吡咯烷二硫代氨基甲酸对大鼠脑缺血后神经功能的影响

Effect of PDTC on nurologic fuction after cerebral ischemia in rat

目的 :观察核因子-κB p50(nuclear factor-κB subunit p50,NF-κB p50)特异性抑制剂吡咯烷二硫代氨基甲酸(pyrrolidine dithiocarbamate,PDTC)抑制NF-κB p50后,炎症因子TNF-α及相关康复蛋白Reelin、突触素、Lingo-1表达的变化,进一步观察大鼠神经功能评分、海马神经细胞形态的变化。方法:本实验的研究对象为雄性健康SD大鼠,随机分为3组:假手术组(S组,n=6)、缺血组(I组,n=18)、PDTC干预组(P组,n=18)。脑缺血模型采用永久性MCAO法。P组的大鼠在造模后30 min进行腹腔注射PDTC(100 mg/kg),每天1次。S组大鼠于第3天处死,I组和P组大鼠分别于1、3、7 d处死。用Longa评分对3组大鼠进行神经功能损害程度进行评估,用Western blot检测大鼠缺血海马NF-κBp50、TNF-α、Reelin、突触素蛋白的表达,用HE染色方法观察3组大鼠海马神经元细胞的形态,用免疫组化法检测Lingo-1在脑组织内的表达情况。结果:1P组及I组大鼠Longa评分较S组均有所升高,术后第1天和第3天P组较I组Longa评分降低,但无统计学意义(P>0.05);在术后第7天,P组较I组显著降低(P<0.05)。2I组大鼠海马神经细胞出现异常,而P组大鼠中,海马神经元细胞的结构基本保持正常。3术后1、3天,I组和P组大鼠缺血海马中NF-κB p50较S组升高(P<0.05),但P组NF-κB p50表达较I组降低(P<0.05);术后7天,3组变化无统计学意义。4术后1、3、7天,I组和P组大鼠TNF-α表达较S组升高,但P组TNF-α较I组降低(P<0.05);5术后第1、3天,P组大鼠缺血海马中突触素和Reelin蛋白的表达明显高于I组(P<0.05),第7天两组差别无统计学意义(P>0.05)。S组大鼠脑组织内可见少量Lingo-1阳性细胞表达,P组大鼠脑组织内Lingo-1阳性细胞数在第7天时明显低于I组。结论:PDTC可改善神经功能,维持海马神经元正常形态,这可能与其抑制NF-κB p50表达,减少TNF-α等相关炎症因子生成及调节Reelin、突触素及Lingo-1表达有关。

Objective：To observe the effects of the NF-κB subunit p50（NF-κB p50）specific inhibitor pyrrolidine dithiocarbamate（PDTC）on the expression of NF-κB p50,inflammatory factor TNF-α and related rehabilitation protein Reelin,synaptophysin and Lingo-1 and to further observe the changes of neurological function scores and the morphology of hippocampal neurons in rats.Methods：Male Sprague-Dawley rats were randomly divided into three groups：sham-operated group（S group）,ischemic control group（I group）and PDTC group（P group）. Middle cerebral artery occlusion（MCAO）was used to create focal cerebral ischemic models.Rats in P group underwent PDTC（100 mg/kg,intraperitoneal injection）after MCAO. Rats were sacrificed on day 3 in S group and on day 1,3 or 7 in I group and P group post-MCAO. Longa scores was used to evaluate the neurological function of rats in three groups.Western blot was used to detect the expression of NF-κB p50,TNF-α,synaptophysin and Reelin. HE staining was used to observe the morphology of hippocampal neurons of rats in three groups and immunohistochemistry was used to detect the number of and Lingo-1-positive cells in three groups. Results：The Longa scores of P group and I group were lower than those of S group;the Longa scores of P group were lower than those of I group on day 1 and 3 post-MCAO without statistical differences（P〉0.05）,but the Longa scores of P group were lower than those of I group on day 7 post-MCAO（P〉0.05）. The hippocampal neurons in I group was abnormal,while the morphology of the hippocampal neurons in P group was generally normal. The expression of NF-κB p50 in ischemic hippocampus was higher in I group and P group than in S group（P〈0.05）,but it was lower in P group than in I group. There was no statistical difference on day 7 post-MCAO in three groups. The expression of TNF-α in I group and P group was higher than that in S group on day1,3 and 7 post-MCAO（P〈0.05）,but it was lower in P group than that in I group（P〈0.05）. The expression of synaptophysin and Reelin in P group was remarkably upregulated than that in I group on day 1 and 3 post-MCAO（P〈0.05）,but without statistical differences on day 7 post-MCAO（P〈0.05）. The expression of Lingo-1-positive cells was low in S group,and it was lower in P group than that in I group on day 7. Conclusion：PDTC can improve neurological function,maintain the relatively normal morphology of the hippocampal neurons,which is possibly relative to inhibiting the expression of NF-κB p50,reduc ing the production of inflammatory factor TNF-α and regulating the expression of Reelin,synaptophysin and Lingo-1.