CLEC-2-dependent activation of mouse platelets is weakly inhibited by cAMP but not by cGMP

Research output: Contribution to journalArticle

External organisations

  • Institute for Physiological Chemistry and Pathobiochemistry; Münster University Hospital; Münster Germany
  • Department of Experimental Medicine; ‘Sapienza’ University of Rome; Rome Italy

Abstract

Background
The activation of platelet CLEC-2 by podoplanin on lymphatic endothelial cells (LECs) has a critical role in prevention of mixing of lymphatic and blood vasculatures during embryonic development. Paradoxically, LECs release cAMP and cGMP-elevating agents, prostacyclin (PGI2) and nitric oxide (NO), respectively, which are powerful inhibitors of platelet activation. This raises the question of how podoplanin is able to activate CLEC-2 in the presence of the inhibitory cyclic nucleotides.

Objectives
We investigated the influence of cyclic nucleotides on CLEC-2 signaling in platelets.

Methods
We used rhodocytin, CLEC-2 monoclonal antibody, LECs and recombinant podoplanin as CLEC-2 agonists on mouse platelets. The effects of the cyclic nucleotide-elevating agents PGI2, forskolin and the NO-donor GSNO were assessed with light transmission aggregometry, flow cytometry, protein phosphorylation and fluorescent imaging of platelets on LECs.

Results
We show that platelet aggregation induced by CLEC-2 agonists is resistant to GSNO but inhibited by PGI2. The effect of PGI2 is mediated through decreased phosphorylation of CLEC-2, Syk and PLCγ2. In contrast, adhesion and spreading of platelets on recombinant podoplanin, CLEC-2 antibody and LECs is not affected by PGI2 and GSNO. Consistent with this, CLEC-2 activation of Rac, which is required for platelet spreading, is not altered in the presence of PGI2.

Conclusions
The present results demonstrate that platelet adhesion and activation on CLEC-2 ligands or LECs is maintained in the presence of PGI2 and NO.

Details

Original languageEnglish
Pages (from-to)550-559
Number of pages10
JournalJournal of Thrombosis and Haemostasis
Volume12
Issue number4
Early online date11 Apr 2014
Publication statusPublished - Apr 2014

Keywords

  • blood platelets, C-type lectin, cyclic nucleotides, lymphangiogenesis, platelet activation