中华急诊医学杂志
中華急診醫學雜誌
중화급진의학잡지
CHINESE JOURNAL OF EMERGENCY MEDICINE
2008年
12期
1280-1284
,共5页
战丽彬%路小光%林海燕%隋华%宫晓洋
戰麗彬%路小光%林海燕%隋華%宮曉洋
전려빈%로소광%림해연%수화%궁효양
内质网应激%神经元凋亡%葡萄糖调节蛋白78%CCAAT/增强子结合蛋白同源蛋白%滋补脾阴方约含药血清
內質網應激%神經元凋亡%葡萄糖調節蛋白78%CCAAT/增彊子結閤蛋白同源蛋白%滋補脾陰方約含藥血清
내질망응격%신경원조망%포도당조절단백78%CCAAT/증강자결합단백동원단백%자보비음방약함약혈청
Endoplasmic reticdum stress%Neuron apoptosis%Glucose regulated protein 78%CCAAT/enhancer-binding protein-homologous protein%Zibu Piyin Recipe(ZBPYR),serum
目的 运用中药血清药理学方法研究滋补脾阴方药含药血清对内质网应激致神经元凋亡作用及其机制.方法 实验中体内实验于辽宁省SPF动物重点实验室完成,体外实验于辽宁省脑疾病研究重点实验室完成.SPF级健康雄性SD大鼠(220～250 g)12只,随机分为对照组和滋补脾阴方药(ZBPYR)组,每组6只,制备空白及ZBPYR含药血清.以N-糖链抑制剂农霉素(tunicamycin,Tm,5压计μml)刺激小鼠神经瘤母细胞(Neuro2a)建立内质网应激模型,各浓度滋补脾阴方药含药血清(5%,10%,15%)为干预组,空白血清组为对照组,采用四甲基偶氮唑盐(MTT)方法观察Neuro2a细胞牛存率;流式细胞技术观察Neuro2a细胞凋亡;蛋白质免疫印迹(Western blotting)法观察葡萄糖调节蛋白78(GRP78)、凋亡促进因子CCAAT/增强子结合蛋白同源蛋白(CHOP)的蛋白表达.统计结果行SNK-q检验.结果 各浓度ZBPYR含药血清预处理组(生存率:5%0.5295±0.0373,10%0.5843±0.0428,15%0.6274±0.0324;凋亡率:5%47.8733±2.8166,10%46.3366±1.2748,15%39.8833±1.0524)与Tm组(牛存率:0.1673±0.0213;凋亡率:62.7050±1.4056)相比,细胞牛存率明显升高(P<0.05);细胞凋亡率显著降低(P<0.05);各浓度ZBPYR含药血清预处理组GRP 78(5%2.1228±0.2251,10%1.3293±0.9443.15%;15%0.0931±0.1168)及CHOP(5%1.1776±0.2927,10%0.7290±0.1708,15%0.6577±0.1883)蛋白表达与Tm组(GRP 78 2.9149±0.5355;CHOP 1.6611±0.2913)相比,表达水平明显下调(P<0.05).结论 ZBPYR能提高Tm刺激后的Neuro2a细胞生存率,抑制细胞凋亡,具有神经保护作用,其机制可能是减轻内质网应激及抑制内质网应激凋亡通路.
目的 運用中藥血清藥理學方法研究滋補脾陰方藥含藥血清對內質網應激緻神經元凋亡作用及其機製.方法 實驗中體內實驗于遼寧省SPF動物重點實驗室完成,體外實驗于遼寧省腦疾病研究重點實驗室完成.SPF級健康雄性SD大鼠(220～250 g)12隻,隨機分為對照組和滋補脾陰方藥(ZBPYR)組,每組6隻,製備空白及ZBPYR含藥血清.以N-糖鏈抑製劑農黴素(tunicamycin,Tm,5壓計μml)刺激小鼠神經瘤母細胞(Neuro2a)建立內質網應激模型,各濃度滋補脾陰方藥含藥血清(5%,10%,15%)為榦預組,空白血清組為對照組,採用四甲基偶氮唑鹽(MTT)方法觀察Neuro2a細胞牛存率;流式細胞技術觀察Neuro2a細胞凋亡;蛋白質免疫印跡(Western blotting)法觀察葡萄糖調節蛋白78(GRP78)、凋亡促進因子CCAAT/增彊子結閤蛋白同源蛋白(CHOP)的蛋白錶達.統計結果行SNK-q檢驗.結果 各濃度ZBPYR含藥血清預處理組(生存率:5%0.5295±0.0373,10%0.5843±0.0428,15%0.6274±0.0324;凋亡率:5%47.8733±2.8166,10%46.3366±1.2748,15%39.8833±1.0524)與Tm組(牛存率:0.1673±0.0213;凋亡率:62.7050±1.4056)相比,細胞牛存率明顯升高(P<0.05);細胞凋亡率顯著降低(P<0.05);各濃度ZBPYR含藥血清預處理組GRP 78(5%2.1228±0.2251,10%1.3293±0.9443.15%;15%0.0931±0.1168)及CHOP(5%1.1776±0.2927,10%0.7290±0.1708,15%0.6577±0.1883)蛋白錶達與Tm組(GRP 78 2.9149±0.5355;CHOP 1.6611±0.2913)相比,錶達水平明顯下調(P<0.05).結論 ZBPYR能提高Tm刺激後的Neuro2a細胞生存率,抑製細胞凋亡,具有神經保護作用,其機製可能是減輕內質網應激及抑製內質網應激凋亡通路.
