阿加曲班对MCAO大鼠不同时间点血小板功能的影响

建立大脑中动脉闭塞(model of middle cerebral artery occlusion,MCAO)大鼠模型,研究阿加曲班对大鼠不同时间脑缺血再灌注损伤血小板功能的影响。将120只健康雄性SD大鼠随机分为假手术组(Sham)、模型组(Model)和阿加曲班给药组(Argatroban),其中Argatroban组分为4组:Argatroban+30 min组、Argatroban+1 h组、Argatroban+3 h组和Argatroban+7 h组,Sham组和Model组40只,其余各组每组10只。线栓法制备大鼠MCAO模型,术中输注Argatroban,剂量为0.3μg·kg^-1·min^-1,完成手术前5 min停止输注。手术完成后,Sham组和Model组分别于术后30 min、1 h、3 h和7 h处死10只取样;Argatroban给药组5个时间点处死取样。检测大鼠血浆血细胞数量,并计数骨髓有核细胞数的变化。采用流式细胞仪检测血小板-白细胞聚集体(platelet-leukocyte aggregates,PLA)表达水平,观察活化部分凝血活酶时间(activated partial thromboplastin time,APTT)、血小板计数变化,ELISA检测血浆D-二聚体和TAT。结果显示,与Sham组相比,Model组大鼠的红细胞(RBC)、白细胞(WBC)、血红蛋白含量(HGB)和中性粒细胞(GR)数都显著升高(p<0.05),且与给药组没有统计学差异。Sham组、Model组和Argatroban给药组的骨髓有核细胞数也没有统计学差异。与Sham组相比,Model组PLA表达水平极显著升高(p<0.01),Argatroban+30 min组PLA表达水平却低于Sham组,但没有统计学差异,Argatroban+1 h组高于Sham组和Argatroban+30 min组(p<0.05),Argatroban+3 h组和Argatroban+7 h组高于Argatroban+1 h组低于Sham组。术后各组的血小板计数均高于Sham组(p<0.05),Argatroban各组的血小板计数要低于Model组(p<0.05),并在30 min时达到最低,30 min以后血小板升高,但是1 h以后血小板不再升高,趋于稳定。Argatroban各组PT和APTT虽略有降低,但是与Sham组和Model组并没有统计学差异。造模后各组TAT、D-二聚体水平显著升高,Argatroban各组TAT水平较模型组明显降低(p<0.05),Argatroban+30 min组水平最低。MCAO模型大鼠血液呈现高凝状态,凝血功能和纤维蛋白溶解功能亢进,血小板活化增强,阿加曲班可对此起到改善作用。

Effects of Agathuban on Platelet Function at Different Time Points in MCAO Rats

To establish a rat model of middle cerebral artery occlusion(MCAO)and study the effect of argatroban on platelet function in rats with cerebral ischemia-reperfusion injury at different time points,120 healthy male Sprague-Dawley rats were randomly divided into sham operation group(Sham),model group(Model)and argatroban administration group(Argatroban).The Argatroban component was divided into four groups:Argatroban+30 min group,Argatroban+1 h group,Argatroban+3 h group and Argatroban+7 h group,40 in Sham group and Model group,and 10 in the other groups.Rat MCAO model was prepared by suture method.Intraoperative injection of Argatroban was performed at a dose of 0.3μg·kg^-1·min^-1,and the infusion was stopped 5 minutes before the operation.After the operation,10 samples were sacrificed at 30 min,1 h,3 h,and 7 h after Sham group and Model;The Argatroban administration group was sacrificed at each time point.Flow cytometry was used to detect the expression level of platelet-leukocyte aggregates(PLA),and the activated partial thromboplastin time(APTT)and platelet count changes were observed.Plasma D-dimer and TAT were detected by ELISA.Compared with the Sham group,the expression level of PLA in the Model group was significantly increased(p<0.01).The expression level of PLA in the Argatroban+30 min group was lower than that in the Sham group,but there was no statistical difference.The Argatroban+1 h group was higher than the Sham group.Sham group and Argatroban+30 min group(p<0.05),Argatroban+3 h group and Argatroban+7 h group were higher than Argatroban+1 h group than Sham group.The platelet count of each group was higher than that of Sham group(p<0.05).The platelet count of each group of Argatroban was lower than that of Model group(p<0.05),and reached the lowest at 30 min,and the platelet increased after 30 min.However,after 1 h,the platelets no longer rise and tend to be stable.Although the PT and APTT of each group of Argatroban were slightly decreased,there was no statistical difference between the Sham group and the Model group.The levels of TAT and D-dimer in each group were significantly increased after modeling,and the levels of TAT in each group of Argatroban were significantly lower than those in the model group(p<0.05),and the lowest in Argatroban+30 min group.The blood of MCAO model is hypercoagulable,coagulation function and fibrinolytic function,platelet activation is enhanced,and argatroban can improve this effect.