caspase-3、caspase-9在醋酸铅诱导NRK细胞凋亡中的作用  

The role of caspase-3 and caspase-9 in apoptosis in NRK cells induced by lead acetate

作　　者：贺秀媛[1] 陈渊[1,2] 李梦[3] 袁莉芸[2] 邬静[2] 李静[1] 常洪涛[1] 郭成志[2] 袁慧[2] 

机构地区：[1]河南农业大学牧医工程学院,河南郑州450002 [2]湖南农业大学动物医学院,湖南长沙410128 [3]华南农业大学兽医学院,广东广州510642

出　　处：《中国兽医学报》2014年第9期1513-1519,共7页Chinese Journal of Veterinary Science

基　　金：河南省科技攻关资助项目(132102110036)

摘　　要：将体外培养的大鼠肾小管上皮细胞NRK分别用0、0.5、1.0、2.0μmol/L醋酸铅作用12h后,MTT法检测不同剂量的醋酸铅对其增殖的抑制率,Annexin V-FITC/PI双染法检测醋酸铅引起的NRK凋亡,罗丹明123检测线粒体膜电位的变化,Western blot检测caspase-3和caspase-9蛋白的表达,以及caspase广谱抑制剂和caspase-9的抑制剂(Z-VAD-FMK和Ac-LEHD-FMK)对醋酸铅诱导细胞凋亡的抑制作用。结果显示,与对照组相比,细胞存活率明显下降(P<0.05或P<0.01),醋酸铅明显抑制NRK细胞增殖,且呈一定剂量效应,IC50为(2.44±0.32)μmol/L;凋亡检测表明,随着醋酸铅浓度的升高,细胞凋亡现象越明显;明显减低线粒体膜电位;激活体caspase-3和caspase-9的表达水平呈剂量依赖性增加;caspase抑制剂能显著抑制醋酸铅诱导的细胞凋亡。结果表明,醋酸铅抑制NRK细胞增殖,可能通过激活caspase依赖性的线粒体信号通路而诱导NRK细胞凋亡。This study aims to investigate whether lead acetate（Pb ion）induces apoptosis and its potential signaling pathway in normal rat kidney epithelial cells（NRK cells）.NRK cells were treated with different concentrations of lead acetate（0,0.5,1.0,2.0μmol/L）for 12 hto determine the cytotoxicity by MTT assay and apoptosis by Hochest 33342 staining and flow cytometry.Mitochondrial membrane potential（ΔΨm）was analyzed by fluorescence spectrophotometer.The expressions and activities of caspase-3and-9were determined by Western blot and calorimetric activity assay kit.After pretreated with caspase inhibitor Z-VAD-FMK and Ac-LEHD-FMK,apoptosis rate of NRK cells was evaluated by flow cytometry.Our results showed that lead acetate inhibited proliferation and viability of NRK cells significantly.The IC50 of lead acetate was（2.44±0.32）μmol/L.The lead acetate changed the mitochondrial membrane potential and caused apoptosis of NRK cell via the caspase-3and-9pathways,which was further verified by the apoptosis of NRK cells blocked by adding the caspase inhibitors.We concluded that lead acetate changed mitochondrial membrane potential,which induced the release of substances in mitochondria,and stimulated the initiation of apoptosis pathway in which caspase-3and-9were involved.

关 键 词：醋酸铅 肾小管上皮细胞 凋亡 线粒体信号通路 

分 类 号：S852.2[农业科学—基础兽医学]