Gastrointestinal (GI) injury is a major cause of acute death after total-body exposure to large doses of ionizing radiation, but the cellular and molecular explanations for GI death remain dubious. To address this issue, we developed a murine abdominal irradiation model. Mice were irradiated with a single dose of X rays to the abdomen, treated with daily s.c. injection of N-acetyl-l-cysteine (NAC) or vehicle for 7 days starting either 4 h before or 2 h after irradiation, and monitored for up to 30 days. Separately, mice from each group were assayed 6 days after irradiation for bone marrow reactive oxygen species (ROS), ex vivo colony formation of bone marrow stromal cells, and histological changes in the duodenum. Irradiation of the abdomen caused dose-dependent weight loss and mortality. Radiation-induced acute death was preceded not only by a massive loss of duodenal villi but also, surprisingly, abscopal suppression of stromal cells and elevation of ROS in the nonirradiated bone marrow. NAC diminished these radiation-induced changes and improved 10- and 30-day survival rates to >50% compared with <5% in vehicle-treated controls. Our data establish a central role for abscopal stimulation of bone marrow ROS in acute death in mice after abdominal irradiation.

The effectiveness of lung radiotherapy is limited by radiation tolerance of normal tissues and by the intrinsic radiosensitivity of lung cancer cells. The chemopreventive agent curcumin has known antioxidant and tumor cell radiosensitizing properties. Its usefulness in preventing radiation-induced pneumonopathy has not been tested previously. We evaluated dietary curcumin in radiation-induced pneumonopathy and lung tumor regression in a murine model. Mice were given 1% or 5% (w/w) dietary curcumin or control diet prior to irradiation and for the duration of the experiment. Lungs were evaluated at 3 weeks after irradiation for acute lung injury and inflammation by evaluating bronchoalveolar lavage (BAL) fluid content for proteins, neutrophils and at 4 months for pulmonary fibrosis. In a separate series of experiments, an orthotopic model of lung cancer using intravenously injected Lewis lung carcinoma (LLC) cells was used to exclude possible tumor radioprotection by dietary curcumin. In vitro, curcumin boosted antioxidant defenses by increasing heme oxygenase 1 (HO-1) levels in primary lung endothelial and fibroblast cells and blocked radiation-induced generation of reactive oxygen species (ROS). Dietary curcumin significantly increased HO-1 in lungs as early as after 1 week of feeding, coinciding with a steady-state level of curcumin in plasma. Although both 1% and 5% w/w dietary curcumin exerted physiological changes in lung tissues by significantly decreasing LPS-induced TNF-α production in lungs, only 5% dietary curcumin significantly improved survival of mice after irradiation and decreased radiation-induced lung fibrosis. Importantly, dietary curcumin did not protect LLC pulmonary metastases from radiation killing. Thus dietary curcumin ameliorates radiation-induced pulmonary fibrosis and increases mouse survival while not impairing tumor cell killing by radiation.

We investigated whether genistein could protect the lung from radiation-induced injury. We hypothesized that genistein would reduce the levels of inflammatory cytokines and ROS after irradiation and therefore lead to reduced DNA damage and functional deficits. Whole lungs of Sprague-Dawley rats were irradiated with 18 Gy at ∼0.5 Gy/min. At 28 weeks a micronucleus assay was used to examine DNA damage and, using immunohistochemical analysis, expression of IL-1α, IL-1β, IL-6, TNF-α and TGF-β, macrophage activation, oxidative stress (8-OHdG) and collagen levels were measured. A TBARS assay was used to measure the level of malondialdehyde. Functional damage was assessed by measuring the breathing rate of the rats over the course of the experiment. The increase in breathing rate after irradiation was damped in rats receiving genistein during the phase of pneumonitis (6–10 weeks), and there was a 50–80-day delay in lethality in this group. Genistein treatment also decreased the levels of the inflammatory cytokines TNF-α, IL-1β and TGF-β and led to a reduction in collagen content, a reduction in 8-OHdG levels, and complete protection against DNA damage measured in surviving rats at 28 weeks after irradiation. These results demonstrates that genistein treatment can provide partial protection against the early (pneumonitis) effects of lung irradiation and reduce the extent of fibrosis, although not sufficiently to prevent lethality at the radiation dose used in this study.

