ABSTRACT
OBJECTIVE Salvia miltiorrhiza bunge contains various active constituents, some of which have been developed as commercially available medicine. Moreover, some other ingredients in Salvia miltiorrhiza play great roles in anti-platelet activity. The aim of the present study was to investi-gate the effects and the underlying mechanism of miltirone,a lipophilic compound of Salvia miltiorrhiza Bunge. METHODS The ability of miltirone to modulate platelet function was investigated by a variety of in vitro and in vivo experiments.Platelet aggregation and dense granule secretion induced by various agonists were measured with platelet aggregometer.Clot retraction and spreading were imaged by digital camera and fl uorescence microscope. Ferric chloride-induced carotid injury model and pulmonary thromboembolism model were used to check miltirone effect in vivo. To elucidate the mechanisms of anti-platelet activity of miltirone,flow cytometry and Western blotting were performed. RESULTS Miltirone (2,4,8 μmol·L-1)was shown to suppress platelet aggregation,dense granule and α granule secretion in a dose-dependent manner. Meanwhile, miltirone inhibited the clot retraction and spreading of washed platelets.It reduced the phosphorylation of PLCγ2,PKC,Akt,GSK3β and ERK1/2 in the down-stream signal pathway of collagen receptor.It also reduced the phosphorylation of Src and FAK in the integrin αⅡbβ3 mediated"outside-in"signaling,while it did not suppress the phosphorylation of β3.In addition, miltirone prolonged the occlusion time and reduced collagen/epinephrine induced pulmonary thrombi. CONCLUSION Miltirone suppresses platelet "inside-out" and "outside-in" signaling by affecting PLCγ2/PKC/ERK1/2, PI3K/Akt and Src/FAK signaling. Therefore, miltirone might represent a potential anti-platelet candidate for the prevention of thrombotic disorders.