Characterization of a novel and potent collagen antagonist, caffeic acid phenethyl ester, in human platelets: In vitro and in vivo studies

Objective: Caffeic acid phenethyl ester (CAPE), which is derived from the propolis of honeybee hives, has been demonstrated to possess multiple pharmacological activities. In the present study, CAPE (6-25 mu M) specifically inhibited collagen-induced platelet aggregation and the ATP release reaction in platelet suspensions. Methods: Platelet aggregation, flow cytometric analysis, immunoblotting, and electron spin resonance (ESR) were used to assess the antiplatelet activity of CAPE. Fluorescein sodium-induced platelet thrombi in mesenteric microvessels of mice were used for an in vivo study. Results: CAPE (15-100 mu M) produced a concentration-related rightward displacement of the collagen concentration-response curve, and the Schild plot gave pA(2) and pA(10) values of 4.28 +/- 0.07 and 3.14 +/- 0.73, respectively, with a slope of -0.83 +/- 0.16, indicating specific antagonism. CAPE (25 mu M) also inhibited platelet aggregation stimulated by the glycoprotein VI agonist, convulxin, and the alpha B-2(1) integrin agonist, aggretin. CAPE (25 mu M) also markedly interfered with FITC-collagen binding to platelet membranes. CAPE (15 and 25 mu M) concentration-dependently inhibited collagen-induced platelet activation accompanied by [Ca+2]i mobilization, phosphomositide breakdown, activation of protein kinase C and mitogen-activated protein kinases (i.e., ERK2, JNK, and p38 MAPK), Akt phosphorylation, and thromboxane A(2) formation. In the ESR study, CAPE (15 and 25 mu M) markedly reduced hydroxyl radical (OH center dot) formation in collagen-activated platelets. In an in vivo study, CAPE (5 mg/kg) significantly prolonged the latency in inducing platelet plug formation in mesenteric venules of mice. Conclusions: The most important findings of this study suggest that CAPE specifically inhibits collagen-induced platelet activation. Thus, CAPE treatment may represent a novel approach to lowering the risk of or improving function in thromboembolism-related disorders. (c) 2007 European Society of Cardiology. Published by Elsevier B.V. All rights reserved.