摘要
目的发展一种可用于评估抗血小板药物对病理性高剪切诱导的血小板活化和聚集抑制效果的微流控芯片分析方法。方法采用软光刻工艺加工聚二甲基氧烷(polydimethylsiloxane,PDMS)-玻璃微通道芯片。将抗凝人外周全血以1500 s^(-1)剪切率流过微通道芯片,应用荧光倒置显微镜拍摄血小板在狭窄通道下游的聚集行为,通过图像分析得到血小板聚集覆盖率、聚集总面积、血栓平均尺寸和血栓数量。收集流过微流控芯片的全血,通过流式细胞技术分析血小板表面活化标志物(P-选择素和PAC-1)的表达,并评估抗血小板药物(阿司匹林、替格瑞洛、替罗非班)对狭窄下游血小板聚集和活化的抑制能力。结果随着输入剪切率的提高,血小板聚集能力逐渐增强。阿司匹林不能抑制病理性高剪切诱导的血小板活化和聚集,替格瑞洛和替罗非班能够显著抑制血小板活化和聚集。结论本研究发展的微流控芯片模型,可用于模拟动脉狭窄血管的局部特征,研究不同剪切力梯度对血小板聚集功能的影响,并在病理性高剪切力条件下评估抗血小板药物对血小板活化和聚集功能的抑制效能。区别于传统的比浊法及血栓弹力图仪等在静止条件下对血小板功能的检测,本研究结果为临床提供了一种更接近动脉血栓性疾病患者体内真实流动环境的体外模型,为经皮冠状动脉介入术后或动脉粥样硬化患者的血小板功能检测提供相关的分析方法。
Objective To develop a microfluidic chip analysis method which can be used to evaluate the inhibitory effect of antiplatelet drugs on platelet activation and aggregation induced by pathological high shear.Methods Polydimethoxane(PDMS)-glass microchannel chip was fabricated by soft lithography.The anticoagulant human whole blood was flowed through the microchannel chip at 1500 s^(-1) shear rate,and the platelet aggregation behavior downstream of the narrow channel was photographed by fluorescence inversion microscope.The platelet aggregation coverage rate,total aggregation area,average size of thrombus and number of thrombus were obtained by image analysis.The whole blood flowing through the microfluidic chip was collected and the expression of platelet surface activation markers(P-selectin and PAC-1)was analyzed by flow cytometry.The 931 inhibitory effect of antiplatelet drugs(aspirin,tirofiban,tirofiban)on platelet aggregation and activation downstream of stenosis was also evaluated.Results With the increase of input shear rate,the ability of platelet aggregation increased gradually.Aspirin could not inhibit platelet activation and aggregation induced by pathological high shear,while tigrel and tirofiban could significantly inhibit platelet activation and aggregation.Conclusions The microfluidic chip model developed in this study can be used to simulate the local characteristics of arterial stenosis and study the effects of different shear force gradients on platelet aggregation,as well as to evaluate the inhibitory effect of antiplatelet drugs on platelet activation and aggregationunder pathological high shear stress.Different from the traditional turbidimetry and thromboelastography in the detection of platelet function under static conditions,this study provides an in vitro model closer to the real flow environment in patients with arterial thrombotic diseases,as well as a related analysis method for the detection of platelet function in patients with percutaneous coronary intervention(PCI)or atherosclerosis.
作者
张天聪
黄小静
高雪梅
宦宣容
何翠
李远
ZHANG Tiancong;HUANG Xiaojing;GAO Xuemei;HUAN Xuanrong;HE Cui;LI Yuan(Central Laboratory,Yongchuan Hospital Affiliated to Chongqing Medical University,Chongqing 402160,China)
出处
《医用生物力学》
CAS
CSCD
北大核心
2023年第5期931-937,共7页
Journal of Medical Biomechanics
基金
国家自然科学基金项目(11702047)
重庆市社会事业与民生保障科技创新专项(cstc2017shmsA130009)
重庆市科卫联合科研项目中青年医学高端人才项目(2023GDRC008)
重庆市博士后研究项目(Xm2017082)。
