Long‐lasting genomic instability following arsenite exposure in mammalian cells: The role of reactive oxygen species

Abstract

Previously, we reported that the progeny of mammalian cells, which has been exposed to sodium arsenite for two cell cycles, exhibited chromosomal instability and concurrent DNA hypomethylation, when they were subsequently investigated after two months of subculturing (about 120 cell generations) in arsenite-free medium. In this work, we continued our investigations of the long-lasting arsenite-induced genomic instability by analyzing additional endpoints at several time points during the cell expanded growth. In addition to the progressive increase of aneuploid cells, we also noted micronucleated and multinucleated cells that continued to accumulate up to the 50th cell generation, as well as dicentric chromosomes and/or telomeric associations and other complex chromosome rearrangements that began to appear much later, at the 90th cell generation following arsenite exposure. The increasing genomic instability was further characterized by an increased frequency of spontaneous mutations. Furthermore, the long-lasting genomic instability was related to elevated levels of reactive oxygen species (ROS), which at the 50th cell generation appeared higher than in stable parental cells. To gain additional insight into the continuing genomic instability, we examined several individual clones isolated at different time points from the growing cell population. Chromosomally and morphologically unstable cell clones, the number of which increased with the expanded growth, were also present at early phases of growth without arsenite. All genomically unstable clones exhibited higher ROS levels than untreated cells suggesting that oxidative stress is an important factor for the progression of genomic instability induced by arsenite.

Dysregulation of DNA methylation induced by past arsenic treatment causes persistent genomic instability in mammalian cells

“…Thereafter, the frequency of aneuploidy among SMA treated cells sharply decreased to the value of the untreated cells after 2 cell generations of subculturing in SMA‐free medium; however, we subsequently observed an increasing frequency of aneuploid cells from 10 to 40 cell generations after the removal of SMA. Thus, the aneuploidy trend seen for the H0.1 and H0.5 HaCaT cells was comparable to that which was previously reported in the ASO V79 cells [Sciandrello et al, ], where the percentage of aneuploid ASO cells first declined to the same level as in untreated cells but gradually increased over the time even in the absence of continued SMA treatment. In contrast, in the HaCaT cells we did not find the persistent gross chromosome aberrations (data not shown) that we previously observed in the ASO V79 cells [Sciandrello et al, ], although we have previously reported preliminary findings on persistent changes in mitotic‐regulatory BubR1 gene expression that could contribute to aneuploidy in these exposed HaCaT cells [Mauro et al, ], and future studies on centromeric repeats, or other mitotic control or centrosomal genes may be interesting to investigate.…”

“…Previously, we reported that V79 Chinese hamster cells underwent early genetic instability when exposed to 10 μM arsenite for 24 hr, and we demonstrated that the descendants of the surviving cells continued to be genetically unstable, showing ongoing gross aneuploidy and structural chromosome changes linked to DNA hypomethylation that persisted for about two months (up to 120 cell generations) of sub‐culturing in arsenite‐free medium [Sciandrello et al, ; Sciandrello et al, ]. This prolonged duration of genomic instability observed in the absence of continuous arsenite exposure suggested underlying epigenetic perturbations, the temporality (persistence) of which required further investigation.…”

“…In order to verify if the progressive and the long‐term aneuploidy that we previously observed in SMA treated V79 cells [Sciandrello et al, ] was also observed in human cells, cytogenetic analyses were performed on HaCaT human keratinocytes exposed to 0.1 and 0.5 μM SMA for 48 h (two cell cycles), then allowed to grow in arsenic‐free medium and tested for up to 40 cell generations (42 days). As shown in Figure , the SMA treatments induced immediate increases of aneuploid HaCaT cells to 8.6 and 10.4% aneuploidy, respectively, vs an average of 2.8% aneuploidy among untreated HaCaT cells that remained constantly low over 40 cell generations.…”

“…Over many years we have extensively studied the effects of arsenic as an inducer of cytogenetic and genomic instability and associated epigenetic reprogramming [Sciandrello et al, ; Sciandrello et al, ; Catanzaro et al, ; Sciandrello et al, ]. Whereas a conventional hypothesis of carcinogenesis postulated that the accumulation of numerous somatic mutations, or a hypermutable phenotype, were drivers of cancer development, they did not offer full explanation for the observations that weakly or non‐mutagenic agents, arsenite for example, also induced cancers.…”

Tanshinone I Activates the Nrf2-Dependent Antioxidant Response and Protects Against As(III)-Induced Lung Inflammation In Vitro and In Vivo

“…There is mounting evidence suggesting that As(III) induces ROS by depleting GSH or by damaging mitochondria (33,47). Therefore, it was proposed that As(III) may indirectly activate the Nrf2 pathway by increasing ROS (36).…”