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 microM) 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 anti-platelet activity of CAPE. Fluorescein sodium-induced platelet thrombi in mesenteric microvessels of mice were used for an in vivo study.


RESULTS
CAPE (15-100 microM) 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 microM) also inhibited platelet aggregation stimulated by the glycoprotein VI agonist, convulxin, and the alpha(2)beta(1) integrin agonist, aggretin. CAPE (25 microM) also markedly interfered with FITC-collagen binding to platelet membranes. CAPE (15 and 25 microM) concentration-dependently inhibited collagen-induced platelet activation accompanied by [Ca(+2)](i) mobilization, phosphoinositide 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 microM) markedly reduced hydroxyl radical (OH) 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.
Hsiao
George
G
Graduate Institute of Pharmacology, Taipei Medical University, Taipei, Taiwan.
Lee
Jie J
JJ
Lin
Kuang H
KH
Shen
Chia H
CH
Fong
Tsorng H
TH
Chou
Duen S
DS
Sheu
Joen R
JR
D000818:Animals; D001681:Biological Assay; D001792:Blood Platelets / Q000378:metabolism*; D002109:Caffeic Acids / Q000494:pharmacology*; D003094:Collagen / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000494:pharmacology; D003435:Crotalid Venoms / Q000494:pharmacology; D003864:Depression, Chemical; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D006801:Humans; D017665:Hydroxyl Radical; D015151:Immunoblotting / Q000379:methods; D038982:Integrin alpha2beta1 / Q000819:agonists; D037181:Lectins, C-Type; D008638:Mesenteric Arteries; D051379:Mice; D010626:Phenylethyl Alcohol / Q000031:analogs & derivatives* / Q000494:pharmacology; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology*; D010980:Platelet Membrane Glycoproteins / Q000819:agonists; D013050:Spectrometry, Fluorescence; D013927:Thrombosis; D013929:Thromboxane B2 / Q000032:analysis / Q000378:metabolism; D014757:Viper Venoms / Q000494:pharmacology
D002109:Caffeic Acids; D003435:Crotalid Venoms; D038982:Integrin alpha2beta1; D037181:Lectins, C-Type; D010975:Platelet Aggregation Inhibitors; D010980:Platelet Membrane Glycoproteins; D014757:Viper Venoms; C098341:platelet membrane glycoprotein VI; C112292:rhodocytin protein, Calloselasma rhodostoma; D017665:Hydroxyl Radical; C028890:convulxin; D013929:Thromboxane B2; D003094:Collagen; C055494:caffeic acid phenethyl ester; D010626:Phenylethyl Alcohol