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CC BY 4.0 · Thromb Haemost 2018; 118(02): 369-380
DOI: 10.1160/TH17-07-0530
Cellular Haemostasis and Platelets
Schattauer GmbH Stuttgart

P2X1 Receptors Amplify FcγRIIa-Induced Ca2+ Increases and Functional Responses in Human Platelets

Zeki Ilkan
1   Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, Leicester, United Kingdom
,
Stephanie Watson
2   Institute of Cardiovascular Sciences, Institute of Biomedical Research Building, University of Birmingham, Birmingham, United Kingdom
,
Steve P. Watson
2   Institute of Cardiovascular Sciences, Institute of Biomedical Research Building, University of Birmingham, Birmingham, United Kingdom
3   Centre of Membrane Proteins and Receptors (COMPARE), Universities of Birmingham and Nottingham, Midlands, UK
,
Martyn P. Mahaut-Smith
1   Department of Molecular and Cell Biology, Henry Wellcome Building, University of Leicester, Leicester, United Kingdom
› Author Affiliations Funding This work was supported by the British Heart Foundation grants PG/11/56, PG/05/014 and RG/13/18/30563. Z.I. was supported by a Medical Research Council Doctoral Training Award.
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Publication History

31 July 2017

18 November 2017

Publication Date:
29 January 2018 (online)

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Abstract

Platelets express key receptors of the innate immune system such as FcγRIIa and Toll-like receptors (TLR). P2X1 cation channels amplify the platelet responses to several major platelet stimuli, particularly glycoprotein (GP)VI and TLR2/1, whereas their contribution to Src tyrosine kinase-dependent FcγRIIa receptors remains unknown. We investigated the role of P2X1 receptors during activation of FcγRIIa in human platelets, following stimulation by cross-linking of an anti-FcγRIIa monoclonal antibody (mAb) IV.3, or bacterial stimulation with Streptococcus sanguinis. Activation was assessed in washed platelet suspensions via measurement of intracellular Ca2+ ([Ca2+]i) increases, ATP release and aggregation. P2X1 activity was abolished by pre-addition of α,β-meATP, exclusion of apyrase or the antagonist NF449. FcγRIIa activation evoked a robust increase in [Ca2+]i (441 ± 33 nM at 30 μg/mL mAb), which was reduced to a similar extent (to 66–70% of control) by NF449, pre-exposure to α,β-meATP or apyrase omission, demonstrating a significant P2X1 receptor contribution. FcγRIIa activation-dependent P2X1 responses were partially resistant to nitric oxide (NO), but abrogated by 500 nM prostacyclin (PGI2). Aggregation responses to bacteria and FcγRIIa activation were also inhibited by P2X1 receptor desensitization (to 66 and 42% of control, respectively). However, FcγRIIa-mediated tyrosine phosphorylation and ATP release were not significantly altered by the loss of P2X1 activity. In conclusion, we show that P2X1 receptors enhance platelet FcγRIIa receptor-evoked aggregation through an increase in [Ca2+]i downstream of the initial tyrosine phosphorylation events and early dense granule release. This represents a further route whereby ATP-gated cation channels can contribute to platelet-dependent immune responses in vivo.

Keywords

bacteria - immunity - inflammation - thrombosis - platelet
 

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