参附注射液对局灶性脑缺血再灌注损伤大鼠血脑屏障通透性及神经功能的影响

Effects of Shenfu injection on the permeability of blood brain barrier and the nervous function of focal cerebral ischemic reperfusion injured rats

目的:观察参附注射液(Shenfu injection,SFI)对大鼠脑局灶性缺血再灌注(Ischemic-reperfusion,IR)损伤后血脑屏障(Blood brain barrier,BBB)通透性和神经功能的影响,评价参附注射液对大鼠局灶性脑缺血再灌注损伤有无保护作用,为参附注射液的临床应用提供参考。方法:清洁级的SD雄性大鼠80只,随机分为A、B、C、D4个组:A组为假手术加生理盐水处理组,B组为假手术加参附注射液处理组,C组为造模加生理盐水处理组,D组为造模加参附注射液处理组。A、C组于麻醉后5 min经尾静脉注射生理盐水10 ml/kg,B、D组在相同时点给予相同剂量的SFI。随后对C、D两组采用Zea Longa线栓法制备大鼠右侧大脑中动脉(Middle cerebral artery,MCA)缺血2 h再灌注24 h模型。A、B两组均不作造模处理,其余处理步骤与C、D两组相同。记录各组麻醉后苏醒时间,缺血2 h及再灌注24 h两时间点观察神经功能缺损情况,于再灌注24 h结束时在每组随机选择5只取右侧额顶叶大脑皮质部分电镜下观察血脑屏障超微结构的情况,其余15只大鼠静脉注射伊文思蓝(Evan’s blue,EB),对比两组伊文思蓝通过血脑屏障进入脑组织的量。结果:各组间比较,体重差异无统计学意义。麻醉后苏醒时间A、B两组短于C、D两组(P<0.05),A、B两组之间无差异,C组长于D组(P<0.05)。缺血2 h神经功能缺损评分(Longa score,Longa评分):A、B两组低于C、D两组(P<0.01),A、B两组之间以及C、D两组之间差异无统计学意义。再灌注24 h后Longa评分:A、B两组低于C、D两组(P<0.01),A、B两组之间差异无统计学意义,C组高于D组(P<0.01)。C组再灌注24 h Longa评分明显高于缺血2 h(P<0.01),D组这两个时点比较,差异无统计学意义。缺血2 h,再灌注24h结束时,A、B两组大鼠脑组织内伊文思蓝含量明显低于C、D两组(P<0.01),A、B两组之间差异无统计学意义,C组明显高于D组(P<0.05)。电镜下,A、B两组大鼠大脑皮质毛细血管周围及管腔未见明显变化。C组大鼠大脑皮质缺血再灌注区毛细血管周围重度水肿,毛细血管管腔明显受压。D组相应区域毛细血管周围轻度水肿,毛细血管轻度受压。结论:参附注射液可减轻局灶性脑缺血再灌注损伤大鼠血脑屏障通透性升高程度,减轻神经功能受损伤程度,发挥脑保护作用。

Objective： To investigate the effects of Shenfu injection （SFI） on the permeability of blood brain barrier （BBB） and the nervous function of focal cerebral ischemic-reperfusion injured rats, and judge if SFI has protective effects for focal cerebral ischemic-reperfusion injured rats to provide theoretical proof for the clinical application of SFI. Methods： Eighty male SD rats were divided into four groups randomly： group A （the sham operated control group）, group B （the sham operated trial group ）, group C （the control group） and group D （the trial group）. The rats in group A and group C received i.v. a bolus of normal saline （NS） for 10 mg/kg 5 min after anesthesia, while those in group B and group D received a bolus of SFI for 10 mg/kg instead. The focal cerebral ischemic-reperfusion injury model was established with thread embolism of middle cerebral artery（MCA ） according to Zea Longa＇ s method in group C and group D. The time for postanesthetic revival in each group was recorded. The Longa score in each group was recorded at 2 h of occlusion and 24 h of reperfusion. The injury to the blood-brain barrier was observed by the content of Evan＇ s blue （EB） in the cerebral tissue, the changes of the structure of BBB were observed with electron microscope at the end of the experiment. Results： The body weight in each group had no statistical difference. The time for revival from anesthesia in group A and group B was shorter than that in group C and group D （P〈0.05）,and that in group C was longer than that in group D（P 〈0.05 ） but there was no difference between group A and group B. After ischemia for two hours, Longa score in group A and group B was lower than that in group C and group D（P〈0.01 ） , and there was no difference either between group A and group B, group C and group D. After reperfusion for 24 h, Longa score in group A and group B was lower than that in group C and group D, there was no difference between group A and group B （P〈0.01）, and Longa score in group C was higher than that in group D（P〈0.01 ）. At the 24th hour of reperfusion, Longa score in group C was higher than that at the 2rid hour of ischemia（P〈 0.01 ）, but there was no difference for group D. After ischemia for 2 h and reperfusion for 24 h, the rats in group A and group B had an evidently lower Evan＇s blue content in the cerebral tissue than that of the rats in group C and group D, and there was no difference between group A and group B, while that of the rats in group C was higher than that of the rats in group D. Under the electron microscope, brain edema and microvessel pressed in group C were more severe compared with those in group D. But there was no significant abnormality in group A and group B. Conclusion： Shenfu injection could reduce blood-brain barrier permeability after focal cerebral ischemic-reperfusion injury and it has significant nervous function protection against ischemic-reperfusion damage.