Tumor radiotherapy with large-field irradiation results in an increase of p53-dependent apoptosis of the radiosensitive hematopoietic stem cells. Proapoptotic PUMA is a transcriptional target of p53. Thus suppression of PUMA expression by gene therapy with the transcription repressor SNAI2 as transgene might be a potential approach for normal tissue protection during radiotherapy. SNAI2 cDNA was cloned in a lentiviral SIN vector in a bicistronic expression cassette followed by a floxed IRES-EMCV linker and EGFP as selection gene. Wild-type p53 TK6 cells were used as the cellular model system. We could demonstrate the significant radioprotective effect of SNAI2 overexpression in a cytotoxicity assay after irradiation with 0–5 Gy compared with untransduced or control vector (inverse oriented SNAI2 cDNA)-transduced cells. Additionally, TK6-SNAI2 compared to TK6-SNAI2inv cells showed a survival advantage in a clonogenic assay after irradiation with 0–3 Gy. Determination of the proportion of sub-G₁ cells in TK6-SNAI2 cells revealed an approximately 50% reduction in apoptosis compared with both control entities. In this study using a bicistronic lentiviral vector, we were able to provide proof of principle that lentiviral overexpression of SNAI2 might be used for radioprotective gene therapy to widen the therapeutic range in radiotherapy.

Exposure to X radiation is associated with a decline in the proliferative activity of the liver, but the molecular mechanism(s) is not well understood. We investigated whether exposure to X radiation is involved in functional changes in the epidermal growth factor (EGF) receptor (EGFR), thereby causing a reduction of EGF-induced DNA synthesis using periportal hepatocytes (PPH) and perivenous hepatocytes (PVH), which differ in their proliferative activity. X radiation dose-dependently decreased DNA synthesis in both subpopulations. The rate of decline in the DNA synthesis was greater in PPH than in PVH, but the zonal difference disappeared after exposure to 10 Gy X radiation. [¹²⁵I]EGF binding studies indicated that high-affinity EGFRs in both subpopulations were down-regulated after X irradiation. Furthermore, EGF-induced EGFR dimerization and phosphorylation at Y1173 in both subpopulations were down-regulated after X irradiation, and the rate of decline was greater in PPH than in PVH. In contrast, phosphorylation at Y845 after EGF treatment was dose-dependently up-regulated after X irradiation in both subpopulations. These results suggest that the X-radiation-related decline in EGF-induced DNA synthesis is caused at least partly by the modification of EGFR function.

Exposure to γ radiation causes a wide variety of biological damages and alterations, including oxidative stress. Among the key cellular components that are exquisitely sensitive to oxidative stress is the transcription factor nuclear factor (NF)-κB, which plays a central role in the activation of various pro-inflammatory genes. Recently, senescence marker protein 30 (SMP30), which has been used as an aging marker, was shown to have an antioxidant property. In the current study, using SMP30 knockout (SMP30^−/−) mice that are vitamin C-deficient, we explored the effect of radiation on the activation of NF-κB and several key pro-inflammatory genes. Six groups of mice were studied. Group 1 mice were not irradiated and were supplemented with vitamin C (2.5 mg/kg/day). Group 2 mice were irradiated and were not supplemented with vitamin C. Group 3, 4 and 5 mice were irradiated with 1, 3 and 5 Gy of γ radiation (⁶⁰Co), respectively, without vitamin C supplementation. The wild-type mice (SMP30 ^/ ) in group 6 were not irradiated or supplemented. At 24 h after irradiation, mice were killed humanely and the kidneys were removed analysis. The results showed that γ radiation induced oxidative stress with corresponding NF-κB activation; this activated NF-κB led to the up-regulation of several major pro-inflammatory mediators such as COX-2, iNOS, VCAM1, ICAM1 and E-selectin in irradiated groups with no vitamin C supplementation. Our data provide molecular insights into mechanisms through which γ radiation enhances oxidative stress-induced inflammation by showing the activation of NF-κB signaling pathway in vitamin C-deficient SMP30^−/− mice. In addition, our present study produced evidence that γ radiation exerts its deleterious action by activating the inflammatory process that are known to be a major risk factor for many chronic diseases. Furthermore, our data revealed vitamin C may play an important protective role in attenuating the adverse γ-radiation-induced adverse effects by suppressing adverse oxidative effects and pro-inflammatory mediators.

Mitochondrial DNA (mtDNA) contains 13 genes that encode proteins of the oxidative phosphorylation complex that are involved in ATP generation. Leber's optic atrophy and Leigh's syndrome are diseases that are caused by point mutations in the mitochondrial genome and that have phenotypes associated with energy deprivation. We hypothesized that energy deficiency from mitochondrial mutations in these cells leads to radiation hypersensitivity. Here we compared mitochondrial gene expression for the 13 mitochondrial protein-coding genes in two mitochondrial mutant cell lines, GM13740 (Leigh's syndrome) and GM10744 (Leber's optic atrophy) and a normal human lymphoblastoid cell line (GM15036) after X irradiation (0–4 Gy) 0 to 24 h postirradiation. Changes in gene expression were compared with cellular radiosensitivity. Statistically significant differences between Leigh's syndrome and normal cells were found in mitochondrial gene expression for all radiation doses and times that were commensurate with changes in radiation sensitivity. The data suggest that Leigh's syndrome cells have an impaired ability to repair radiation-induced DNA damage that results in radiation hypersensitivity. This may be attributable to mitochondrial dysfunction from reductions in mitochondrial gene expression and ATP generation, since Leigh's optic atrophy cells exhibit a mutation in the ATPase6 gene, which is an important component of Complex V of ATP synthase. In contrast, the mutation of the Leber's cells conferred radioresistance, which might be attributed to the mutation in the ND4 gene in the mitochondrial genome. The altered sensitivity of mitochondrial mutant cells to ionizing radiation can lead to decreased DNA repair, which may put individuals with mtDNA mutations at greater risk for cancer and other diseases.

