine, State University of New York, Stony Brook, New York, USA

,

Varsha Sitaraman

¹Department of Medicine, State University of New York, Stony Brook, New York, USA

²Program in Genetics, State University of New York, Stony Brook, New York, USA

,

Patrick Hearing

²Program in Genetics, State University of New York, Stony Brook, New York, USA

³Dept. Microbiology and Molecular Genetics, State University of New York, Stony Brook, New York, USA

,

Jolyon Jesty

¹Department of Medicine, State University of New York, Stony Brook, New York, USA

,

Wadie F. Bahou

¹Department of Medicine, State University of New York, Stony Brook, New York, USA

²Program in Genetics, State University of New York, Stony Brook, New York, USA

› Author Affiliations

Abstract

Summary

Ectopic delivery of factor VIII (FVIII) to megakaryocytes (Mk) represents a viable approach for localized tenase generation by concentrating the FVIIIa/FIXa enzymecofactor complex onto activated platelet membranes. We utilized a core rat platelet factor 4 (PF4) promoter for Mk/platelet-restricted expression of human B-domain-deleted (hBDD) FVIII within the background of a haemophilia A mouse (rPF4/hBDD/FVIII^-/-). Platelets from rPF4/hBDD/FVIII^-/- mice contained ∼122 mU FVIII:C/1x10⁹ platelets/ml with no detectable plasmatic FVIII:C,and with no effect on α-granule-derived platelet factorV/Va function.Paired tenase assays (± thrombin) confirmed that platelet (pt) FVIII (unlike platelet FV) required thrombin cleavage for complete activation. rPF4/hBDD/FVIII^-/- mice exposed to a thrombocytotic stimulus (thrombopoietin, TPO) demonstrated a statistically-significant 66% reduction in molar ptFVIII activity with a non-significant reduction in total ptFVIII biomass. Decreased molar ptFVIII concentration correlated with loss of phenotypic correction as evaluated using a haemostatic tail-snip assay. Comparative studies using a transgenic mouse expressing human amyloid-β-precursor protein (hAβPP) from the rPF4 promoter confirmed diminished hAβPP expression without affecting endogenous α-granule PF4, establishing generalizability of these observations.While Mk/plateletreleased ptFVIII (unlike pFV) is proteolytically inactive, we also conclude that thrombocytotic stimuli negatively affect ptFVIII bioavailability and phenotypic efficacy, results which correlate best with molar ptFVIII concentration, and not systemically available ptFVIII.

Keywords

Animal models - gene targeting - coagulation factors

Full Text

References

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