DNA microarray analysis for HepG2 cells exposed to arsenic trioxide. Homo sapiens

DNA microarray analysis revealed that the genes related to cell cycle regulation were significantly induced at non- and sub-lethal concentrations (i.e., 0.05-6 µM), while the common feature of heavy metal toxicity such as oxidative damage and following increase in glutathione, heat shock proteins, and metallothionein were confirmed at high concentrations (i.e., 6-40 µM). The concentration dependent modulation of gene expression (induction of cell cycle genes, induction of cell cycle arrest genes and apoptotic genes) following exposure to arsenic was further supported by acceleration of cell proliferation, ROS generation, and cytotoxicity. Furthermore, three cell cycle genes (i.e., CDC25B, UBE2C, and PTTG1) were proposed as marker genes of inorganic arsenic exposure. Those results indicated the potential pro-carcinogenic actions of inorganic arsenic occur in environmentally relevant exposures (as low as 0.07 µM). Overall design: In this study, we examined the gene expression alteration in HepG2 cells exposed to arsenic trioxide (5 nM to 40 µM as As2O3 for 48 hours) by DNA microarray with 8795 human genes. HepG2 cells were also exposed to ε-caprolactam as a reference of non carcinogen since it is the only chemical categorized in IARC’s group 4 (Probably Not Carcinogenic). MQ water was used as control. For replicate, three dishes were prepared for each sample and individually treated in parallel.

DNA微阵列（DNA microarray）分析显示，在非致死与亚致死浓度（即0.05~6 µM）条件下，细胞周期调控相关基因被显著诱导；而在高浓度（即6~40 µM）下，可观察到重金属毒性的典型特征，包括氧化损伤、谷胱甘肽水平升高、热休克蛋白及金属硫蛋白表达上调。砷暴露后基因表达的浓度依赖性调控（涵盖细胞周期基因、细胞周期阻滞基因与凋亡基因的诱导），进一步通过细胞增殖加速、活性氧（Reactive Oxygen Species, ROS）生成及细胞毒性得到验证。此外，3个细胞周期基因（即CDC25B、UBE2C及PTTG1）被提议作为无机砷暴露的标志物基因。上述研究结果表明，无机砷的潜在促癌作用可发生于环境相关的暴露浓度下（最低可至0.07 µM）。实验整体设计：本研究采用搭载8795个人类基因的DNA微阵列，检测了三氧化二砷（以As₂O₃计，浓度范围5 nM至40 µM）暴露48小时后的HepG2细胞基因表达变化。同时，本研究以ε-己内酰胺（ε-caprolactam）作为非致癌物参照进行处理——因其是唯一被国际癌症研究机构（International Agency for Research on Cancer, IARC）归为第4组（极可能非致癌物）的化学物质；以超纯水（MQ water）作为空白对照。每个样本设置3个培养皿作为生物学重复，并行开展独立处理。