Kinetic Studies of Intravascular Thrombus Formation Using Newly Developed Thrombus Model in Microvessels

在我们的实验室中发现，在激发光照射下结合荧光染料在微血管中可诱导血小板血栓形成。我们已经报道了这种新的血小板血栓模型对体内血液凝固过程的研究和新开发的抗血栓药物的评价是有用的。然而，到目前为止，机制的细节尚不清楚。本研究采用该血栓形成模型，在大鼠肠系膜微血管中检测血小板血栓形成动力学。血流速度在血栓形成后不久就开始增加，并随着血栓的增大而逐渐增加，尤其是在径向上。在达到几乎恒定的狭窄水平后，速度急剧下降。这个临界区域狭窄几乎是一个恒定的值，即75- 95%的管腔面积。其他实验也对几种药物的抗血栓作用进行了评价。前列腺素（PGE1, PGD2, PGI2）抑制血小板血栓的生长呈剂量依赖性。由于激发的彩色敏化剂（染料）预计会与氧相互作用产生活性氧，因此研究了活性氧对血栓形成的影响。超氧自由基和单线态氧清除剂对延长血栓形成时间有显著作用。此外，从电镜观察来看，粘附在内皮细胞上的白细胞可能与血栓形成有关。这些结果表明，光化学反应产生的活性氧在诱导内皮细胞、血小板和/或白细胞的细胞膜损伤中起决定性作用。由于这些损伤，血小板似乎开始粘附在内皮细胞上并相互聚集。

It was found in our laboratory that the platelet thrombi were inducible in microvessels by the irradiation of exciting light in combination with the intravascular administration of fluorescent dye. We have reported that this new platelet thrombus model is useful for the studies on blood coagulation processes in vivo and the evaluation of newly developed antithrombotic agents. However, the details of the mechanisms are not clear so far. In this research project, the kinetics of the platelet thrombus formation was examined in microvessels of the rat mesentery, using this thrombus formation model. The blood flow velocity began to increase soon after the initiation of thrombus formation and continued to increase progressively with the growth of the thrombus particularly in the radial direction. The velocity precipitously dropped after reaching an almost constant level of stenosis. This critical area stenosis had almost a constant value, i.e., 75-95 % of the lumen area. The other experiments were also carried out to evaluate the antithrombotic effects of a few drugs. The growth of platelet thrombi was inhibited dose-dependently by the administrations of prostaglandins (PGE1, PGD2, PGI2). Since excited colored sensitizers (dyes) are expected to interact with oxygen to produce active oxygens, the effects of active oxygens on the thrombus formation were examined. The scavengers of superoxide radiacal and singlet oxygen had significant effects to prolong the thrombus formation times. Furthermore, from the electron micoscopic observations, the leucocytes adhered to the endothelial cells were supposed to have relevance to the thrombogenesis. It was suggested from these results that the active oxygens produced by photochemical reaction play decisive roles to induce damage of the cell membranes of the endothelium, platelet and/or leucocyte. Due to these damages the platelets seemed to begin to adhere to the endothelium and aggregate each other.