Toll-like receptor 2 stimulation of platelets is mediated by purinergic P2X1-dependent Ca2+ mobilisation, cyclooxygenase and purinergic P2Y1 and P2Y12 receptor activation

Hanna Kälvegren; Caroline Skoglund; Christian Helldahl; Maria Lerm; Magnus Grenegård; Torbjörn Bengtsson

doi:10.1160/TH09-07-0442

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2010; 103(02): 398-407
DOI: 10.1160/TH09-07-0442

Platelets and Blood Cells

Schattauer GmbH

Toll-like receptor 2 stimulation of platelets is mediated by purinergic P2X1-dependent Ca²⁺ mobilisation, cyclooxygenase and purinergic P2Y1 and P2Y12 receptor activation

Hanna Kälvegren

¹Division of Drug Research, Department of Medical and Health Sciences, Cardiovascular Inflammation Research Centre (CIRC), Faculty of Health Sciences, Linköping University, Linköping, Sweden

³Division of Clinical Medicine, Department of Biomedicine, Örebro University, Örebro, Sweden

,

Caroline Skoglund

¹Division of Drug Research, Department of Medical and Health Sciences, Cardiovascular Inflammation Research Centre (CIRC), Faculty of Health Sciences, Linköping University, Linköping, Sweden

,

Christian Helldahl

¹Division of Drug Research, Department of Medical and Health Sciences, Cardiovascular Inflammation Research Centre (CIRC), Faculty of Health Sciences, Linköping University, Linköping, Sweden

,

Maria Lerm

²Division of Medical Microbiology, Department of Clinical and Experimental Medicine, Faculty of Health Sciences, Linköping University, Linköping, Sweden

,

Magnus Grenegård

¹Division of Drug Research, Department of Medical and Health Sciences, Cardiovascular Inflammation Research Centre (CIRC), Faculty of Health Sciences, Linköping University, Linköping, Sweden

,

Torbjörn Bengtsson

¹Division of Drug Research, Department of Medical and Health Sciences, Cardiovascular Inflammation Research Centre (CIRC), Faculty of Health Sciences, Linköping University, Linköping, Sweden

³Division of Clinical Medicine, Department of Biomedicine, Örebro University, Örebro, Sweden

› Author Affiliations

Abstract

Summary

Toll-like receptor 2 (TLR2), which recognise and respond to conserved microbial pathogen-associated molecular patterns, is expressed on the platelet surface. Furthermore, it has recently been shown that the TLR2/1 agonist Pam3CSK4 stimulates platelet activation. The aim of the present study was to clarify important signalling events in Pam3CSK4-induced platelet aggregation and secretion. Platelet interaction with Pam3CSK4 and the TLR2/6 agonist MALP-2 was studied by analysing aggregation, ATP-secretion, [Ca²⁺]i mobilisation and thromboxane B2 (TxB2) production. The results show that Pam3CSK4 but not MALP-2 induces [Ca²⁺]i increase, TxB2 production, dense granule secretion and platelet aggregation. Preincubation of platelets with MALP-2 inhibited the Pam3CSK4-induced responses. The ATP-secretion and aggregation in Pam3CSK4-stimulated platelets was impeded by the purinergic P2X1 inhibitor MRS 2159, the purinergic P2Y1 and P2Y12 antagonists MRS 2179 and cangrelor, the phospholipase C inhibitor U73122, the calcium chelator BAPT-AM and aspirin. The calcium mobilisation was lowered by MRS 2159, aspirin and U73122 whereas the TxB2 production was antagonised by MRS 2159, aspirin and BAPT-AM. When investigating the involvement of the myeloid differentiation factor-88 (MyD88) -dependent pathway, we found that platelets express MyD88 and interleukin 1 receptor-associated kinase (IRAK-1), which are proteins important in TLR signalling. However, Pam3CSK4 did not stimulate a rapid (within 10 minutes) phosphorylation of IRAK-1 in platelets. In conclusion, the results show that Pam3CSK4-induced platelet aggregation and secretion depends on a P2X1-mediated Ca²⁺ mobilisation, production of TxA2 and ADP receptor activation. The findings in this study further support a role for platelets in sensing bacterial components.

Keywords

Infection - purinergic P2X1 receptor - atherosclerosis - MALP-2 - Pam3CSK4 - platelet - MyD88 - IRAK-1

Full Text

References

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