Thromb Haemost 2015; 113(05): 1109-1120
DOI: 10.1160/TH14-09-0762
Cellular Signalling and Proteolysis
Schattauer GmbH

Platelet adhesion to podoplanin under flow is mediated by the receptor CLEC-2 and stabilised by Src/Syk-dependent platelet signalling

Leyre Navarro-Núñez
1  Centre for Cardiovascular Sciences, Institute for Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK
,
Alice Y. Pollitt
1  Centre for Cardiovascular Sciences, Institute for Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK
,
Kate Lowe
1  Centre for Cardiovascular Sciences, Institute for Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK
,
Arusa Latif
1  Centre for Cardiovascular Sciences, Institute for Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK
,
Gerard B. Nash
1  Centre for Cardiovascular Sciences, Institute for Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK
,
Steve P. Watson
1  Centre for Cardiovascular Sciences, Institute for Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK
› Author Affiliations
Financial support: Study supported by the British Heart Foundation (PG/11/119, CH/03/003) and the Wellcome Trust (088410).
Further Information

Correspondence to:

Leyre Navarro-Núñez and Steve P. Watson
Centre for Cardiovascular Sciences
Institute for Biomedical Research
College of Medical and Dental Sciences
University of Birmingham, B15 2TT, United Kingdom
Phone: +44 1214158678   
Fax: +44 1214158817   
Email: [email protected]   

Publication History

Received: 12 September 2014

Accepted after major revision: 06 January 2015

Publication Date:
24 November 2017 (online)

 

Summary

Platelet-specific deletion of CLEC-2, which signals through Src and Syk kinases, or global deletion of its ligand podoplanin results in bloodfilled lymphatics during mouse development. Platelet-specific Syk deficiency phenocopies this defect, indicating that platelet activation is required for lymphatic development. In the present study, we investigated whether CLEC-2-podoplanin interactions could support platelet arrest from blood flow and whether platelet signalling is required for stable platelet adhesion to lymphatic endothelial cells (LECs) and recombinant podoplanin under flow. Perfusion of human or mouse blood over human LEC monolayers led to platelet adhesion and aggregation. Following α∥bβ3 blockade, individual platelets still adhered. Platelet binding occurred at venous but not arterial shear rates. There was no adhesion using CLEC-2-deficient blood or to vascular endothelial cells (which lack podoplanin). Perfusion of human blood over human Fc-podoplanin (hFcPDPN) in the presence of monoclonal antibody IV.3 to block FcγR∥A receptors led to platelet arrest at similar shear rates to those used on LECs. Src and Syk inhibitors significantly reduced global adhesion of human or mouse platelets to LECs and hFcPDPN. A similar result was seen using Syk-deficient mouse platelets. Reduced platelet adhesion was due to a decrease in the stability of binding. In conclusion, our data reveal that CLEC-2 is an adhesive receptor that supports platelet arrest to podoplanin under venous shear. Src/Syk-dependent signalling stabilises platelet adhesion to podoplanin, providing a possible molecular mechanism contributing to the lymphatic defects of Syk-deficient mice.


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Conflicts of interest

None declared.


Correspondence to:

Leyre Navarro-Núñez and Steve P. Watson
Centre for Cardiovascular Sciences
Institute for Biomedical Research
College of Medical and Dental Sciences
University of Birmingham, B15 2TT, United Kingdom
Phone: +44 1214158678   
Fax: +44 1214158817   
Email: [email protected]