不同类型栓子所致急性缺血大鼠脑组织形态学变化及其蛋白表达差异的研究

目的 探讨不同类型栓子所致急性缺血大鼠脑组织形态学变化及其蛋白表达的差异.方法 健康无特定病原体雄性Sprague-Dawley(SD)大鼠120只.粉碎胆固醇晶体,制备胆固醇栓子.取大鼠自体动脉血,凝固烘干后,制成血栓栓子.栓子直径为60～100μm.采用改进的微栓子栓塞法(少量多次注射栓子代替传统的一次性注射)制作...

Histomorphological and protein expression changes in acute ischemic stroke rats induced by different types of emboli

Objective To investigate the histomorphological changes and protein expression differences of acute ischemic stroke rats caused by different types of emboli.Methods A total of 120 healthy specific pathogen free male Sprague-Dawley(SD)rats were used.Cholesterol emboli was prepared by crushing cholesterol crystals.Autologous arterial blood was taken from rats,coagulated and dried to make thromboemboli.The diameter of embolus ranged 60-100μm.The rat model of acute ischemia was made by the improved microembolus embolization method(multiple injections of small number of emboli instead of the traditional one-time injection).The Longa behavioral score of 1 to 4 was a successful modeling with a total of 105 rats successfully modeled.The 105 rats were randomly divided into cholesterol embolus group(n=35),thromboembolus group(n=35)and sham operation group(n=35).Longa behavioral score,edema rate and ischemic area were compared between 8 rats in cholesterol embolus group and 8 rats in thromboembolus group at 6 and 24 hours after successful establishment of ischemic model.Evans blue and triphenyltetrazolium chloride staining was performed in the ischemic cortical tissue of 8 rats in each of the three groups to assess blood-brain barrier injury and observe the ischemic area of cortical tissue.Twenty four hours after the establishment of ischemic mode,3 rats in cholesterol embolus group and 3 rats in thromboembolus group were used to detect the difference of protein expression in cerebral ischemic area by tandem mass spectrometry and library search identification.Proteins with fold-change>1.2 or<0.83(P<0.05)in the two groups were considered to be differential proteins,and the differentially expressed proteins were analyzed by gene ontology(GO)and Kyoto encyclopedia of genes and genomes(KEGG)enrichment.Results(1)The results of Logna score and edema infarct size were 0 in sham operation group.At 6 and 24 hours after the establishment of the rat cerebral ischemia model,compared with the thromboembolus group,the ischemic area of the rats in the cholesterol embolus group was smaller(6 h:[9.7±2.3]%vs.[21.1±7.6]%,t=-4.080;24 h:[19.4±3.7]%vs.[24.8±4.2]%,t=-2.741;both P<0.05),while there were no significant differences in Longa behavioral scores and edema rates(both P>0.05).(2)The concentrations of Evans blue were(1.26±0.07),(1.47±0.14)and(1.53±0.15)μg/g at 6 hours after the establishment of the rat model in the sham,cholesterol embolus and thromboembolus groups,respectively;the concentrations of Evans blue were(1.25±0.05),(1.56±0.20)and(1.59±0.20)μg/g at 24 hours after the establishment of the rat model.The differences in Evans blue concentrations were statistically significant(F values were 10.826 and 10.026,both P<0.01).Further pairwise comparative analysis showed that compared with the sham group,the Evans blue concentrations in the cerebral cortical tissues of the cholesterol embolus group and thromboembolus group were increased(both P<0.05);compared with the cholesterol embolus group,there was no significant difference in Evans blue concentration in the cerebral cortical tissue of the thromboembolus group(all P>0.05).(3)The results of protein profiling showed that a total of 15 differentially expressed proteins that met the fold expression criteria of this experiment were identified.Compared with the thromboembolic group,there were 12 up-regulated proteins(vitronectin,etc.)and 3 down-regulated proteins(foveolar protein,etc.)in cholesterol emboli.GO enrichment analysis showed that the differentially expressed proteins mainly focused on phospholipid metabolic processes and functioned to bind to cytokine receptors.KEGG pathway enrichment analysis showed that coagulation versus complement pathway was the main concentrated pathways of differentially expressed proteins.Conclusion In the ischemic rat model,ischemic area of the cholesterol embolus group was smaller than that of the thromboembolus group,and there was a significant difference in protein expression between the two groups.