参附注射液对心肺复苏后大鼠神经细胞的保护作用

Study on the protective effects of Shenfu injection on neurocyte in rats after cardiopulmonary resuscitation

目的:探讨参附注射液对心肺复苏后大鼠脑神经细胞凋亡及B细胞淋巴瘤/白血病-2基因(Bcl-2)、Bcl相关x蛋白(Bax)、核转录因子κB(NF-κB)等蛋白表达的影响,为防治神经细胞的凋亡提供依据。方法:90只Sprague Dawley大鼠随机分为假手术组、模型组、参附治疗组,各30只。采用窒息合并冰氯化钾致大鼠心跳骤停-心肺复苏模型。运用免疫组化观察复苏后神经细胞中NF-κB、Bcl-2、Bax基因的蛋白表达,采用末端标记技术(TUNEL)检测各组神经细胞的凋亡情况;并在电镜下观察神经细胞超微结构。结果:参附治疗组复苏后各时相点Bcl-2表达明显增强(P<0.01),而Bax表达无明显差异(P>0.05),NF-κB的表达明显降低(P<0.05)。Bcl-2阳性表达率在复苏后24h达到高峰,参附治疗组阳性表达率明显高于模型组(P<0.01)。各时相点参附治疗组Bax表达与复苏模型组比较无显著差异(P>0.05)。在心肺复苏后的不同时段神经细胞确实有凋亡发生,参附治疗组各时相点凋亡细胞阳性指数均明显低于复苏模型组(P<0.05);而假手术组无明显凋亡现象发生(P<0.01)。超微结构显示参附治疗组神经细胞损伤较模型组明显减轻,假手术组神经细胞超微结构正常。结论:细胞凋亡参与了心肺复苏后大鼠神经细胞的损伤,参附注射液可降低NF-κB活性,上调Bcl-2蛋白表达,改善神经细胞的超微结构,抑制细胞凋亡的发生,对神经细胞具有明显的保护作用。

Objective To study the protective effect of Shenfu（SF） injection on neurocyte apoptosis induced by neurocyte ischemia/reperfusion injury after cardiopulmonary resuscitation （CPR） in rats and the expressions of B-cell lymphoma/leukemia-2 gene （Bcl-2）, Bcl-associated x protein （Bax）and nuclear factor-κB （NF-κB） for providing the explanation of neurocyte apoptosis protection after CPR. Methods Cardiac arrest was induced by asphyxia and ice-cold 0.5 mmol/L KCl in rats and resuscitation was begun five minutes afterwards. Ninety Spraque Dawley rats were randomly divided into 3 groups： sham control group （n=30）, resuscitation model group （n=30）, and SF injection treatment group （SF group, n=30）. Bcl-2 and Bax expression were determined by immunocytoehemistry at 3, 6, 12, 24, 48 h after resuscitation. The presence of apoptotic neurocyte was determined by the terminal deoxynucleotidyl transferase mediated dUTP nick end labeling method （TUNEL）, and the neurocyte ultrastructure was examined by electron microscopy. Results Bcl-2 were actively expressed with peak value at 24 h following CPR, the expression positive rate was significantly higher in SF group than in resuscitation model group （P〈0.01）. Bax expression positive rate was not significantly different between SF group and resuscitation model group （P〉0.05）; expression of NF-κB was significantly lower in SF group than in resuscitation model group （P〈0.05）. Neuroeyte apoptosis was existent in different phases after CPR, the apoptotic index was significantly lower in SF group than in resuscitation model group （P〈0.05）; no obvious apoptosis was seen in sham control group （P〈0.01）. Uhrastrueture examination revealed that SF group had milder injury than resuscitation model group; uhrastructure was normal in sham control group. Conclusions Neurocyte apoptosis was involved in neurocyte injury after CPR in rats. SF injection could inhibit NF-κB, increase Bcl-2 expression, improve neurocyte uhrastructure, inhibit neurocyte apoptosis. SF injection has significant protective effect on neurocyte after CPR.