Cr(Ⅵ)对肝细胞线粒体ATP酶6和ATP酶8基因表达的影响以及与能量代谢障碍的关系  

Effect of Cr(Ⅵ) on mitochondrial ATPase 6 and ATPase 8 genes expression and its relation with dysfunction of energy metabolism in hepatocytes

作　　者：邹悦[1] 钟才高[1] 曾明[1] 刘新民[1] 肖芳[1] 李鹏[1] 杨渊[1] 

机构地区：[1]中南大学公共卫生学院卫生毒理学系,湖南长沙410078

出　　处：《中国药理学与毒理学杂志》2012年第6期853-859,共7页Chinese Journal of Pharmacology and Toxicology

基　　金：国家自然科学基金(30972511)~~

摘　　要：目的探讨Cr(Ⅵ)对肝细胞线粒体ATP酶6和ATP酶8基因表达水平与能量代谢的影响及其相互联系。方法用Cr(Ⅵ)2,8和32μmol·L-1分别处理体外培养的人胚L-02肝细胞24 h后,用细胞总RNA提取试剂盒分离RNA,线粒体ATP酶6和ATP酶8 mRNA表达水平用逆转录-荧光定量聚合酶链反应(qRT-PCR)测定;细胞ATP含量、线粒体呼吸链复合体酶Ⅴ活性与细胞活性氧(ROS)含量分别用化学发光法、紫外分光光度法和荧光分光光度法检测。结果与正常对照组相比,Cr(Ⅵ)2,8和32μmol·L-1使ROS显著升高(P<0.05);与正常对照组相比,Cr(Ⅵ)2μmol·L-1可使ATP酶6和ATP酶8基因表达水平增高(P<0.05),Cr(Ⅵ)8和32μmol·L-1使之明显降低(P<0.05);而呼吸链复合体酶Ⅴ活性及细胞内ATP含量均随Cr(Ⅵ)浓度的增加而显著降低(P<0.05)。ATP酶6和ATP酶8基因表达与呼吸链复合体酶Ⅴ活性及细胞内ATP含量呈正相关(r=0.858,0.795,0.809,0.766,P<0.01),与ROS水平呈负相关(r=-0.738,-0.801,P<0.01)。结论 Cr(Ⅵ)能诱导肝细胞ATP酶6和ATP酶8基因表达水平改变,导致线粒体呼吸链复合体酶Ⅴ活性及细胞内ATP含量降低。OBJECTIVE To explore the correlation between mitochondrial DNA ATPase 6 and ATPase 8 genes expression and dysfunction of energy metabolism in L-02 hepatocytes treated with hexavalent chromium[ Cr（VI）]. METHODS L-02 hepatocytes were treated with Cr（VI） 2, 8 and 32 μmol· L-1, respectively, for 24 h and then harvested. Total RNA was extracted from L-02 hepatocytes using RNA extraction kit. The quantitative reverse transcription polymerase chain reac- tion （qRT-PCR） was applied to detect the mRNA levels of ATPase 6 and ATPase 8 genes. The con- tent of ATP was measured by bioluminescence technique. The activity of mitochondrial respiratory chain complex V and the level of cellular ROS were determined by ultraviolet spectrophotometry and fluorometric methods, respectively. RESULTS Compared with normal control group, ROS level significantly increased in Cr （ VI ） 2, 8 and 32 μmol· L-1 groups. Compared with normal control group, the expression levels of ATPase 6 and ATPase 8 genes in Cr（VI） 2 μmol·L-1 group firstly increased （ P 〈 0.05 ）, then they gradually decreased in Cr（ VI ） 8 and 32 μmol· L -1 groups, while the activity of mitochondrial respiratory chain complex V and cellular ATP level significantly de- creased （P 〈 0.05 ）. The relative analysis showed the complex V activity was positively correlated with ATPase 6 and ATPase 8 genes expression levels （ r = 0. 858,0. 809, P 〈 0.01 ）, and the cellu- lar ATP level was yet positively correlated with ATPase 6 and ATPase 8 genes expression levels （ r =- 0. 795, 0. 766, P 〈 0.01 ）. But ROS level was negatively correlated with ATPase 6 and ATPase 8 genes expression levels （r = -0. 738, -0. 801, P 〈0.01）. CONCLUSION Cr（VI） can induce expression changes in mitochondrial encoding ATPase genes, and this might result in the decrease in the complexV activity and ATP synthesis.

关 键 词：铬 肝细胞 线粒体 能量代谢 腺苷三磷酸酶类 

分 类 号：R114[医药卫生—卫生毒理学]