[en] Full text: Genomic instability is a hallmark of tumorigenic progression, and a similar phenotype is also observed in a high fraction (10 - 50%) of cells that survive exposure to ionizing radiation. In both cases unstable clones are characterized by non-clonal chromosomal rearrangements, which are indicative of a high rate of genetic change during the outgrowth of an unstable parental cell. We postulate that the remarkably high frequency of radiation-induced genomic instability is incompatible with a mutational mechanism for a specific gene, or even a large family of genes. Rather, we hypothesize that a major portion of instability is attributable to the formation of chromosomal rearrangement junction sequences that act as de novo chromosomal breakage hotspots. We further suggest that critical target sequences, which represent at least 10% of the genome and include repetitive DNA sequences such as those found in centromeric heterochromatin, can be involved in breakage and rearrangement hotspots that drive persistent genomic instability and karyotypic heterogeneity. Since chromosomal damage is induced even by low dose radiation exposure, we hypothesize that this phenotype can be efficiently induced at doses that are relevant to environmental, occupational, or medical exposure. In the present study, TK6 human B-lymphoblastoid cells were irradiated with 0, 10, 20 and 200cGy, in order to provide a set of data points for single, low dose exposures. Independent clones were analyzed karyotypically approximately 40 generations after radiation exposure. Preliminary results suggest that the fraction of clones exhibiting genomic instability after 20 cGy (0.16) is similar to and statistically indistinguishable from the fraction of unstable clones following 200 cGy (0.2) exposure. These findings support the hypothesis that instability following radiation, and perhaps also in cancer, primarily reflects non-mutational mechanisms

[en] Complete text of publication follows. Genomic instability has been demonstrated in the progeny of irradiated cells and unirradiated bystander cells. Bystander responses are thought to depend on the activation of cellular communication processes. In this study we examine one such mediator of cellular communication, the pro-inflammatory cytokine tumor necrosis factor alpha (TNF-α) TNF-α is known to increase in expression following ionizing radiation (IR) exposure. Upon binding to its cellular receptors, TNF-α initiates a signaling cascade mediated by reactive oxygen species (ROS) that can activate sequestered NF-κB, thus initiating a pro-inflammatory and antiapoptotic pathway. NF-κB can in turn upregulate TNF-α expression, which when secreted can induce subsequent autocrine and paracrine stimulation of TNF-α and NF-κB. We speculate that this increase in TNF-α signaling and concomitant ROS generation has a mechanistic role in the initiation of genomic instability and a potential involvement in producing bystander responses. Genomic instability is induced by IR in a non-dose-dependent manner. Previous investigation by our group using primary human vascular endothelial cells has shown that both low (0.1 Gy) and high (2 Gy) doses of IR raise levels of secreted TNF-α in a non-linear manner, that both immediate genetic damage and delayed chromosomal instability can be induced at similar levels following treatment with either 0.1-10 ng/mL TNF-α or 0.1 or 2 Gy IR, and that this immediate damage was abrogated by pre-incubation with antioxidants. The current study is therefore focused on the mechanism responsible for this TNF-α-induced instability, and whether TNF-α is a signaling mediator of bystander-induced responses. TNF-α suppressors are added to either directly irradiated or bystander cell cultures exposed to low or high doses of low-LET radiation, and the results are compared to cells pre-treated with antioxidants. Cellular damage is assessed by cell survival, the comet assay, formation of damage-induced foci, and presence of delayed chromosomal instability. Preliminary results indicate that: 1) suppressing TNF-a prevents immediate genetic damage in directly irradiated cells, similar to antioxidant treatment, 2) antioxidants but not TNF-a suppression can abrogate delayed chromosomal instability in directly irradiated cells, and 3) the suppression of either TNF-α or nitric oxide in bystander cells increases survival and protects against immediate genetic damage after exposure to medium from cells irradiated with 2 Gy.