Persistent genomic instability by arsenic exposure in V79 Chinese hamster cells

Abstract

Previously, we demonstrated that acute treatment with arsenic leads mammalian cells to exhibit persistent chromosomal instability and DNA hypomethylation, by performing investigations after about 2 months of subculturing. In order to evaluate quantitatively the continuing instability during the expanded growth, we carried out cytogenetic, morphologic and molecular analyses immediately after exposure and every week up to 112 cell generations. Briefly, V79 Chinese hamster cells were treated with 10 µM sodium arsenite (SA) for 24h; at the end of exposure, mitotic rounded-up cells were harvested by shake-off and allowed to grow in drug-free medium. The instability markers, micronucleated and multinucleated cells as well as aneuploid cells, seen just after treatment, reappeared since 60th cell generation. Metaphases with dicentric chromosomes or telomeric associations characterized cell population since 90th generation. To gain insight into the mechanism involved in perpetuating the unstable phenotype, groups of clones, stable and unstable, were analysed also for telomerase activity by TRAP assay and for levels of reactive oxygen species (ROS) measured by ability to oxidize fluorogenic dye. Some of the isolated unstable clones, also bearing chromosomal end-to-end fusions, maintained telomerase activity and were capable to proliferate accumulating genomic instability as well as transformed phenotype and spontaneous increased gene mutation oxidative stress associated. Furthermore these clones showed altered DNA methylation pattern. On the whole, these results raise the possibility that cell variants induced by the short-term exposure to arsenic probably gain a selective advantage when they are able to epigenetically reprogram their genome and proliferate in an error-prone mode.