Reduced Apoptosis and Increased Deletion Mutations at Aprt Locus In vivo in Mice Exposed to Repeated Ionizing Radiation

Abstract

Exposure to ionizing radiation (IR) is a risk factor for carcinogenesis because it is a mutagen. However, a single 4-Gy whole body X-ray exposure only induced a modest increase of mutations at the Aprt reporter gene locus in mouse T cells. Intriguingly, when the same dose of IR was given in a fractionated protocol (1 Gy  4 at weekly intervals), there was a strong induction of Aprt mutations in T cells. Many of these were mutations that arose via interstitial deletions inclusive of Aprt or by intragenic deletions. We hypothesized that the weekly fractionated X-ray exposures select for somatic cells with reduced p53 expression and/or reduced apoptosis, which, in turn, may have facilitated the accumulation of interstitial deletions, as in p53-deficient mice. We indeed found that splenocytes of mice with three previous exposures (1 Gy  4 in total) were more resistant to X-ray-induced apoptosis than those of mice exposed to X-rays for the first time (1 Gy total). Thus, repeated X-ray radiation selects for reduced apoptosis in vivo. However, this reduced apoptosis is p53-independent, because p53 induction and the up-regulation of genes downstream of p53, such as Bax and p21, were similar between the 1-Gy and 1 Gy  4 groups. Reduced apoptosis probably allows the generation of more mutations, particularly deletion mutations. Because both reduced apoptosis and increased somatic mutation are risk factors for carcinogenesis, they may contribute to the paradigm in which different radiation exposure schemes are varied in their efficiency in inducing lymphomagenesis. [Cancer Res 2007;67(5):1910-7]

Homologous Recombination Conserves DNA Sequence Integrity Throughout the Cell Cycle in Embryonic Stem Cells

“…Expression of the RAD51 gene was measured with a quantitative real-time polymerase chain reaction (PCR) assay as previously reported [38]. Expression level was calculated as a ratio of the mRNA level for a given gene relative to the mRNA level for glyceraldehyde-3-phosphate dehydrogenase (GAPDH).…”

“…Nuclear protein extracts were prepared as described [38]. Equivalent concentrations of nuclear extracts were electrophoresed with sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels and transferred onto polyvinylidene difluoride membranes.…”

Studies of thioguanine‐resistant lymphocytes induced by in vivo irradiation of mice

“…The occurrence of dosedependent radiation mutagenesis in mice has been demonstrated in a number of studies [Dempsey and Morley 1986;Lorenz et al, 1990Lorenz et al, , 1993Lorenz et al, , 1994Grdina et al, 1992;Kataoka et al, 1992Kataoka et al, , 1993Gaziev et al, 1995;Klarmann et al, 1995;Dobrovolsky et al, 2002;Liang et al, 2007]. However, the more extensive evaluation of time elapsed after acute g irradiation revealed that mutant frequency dependence on dose shifted between 3 and 5 weeks and 10-53 weeks, from being clearly quadratic at the peak of mutant recovery in weeks 3-5 to a statistically strong but less-defined dose relationship in the later time frame.…”

X-rays induce distinct patterns of somatic mutation in fetal versus adult hematopoietic cells

“…PCR amplification of Hprt indicated that intragenic deletions in Hprt were significantly induced by X-irradiation (Nguyen, Liang and Tischfield, unpublished data). Molecular analyses of Aprt mutants also demonstrated that intragenic alterations in Aprt (base substitutions and deletions) but not large chromosome alterations were induced by 1 Gy X-ray [8]. However, the putative induction in intragenic alterations (3.2 × 10 −6 ) at Aprt was not enough to produce a significant increase in overall Aprt MF due to the high spontaneous Aprt MF (24 × 10 −6 ).…”

“…Specifically, 1 Gy X-ray increased the Hprt MF by 3.3-fold as compared to unirradiated controls (11 × 10 −6 v.s. 3.3 × 10 −6 , in irradiated mice and controls, respectively) [8]. PCR amplification of Hprt indicated that intragenic deletions in Hprt were significantly induced by X-irradiation (Nguyen, Liang and Tischfield, unpublished data).…”