on January 8, 2004
Published online before print January 8, 2004, doi: 10.1161/01.ATV.0000116219.09040.8c
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Submitted on November 19, 2003
Accepted on December 9, 2003
Membrane Potential-Dependent Inhibition of Platelet Adhesion to Endothelial Cells by Epoxyeicosatrienoic Acids
Florian Krötz *;
From the Institute of Physiology (F.K., T.R., T.G., U.P.), Clinic of Anaesthesiology (M.A.B.), and Cardiology Division (H.-Y.S.), Medizinische Poliklinik-Innenstadt, Ludwig-Maximilians-University, Munich, Germany; and the Department of Pharmacology and Toxicology (K.N., W.B.C.), Medical College of Wisconsin, Milwaukee, Wisc.
* To whom correspondence should be addressed. E-mail: fkroetz{at}lmu.de.
Objective--Epoxyeicosatrienoic acids (EETs) are potent vasodilators produced by endothelial cells. In many vessels, they are an endothelium-derived hyperpolarizing factor (EDHF). However, it is unknown whether they act as an EDHF on platelets and whether this has functional consequences.
Methods and Results--Flow cytometric measurement of platelet membrane potential using the fluorescent dye DiBac4 showed a resting potential of -58±9 mV. Different EET regioisomers hyperpolarized platelets down to -69±2 mV, which was prevented by the non-specific potassium channel inhibitor charybdotoxin and by use of a blocker of calcium-activated potassium channels of large conductance (BKCa channels), iberiotoxin. EETs inhibited platelet adhesion to endothelial cells under static and flow conditions. Exposure to EETs inhibited platelet P-selectin expression in response to ADP. Stable overexpression of cytochrome P450 2C9 in EA.hy926 cells (EA.hy2C9 cells) resulted in release of EETs and a factor that hyperpolarized platelets and inhibited their adhesion to endothelial cells. These effects were again inhibited by charybdotoxin and iberiotoxin.
Conclusions--EETs hyperpolarize platelets and inactivate them by inhibiting adhesion molecule expression and platelet adhesion to cultured endothelial cells in a membrane potential-dependent manner. They act as an EDHF on platelets and might be important mediators of the anti-adhesive properties of vascular endothelium.
Key words: epoxyeicosatrienoic acids • platelet adhesion • membrane potential • potassium channels • EDHF
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