腺苷处理后犬脊髓缺血再灌注损伤中神经元凋亡及相关蛋白表达的变化

The neuronal apoptosis and related gene expression changes after adenosine treatment in canine spinal cord ischemiareperfusion injury experiments

目的探讨腺苷处理后犬脊髓缺血再灌注损伤实验中脊髓神经元凋亡及其相关蛋白Bax、Bcl-2的表达变化，证实腺苷对犬脊髓缺血再灌注的保护作用。方法16只健康成年杂种犬，雌雄不拘，体质量10—20妇。随机均分为两组，采用胸主动脉、锁骨下动脉阻断法造成脊髓缺血模型（75min）。单纯缺血再灌注组，阻断前30min颈静脉泵人50rnl生理盐水，300ml／h，10min泵完；阻断结束时再以相同的流速和时间泵完（对照组）。阻断前30min颈静脉泵入50m1腺苷溶液（0．4mg／ml，50ml），阻断结束时再以相同的流速相同浓度和时间泵完（实验组）。术后继续笼内饲养，采用Tarlov下肢神经功能评分系统评估6、24、48h神经运动功能评分。再灌注48h各组分别处死实验动物，迅速取出L2-L5及骶段脊髓。采用Tunel法进行细胞凋亡检测，免疫组化法检测Bax、Bcl-2的表达。结果实验组犬运动功能评分24、48h高于对照组，差异有统计学意义（P〈0．05）。实验组脊髓神经元凋亡量明显低于对照组（P〈0．05），较对照组Bax表达减少和Bcl一2表达增多。结论腺苷处理后可减少犬脊髓缺血再灌注后神经元凋亡，对脊髓缺血再灌注损伤具有明显的保护作用。其保护作用可能与凋亡蛋白Bax表达减少和凋亡抑制蛋白Bcl·2表达增多有关。

Objective To investigate the protective effect of adenosine on spinal cord Isehemia/Reperfusion through the observation of the neurons＇apoptosis and the change of Bax and Bel-2. Methods 16 healthy adult mongrel canines of either sex, weighted 10-20kg , were randomly divided into two groups （n = 8 ）. Creating the models of spinal cord Isehemia/Reperfusion by thoracic aorta and subclavian artery occlusion. Control group： i. e the simple Ischemia/Repeffusion group. Pumping 50 rul saline through the jugular vein 30 minutes before blocking the artery, and the rate is 300ml / h, pumping time 10 minutes. Then pump another 50ml saline right after blocking at the same rate. Experimental group ： 20mg adenosine were dissolved into 50ml saline and was pumped 30min before blocking the artery in the same way. Still pump another 50ml adenosine solution of the same con- centration right after blocking. After the operation, canines were continued to feed with free access to food and water. Recording the motor function score at 6 h, 24 h, 48 h after reperfusion （ see Taylor motor function scoring system）. All 8 canines were killed after 48 hours of reperfusion. L2-L5 and the sacral spinal cord were taken out as quickly as possible. The Tunel method was a- dopted to detect neurons＇apoptosis, and the immunohistochemical method to detect the expression changes of Bax and Bcl-2 suits In the experimental group, the 24 h, 48 h motor function scores were significantly higher than that in the control group （ P 〈 O. 05 ） . The neurons＇apoptosis was significantly lower than that of the control group （P 〈 0. 05）. Compared with the control group, the expression of the apoptotic protein Bax reduced while the apoptosis-inhibited protein Bcl-2 increased. Conclusion The neurons＇apoptosis can be reduced significantly through the adenosine treatment and adenosine has a apparently protective effect on canines＇spinal cord Ischemia/Reperfusion injury. The effect might be related to Bax and Bcl-2.