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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 investigate the effects of Cr (Ⅵ) on the gene expression of mitochondrial ATPase 6 and ATPase 8 and its energy metabolism in hepatocytes. Methods The human embryonic L-02 hepatocytes cultured in vitro were treated with Cr (Ⅵ) 2, 8 and 32 μmol·L-1 for 24 h. RNA, mitochondrial ATPase 6 and ATPase 8 mRNA The levels of ATP, mitochondrial respiratory chain complex enzyme activity and reactive oxygen species (ROS) content were determined by chemiluminescence, ultraviolet spectrophotometry and Fluorescence spectrophotometry. Results Compared with the normal control group, Cr (Ⅵ) 2, 8 and 32 μmol·L-1 significantly increased ROS (P <0.05). Cr (Ⅵ) 2 μmol·L- (P <0.05), and the levels of ATPase 6 and ATPase 8 (P <0.05), Cr (Ⅵ) 8 and 32μmol·L-1 decreased significantly Content decreased with the increase of Cr (Ⅵ) concentration (P <0.05). There was a positive correlation between the expression of ATPase 6 and ATPase 8 and the activity of respiratory chain complex Ⅴ and the content of intracellular ATP (r = 0.858,0.795,0.809,0.766, P <0.01) 0.738, -0.801, P <0.01). Conclusion Cr (Ⅵ) can induce changes of the expression of ATPase 6 and ATPase 8 in hepatocytes, resulting in the decrease of enzyme V activity and intracellular ATP content in mitochondrial respiratory chain complex.