The Fibrinogen γ Chain Dodecapeptide Inhibits Agonist-induced Aggregation of Rabbit Platelets and Fibrinogen Binding to Rabbit Glycoprotein IIb-IIIa

Margaret L. Rand; Marian A. Packham; Desirée M. Taylor; Erik L. Yeo; Cynthia H. Gemmell; Sonali Patil; Stephen C.-T. Lam

doi:10.1055/s-0037-1614899

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 1999; 82(06): 1680-1686
DOI: 10.1055/s-0037-1614899

Rapid Communication

Schattauer GmbH

The Fibrinogen γ Chain Dodecapeptide Inhibits Agonist-induced Aggregation of Rabbit Platelets and Fibrinogen Binding to Rabbit Glycoprotein IIb-IIIa

Authors

Margaret L. Rand

¹From the Departments of Biochemistry and The Hospital for Sick Children, Toronto, Canada

³Division of Haematology/Oncology, The Hospital for Sick Children, Toronto, Canada
Marian A. Packham

¹From the Departments of Biochemistry and The Hospital for Sick Children, Toronto, Canada
Desirée M. Taylor

¹From the Departments of Biochemistry and The Hospital for Sick Children, Toronto, Canada
Erik L. Yeo

²Medicine, University of Toronto and The Hospital for Sick Children, Toronto, Canada
Cynthia H. Gemmell

²Medicine, University of Toronto and The Hospital for Sick Children, Toronto, Canada
Sonali Patil

⁴Department of Pharmacology, University of Illinois, Chicago, IL, USA
Stephen C.-T. Lam

⁴Department of Pharmacology, University of Illinois, Chicago, IL, USA

Abstract

Summary

The HHLGGAKQAGDV (H₁₂) sequence at the carboxyl termini of the γ chains and the RGD sequences in the Aα chains of human fibrinogen are potential recognition sites for the binding of soluble fibrinogen to glycoprotein IIb-IIIa (GPIIb-IIIa) on activated human platelets. Thus, addition of either H₁₂ or RGD-containing peptides inhibits aggregation of and fibrinogen binding to human platelets. In contrast, we reported previously that RGDS had relatively little inhibitory effect on these functions of rabbit platelets. In the present study, we found that H₁₂ inhibited ADP- and thrombin-induced aggregation of rabbit platelets in a dose-dependent manner. Specificity was demonstrated by the failure of the variant HHLGGAKQAGEV peptide to inhibit ADP-induced aggregation. Furthermore, flow cytometric analyses demonstrated that H₁₂ inhibited the binding of FITC-fibrinogen to ADP-activated rabbit platelets in a dose-dependent manner. To examine the direct interaction of H₁₂ with rabbit GPIIb-IIIa, we performed affinity chromatography by applying an octylglucoside extract of rabbit platelet proteins onto an affinity matrix containing the fibrinogen γ chain sequence. Proteins of ∼135 kDa and ∼95 kDa were specifically eluted by soluble H₁₂, and the 95 kDa protein band was immunoblotted by anti-LIBS1, a monoclonal antibody against human GPIIIa. In control samples, no detectable protein from rabbit platelet lysates was eluted from an RGD affinity matrix by GRGDSP. Collectively, our results demonstrated that H₁₂ inhibits aggregation of and fibrinogen binding to rabbit platelets by directly interacting with rabbit GPIIb-IIIa. These findings suggest that rabbit platelets would serve as a suitable thrombosis model for testing the efficacy of peptide mimetics derived from H₁₂.

Keywords

Rabbit platelets - fibrinogen - GPIIb-IIIa - gamma chain

Full Text

References

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