In this study, we attempted to clarify the influence of the DNA repair genes RAD54 and KU70, components of the homologous recombination (HR) and non-homologous end-joining (NHEJ) pathways, respectively, on apoptosis induced by 1 Gy (low-dose) and 5 Gy (high-dose) irradiation. All experiments were performed using chicken B-lymphocyte DT40 cells and the DNA repair-deficient cell lines KU70^−/−, RAD54^−/− and KU70^−/−/RAD54^−/−. Morphological changes were detected by fluorescence methods, and the sub-G₁ fraction and the activated caspases in DT40 cells were analyzed by flow cytometry. Irradiation with 1 Gy significantly increased the level of apoptosis in cells with the defective DNA repair genes, with the maximum apoptosis occurring in double mutant cells, KU70^−/−/RAD54^−/−, demonstrating that 1 Gy is enough to induce apoptosis in DNA repair-deficient DT40 cells, and that KU70 and RAD54 must have almost the same role in low-dose radiation-induced apoptosis. After 5 Gy, fast induction of apoptosis, within 2 h, was seen in both wild-type cells and RAD54^−/− cells, indicating that functional KU70 must be important for the rescue of the cells from the induction of fast apoptosis.

We sought to reduce tumor hypoxia by topical application of a vasodilator, benzyl nicotinate (BN), and investigated its effect on the growth of tumors irradiated at times when tumor pO₂ increased. EPR oximetry was used to follow the changes in the tissue pO₂ of subcutaneous radiation-induced fibrosarcoma (RIF-1) tumors during topical applications of 1.25–8% BN formulations for 5 consecutive days. The RIF-1 tumors were hypoxic with a tissue pO₂ of 4.6–7.0 mmHg. A significant increase in tumor pO₂ occurred 10–30 min after BN application. The formulation with the minimal BN concentration that produced a significant increase in tumor pO₂ was used for the radiation study. The tumors were irradiated (4 Gy × 5) at the time of the maximum increase in pO₂ observed with the 2.5% BN formulation. The tumors with an increase in pO₂ of greater than 2 mmHg from the baseline after application of BN on day 1 had a significant growth inhibition compared to the tumors with an increase in pO₂ of less than 2 mmHg. The results indicate that the irradiation of tumors at the time of an increase in pO₂ after the topical application of the 2.5% BN formulation led to a significant growth inhibition. EPR oximetry provided dynamic information on the changes in tumor pO₂, which could be used to identify responders and non-responders and schedule therapy during the experiments.

Exposure to radioactive iodine (¹³¹I) from atmospheric nuclear tests conducted in Nevada in the 1950s may have increased thyroid cancer risks. To investigate the long-term effects of this exposure, we analyzed data on thyroid cancer incidence (18,545 cases) from eight Surveillance, Epidemiology, and End Results (SEER) tumor registries for the period 1973–2004. Excess relative risks (ERR) per gray (Gy) for exposure received before age 15 were estimated by relating age-, birth year-, sex- and county-specific thyroid cancer rates to estimates of cumulative dose to the thyroid that take age into account. The estimated ERR per Gy for dose received before 1 year of age was 1.8 [95% confidence interval (CI), 0.5–3.2]. There was no evidence that this estimate declined with follow-up time or that risk increased with dose received at ages 1–15. These results confirm earlier findings based on less extensive data for the period 1973–1994. The lack of a dose response for those exposed at ages 1–15 is inconsistent with studies of children exposed to external radiation or ¹³¹I from the Chernobyl accident, and results need to be interpreted in light of limitations and biases inherent in ecological studies, including the error in doses and case ascertainment resulting from migration. Nevertheless, the study adds support for an increased risk of thyroid cancer due to fallout, although the data are inadequate to quantify it.

