Cr（Ⅵ）在细胞p53封闭条件下的线粒体损伤应-卫生毒理学专业毕业论文.docx

PFT-a+Cr(VI)染毒组与PFT-0【+Z．VAD．fmk+Cr(VI)染毒组细胞Bcl．2和 PFT-a+Cr(VI)染毒组与PFT-0【+Z．VAD．fmk+Cr(VI)染毒组细胞Bcl．2和 Bax基因表达减少，加入NAC后均出现相应基因表达的增多。 6． 细胞核DNA损伤： Cr(VI)、PFT-a+Cr(VI)与 PFT．0c+Z．VAD．fmk+Cr(VI)染毒组细胞均出现凋亡典型的DNA梯带降 解现象，加入NAC后未见明显保护作用。 结论：p53封闭条件下，20I_tmol／L C“Ⅵ)能明显引起L．02肝细胞 氧化应激，导致线粒体膜损伤、cyt C与凋亡诱导因子(AIF)释放与 细胞DNA损伤；同时在抑带ljcaspase条件下，受试细胞仍然表现凋亡 现象。结果提示，Cr(Ⅵ)可能绕过p53／caspase环节直接通过损伤细胞 线粒体而诱导凋亡。 关键词： 六价铬，L．02肝细胞，线粒体损伤，PFT-et II Mitochondrial Mitochondrial damage induced by hexavalent chromium under the condition of p53 closed in L··02 hepatocytes ABSTRACT objective：To explore the mitochondria damage induced by hexavalent chromium(Cr(VI)under the condition of p53 closed in L一02 hepatocytes，and to further investigate the pathway of apoptosis induced independently by mitochondria damage in L-02 hepatocytes treated with Cr(VI)． Methods：L-02 hepatocytes were cultured，and MTT test was adopted to test the viability of L一02 hepatocytes treated with PFT-aalone and／or with Cr(VI)．The detectation of p5 3 protein level in nuclei of L-02 hepatocytes was performed to verify the inhibitive effect of PFT-a on p5 3， in order to select the exposure concentrations of PFT-a and Cr(VI)．The concentrations of PFT-a and Cr(VI)were both designed at 201xmol／L． After cells were treated with PFT-a for 4hrs，and then treated with Cr(VI) for 24hrs． The cultured L-02 hepatocytes were randomly divided into six groups，namely control group，Cr(VI)alone，PFT-a alone，PFT-a+Cr(VI)， PFT-仪+Z．VAD．fmk+Cr(VI)，and PFT-叶NAC+C“Ⅵ)．The activity of SOD and GST and the levels of MDA and GSH were detected by the colorimetric method．The mitochondrial permeability transition pore(PTP) and the mitochondnal transmembrane potential(Arm)were determined by the fluorospectrophotometric method．卟e protein expression levels of p53，cytochrome C and apoptosis-inducing factor(AIF)were assayed with the western blotting．The gene expressions of Bax and Bcl一2 were detected with the reverse transcription polymerase chain reaction(RT-P