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Platelet adhesion to human brain microvascular endothelial cells in vitro

血小板对人脑微血管内皮细胞的体外粘附

基本信息

批准号：
12670617
负责人：
TANAHASHI Norio
金额：
$ 2.18万
依托单位：
Keio University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12670617/
关键词：
platelet adhesion endothelial cell Argatroban Beraprost GPIIb IIIb antagonist beraprost humanbrain microvascular endothelial cell thrombin platelet aggregation

项目摘要

We examined whether activated platelets adhere to human Brain microvascular endothelial cells (HBEC) in vitro and whether argatroban (selective thrombin inhibitor) or GPIIb/IIIa antagonist (TAK-029) ameliorates the adhesion of activated platelets to HBEC in vitro using VEC-DIC microscopy. In the control group (N=5), HBECs were cultured on a coverglass and put in the observation chamber of VEC-DIC microscopy. Then, platelet rich plasma (PRP) was superfused with an infusion pump at shear rates (10/sec) for 30 min and platelet adhesion to HBEC was observed. In the ADP group (N=9), PRP with ADP (2μM) was superfused for 30 min and washed out. In the argatroban group (N=9) PRP with ADP (2μM) and argatroan (5μg/ml) was superfused and platelet adhesion to HBEC was observed. In the TAK-029 group (N=9), PRP with ADP (2μM) plus TAK-029 (GPIIb/IIIa antagonist; 1μM) was superfused. In the control group, platelet adhesion to HBEC was rarely seen. In the ADP group, platelets adhered to HBEC in all experiments and microaggregates of platelets were seen. In the argatroban and TAK-029 groups, platelet adhesion to HBEC was clearly suppressed. The average number of platelets adhering and aggregating to HBEC was 0.3 ± 0.6/900 μm^2 in the control group, 25.5 ± 11.3/ 900μm^2 (P<0.01, vs control) in the ADP group, 1.8 ± 1.8/ 900 μm^2 in the argatroban group (p<0.01, vs ADP) and 1.6 ± 1.8/ 900 μm^2. The above results showed argatroban and TAK-029 ameliorated adhesion and aggregated pileup of activated platelets to HBEC at a low-flow state in vitro. The above results showed adhesion of ADP-activated platelets to HBEC under flow in vitro is mediated via GPIIb/IIIa. The results also suggests that thrombin produced by platelet activation makes HBEC procoagulant and is the most likely candidate to subsequently induce platelet adhesion to HBEC.

我们使用VEC-DIC显微镜检查了体外活化血小板是否粘附于人脑微血管内皮细胞（HBEC），以及阿加曲班（选择性凝血酶抑制剂）或GPIIb/IIIa拮抗剂（TAK-029）是否改善体外活化血小板与HBEC的粘附。对照组（N=5）将HBECs培养于盖玻片上，置于VEC-DIC显微镜观察室中。然后，用输注泵以剪切速率（10/sec）灌注富血小板血浆（PRP）30 min，观察血小板与HBEC的粘附。ADP组（N=9），灌流含ADP（2μM）的PRP 30 min后冲洗。阿加曲班组（N=9）灌流含ADP（2μM）和阿加曲安（5μg/ml）的PRP，观察血小板与HBEC的粘附。在TAK-029组（N=9）中，灌流PRP + ADP（2μM）+TAK-029（GPIIb/IIIa拮抗剂; 1μM）。在对照组中，很少见到血小板粘附于HBEC。ADP组在所有实验中均可见血小板粘附于HBEC，并可见血小板微聚集体。阿加曲班和TAK-029组中，血小板与HBEC的粘附明显受到抑制。粘附和聚集到HBEC上的血小板平均数量在对照组为0.3 ± 0.6/900 μm^2，ADP组为25.5 ± 11.3/ 900μm^2（与对照组相比，P<0.01），阿加曲班组为1.8 ± 1.8/ 900 μm^2（与ADP相比，P<0.01），ADP组为1.6 ± 1.8/ 900 μm^2。上述结果表明，阿加曲班和TAK-029在体外改善了低流量状态下活化血小板与HBEC的粘附和聚集堆积。上述结果表明，ADP活化的血小板在体外流动条件下与HBEC的粘附是通过GPIIb/IIIa介导的。结果还表明，血小板活化产生的凝血酶使HBEC促凝，是最有可能的候选人，随后诱导血小板粘附HBEC。