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.
出处
《中国药理学与毒理学杂志》
CAS
CSCD
北大核心
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)~~
关键词
铬
肝细胞
线粒体
能量代谢
腺苷三磷酸酶类
chromium
hepatocytes
mitochondria
energy metabolism
adenosine triphophatases