Rats were exposed once by inhalation to plutonium-239 dioxide (²³⁹PuO₂), resulting in chronic α-particle irradiation of the lung, and exposed chronically to cigarette smoke to examine carcinogenic interactions between the two exposures. F344 rats were exposed to ²³⁹PuO₂ to achieve an initial lung burden of 0.5 kBq and then exposed 6 h/day, 5 days/week to cigarette smoke at 100 or 250 mg particulate matter/m³ for up to 30 months. Exposure to cigarette smoke increased the cumulative radiation dose to lung by slowing the clearance of ²³⁹PuO₂. ²³⁹PuO₂ alone did not affect survival, but the higher cigarette smoke exposure shortened survival in females. Combined exposure to ²³⁹PuO₂ and cigarette smoke acted synergistically to shorten survival in both genders. The combined effects of cigarette smoke and ²³⁹PuO₂ were approximately additive for lung hyperplasia and adenomas but were strongly synergistic for carcinomas. Differences between observed incidences and incidences predicted by survival-adjusted models accounting for increased radiation dose revealed a substantial component of synergy for carcinomas above that attributable to the radiation dose effect. The synergy for malignant lung tumors is consistent with findings from uranium miners and nuclear weapons production workers. These results bolster confidence in the epidemiological findings and have implications for risk assessment.

A Monte Carlo simulation model for DNA repair via the non-homologous end-joining pathway has been developed. Initial DNA damage calculated by the Monte Carlo track structure code PARTRAC provides starting conditions concerning spatial distribution of double-strand breaks (DSBs) and characterization of lesion complexity. DNA termini undergo attachment and dissociation of repair enzymes described in stochastic first-order kinetics as well as step-by-step diffusive motion considering nuclear attachment sites. Pairs of DNA termini with attached DNA-PK enter synapsis under spatial proximity conditions. After synapsis, a single rate-limiting step is assumed for clean DNA ends, and step-by-step removal of nearby base lesions and strand breaks is considered for dirty DNA ends. Four simple model scenarios reflecting different hypotheses on the origin of the slow phase of DSB repair have been set up. Parameters for the presynaptic phase have been derived from experimental data for Ku70/Ku80 and DNA-PK association and dissociation kinetics. Time constants for the post-synaptic phase have been adapted to experimental DSB rejoining kinetics for human fibroblasts after ¹³⁷Cs γ irradiation. In addition to DSB rejoining kinetics, the yields of residual DSBs, incorrectly rejoined DSBs, and chromosomal aberrations have been determined as a function of dose and compared with experimental data. Three of the model scenarios obviously overestimate residual DSBs after long-term repair after low-dose irradiation, whereas misrejoined DSBs and chromosomal aberrations are in surprisingly good agreement with measurements.

Single crystals of 5-methylcytosine hemihydrate and 5-methylcytosine hydrochloride were X-irradiated and studied at 10 K and at higher temperatures using X- and K-band EPR, ENDOR and EIE spectroscopy. In the hemihydrate crystals, four radicals were identified at 10 K, one of them being the recently reported N1-deprotonated one-electron oxidation product (Krivokapić et al., J. Phys. Chem. A 113, 9633–9640, 2009). The other radicals were the 3αH radical and the C6 and C5 H-addition radicals (the 5-yl and 6-yl radicals, respectively). After irradiation at 295 K, only the 3αH and the 5-yl radicals were observed. In the hydrochloride crystals, at least seven different radicals were present after irradiation at 10 K. These were the N1-deprotonated one-electron oxidation product, the 3αH radical, three different one-electron reduction products, and the 5- and 6-yl radicals. DFT calculations were used to assist in assigning the observed couplings. The 3αH and 5-yl radicals were dominant after thermal annealing to room temperature. In neither crystal system did the N1-deprotonated oxidation product transform into the 3αH radical upon warming. The radical yield was significantly greater after irradiation at 300 K compared to that after irradiation at 10 K followed by warming to 300 K and was also considerably greater in the hydrochloride crystals than in the hemihydrate crystals.

This paper evaluates the global vitamin D effective UV (UV_vitd) irradiances under cloudy conditions at a subtropical, southern hemisphere site. The UV_vitd irradiances were analyzed on a horizontal plane and sampled at 5-min intervals over 18 months so that a wide range of parameters including cloud conditions, solar zenith angles (SZA) and ozone levels were taken into account. Cloud modification factors were determined from the influence of clouds on the global broadband solar radiation, and these were applied to the cloud-free vitamin D effective UV irradiance to evaluate the UV_vitd irradiances on a horizontal plane for cloudy conditions. For vitamin D effective UV irradiance, cloud modification factors were found to range from 0.9 to 1.0 for no cloud and 0.4 to 0.5 for 8 octa of cloud cover. SZA played a minimal role in this variation. A comparison of the measured and calculated UV_vitd irradiances for the 2004 data set in the range of SZA of 70° or less provided an R² value of 0.90. The output of the model was compared to data measured during the first 6 months of 2005 for an SZA of 70° or less and provided an R² value of approximately 0.82.