Cadmium chloride (CdCl2) is a known genotoxic carcinogen, with a mechanism of action thought to partly involve the generation of reactive oxygen species (ROS). We applied here a multi-endpoint approach in vitro to explore the impact of CdCl2 on both the genome and on wider cell biology pathways relevant to cancer. Multi-endpoint approaches are believed to offer greater promise in terms of understanding the holistic effects of carcinogens in vitro. This richer understanding may help better classification of carcinogens as well as allowing detailed mechanisms of action to be identified. We found that CdCl2 caused DNA damage (micronuclei; MN) in both TK6 and NH32 cells in a dose dependent manner after 4 hours exposure (plus 23 hours recovery), with lowest observable effect levels (LOELs) for MN induction of 1μM (TK6) and 1.6μM (NH32). This DNA damage induction in TK6 cells was ROS dependent as pre-treatment with the antioxidant N Acetyl Cysteine (1mM), abrogated this effect. However, DCFDA was not capable of detecting the ROS induced by CdCl2. The use of NH32 cells allowed an investigation of the role of p53 as they are a p53 null cell line derived from TK6. NH32 showed a 10-fold increase in MN in untreated cells and a similar dose dependent effect after CdCl2 treatment. In TK6 cells, CdCl2 also caused activation of p53 (accumulation of total and phosphorylated p53), imposition of cell cycle checkpoints (G2/M) and intriguingly the production of smaller and more eccentric (elongated) cells. Overall, this multi-endpoint study suggests a carcinogenic mechanism of CdCl2 involving ROS generation, oxidative DNA damage and p53 activation, leading to cell cycle abnormalities and impacts of cell size and shape. This study shows how the integration of multiple cell biology endpoints studied in parallel in vitro can help mechanistic understanding of how carcinogens disrupt normal cell biology.

中文翻译：

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氯化镉诱导的基因毒性的多端点分析显示活性氧和 p53 激活在 DNA 损伤诱导、细胞周期不规则和细胞大小畸变中的作用


氯化镉 (CdCl2) 是一种已知的基因毒性致癌物，其作用机制被认为部分涉及活性氧 (ROS) 的产生。我们在体外应用多端点方法来探索 CdCl2 对基因组以及与癌症相关的更广泛细胞生物学途径的影响。多端点方法被认为在了解体外致癌物的整体影响方面提供了更大的希望。这种更丰富的理解可能有助于更好地对致癌物进行分类，并确定详细的作用机制。我们发现，暴露 4 小时（加上 23 小时恢复）后，CdCl2 以剂量依赖性方式对 TK6 和 NH32 细胞造成 DNA 损伤（微核；MN），1μM (TK6) 诱导 MN 的可观察效应水平 (LOEL) 最低和 1.6μM (NH32)。 TK6 细胞中的这种 DNA 损伤诱导是 ROS 依赖性的，因为用抗氧化剂 N 乙酰半胱氨酸 (1mM) 进行预处理，消除了这种效应。然而，DCFDA 无法检测 CdCl2 诱导的 ROS。使用 NH32 细胞可以研究 p53 的作用，因为它们是源自 TK6 的 p53 无效细胞系。 NH32 在未处理的细胞中显示 MN 增加 10 倍，并且在 CdCl2 处理后具有类似的剂量依赖性效应。在 TK6 细胞中，CdCl2 还引起 p53 的激活（总 p53 和磷酸化 p53 的积累）、施加细胞周期检查点 (G2/M) 以及有趣的是产生更小、更偏心（拉长）的细胞。总体而言，这项多终点研究表明 CdCl2 的致癌机制涉及 ROS 生成、氧化 DNA 损伤和 p53 激活，导致细胞周期异常以及细胞大小和形状的影响。 这项研究表明，体外平行研究的多个细胞生物学终点的整合如何有助于从机制上理解致癌物如何破坏正常细胞生物学。