Dietary phytoestrogens and their synthetic structural analogues as calcium channel blockers in human platelets

Phytoestrogens have been shown to inhibit platelet activation by blocking platelet calcium channels. This study examined the effect of several synthetic derivatives of trans-resveratrol, genistein, and daidzein on platelet free intracellular calcium ([Ca2+](i)) elevation in thrombin-activated platelets and the possible mechanisms of this inhibitory effect. Studies were conducted on fresh human platelets from healthy volunteers. The fluorescent dye fura-2 was used to monitor [Ca2+](i) in platelets. At 10 muM trans-resveratrol, triacetyl-trans-resveratrol, and trimethoxy-trans-resveratrol produced, respectively, 57 +/- 4%, 40 +/- 4%, and 21 +/- 1% inhibition; genistein, acetylgenistein, and dihydrogenistein produced 51 +/- 10%, 26 +/- 7%, and 16 +/- 2% inhibition, respectively; daidzein and diacetyldaidzein produced 56 +/- 5% and 45 10% inhibition of thrombin-induced [Ca2+](i) elevation. The inhibitory effect was immediate and appeared to directly affect the calcium influx channels. Phytoestrogen action on [Ca2+](i) did not cause alteration in nitric oxide signaling. Tyrosine phosphorylation was not involved in the inhibition of [Ca2+](i) elevation by phytoestrogens, because the percent inhibition produced by the tyrosine kinase inhibitor genistein and its inactive analogue daidzein on thrombin-induced and thapsigargin-induced [Ca2+](i) elevation was not significantly different for either compound at any concentration tested. Structure-activity relationship studies on this limited set of compounds reveal the requirements for the stilbene pharmacophore for the calcium-blocking activity.