Likhtarov, I., Kovgan, L., Vavilov, S., Chepurny, M., Bouville, A., Luckyanov, N., Jacob, P., Voillequé, P. and Voigt, G. Post-Chornobyl Thyroid Cancers in Ukraine. Report 1: Estimation of Thyroid Doses. Radiat. Res. 163, 125–136 (2005).

About 1.8 EBq of ¹³¹I was released into the atmosphere during the Chornobyl accident that occurred in Ukraine on April 26, 1986. More than 10% of this activity was deposited on the territory of Ukraine. Beginning 4–5 years after the accident, an increase in the incidence of thyroid cancer among children, believed to be caused in part by exposure to ¹³¹I, has been observed in different regions of Ukraine. A three-level system of thyroid dose estimation was developed for the reconstruction of thyroid doses from ¹³¹I for the entire population of Ukrainian children aged 1 to 18 at the time of accident: (1) At the first level, individual doses were estimated for the approximately 99,000 children and adolescents with direct measurements of radioactivity in the thyroid (so-called direct thyroid measurements) performed in May–June of 1986; (2) at the second level, group doses by year of age and by gender were estimated for the population of 748 localities (with 208,400 children aged 1–18 in 1986) where direct thyroid measurements of good quality were performed on some of the residents; and (3) at the third level, group doses by age and by gender were estimated for the population of the localities where no thyroid measurements were made in 1986. The third-level doses were then aggregated over the population of each oblast. Data, models and procedures required for each level of thyroid dose estimation are described in the paper. At the first level, individual doses were found to range up to 27,000 mGy, with geometric and arithmetic means of 100 and 300 mGy, respectively. At the second level, group doses were found to be highest for the younger children (aged 1 to 4 years); doses for the older children (aged 16 to 18 years) were 3.5 times smaller. At the third level, average population-weighted doses were found to exceed 35 mGy in the five northern oblasts closer to the Chornobyl reactor site; to be in the 14- to 34-mGy range in seven other oblasts, Kyiv city and Crimea; and to be less than 13 mGy in all other oblasts.

Matsuu, M., Shichijo, K., Ikeda, Y., Ito, M., Naito, S., Okaichi, K., Nakashima, M., Nakayama, T. and Sekine, I. Sympathetic Hyperfunction Causes Increased Sensitivity of the Autonomic Nervous System to Whole-Body X Irradiation. Radiat. Res. 163, 137–143 (2005).

Although the etiology of radiation sickness is still unknown, disturbance of the autonomic nervous system is suggested to be a factor. This study was designed to compare the radiosensitivity of spontaneously hypertensive rats possessing sympathetic hyperfunction and control Wistar-Kyoto rats, and to analyze the effects of radiation on the autonomic nervous system in both strains. After a 7.5-Gy dose of whole-body X irradiation, the blood pressure decreased significantly at 8 h and 2 days in the spontaneously hypertensive rats, but not in the Wistar-Kyoto rats. Epinephrine levels in the adrenal gland of spontaneously hypertensive rats decreased at 4, 8 and 24 h, unlike the Wistar-Kyoto rats. Radiation evoked a stronger increase in norepinephrine in the jejunum and colon of spontaneously hypertensive rats than in Wistar-Kyoto rats. Acethylcholine levels in the jejunum of spontaneously hypertensive rats decreased, in contrast to the increase in Wistar-Kyoto rats within 24 h after irradiation. The survival rate of spontaneously hypertensive rats was lower than that of Wistar-Kyoto rats and weight loss, appetite loss and morphological changes in the jejunum were greater in spontaneously hypertensive rats than in Wistar-Kyoto rats after irradiation. These results indicated that X irradiation caused greater activities in autonomic nervous function and severe radiation injury in spontaneously hypertensive rats. Sympathetic hyperfunction may be associated with a higher sensitivity to radiation, including radiation injury and radiation sickness.

Van der Meeren, A., Monti, P., Vandamme, M., Squiban, C., Wysocki, J. and Griffiths, N. Abdominal Radiation Exposure Elicits Inflammatory Responses and Abscopal Effects in the Lungs of Mice. Radiat. Res. 163, 144–152 (2005).

An inflammatory reaction is a classical feature of radiation exposure and appears to be a key event in the development of the acute radiation syndrome. We have investigated the radiation-induced inflammatory response in C57BL6/J mice after total abdominal or total-body irradiation at a dose of 15 Gy. Our goal was to determine the radiation-induced inflammatory response of the gut and to study the consequences of abdominal irradiation for the intestine and for the lungs as a distant organ. A comparison with total-body irradiation was used to take into account the hematopoietic response in the inflammatory process. For both irradiation regimens, systemic and intestinal responses were evaluated. A systemic inflammatory reaction was found after abdominal and total-body irradiation, concomitant with increased cytokine and chemokine production in the jejunum of irradiated mice. In the lungs, the radiation-induced changes in the production of cytokines and chemokines and in the expression of adhesion molecules after both abdominal and total-body irradiation indicate a possible abscopal effect of radiation in our model. The effects observed in the lungs after irradiation of the abdomino-pelvic region may be caused by circulating inflammatory mediators consequent to the gut inflammatory response.

Ina, Y., Tanooka, H., Yamada, T. and Sakai, K. Suppression of Thymic Lymphoma Induction by Life-Long Low-Dose-Rate Irradiation Accompanied by Immune Activation in C57BL/6 Mice. Radiat. Res. 163, 153–158 (2005).

