远志皂苷对β淀粉样蛋白片段1-40诱导PC12细胞凋亡的抑制作用  

Protection of tenuigenin against apoptosis of PC12 cells induced by amyloid beta-protein fragment 1-40

作　　者：杨贤志[1] 陈勤[1] 陈庆林[1] 金蓓蓓[1] 叶海燕[1] 

机构地区：[1]安徽大学生命科学学院安徽省中药研究与开发重点实验室,安徽合肥230039

出　　处：《中国药理学与毒理学杂志》2013年第3期379-384,共6页Chinese Journal of Pharmacology and Toxicology

基　　金：安徽省教育厅自然科学研究项目(KJ2009A116);安徽大学大学生科研训练计划项目(201135);安徽大学研究生学术创新研究项目(2010121)~~

摘　　要：目的探讨远志皂苷抑制β淀粉样蛋白片段1-40(Aβ1-40)诱导的PC12细胞凋亡的作用机制。方法采用聚集状的Aβ1-40 25μmol·L-1诱导PC12细胞凋亡,然后将处理后的PC12细胞分为Aβ1-40模型组和远志皂苷50,100和200μmol·L-1组,同时设正常细胞对照组。采用MTT比色法检测细胞存活率;膜联蛋白-Ⅴ和PI双染法检测细胞凋亡率;免疫细胞化学法检测细胞凋亡基因Bcl-2和Bax及细胞色素c(Cyt c)表达阳性的细胞百分率;Western印迹法检测PC12细胞中Cyt c的表达水平。结果与正常对照组比较,Aβ1-40模型组PC12细胞的存活率明显降低(P<0.01),为(31±7)%;Bcl-2阳性表达细胞率降低(P<0.01),为(23.9±1.9)%;Bax和Cyt c阳性表达细胞率升高(P<0.01),分别为(79.0±3.7)%和(49.2±3.6)%,Bcl-2/Bax阳性表达细胞比值为0.30。与模型对照组比较,远志皂苷50,100和200μmol·L-1作用24 h后,细胞存活率分别升高至(51±13)%,(64±7)%和(84±10)%(P<0.01);Bcl-2阳性率升高至(38.7±0.9)%,(53.7±1.6)%和(60.3±0.8)%(P<0.01),Bax阳性率分别降低为(60.8±1.9)%,(41.5±2.2)%和(32.7±1.4)%(P<0.01),Bcl-2/Bax比值亦分别上升为0.64,1.29和1.84;Cyt c阳性率分别降低至(45.4±3.4)%,(30.2±2.2)%和(27.5±1.0)%(P<0.05,P<0.01)。与正常对照组比较,模型组PC12细胞凋亡率和Cyt c蛋白表达水平亦明显升高(P<0.01);远志皂苷50,100和200μmol·L-1作用24h,PC12细胞凋亡率和Cyt c表达水平较模型组均降低(P<0.01)。结论远志皂苷对Aβ1-40诱导的PC12细胞凋亡具有明显的抑制作用,其作用机制可能是抑制Bax和Cyt c表达,增加Bcl-2表达和Bcl-2/Bax比值,从而阻断内源性细胞凋亡通路。OBJECTIVE To investigate the protective mechanism of tenuigenin（TEN） on the apoptosis of PC12 cells induced by amyloid beta-protein 1-40（Aβ1-40） in vitro.METHODS Aggregated Aβ1-40 25 μmol·L-1 which induced the apoptosis of PC12 cells was used to establish Alzheimer′s disease neuronal cell model.These model neurons were divided into Aβ1-40 model group and TEN 50,100 and 200 μmol·L-1 groups.At the same time,normal cell control group was established without Aβ1-40 pretreatment.The survival rate of PC12 cells was detected by MTT assay.The apoptosis rate of PC12 cells was detected by flow cytometery with Annexin-Ⅴ/PI double staining.The rates of positive cells expressed Bcl-2,Bax and cytochrome c（Cyt c） were observed by immunocytochemical method.The expression level of Cyt c was detected through Western blotting analysis.RESULTS Compared with normal control group,survival rate of PC12 cell decreased to（31±7）%（P〈0.01）,the positive rate of Bcl-2 was declined to（23.9±1.9）%（P〈0.01）,the positive rate of Bax and Cyt c increased to（79.0±3.7）% and（49.2±3.6）%（P〈0.01）,respectively,and the ratio of Bcl-2/Bax was 0.30 in Aβ1-40 model group.Compared with model group,after 24 h incubation of with PC12 cells TEN 50,100 and 200 μmol·L-1,PC12 cell survival rate increased to（51±13）%,（64±7）% and（84±10）%（P〈0.01）,respectively;the positive rate of Bcl-2 increased to（38.7±0.9）%,（53.7±1.6）% and（60.3±0.8）%（P〈0.01）,respectively;the positive rate of Bax was declined to（60.8±1.9）%,（41.5±2.2）% and（32.7±1.4）%（P〈0.01）,and the ratio of Bcl-2/Bax increased to 0.64,1.29 and 1.84;the positive rate of Cyt c decreased to（45.4±3.4）%,（30.2±2.2）% and（27.5±1.0）%（P〈0.05,P〈0.01）.Compared with normal control group,the apoptosis rate in PC12 cells and expression level of Cyt c protein were also obviously elevated（P〈0.01）;and after 24 h incubation of TEN 50,100 and 200 μmol·L-1 with PC12 cells,the apoptosis

关 键 词：远志皂苷 β淀粉样蛋白片段1-40 PC12细胞 细胞凋亡 细胞色素C 

分 类 号：R285[医药卫生—中药学]