三七总皂苷对ox-LDL诱导血小板凋亡的保护作用及机制研究

The Protective Effect and Mechanism of Panax notoginseng Saponins on ox-LDL-Induced Platelet Apoptosis

目的:探讨三七总皂苷(PNS)对体外实验中氧化型低密度脂蛋白(ox-LDL)诱导的血小板凋亡的抑制作用及潜在的分子机制。方法:将健康人纯化血小板分别与不同浓度的PNS(1、10、100μg/mL)共同孵育40 min,然后用50μg/mL ox-LDL刺激血小板60 min后,用流式细胞术检测血小板凋亡的指标包括血小板线粒体膜电位(Δψ_(m))水平和磷脂酰丝氨酸(PS)的暴露水平;用活性氧(ROS)检测试剂盒测定胞内总ROS水平和超氧化物水平;用蛋白质印迹法检测血小板Src、Syk蛋白的磷酸化水平以及Caspase-3和Caspase-9活化水平。结果:与对照组相比,PNS各剂量组显著降低ox-LDL诱导的血小板Δψ_(m)去极化(P<0.01),PNS中、高剂量组能抑制ox-LDL诱导的血小板表面PS的暴露水平(P<0.05)、总ROS水平和超氧化物水平(P<0.05)、Caspase-3和Caspase-9的活化水平(P<0.05)以及Src、Syk蛋白的磷酸化水平(P<0.05)。结论:PNS可能通过抑制Src/Syk/ROS信号通路发挥抗ox-LDL诱导血小板凋亡的作用。

Objective:To explore the effect of Panax notoginseng saponins(PNS)on oxidized low-density lipoprotein(ox-LDL)-induced platelet apoptosis and its potential molecular mechanism in vitro.Methods:Human gel-filtered platelets were pre-treated with different concentrations of PNS(0,1,10,100μg/mL)for 40 minutes,and then stimulated with 50μg/mL ox-LDL for 60minutes.Subsequently,indicators of platelet apoptosis including platelet mitochondrial membrane potential(Δψ_(m))levels and phosphatidylserine(PS)exposure levels were detected by flow cytometry.Reactive oxygen species(ROS)levels and superoxide levels were measured by commercial ROS assay kit.The phosphorylation levels of Src and Syk proteins as well as the activation levels of Caspase-3 and Caspase-9 were detected by Western blotting.Results:Compared with the control group,ox-LDL-induced depolarization of plateletsΔψ_(m)was significantly reduced in each dose group of PNS(P<0.01),ox-LDL-induced exposure level of platelet surface PS(P<0.05),total ROS levels and superoxide levels(P<0.05),Caspase-3 and Caspase-9 activation levels(P<0.05),and phosphorylation levels of Src and Syk proteins(P<0.05)were inhibited in the medium and highdose PNS groups.Conclusion:PNS may play the role of anti ox-LDL-induced platelet apoptosis by inhibiting Src/Syk/ROS signalling pathway.