The Antiplatelet Activity Of Chrysin And Its Antithrombotic Effect

Part Ⅰ The effects of chrysin on mouse platelet functionAims:Inappropriate activation of platelets under some pathological conditions leads to thrombosis, which result in myocardial infarction or stroke. Chrysin has been reported to possess antiplatelet activity. However, it is not clear whether it has anti-thrombus effect or not, and the mechanism by which it inhibits platelet function. As platelets play pivotal roles in cardiovascular diseases, we studied the effect of chrysin on platelet activities/aggregation/spreading and underlying mechanisms.Methods:The effects of chrysin on agonist-induced platelet aggregation, granule secretion, integrin-αIIbβ3activation, platelet adhesion to immobilized collagen and fibrinogen, and clot retraction were examined. In order to reveal the mechanism by which chrysin exerts its effects, PI3K/Akt and MAPK expressions, and levels of protein tyrosine phosphorylation of the GPVI signaling pathway in the presence or absence of chrysin were inspected.Results:Chrysin dose-dependently inhibited platelet aggregation and granule secretion induced by collagen and thrombin, and inhibited platelet aggregation induced by ADP, thrombin and U46619. Pretreatment of platelet with chrysin markedly reduced number of adherent platelets and single platelet spreading area on immobilized collagen and fibrinogen, and also reduced clot retraction. Chrysin attenuated not only the activation of Syk, PLCy2and PKC, but also the phosphorylation of Akt and ERKl/2in platelets stimulated by collagen. Chrysin and LY294002demonstrated an additive inhibition on aggregation when combined.Conclusions:This study demonstrates for the first time that chrysin possesses antithrombosis activity, which may be mediated by inhibiting the PI3K/Akt-GSK3β/eNOS and Syk-PLCy2-PKC cascades. Part Ⅱ The effects of chrysin on human platelet functionAims:To further confirm the effect of chrysin on human platelet and try to find the mechanism of antiplatelet effect.Methods:The effects of chrysin on agonist-induced platelet aggregation, granule secretion, integrin-αⅡbβ3activation, platelet adhesion to immobilized fibrinogen, and clot retraction were examined. In order to reveal the mechanism by which chrysin exerts its effects, PI3K/Akt and MAPK expressions, and levels of protein tyrosine phosphorylation of the GPVI signaling pathway or platelet spreading mediated outside-in signaling pathway in the presence or absence of chrysin were inspected.Results:Chrysin dose-dependently inhibited platelet aggregation by ADP, collagen, thrombin and U46619and chrysin inhibited granule secretion induced by collagen. Pretreatment of platelet with chrysin markedly reduced number of adherent platelets and spreading area on immobilized fibrinogen, and also reduced clot retraction. Chrysin attenuated not only the activation of PLCy2and PKC, but also the phosphorylation of Akt and ERKl/2in platelets stimulated by collagen. Chrysin and LY294002/PP2demonstrated an additive inhibition on aggregation when combined. Chrysin could inhibit platelet FcyRIIa, Akt and GSK3β phosphorylation of platelet spreading mediated outside-in signaling moleculars.Conclusions:This study demonstrates that chrysin possesses antiplatelet activity, which may be mediated by inhibiting the PI3K/Akt-GSK3β and SFKs-PLCγ2-PKC cascades and inhibits "inside-ouf" and "outside-in"signaling pathway。 PartⅢ The effects of chrysin on thrombosis and coagulation system in vivoAims:To further observed the effect of chrysin on platelet in vivo.Methods:With collagen-epinephrine-induced acute pulmonary thrombus formation mouse model, the effect of chrysin on thrombosis in vivo was also investigated. Chrysin effects in ex vivo coagulation time, platelet aggregation and bleeding time in mice were also determined.Results:In collagen-epinephrine-induced acute pulmonary thrombus mouse model, chrysin, at80mg/kg, significantly attenuated thrombus formation and mortality. Chrysin significantly and dose-dependently, inhibited collagen-induced platelet aggregation. Chrysin extends bleeding time in mice but it did not affect the coagulation time.Conclusions:Chrysin inhibits thrombosis formation by affecting the functions of platelet.