参附注射液对新生大鼠缺氧缺血性脑损伤干预作用的实验研究

Effects of Shenfu injection on hypoxic-ischemic brain damage:experiment with neonatal rats

目的研究参附注射液对新生大鼠缺氧缺血性脑损伤(HIBD)的干预作用。方法建立新生大鼠HIBD模型,把实验分成两部分(1)观察缺氧缺血后3、7、14和28d各组新生大鼠的体重变化以及各组28d内生存率。(2)流式细胞术测量缺氧缺血前2h,缺氧缺血后2h、12h、24h、3d、7d、14d和28d各时间点新生大鼠海马CA1区神经元的凋亡率,每组实验随机分为4组(每组20只)假手术组,生理盐水对照组,参附预处理组,参附治疗组。结果缺氧缺血后3、7、14和28d,生理盐水对照组大鼠体重的增长均明显小于假手术组(7d8.8g±2.1gvs14.0g±2.9g,均P<0.01),且明显小于参附预处理组(7d11.7g±3.3g,P<0.01或P<0.05),缺氧缺血后7、14、28d生理盐水对照组大鼠体重的增长均明显小于参附治疗组(7d10.9g±2.7g,P<0.01或P<0.05)。各组的体重明显小于假手术组。生理盐水对照组存活率为60%(12/20),参附预处理组存活率为90%(18/20),参附治疗组存活率为85%(17/20),生理盐水对照组存活率明显低于其他各组(P<0.05),其余各组存活率差异无统计学意义(均P>0.05)。缺氧缺血后2h生理盐水对照组、参附治疗组、参附预处理组海马神经元凋亡率开始升高,于缺氧缺血后24h凋亡率达到峰值后开始下降,缺氧缺血后14d降到正常。参附治疗组和参附预处理组海马神经元凋亡情况明显好于生理盐水对照组(24h16.0%±4.2%vs11.9%±2.3%vs18.1%,P<0.05或P<0.01)。结论参附注射液减少了新生大鼠缺氧缺血后神经元凋亡的发生,改善了新生大鼠HIBD后的生长发育,提高了其生存率。

Objective To investigate the effect of Shenfu （ginseng and aconite root） injection on hypoxic-ischemic brain damage （HIBD）. Methods Sixty 7-day-old Sprague-Dawley rats undergoing ligation of left common carotid artery and then put into a container with 8% O2 and 92% N2 for 2 h so as to establish HIBD models, were randomly divided into 3 groups ： Shenfu injection pretreatment group （ since d days before the experiment Shenfu injection 10 ml/kg was injected intraperitoneally once a day for 4 days）, Shenfu injection treatment group [ Shenfu injection 10 ml/kg was injected intraperitoneally immediately after hypoxic-ischemia （HI） insult once a day for 7 days], and control group （normal saline 10 ml/mg was injected intraperitoneally immediately after HI insult once a day for 7 days）. Twenty neonatal rats underwent sham operation as control group. The d groups were further divided into subgroups of 6 rats according to the time points： 2 hours before and 2 hours, 12 hours, 24 hours, 3 days,7 days, ld days, and 28 days after HI insult. 3, 7, 14, and 28 days after the HI insult the body weight was observed and the survival rate was observed 28 d after the HI insult. At different time points the rats of different subgroups were killed and their brains were taken out. Flow cytometry was used to calculate the neuron apoptosis rate in the hippocampal CA1 region. Results The body weight increase levels 3, 7, 14 and 28 days after HI insult of the control group were all significantly less than those of the sham operation group （ e. g 7 days ：8.8 g±2. 1 g vs 14.0 g ± 2.9 g, all P 〈 0.01 ） and the body weight increase levels 3, 7, 14, and 28 days after HI insult of the control group were all significantly less than those of the Shenfu injection pretreatment group （ e. g 7 d ： 11.7 g ± 3.3 g） and Shenfu injection treatment group （ e. g 7 d ： 10.9 g±2.7 g,P 〈 0.01 or P 〈 0.05）. The survival rate 28 d after HI insult of the control group was 60%, significantly less than those of other groups （ all P 〈 0.05）, and there was no significant difference in the survival rate among the other groups （ all P 〉 0.05 ）.Compared with the sham operation group the neuron apoptosis rates of the hippocampal CA1 region of the Shenfu injection treatment group and Shenfu injection pretreatment group began to increase 2 hours after HI insult, peaked 24 hours after, then gradually decreased, and recovered to normal 14 days after. The neuron apoptosis rates 2, 12, 24, 72 hours, and 7 days after HI insult of the Shenfu injection pretreatment group were all significantly lower than those of the control group （ e. g24 hours ： 16.0% ± 4.2% vs 11.9% ± 2.3% vs 18.1% ,P 〈0.01 or P 〈0.05） , and the neuron apoptosis rates 72 hours and 7 days after HI insult of the Shenfu injection treatment group were significantly lower than those of the control group. Conclusion Shenfu injection can enhance the physical development and elevate the survival rate of neonatal rats with HI insult, and significantly prevents apoptosis of the hippocampus neurons from HI insult.