Arsenite induces delayed mutagenesis and transformation in human osteosarcoma cells at extremely low concentrations

Abstract

Arsenite is a human multisite carcinogen, but its mechanism of action is not known. We recently found that extremely low concentrations (</=0.1 microM) of arsenite transform human osteosarcoma TE85 (HOS) cells to anchorage-independence. In contrast to other carcinogens which transform these cells within days of exposure, almost 8 weeks of arsenite exposure are required for transformation. We decided to reexamine the question of arsenite mutagenicity using chronic exposure in a spontaneous mutagenesis assay we previously developed. Arsenite was able to cause a delayed increase in mutagenesis at extremely low concentrations (</=0.1 microM) in a dose-dependent manner. The increase in mutant frequency occurred after almost 20 generations of growth in arsenite. Transformation required more than 30 generations of continuous exposure. We also found that arsenite induced gene amplification of the dihydrofolate reductase (DHFR) gene in a dose-dependent manner. Since HOS cells are able to methylate arsenite at a very low rate, it was possible that active metabolites such as monomethylarsonous acid (MMA(III)) contributed to the delayed mutagenesis and transformation in these cells. However, when the assay was repeated with MMA(III), we found no significant increase in mutagenesis or transformation, suggesting that arsenite-induced delayed mutagenesis and transformation are not caused by arsenite's metabolites, but by arsenite itself. Our results suggest that long-term exposure to low concentrations of arsenite may affect signaling pathways that result in a progressive genomic instability.

ATF4-Dependent Oxidative Induction of the DNA Repair Enzyme Ape1 Counteracts Arsenite Cytotoxicity and Suppresses Arsenite-Mediated Mutagenesis

“…An improved comet assay using Fpg glycosylase as a probe provided new evidence that arsenite levels equivalent to those associated with clinical pathology (1 to 500 nM) were indeed able to cause rapid (1-to 6-h) formation of DNA lesions, such as 8-oxoguanine and AP sites (55,59,69). For arsenic mutagenesis and tumorigenesis, Mure et al (45) showed that human osteosarcoma TE85 cells cultured with 100 nM sodium arsenite for 20 generations (3 weeks) increased the mutation frequency at the HPRT locus three-to fivefold, and after 30 generations (5 weeks), phenotypically transformed cells could be detected. Mutations were induced by arsenite but not by methyl arsenic derivates (45).…”

Altered iron homeostasis involvement in arsenite-mediated cell transformation

“…Control parental HOS cells were cultured in α-MEM medium containing 5 or 10% FBS. Two types of As-treated cells were generated by incubating cells with a low dose of arsenite (0.1 µM), as previously described [13,14]. Cells designated here as As-8w-HOS were cells that were chronically exposed to NaAsO 2 for 8 weeks.…”

“…As previously described, HOS cell transformation occurred only after 8 weeks of low-dose arsenite treatment [13,14]. Ferritin levels in the parental HOS cells were measured as a function of time of exposure to 0.1 µM arsenite.…”

“…Other studies show that arsenite is a complete transplacental carcinogen in mice [6], while dimethylarsinic acid, a major metabolite of arsenic in most mammals, including humans, causes bladder cancer in F344 rats [7]. In contrast to its weak mutagenicity, arsenite induces cell transformation of various types of cells to a more malignant phenotype, such as Syrian hamster embryo cells [8], mouse embryo fibroblasts [9], mouse epidermal JB6 C141 cells [10], rat liver-derived TRL 1215 cells [11], normal human immortalized prostate epithelium RWPE-1 cells [12], as well as human osteogenic sarcoma (HOS) cells [13,14]. Many mechanisms involving arsenic-mediated cell transformation have been proposed.…”

“…Using a previously developed model of a low-dose and long-term exposure to arsenite [14], we obtained transformed human osteogenic sarcoma (As-T-HOS) cells, which have exhibited anchorage independence [13]. We found that arsenite can affect iron homeostasis through an abnormal iron regulation mechanism.…”