参芪脑保含药血清在体外培养PC12细胞抗凋亡作用中的量效分析  

Dose-dependent effect of serum containing Senqi Naobao on anti-apoptosis of PC12 cells cultured in vitro

作　　者：崔旭[1] 尹岭[2] 杜卫[2] 韩志涛[1] 房征宇[1] 

机构地区：[1]解放军总医院老年医学研究所神经生物学室,北京市100853 [2]解放军总医院老年医学研究所神经信息室,北京市100853

出　　处：《中国临床康复》2006年第7期22-24,共3页Chinese Journal of Clinical Rehabilitation

基　　金：全军医药卫生科研基金"九五"重点课题(95Z058)~~

摘　　要：目的:观察在经典药方补阳还五汤基础上加减而成的参芪脑保对体外培养的正常PC12细胞增殖和对抗活性氧(100μmol/L过氧化氢)引起的细胞凋亡作用,分析该作用的量效关系。方法:实验于2002-09/2003-04在解放军总医院老年医学研究所老年医学实验室完成。①含药血清制备:选用3月龄SD大鼠,随机分为4组:对照组,参芪脑保6.4,3.2,1.6g/kg组,每组12只。参芪脑保6.4,3.2,1.6g/kg组:将参芪脑保颗粒剂(成分:黄芪、丹参、当归尾、桃仁、红花、川芎、赤芍、地龙,10g/包,由解放军总医院制剂室提供,批号000831)按6.4,3.2,1.6g/kg剂量溶于2mL蒸馏水中灌胃;对照组:给予等量蒸馏水。各组连续灌胃7d后腹主动脉取血分离含药血清。②参芪脑宝对正常细胞增殖的影响:采用大鼠PC12细胞作为体外神经细胞模型。将对数生长期细胞接种于96孔培养板,将细胞随机分为4组:对照组和参芪脑保6.4,3.2,1.6g/kg组,分别加入含体积分数0.1各组相应含药血清的DMEM培养液,分别于培养24,60和86h进行细胞计数。③参芪脑保抗活性氧引起的细胞凋亡作用:将对数生长期细胞培养24h。将细胞随机分为5组:对照组、过氧化氢损伤组和过氧化氢+参芪脑保6.4,3.2,1.6g/kg组,除对照组外,其余各组均用含过氧化氢(终浓度100μmol/L)和20g/LB27的无血清DMEM培养基处理30min,过氧化氢损伤组和氧化损伤组改用含体积分数0.1的对照组血清培养基,参芪脑保各剂量组改用相应的含药血清培养基继续培养48h。④采用四氮唑蓝比色法测定正常细胞增殖和活性氧损伤后细胞存活数;采用流式细胞仪和DNAladder法观察细胞凋亡及参芪脑保抗凋亡作用及其剂量效应关系。⑤多组间差异采用方差分析,根据方差齐性结果,采用非配对t检验作两组间比较。结果:①参芪脑保对正常细胞增殖的影响:与对照组相比,参芪脑保6.4,3.2,1.6g/kg组在培养24,48和86h时均能明显�AIM： To study the effects of senqi naobao on normal cell proliferation and the protection against apoptosis induced by reactive oxygen species （100 μmol/L H2O2） in cultured PC12 cells. METHODS： The experiment was completed in the Department of Gerontology, Institute of Gerontology and Geriatrics, General Hospital of Chinese PLA from September 2002 to April 2003. ① Preparation of blood serum： Forty-eight SD rat with 3-month old were randomly divided into four groups： control group, senqi naobao 6.4 g/kg, 3.2 g/kg and 1.6 g/kg groups with 12 in each group. Senqi naobao 6.4 g/kg, 3.2 g/kg and 1.6 g/kg groups： Senqi naobao （component： huangqi, danshen, dangguiwei, taoren, honghua, chuanxiong, chishao and dilong, 10 g/bag, provided by Agent Department of General Hospital of Chinese PLA, batch number： 000831） was dissolved into 2 mL distilled water as the dosage of 6.4 g/kg, 3,2 g/kg and 1.6 g/kg for perfusion. Control group： Rats in control group were treated with the same volume of saline. The animals were anesthetized after 7 days by daily stomach perfusion and the sera containing the drug component were isolated from the major artery. ② Effect of senqi naobao on proliferation of normal cell： The rat pheochromocytoma cell line （PC12） was used as the neuronal cell model in vitro. Cells＇ were planted in 96-well culture plate, and were randomly divided into 4 groups： control group and senqi naobao 6.4 g/kg, 3.2 g/kg and 1.6 g/kg groups. 0.1 volume of related serum was added into DMEM culture medium of each group. Numbers were counted at 24, 60 and 86 hours respectively. ③ Effect of senqi naobao on apoptosis induced by active oxygen： Cells growing in the logarithm proliferation period were cultured for 24 hours. The cells were randomly divided into 5 groups： control group （control group sera）, ROS damage group （H2O2 ＋control group sera）, senqi naobao large middle and small dose groups （H2O2 ＋ senqi naobao 6.4 g/kg, 3.2 g/kg and 1.6 g/kg group sera）. Except con

关 键 词：中药学 细胞凋亡 活性氧 补阳还五汤/投药和剂量 

分 类 号：R243[医药卫生—中医临床基础]