Cardiovascular Research Advance Access first published online on August 28, 2009
This version [Corrected Proof] published online on October 4, 2009
Cardiovascular Research, doi:10.1093/cvr/cvp301
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A sulfaphenazole-sensitive EDHF opposes platelet–endothelium interactions in vitro and in the hamster microcirculation in vivo
1 Cardiology Division, Medizinische Poliklinik, Ludwigs-Maximilians-Universität, Ziemssenstr. 1, Munich 80336, Germany
2 Medizinische Poliklinik - Innenstadt, the Institute of Anesthesiology, Ludwig-Maximilians-Universität, Munich, Germany
3 Walter-Brendel Centre of Experimental Medicine, Ludwig-Maximilians-Universität, Munich, Germany
* Corresponding author. Tel: +49 89 218075 384, Fax: +49 89 218075 378, Email: florian.kroetz{at}med.uni-muenchen.de
Aims: A CYP2C9-dependent endothelium-derived hyperpolarizing factor (EDHF) controls blood flow in many microvascular beds of various species by targeting vascular smooth muscle potassium channels. Since platelets express the same channels, we tested whether EDHF hyperpolarizes platelets and exerts an antithrombotic function in vivo.
Methods and results: Interaction of injected human platelets with the arteriolar wall (platelet-vessel wall interaction, PVWI) was assessed by intravital microscopy in skin muscle of awake hamsters. To understand the mechanisms of EDHF-induced platelet inhibition, we studied whether cultured human umbilical vein endothelial cells overexpressing CYP2C9-mRNA in vitro released a factor that could hyperpolarize human platelets. Under control conditions, there was no firm adhesion of platelets to the arteriolar wall, but temporary PVWI occurred. Local superfusion of the CYP2C9 inhibitor sulfaphenazole, at doses known to block EDHF-dependent dilations, significantly augmented PVWI, as did inhibition of NO synthase. Inhibition of both factors exerted additive effects on PVWI. Likewise, firm adhesion of a small fraction of platelets was observed. The prothrombotic effects of CYP2C9 inhibition in vivo were reversed by exogenous superfusion with 11,12-epoxyeicosatrienoic acids. Hyperpolarization reduced platelet adhesion to endothelial cells under static conditions in vitro and was dependent on calcium-activated potassium channels. The factor also reduced ADP-induced expression of platelet P-selectin, indicating reduction of platelet activity.
Conclusion: The arteriolar endothelium in vivo continuously releases a CYP2C9-derived EDHF. This EDHF exerts its effects by hyperpolarization of platelets through activation of KCa channels and reduction of platelet adhesion molecule expression, indicating that hyperpolarization reduces platelet activation. This demonstrates that EDHF is part of the antithrombotic properties of healthy endothelium in vivo.
KEYWORDS EDHF; Platelets; Thrombosis; Microcirculation
Time for primary review: 29 days