五氯酚对稀有鮈鲫卵黄蛋白原及p53的诱导效应  

Induction Effects of Pentachlorophenol on Vitellogenin and p53 in Chinese Rare Minnow(Gobiocypris rarus)

作　　者：熊力[1] 马永鹏[2] 张晓峥[1] 金帮明[1] 李伟[1] 苏永良[1] 毛思予[1] 刘堰[1] 

机构地区：[1]西南大学生命科学学院,淡水鱼类资源与生殖发育教育部重点实验室,三峡库区生态环境教育部重点实验室,水产科学重庆市市级重点实验室,重庆400715 [2]重庆市第三人民医院检验科,重庆市临床检验中心,重庆400016

出　　处：《环境科学》2012年第6期1858-1864,共7页Environmental Science

基　　金：国家自然科学基金项目(21147002);国家重点基础研究发展规划(973)项目(2012CB723205;2010CB134405);重庆市自然科学基金项目(CSTC.2009BB1131)

摘　　要：采用中国特有鱼种稀有鮈鲫为实验动物,研究五氯酚(PCP)对其肝脏卵黄蛋白原(VTG)、VTG基因及抑癌基因p53的诱导效应,评价PCP的内分泌干扰作用,并从蛋白和基因表达水平上筛选PCP检测的敏感生物标志物.1.5、15、40、80、120、150、160μg.L-1PCP暴露稀有鮈鲫,同时设置空白、溶剂对照和17α-雌二醇(EE2)阳性对照,运用SDS-PAGE,ELISA法检测经PCP暴露后稀有鮈鲫肝脏VTG蛋白的表达差异;克隆稀有鮈鲫新的VTG和p53基因序列片段,运用实时荧光定量PCR方法检测PCP暴露对基因表达的影响.结果表明,40、80、120、160μg.L-1PCP均能诱导雌雄稀有鮈鲫肝脏VTG蛋白的表达,具有显著的浓度-效应关系;1.5、15、150μg.L-1PCP暴露后,雄性稀有鮈鲫肝脏VTG和p53基因表达均明显升高,具有显著的浓度-效应和时间-效应关系.证明PCP具有雌激素效应,稀有鮈鲫肝脏的VTG蛋白、VTG及p53基因可作为PCP检测的敏感分子生物标志物.Taking the Chinese rare minnow(Gobiocypris rarus) as the test animal,the studies were designed to investigate induction effects of pentachlorophenol(PCP) on vitellogenin(VTG) protein,VTG gene and tumor suppressor p53 gene in the liver of Gobiocypris rarus.The endocrine disrupting of PCP was evaluated by detecting VTG,and sensitive biomarkers of PCP were screened at both protein and mRNA levels.Gobiocypris rarus were exposed to PCP at 1.5,15,40,80,120,150,160 μg·L-1 respectively,while setting blank,solvent control and 17α-ethynylestradiol(EE2) as positive control.Using sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE),enzyme-linked immunosorbent assay(ELISA),VTG protein expression differences were detected in the liver of Gobiocypris rarus after exposure to PCP.Cloning the VTG and p53 gene new fragments of Gobiocypris rarus based on conserved regions,mRNA expression levels of VTG and p53 gene in the liver of Gobiocypris rarus were determined by quantitative real-time PCR assay after PCP treatment.The results showed that 40,80,120,160 μg·L-1 PCP induced the liver of male and female Gobiocypris rarus to produce VTG protein,and had a significant concentration effect.VTG and p53 mRNA levels significantly increased in the liver of Gobiocypris rarus after exposure to 1.5,15,150 μg·L-1 PCP,and had remarkable concentration and time effects.The studies suggested that PCP had estrogenic effects,and VTG protein,VTG and p53 gene in the liver of Gobiocypris rarus could be used as candidate sensitive biomarkers for detecting PCP.

关 键 词：五氯酚 稀有鮈鲫 VTG P53 分子生物标志物 

分 类 号：X171.5[环境科学与工程—环境科学]