黄芪多糖对孕鼠鞘内注入布比卡因脊神经毒性的保护作用

Protective Effect of Astragalus Polysaccharides on Neurotoxicity of Intrathecal Bupivacaine in Pregnant Rats

目的探讨黄芪多糖是否可以减轻布比卡因对孕鼠脊神经毒性损伤反应及其保护机制。方法健康雌性妊娠17日SD大鼠,体质量350～400g,取鞘内置管成功孕鼠30只,随机分成5组(n=6)。对照组(N组),2%和4%布比卡因组(2B组和4B组),2%和4%布比卡因黄芪多糖治疗组(2H组和4H组)。N组:鞘内注射生理盐水30μl,腹腔注入生理盐水;2B组或4B组:鞘内注入2%或4%布比卡因30μl,腹腔注入生理盐水;2H组或4H组:鞘内注入2%或4%布比卡因30μl,腹腔注入黄芪多糖(25mg/kg,生理盐水稀释成2ml,连续注药4d)。鞘内注药后10min,20min,30min,1h,2h,4h,1d,2d,3d和4d测定甩尾反应潜伏期(TFL),用最大效应百分比MPE表示,并进行下肢运动功能(MF)评分。4d后各组大鼠行断颈处死,分离背根神经节,Westernblot分析Caspase-9蛋白表达水平和病理学观察。结果各组MPE比较差异有统计学意义(P<0.05);鞘内注药后10min,2B组、4B组、2H组和4H组MPE均达到峰值,2B组和4B组于2d恢复至基线水平,而2H组和4H组于1d恢复至基线水平;在鞘内注药1d,与4B组比较,2H组和4H组MPE降低(P<0.05)。注入布比卡因后,各组MF评分较N组均明显升高(P<0.05);2B组和2H组于注药后4h恢复正常,而4B和4H组于1d恢复正常。与N组比较,2B组,4B组,2H组和4H组Caspase-9蛋白表达均上调(P<0.05);与2B组比较,2H组Caspase-9蛋白表达降低(P<0.05)。病理学结果:2B组和4B组有部分神经细胞萎缩、空泡化;2H组和4H组神经细胞萎缩、空泡化数量明显少于2B组和4B组。结论黄芪多糖对孕鼠布比卡因脊神经毒性损伤反应具有一定保护作用,其机制可能与抑制Caspase-9蛋白表达,抗神经细胞凋亡有关。

Objective To investigate the protective effect of Astragalus polysaccharides（APS）on intrathecal（IT）bupivacaine neurotoxicity in pregnant rats and its mechanism.MethodsThirty late pregnant rats weighting 350-400 g were used in this study.All the rats in which PE-10 catheter were successfully placed without complication were divided into 5 groups randomly（n=6 each）：normal pregnant groups（group N）,2% and 4% bupivacaine groups（group 2B,group 4B）,2% and 4% bupivacaine APS treated group（group 2H,group 4H）.Group N：received normal saline 30 μl IT and intraperitoneal injection with normal saline 2 ml.Group 2B or 4B：received 2% or 4% bupivacaine 30 μl IT and intraperitoneal injection with normal saline 2 ml.Group 2H or 4H：received 2% or 4% bupivacaine 30μl IT and intraperitoneal injection with APS（25 mg/kg,normal saline diluted to 2 ml,continuous injection 4 d）.Tail-flick test and motor function were performed at 10min,20 min,30 min,1 h,2 h,4 h,1 d,2 d,3 d and 4 d after IT administration.TF latent period was converted to the percent of maximal possible effect（MPE）.Four days later,the animals were sacrificed and the DRG were removed for determination of caspase-9 protein expression by western blotting and the pathological changes were observed with light microscope.ResultsAfter the IT 10min,2B,4B,2H and 4H group MPE reached the peak.MPE in group 2B and 4B returned to the baseline values after 2 d,in group 2H and 4H returned the baseline values after IT 1 d.In each group MF score was significantly increased after IT bupivacaine.Group 2B and 2H returned to normal after IT 4 h,then group 4B and 4H returned to normal after IT 1 d.Rats in group 2B,4B,2H and 4H significantly increase the expression of caspase-9 in DRG compared with those in group N.Compared with group 2B,rats in group 2H decreased significantly the expression of caspase-9 in DRG.Pathological results showed the neuroval injury in 2B and 4B group was more serious than in 2H and 4H group.Conclusion APS has neuron protection against neurotoxicity of intrathecal bupivacaine in pregnant rats,which is related with down-regulation of caspase-9 expression.