Tanahashi N, Fukuuchi Y, Tomita M, Yokoyama M, Tomita Y, Inoue K, Schiszler I
Department of Neurology, School of Medicine, Tokyo, Japan.
Neurosci Lett. 1999 Oct 29;274(3):199-202. doi: 10.1016/s0304-3940(99)00703-x.
Employing video-enhanced contrast (VEC) microscopy and perfusion systems, we examined whether platelets adhere directly to human brain microvascular endothelial cells (HBEC) in vitro after thrombin treatment and whether adenosine diphosphate (ADP) or thromboxane A2-stimulated platelets adhere directly to HBEC at a low flow state in vitro. HBECs were cultured on a coverglass and put in the observation chamber of VEC microscopy. Following pretreatment with human alpha-thrombin 1.0 units/ml (n = 8) for 20 min, thrombin was thoroughly washed out. Platelet rich plasma (PRP) was perfused over HBEC at a low shear rate of 10 s(-1) for 30 min. Platelets adhered directly to thrombin-treated HBEC. Activated platelets by ADP (2 microM, n = 8) or thromboxane A2 (U-46619 10 microM, n = 5) were perfused over HBEC for 30 min and washed out. Platelets also adhered directly to HBEC. However, platelets did not adhere to HBEC when PRP only (n = 6) was perfused over HBEC for 30 min and washed out. Platelet adhesion directly to HBEC following thrombin treatment or platelet activation may play a pivotal role in secondary thrombus formation and microcirculatory disturbance in the ischemic brain.
我们采用视频增强对比（VEC）显微镜和灌注系统，研究了凝血酶处理后血小板在体外是否直接黏附于人脑微血管内皮细胞（HBEC），以及二磷酸腺苷（ADP）或血栓素A2刺激的血小板在体外低血流状态下是否直接黏附于HBEC。将HBEC培养在盖玻片上，并置于VEC显微镜的观察室中。用1.0单位/毫升的人α-凝血酶（n = 8）预处理20分钟后，彻底洗去凝血酶。以10秒-1的低剪切速率将富含血小板血浆（PRP）灌注到HBEC上30分钟。血小板直接黏附于经凝血酶处理的HBEC。用ADP（2微摩尔，n = 8）或血栓素A2（U-46619 10微摩尔，n = 5）激活的血小板灌注到HBEC上30分钟后洗去。血小板也直接黏附于HBEC。然而，仅将PRP（n = 6）灌注到HBEC上30分钟后洗去时，血小板不黏附于HBEC。凝血酶处理或血小板激活后血小板直接黏附于HBEC可能在缺血性脑的继发性血栓形成和微循环紊乱中起关键作用。
