The Anti-platelet Effect Of Loureirin A/B And Its Mechanism

Part ⅠThe Antiplatelet Effects of Loureirin A/BObjective: Dragon’s blood, the Chinese traditional drug, has the effect of promoting bloodcirculation to remove blood stasis, eliminating edema and relieving pain. Modernresearches found that dragon’s blood could activate blood and stop bleeding, ease pain,exhibit anti-inflammatory and antiplatelet effect. The flavonoids are the mainantithrombotic compound in dragon’s blood containing Lourerin A and Loureirin B. TheCollege of Life Science in Shanghai University found that Lourerin A and Loureirin Bcould highly inhibit ADP (adenosine diphosphate) induced human platelet aggregation，upto80%, but we still don’t know the effects of Lourerin A and Loureirin B onother agonistsinduced platelet aggregation such as collagen, CRP (collagen related peptide) and thrombinor other platelet functions as well as the mechanism. Our researches mainly discussed theantiplatelet effect of Lourerin A and Loureirin B together with their mechanism.Methods: We detected different concertrations of Lourerin A and Loureirin B on mouseplatelet aggregation, P-selectin expression and human platelet spreading on immbolisedfibrinogen with platelet aggregational and secretional detector, flow cytometry andfluorescence microscope.Results: Both of Lourerin A and Loureirin B could apparently inhibite mouse plateletaggregation induced by collagen, CRP, ADP and thrombin, both in a dose-dependent way. Lourerin A could strongly reduce collagen induced mouse platelet aggregation,100μMLourerin A decrease platelet aggregation by84.8%，but weak in turn on CRP, ADP andthrombin stimulated platelet aggregation. The effect of Loureirin B on collagen and CRPstimulated platelet aggregation were similar to the effect of Lourerin A on collagen inducedplatelet activation, equally weak in ADP and thrombin stimulated platelet aggregation.Meanwhile, both Lourerin A and Loureirin B inhibited thrombin induced platelet P-selectinexpression, while in CRP stimulated P-selectin expression, only Lourerin A not Loureirin Bcould apparently inhibit the process. Additionally, Lourerin A could remarkably inhibitmouse platelet ATP secretion and human platelet’s spreading on immbolised fibrinogen.Conclusion: Lourerin A and Loureirin B apparently inhibit collagen, CRP, ADP andthrombin stimulated platelet aggregation, Lourerin A and Loureirin B also remarkablyreduce thrombin stimulated platelet P-selectin expression, additionally, Lourerin Adramatically affects mouse platelet ATP secretion and human platelet spreading onimmbolised fibrinogen. Part ⅡThe Antiplatelet Mechanism of Lourerin A/BObjective: To illustrate the antiplatelet mechanism of Lourerin A and Loureirin B.Methods: Incubate washed platelet with Lourerin A and Loureirin B for5min, stimulatewith collagen, and then split it. Western blot detecte the phosphorylation of platelet Akt（Ser473/474）, Src（Tyr416）and JNK（Thr183/Tyr185）, and to know the Aktphosphorylation change at different time after giving agonist as well as its phosphorylationwhen giving Lourerin A together with pan PI3K inhibitor LY294002.Results: Lourerin A and Loureirin B could apparently inhibit platelet Akt（Ser473/474）phosphorylation when stimulated with collagen, weakly affect Src （Tyr416）phosphorylation, and hardly have any effect on JNK（Thr183/Tyr185） phosphorylation,both of Lourerin A and Loureirin B have similar effect. Akt phosphorylation appeared to betime-dependent but giving Loureirin A dramatically abrogated the process.Additionanly,pan PI3K inhibitor LY294002could enhance its inhibiting role.Conclusion: Lourerin A and Loureirin B may inhibit platelet activation by affectingPI3K/Akt signaling.