Antiplatelet aggregation of naringenin via cyclic nucleotide signaling: in vitro studies / 中国病理生理杂志

ABSTRACT

AIM: To investigate effect of naringenin on ADP-induced platelet aggregation and its possible mechanism.METHODS: The levels of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) were measured in the platelets with ADP stimulation using ELISA in the presence or absence of different concentrations of naringenin.The effect of naringenin at different concentrations on the change of phosphodiesterase (PDE) activity was measured by high efficiency liquid chromatography.The effects of naringenin at different concentrations on phosphorylation of vasodilator-stimulated phosphoprotein (VASP) at positions Ser157 and Ser239 in washed platelets with ADP stimulation were analyzed by Western blot.The phosphorylation of VASP at Ser239 was also analyzed in the presence of protein kinase A (PKA), protein kinase G (PKG), or protein kinase C (PKC) inhibitors before incubation with naringenin.The platelet aggregation was measured in the presence of PKA or PKG inhibitors before incubation with naringenin.RESULTS: Naringenin elevated cGMP levels significantly but not cAMP levels in the platelets with ADP stimulation in a dose-dependent manner.Naringenin inhibited PDE activity.Naringenin increased the phosphorylation of VASP at Ser239 in a dose-dependent manner in the platelets with ADP stimulation but only modest changes in the phosphorylation at position Ser157.The phosphorylation level of VASP at Ser239 position was inhibited when the platelets were treated with PKA inhibitor before incubation with naringenin.Incubation of platelets with neither PKG nor PKC inhibitors before treatment with naringenin affect the phosphorylation of VASP at Ser239.Pretreatment with PKA inhibitor but not PKG inhibitor significantly reversed the antiplatelet aggregation by naringenin in ADP-stimulated platelets.CONCLUSION: Naringenin may inhibit platelet activation through the elevation of cGMP-and PKA-mediated VASP phosphorylation.