【作者】 刘振寰； 潘佩光； 赵勇； 柴铁劬； 唐纯志； 王琴玉； 杨君军； 林锦泉；

【机构】 广州中医药大学附属南海妇产儿童医院小儿神经康复科； 广州中医药大学针灸推拿学院实验室；

【摘要】 目的探讨针刺对新生大鼠缺氧缺血性脑损伤(HIBD)脑神经元调亡及神经生长因子表达的作用及其机制。方法:①实验于2005年8月至2005年10月在广州中医药大学实验动物中心清洁级实验室及针灸推拿学院实验室完成。选用85只7日龄清洁级SD大鼠,参照Rice法并加以改进,将大鼠结扎左侧颈总动脉后恢复2．h,置于透明密闭容器中,并入37℃恒温水中以1L/min 的速度通入低氧气体(体积浓度为8％的氧气和92％的氮气),2．5h后将动物取出,将存活者继续保温1h时后作行为测定,翻身不能、平衡异常或左旋者视为成功的模型。本实验成功制作HIBD动物模型72只,随机分为3组,A组为缺氧缺血+针刺Ⅰ组,B组为缺氧缺血+针刺Ⅱ组,C组为缺氧缺血对照组。另设假手术组D组。每组24只。②选用大鼠百会、患侧颞Ⅰ针、内关、曲池、足三里、涌泉,A组于造模后24小时开始针刺,前7d仅针四肢部穴位,第8天开始加针头部穴位, B组造模后第8天开始针刺,头、体针同步。两组均以针灸针先在内关、涌泉速刺微出血,不留针, 然后针头部及曲池、足三里。百会、颞Ⅰ针及曲池、足三里接G6805-Ⅱ电针仪,连续波,频率5- 10HZ,持续10min。针刺均为每日1次,A组连续针刺20d,B组连续针刺13d,假手术组干预措施与A组相同,至缺血缺氧后第21天处死取材。分别采用TUNEL法及免疫组化方法测定针刺对HIBD 大鼠额页与海马区组织神经元凋亡及海马区神经生长因子(NGF)表达,用图像分析仪在光镜下(10 ×40)计算神经细胞凋亡数目及NGF免疫阳性细胞表达数目。组间差异用方差分析。结果四组左侧海马CA1区神经元及左侧大脑额页皮质神经元凋亡数差异有统计学意义,其中D<A<B<C (P<0．05);四组左侧海马CA1区神经元NGF阳性数差异有显著统计学意义,其中D<C<B<A (P<0．001)。结论针刺对鼠HIBD神经元确有保护作用,可增强脑内NGF生物活性。更多还原

【Abstract】 Objective To study the protective effect and the mechanism of Acupuncture on hypoxic- ischemic brain damage (HIBD) in newborn rats. Method ① The experiment was complete at the clean degree laboratory of experimental animal center of Guangzhou university of TCM and the laboratory of acupuncture and massage college. 85 clean conventional SD rats, aged 7 days, were selected. According to Rice method which was improved , the rats was operated by ligation of left common carotid artery and recover for 2 hours. Then they were put in hyaline airtight vessel in the homeothermal water of 37℃. Hypoxia gas(inhaling 8% O 2 and 92% N2) was infused to the vessel at the speed of 1L/min. The animal were took out after 2.5 hours. The survival act were determined after incubation of 1 hour. The successful model were evaluated by the act of inability of roll over, abnormal balance or turning left. 72 successful HIBD model were divided randomly to 3 groups. Group A was HIBD+acupuncture I group. Group B was HIBD+acupuncture II group. Group C was the control group of HIBD. Sham-operated group was set up to group D in addition. The number of the rats in each group is 24. ② The acupoints of Baihui, I needle of temple in the affected lateral, Neiguan, quchi, Zusanli, Yongquan were selected. Group A were acupunctured at 24 hours after the models were made. Only four extremities were acupunctured in initial 7 days. The head-acupoints were acupunctured in addition at 8th day. Group B began to be acupunctured at 8th day after the models were made. Head and body acupuncture were operated at the same time. In both of the two group, Neiguan and Yongquan were acupunctured rapidly with little blood and without retention, then head-acupoints and Quchi, Zusanli. Baihui, I needle of temple and Quchi, Zusanli was connected by G6805-II electro-acupuncture equipment, continuous wave and 5-10Hz frequency were used, continued 10 minutes. The acupuncture operated once a day. Group A continued 20 days. Group B continued 13 days. Sham-operated group was interfered as same as group A. The rats were put to death 21 days after the HIBD. Neuronal apoptosis in the temple and hippocampus and expression of NGF in the hippocampus of HIBD rat were observed by TUNEL assay and immunohistochemistry staining assay respectively. The cell number of neuronal apoptosis and expression of NGF was counted by the equipment of picture analysis at the light microscope (10×40). The difference between the groups was analyzed by analysis of variance. Result The difference of cell number of neuronal apoptosis in the CA1 zone of the left hippocampus and in the left temple of all the four groups has significance in statistics. The difference of expression of NGF in the CA1 zone of hippocampus of all the four groups has obvious significance in statistics Conclusion The results verified the protective efficacy of electroacupuncture on HIBD, which has relation to enhance the activity of the expression of NGF更多还原