Differential integrin activity mediated by platelet collagen receptor engagement under flow conditions

Nicholas Pugh; Ben D. Maddox; Dominique Bihan; Kirk A. Taylor; Martyn P. Mahaut-Smith; Richard W. Farndale

doi:10.1160/TH16-12-0906

Thrombosis and Haemostasis, Table of Contents

CC BY-NC-ND 4.0 · Thromb Haemost 2017; 117(08): 1588-1600
DOI: 10.1160/TH16-12-0906

Cellular Haemostasis and Platelets

Schattauer GmbH

Differential integrin activity mediated by platelet collagen receptor engagement under flow conditions

Nicholas Pugh

¹Department of Biochemistry, University of Cambridge, Cambridge, UK

²Department of Biomedical and Forensic Sciences, Anglia Ruskin University, Cambridge, UK

,

Ben D. Maddox

¹Department of Biochemistry, University of Cambridge, Cambridge, UK

,

Dominique Bihan

¹Department of Biochemistry, University of Cambridge, Cambridge, UK

,

Kirk A. Taylor

²Department of Biomedical and Forensic Sciences, Anglia Ruskin University, Cambridge, UK

,

Martyn P. Mahaut-Smith

³Department of Molecular and Cell Biology, University of Leicester, Leicester, UK

,

Richard W. Farndale

¹Department of Biochemistry, University of Cambridge, Cambridge, UK

› Author Affiliations

Abstract

Summary

The platelet receptors glycoprotein (Gp)VI, integrin α₂β₁ and GpIb/V/IX mediate platelet adhesion and activation during thrombogenesis. Increases of intracellular Ca²⁺ ([Ca²⁺]_i) are key signals during platelet activation; however, their relative importance in coupling different collagen receptors to functional responses under shear conditions remains unclear. To study shear-dependent, receptor-specific platelet responses, we used collagen or combinations of receptor-specific collagen-mimetic peptides as substrates for platelet adhesion and activation in whole human blood under arterial flow conditions and compared real-time and endpoint parameters of thrombus formation alongside [Ca²⁺]_i measurements using confocal imaging. All three collagen receptors coupled to [Ca²⁺]_i signals, but these varied in amplitude and temporal pattern alongside variable integrin activation. GpVI engagement produced large, sustained [Ca²⁺]_i signals leading to realtime increases in integrins α₂β₁− and α_IIbβ₃-mediated platelet adhesion. α_IIbβ₃-dependent platelet aggregation was dependent on P₂Y₁₂ signalling. Co-engagement of α₂β₁ and GpIb/V/IX generated transient [Ca²⁺]_i spikes and low amplitude [Ca²⁺]_i responses that potentiated GpVI-dependent [Ca²⁺]_i signalling. Therefore α₂β₁ GpIb/V/IX and GpVI synergise to generate [Ca²⁺]_i signals that regulate platelet behaviour and thrombus formation. Antagonism of secondary signalling pathways reveals distinct, separate roles for α_IIbβ₃ in stable platelet adhesion and aggregation.

Supplementary Material to this article is available online at www.thrombosis-online.com.

Keywords

Platelet glycoproteins - collagen receptors - signal transduction - thrombosis - fluid shear

Full Text

References

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