목적 운용중약혈청약이학방법연구자보비음방약함약혈청대내질망응격치신경원조망작용급기궤제.방법 실험중체내실험우요녕성SPF동물중점실험실완성,체외실험우요녕성뇌질병연구중점실험실완성.SPF급건강웅성SD대서(220～250 g)12지,수궤분위대조조화자보비음방약(ZBPYR)조,매조6지,제비공백급ZBPYR함약혈청.이N-당련억제제농매소(tunicamycin,Tm,5압계μml)자격소서신경류모세포(Neuro2a)건립내질망응격모형,각농도자보비음방약함약혈청(5%,10%,15%)위간예조,공백혈청조위대조조,채용사갑기우담서염(MTT)방법관찰Neuro2a세포우존솔;류식세포기술관찰Neuro2a세포조망;단백질면역인적(Western blotting)법관찰포도당조절단백78(GRP78)、조망촉진인자CCAAT/증강자결합단백동원단백(CHOP)적단백표체.통계결과행SNK-q검험.결과 각농도ZBPYR함약혈청예처리조(생존솔:5%0.5295±0.0373,10%0.5843±0.0428,15%0.6274±0.0324;조망솔:5%47.8733±2.8166,10%46.3366±1.2748,15%39.8833±1.0524)여Tm조(우존솔:0.1673±0.0213;조망솔:62.7050±1.4056)상비,세포우존솔명현승고(P<0.05);세포조망솔현저강저(P<0.05);각농도ZBPYR함약혈청예처리조GRP 78(5%2.1228±0.2251,10%1.3293±0.9443.15%;15%0.0931±0.1168)급CHOP(5%1.1776±0.2927,10%0.7290±0.1708,15%0.6577±0.1883)단백표체여Tm조(GRP 78 2.9149±0.5355;CHOP 1.6611±0.2913)상비,표체수평명현하조(P<0.05).결론 ZBPYR능제고Tm자격후적Neuro2a세포생존솔,억제세포조망,구유신경보호작용,기궤제가능시감경내질망응격급억제내질망응격조망통로.
Objective To explore the inhibitory effects of Zibu Piyin Recipe(ZBPYR)serum on neuron apoptosis induced by tunieamyein(Tm,5 μg/ml)and its mechamsm in vitro by using sero-pharmacological method.Method Totally 12 healthy adult male SD rats(220～250 g)(SPF)were divided randomly into control group and ZBPYR group,6 in each group,then the blank and ZBPYR serum were prepared.The mouse.neuroblastoma cell line Neum2a cells were treated with Tunicamycin(Tin,an inhibitor of N-glycoslytion)to establish the endoplasmic reticulum(ER)stress model.The cells treated by ZBPYR aerum of different concentrations were interventional groups,and the cells treated by blank serum were control group.The viability of Neuro2a cells was meusurcdd by MTT assay.Flow cytometry wus applied to observe the apoptosis of Neuro2a cells.Western blotting was utilized to detect the protein expressions of two molecules,ER molecular chaperone-ucose regulated protein 78(CRP78)and transcriptional factor CCAAT/enhancer-binding protein-homologous protein(CHOP).The results were analyzed by sNK-q test.Results Compared to Tm group(cell viability 0.1673±0.0213,apoptotic rate 62.7050±1.4056),The cell viability of interventional groups(5%0.5295±0.0373,10%0.5843±0.0428,15%0.6274±0.0324)increased significantly(P<0.05);and the apoptotic rate(5%47.8733±2.8166,10%46.3366±1.2748,15%39.8833±1.0524)reduced significantly(P<0.05).The protein expressions of GRP 78(5%2.1228±0.2251,10%1.3293±0.9443,15%;15%0.0931±0.1168)and CHOP(5%1.1776±0.2927,10%0.7290±0.1708,15%0.6577±0.1883)of interventional groups reduced significantly compared with Tm group(GRP78 2.9149±0.5355;CHOP 1.6611±0.2913)P<0.05.Condusions ZBPYR serurn could increase the cell viability of Neuro2a cells treated with Tm and inhibit cell apoptosis.Thereby it may have neuroprotective effects,and the mechanism may be associated with the inhibition of ER stress and apoptosis pathway.
