三七总皂苷抑制剪切诱导血小板聚集及血栓形成的生物力药理学分析  

Biomechanopharmacological Study of Panax notoginseng Saponins on High Shear-induced Platelet Aggregation and Thrombosis

作　　者：王贻琳 李佳 刘陆 宫平 许静 廖福龙[1] 游云[1] WANG Yilin;LI jia;LIU Lu;GONG Ping;XU Jing;LIAO Fulong;YOU Yun(Institute of Chinese Materia Medica,China Academy of Chinese Medical Sciences,Beijing 100700,China)

机构地区：[1]中国中医科学院中药研究所,北京100700

出　　处：《中国实验方剂学杂志》2024年第23期111-120,共10页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金面上项目(82274244);中国中医科学院科技创新工程项目(CI2021A04616);中国中医科学院科技创新团队项目(CI2021B015)。

摘　　要：目的:从剪切诱导压电型机械敏感离子通道组件1(Piezo1)介导的钙离子信号通路探讨三七总皂苷(PNS)干预血小板聚集和血栓形成的作用机制。方法:采用Bioflux1000z微流控装置,对血小板施加生理(500 s^(-1))及病理高剪切率(12000 s^(-1)),或在生理剪切率(500 s^(-1))下使用Piezo1激动剂Yoda1刺激血小板活化。流式细胞术检测剪切诱导血小板钙离子内流,血小板与血管性血友病因子(vWF)结合;酶联免疫吸附测定法检测剪切诱导血小板vWF释放;微流通道法检测高剪切下vWF介导的血小板聚集及整合素αⅡbβ3活化。建立了高剪应力联合内皮损伤致小鼠动脉血栓模型,采用超声多普勒血流仪监测血栓反复形成及脱落而导致的周期性血流降低(CFR),流式细胞术检测血小板与血浆vWF结合,评价PNS对高剪切诱导的体内动脉血栓形成的影响。结果:在体外微流控实验中,PNS以浓度依赖方式降低病理高剪切率(12000 s^(-1))或Yoda1诱导的血小板钙离子内流、血小板与vWF结合、vWF介导的血小板与纤维蛋白原结合及血小板α颗粒vWF的释放。在高剪切诱导的小鼠动脉血栓模型中,PNS能够显著降低颈总动脉的周期性血流降低及血管闭塞时间,抑制血小板与vWF的结合。结论:PNS可抑制高剪切诱导的Piezo1介导的钙离子内流,进而影响血小板与vWF结合及整合素αⅡbβ3激活,改善病理高剪切诱导的血小板聚集和动脉血栓形成。Objective:To investigate the mechanisms of Panax notoginseng saponins(PNS)in inhibiting high shear-induced platelet aggregation and thrombosis via the Piezo1-mediated calcium signaling pathway.Method:Bioflux1000z was used for the microfluidic assay,where platelets were stimulated with physiological shear rate(500 s^(-1)),pathological shear rate(12000 s^(-1)),or Piezo1 agonist Yoda1 under the physiological shear rate(500 s^(-1)).The shear-induced platelet calcium influx and the binding of platelet with von Willebrand factor(vWF)were measured by flow cytometry.Enzyme-linked immunosorbent assay(ELISA)was employed to measure the vWF release from platelets.The microfluidic channels were used to determine the vWFmediated platelet aggregation and integrinαⅡbβ3 activation.A mouse model of arterial thrombosis induced by high shear stress combined with endothelial injury was established.The ultrasonic Doppler flow meter was used to monitor the cyclic flow reduction(CFR)caused by the repeated formation and shedding of thrombi,and flow cytometry was employed to examine platelet-vWF binding,on the basis of which the effect of PNS on high shear-induced arterial thrombosis was evaluated.Result:The microfluidic assay showed that PNS decreased the high shear rate(12000 s^(-1))or Yoda1-induced calcium influx,platelet-vWF binding,vWF-mediated plateletfibrinogen binding,and vWF release from platelet alpha-granules in a dose-dependent manner.In the mouse model of high shear-induced thrombosis,PNS markedly reduced the CFR and occlusion time of the common carotid artery and inhibited platelet-vWF binding.Conclusion:PNS can mitigate pathological shear-induced platelet aggregation and arterial thrombosis via influencing Piezo1/GPIbα-vWF signaling.

关 键 词：动脉血栓 剪应力 血小板 Piezo1 血管性血友病因子(vWF) 三七总皂苷 

分 类 号：R284.2[医药卫生—中药学] R285[医药卫生—中医学] R289R287R22R2-031R33R24