As_2O_3诱导肿瘤细胞凋亡依赖H_2O_2途径

Solid tumor cell apoptosis induced by arsenic trioxide is related to generation of reactive oxygen species

目的:探讨三氧化二砷(arsenic trioxide,As2O3)对人乳头瘤病毒(HPV)阳性宫颈癌HeLa细胞和HPV阴性胰腺癌AsPC-1细胞的凋亡诱导作用与细胞内H2O2水平的关系。方法:采用不同浓度的As2O3处理HeLa细胞和AsPC-1细胞不同时段,显微镜下观察细胞的凋亡,噻唑蓝(MTT)法测定细胞生长抑制情况;不同浓度的As2O3处理细胞2、5、8、12、24h后,用DCFH-DA标记检测细胞内的H2O2水平。结果:2μmol/L As2O3处理HeLa细胞48h后细胞的生长受到明显抑制,表现出细胞凋亡的特征,随着As2O3浓度的增加和作用时间的延长效果更加明显。而1μmol/L As2O3处理AsPC-1细胞24h即呈现明显的生长抑制和凋亡特征。As2O3处理HeLa细胞2h细胞内H2O2的水平比对照组升高(10μmol/L组达51·30%),持续到8h达到高峰(升高84·19%),12h后下降,24h水平与对照组接近,并有明显的剂量依赖性。As2O3处理AsPC-1细胞2h后,细胞内H2O2的水平也明显升高(10μmol/L组达79%),持续到5h达到高峰(增高169%),且该细胞内H2O2水平升高比HeLa细胞更明显,12h之后的变化情况与HeLa细胞类似。结论:As2O3诱导HeLa细胞和AsPC-1细胞凋亡与细胞内的H2O2水平改变有关,而与HPV的感染无明显相关性。H2O2积累是As2O3诱导细胞凋亡途径中的早期事件,H2O2可能在As2O3诱导肿瘤细胞凋亡途径中扮演类似第二信使的作用。

AIM： To study the effects of As2O3 on the growth of human cervical carcinoma cell line, HeLa cells, human pancreatic carcinoma cell line, AsPC- 1 cells, and the generation of reactive oxygen species （ROS） in the cells. METHODS： HeLa cells and AsPC- 1 cells were treated with various concentrations of As2O3. The effects of As2O3 on HeLa cells and AsPC - 1 cells survival and apoptosis were determined by MTT assay and light microscope, and cellular ROS was also measured by fluorometer. RESULTS： After being treated with 2μmol/L As2O3 for 48 h, the survival of HeLa cells was deceased, a marked apoptosis characteristic was observed in time- and dose- dependent manner. However, the survival of AaPC - 1 cells markedly decreased after being treated with even 1μmol/L As2O3 for 24 h. When we measured the cellular hydrogen peroxide （H2O2） level, we we lound that the H2O2 level in HeLa cells started to rise after being incubated with As2O3 for 2 h, and sustained increased, reached the peak of H2O2 level at 8 h, then it began to decrease. While the H2O2 level in AsPC - 1 cells started to rise at 2 h, reached the highest point at 5 h, after which it began to descend. At 12 h, the H2O2 level in HeLa cells and AsPC - 1 cells were similar to that of the control group. CONCLUSION： The molecular mechanisms underlying As2O3 - induced apoptosis in cells derived from the solid tumors may be related to the cellular ROS level.