一种用于溶栓研究的大鼠血栓栓塞性卒中模型的改进

Improvement of a rat thromboembolic stroke model for thromboly sis study

目的建立并验证一种改进的大鼠血栓栓塞性卒中模型。方法60只Sprague-Dawley大鼠取股动脉血与凝血酶混匀后注入PE-50导管制备体外血栓，通过右侧颈外动脉插入导管并将栓子注入颈内动脉，建立血栓栓塞性脑缺血模型。30只大鼠随机分为大量栓子组（12个栓子，n=10）、中等量栓子组（10个栓子，n=10）和少量栓子组（8个栓子，n=10），注入栓子后2h进行神经功能评分，比较各组模型制作成功率；注入栓子后24h处死大鼠取脑进行2,3，5-氯化三苯基四氮唑染色，评价梗死后出血、梗死体积、出血发生率和死亡率。选择模型制作成功率高的栓子组随机分为生理盐水组（n=12）和重组组织型纤溶酶原激活剂（recombinant tissue-type plasminogen activator，rtPA）组（n=12），分别在栓子注入后3h给予生理盐水和rtPA，血栓注入前以及血栓注入后2、6、12和24h分别进行神经功能评分；栓子注入后24h处死大鼠取脑进行2,3，5-氯化三苯基四氮唑和伊文思蓝染色，评价出血发生率、梗死体积、水肿程度以及血脑屏障通透性。结果少量栓子组仅40％出现神经功能缺损，梗死体积仅为（10．54±2．82）％。中等量栓子组与大量栓子组成功率分别为80％和100％，均显著性高于少量栓子组（P=0．011），梗死体积亦显著性大于少量栓子组（F=40．897，P=0．000）。大量栓子组给予rtPA后大鼠平均生存时间少于24h，故选择中等馈栓子组进行rtPA溶栓效果研究。rtPA组梗死体积和神经功能评分均较生理盐水组显著改善（t=7．728，P=0．000），而两组间出血发生率、脑水肿程度和血脑屏障通透性无显著性差异。结论经过改良的注入10个栓子的血栓栓塞性脑缺血模型的稳定性和可重复性良好，且经溶栓后神经功能显著改善，适用于脑缺血病理生理学和溶栓治疗的实验研究。

Objective To establish and validate a modified rat thromboembolic stroke model. Methods After taking femoral arterial blood and mixing it with thrombin, they were injected into PE-50 catheter for preparing in vitro thrombosis in 60 Sprague-Dawley rats. A thromboembolic cerebral ischemia model induced by catheterization of the risht enternal carotid artery and the small blood clot emboli were injected into the internal carotid arteries. Thirty rats were randomly divided into a large number of emboli group （n = 10 with 12 emboli）, a median number of emboli group （n = 10 with 10 emboli） and a small number of emboli group （n = 10 with 8 emboli）. Two hours after embolus injection, the neurological deficit score was performed and the success rate of the model was compared in all groups. Twenty-four hours after embolus injection, the rats were sacrificed and the brains were removed for 2, 3, 5-triphenyltetrazolium chloride staining. The hemorrhage, infarct volume, bleeding incidence and mortality after cerebral infarction were evaluated. The high success rates of the modeling in the emboli groups were selected and they were randomly divided into either a normal saline group （n = 12） or a recombinant tissue-type plasminogen activator （rtPA） group （n = 12）. The rats were given normal saline and rtPA at 3 hours after embolus injection. Before embolus injection and 2, 6, 12 and 24 hours after embolus injection, the neurological scores were performed respectively; 24 hours after embolus injection, the rats were sacrificed and the brains were removed for 2,3,5-triphenyltetrazolium chloride staining. The hemorrhage rate, infarction size, degree of cerebral edema, and blood-brain barrier permeability were evaluated. Results Only 40% of rats had neurological deficits in the small number of emboli group, and the infarct volume was only 10.54 ±2. 82%. The success rates in the median and large number of emboli groups were 80% and 100% respectively. They were all significantly higher than those in the small number of emboli group （P = 0. 011 ）. The infarct volume was also significantly greater than that in the small number of emboli group （F = 40. 897, P = 0. 000）. After administration of rtPA, the mean survival time of the rats in the large number of emboli group was less than 24 hours, so the median number of emboli group was selected to study the thrombolytic effect of rtPA. The infarct volume and neurological function score in the rtPA group were improved significantly compared to the normal saline group （t = 7. 728, P = 0. 000）, while there were no significant differences in the hemorrhage rate, degree of brain edema and blood-brain barrier permeability between the 2 groups. Conclusions The stability and reproducibility were good in the modified thromboembolic cerebral ischemia model injected with 10 emboli, the neurological fimction was improved significantly after thrombolysis, and it was applicable to the experimental study of pathophysiology of cerebral ischemia and thrombolytic therapy.