The induction of thymic lymphomas by whole-body X irradiation with four doses of 1.8 Gy (total dose: 7.2 Gy) in C57BL/6 mice was suppressed from a high frequency (90%) to 63% by preirradiation with 0.075 Gy X rays given 6 h before each 1.8-Gy irradiation. This level was further suppressed to 43% by continuous whole-body irradiation with ¹³⁷Cs γ rays at a low dose rate of 1.2 mGy/h for 450 days, starting 35 days before the challenging irradiation. Continuous irradiation at 1.2 mGy/h resulting in a total dose of 7.2 Gy over 258 days yielded no thymic lymphomas, indicating that this low-dose-rate radiation does not induce these tumors. Further continuous irradiation up to 450 days (total dose: 12.6 Gy) produced no tumors. Continuously irradiated mice showed no loss of hair and a greater body weight than unirradiated controls. Immune activities of the mice, as measured by the numbers of CD4 T cells, CD40 B cells, and antibody-producing cells in the spleen after immunization with sheep red blood cells, were significantly increased by continuous 1.2-mGy/h irradiation alone. These results indicate the presence of an adaptive response in tumor induction, the involvement of radiation-induced immune activation in tumor suppression, and a large dose and dose-rate effectiveness factor (DDREF) for tumor induction with extremely low-dose-rate radiation.

Kubota, T., Yoshikai, Y., Tamura, Y., Mishima, Y., Aoyagi, Y., Niwa, O. and Kominami, R. Comparison of Properties of Spontaneous and Radiation-Induced Mouse Thymic Lymphomas: Role of Trp53 and Radiation. Radiat. Res. 163, 159–164 (2005).

Mouse thymic lymphomas are readily induced by radiation and also arise without irradiation when the mice are null in Trp53 functions. In the present study, spontaneous thymic lymphomas in Trp53^−/− mice were compared to those arising in irradiated Trp53 ^/− mice, revealing three features characteristic of the spontaneous lymphomas. (1) Mp53D2, a Trp53 modifier that affects the latent period of radiogenic thymic lymphomas in Trp53 ^/− mice, had no effect on the development of spontaneous lymphomas. (2) A sex difference in the latency was found. (3) A marked difference was noted in the frequency of allelic loss at the Ikaros gene on chromosome 11, encoding a transcription factor required for normal lymphocyte development and differentiation; 2% in the lymphomas of Trp53^−/− mice and 78% in the radiogenic lymphomas of Trp53 ^/− mice, suggesting that loss of Trp53 may reduce the requirement for the loss of Ikaros for lymphomagenesis. Furthermore, allelic loss analysis on chromosome 19 localized a region that may harbor an unknown tumor suppressor gene. These results suggest intricate steps of lymphomagenesis influenced by the presence or absence of Trp53.

Nayak, V. and Uma Devi, P. Protection of Mouse Bone Marrow against Radiation-Induced Chromosome Damage and Stem Cell Death by the Ocimum Flavonoids Orientin and Vicenin. Radiat. Res. 163, 165–171 (2005).

In a previous study, orientin and vicenin, the water-soluble plant flavonoids, protected mice against radiation lethality (Uma Devi et al., Radiat. Res. 151, 74–78, 1999). To study bone marrow protection, adult Swiss mice were exposed to 0–6 Gy ⁶⁰Co γ rays 30 min after an intraperitoneal injection of 50 μg/ kg body weight of orientin/vicenin. Chromosomal aberrations in bone marrow were studied at 24 h postirradiation. Stem cell survival was studied using the exogenous spleen colony (CFU-S) assay. Radiation produced a dose-dependent increase in aberrant cells as well as in the yield of the different types of aberrations (breaks, fragments, rings and dicentrics) and a decrease in CFU-S. Pretreatment with either flavonoid significantly reduced the aberrant cells and different aberrations and increased the number of CFU-S compared to the respective radiation-alone groups. The dose modification factors for 50% reductions in the number of CFU-S were 1.6 for orientin and 1.7 for vicenin. The present finding that very low nontoxic doses of orientin and vicenin provide efficient protection against bone marrow damage at clinically relevant radiation doses suggests their potential for protection of normal tissues in radiotherapy.

Green, L. M., Bianski, B. M., Murray, D. K., Rightnar, S. S. and Nelson, G. A. Characterization of Accelerated Iron-Ion-Induced Damage in Gap Junction-Competent and -Incompetent Thyroid Follicular Cells. Radiat. Res. 163, 172–182 (2005).

Early- and late-passage cultures of Fischer rat thyroid cells differ in their growth properties and gap junction competency. Previous studies comparing early- and late-passage cultures exposed to γ rays and proton beams revealed that differences in growth rate did not influence their responses; however, the presence of connexin 32 gap junctions conferred resistance to γ radiation. To further assess differences in radiation quality, suspension cultures of early- and late-passage cells were exposed to accelerated iron ions, and their comparative biological responses were measured. The iron-ion-irradiated cells displayed sustained levels of incorporated dUTP, reflecting persistent DNA damage. These results were supported by the frequency of chromosomal damage measured by micronucleus formation. Iron-ion irradiation induced micronuclei at a rate of eight per gray per 100 binucleated cells scored in early-passage cells and nine per gray per 100 binucleated cells scored in late-passage cells. Relative to photons, the calculated radiobiological effectiveness for frequency of micronuclei was 5.7 and 6.4 for the early- and late-passage cultures, respectively (P > 0.05). Levels of apoptosis fluctuated as a function of dose, and modest increases above basal levels persisted throughout the 48-h period. The comparison of retained follicular structures revealed differences in the α components of the linear-quadratic dose–response curves (0.60 Gy⁻¹ for early-passage and 0.71 Gy⁻¹ for late-passage cultures, P < 0.014). Cell cycle phase redistribution resulted in a G₂ arrest (P < 0.001) for both early- and late-passage cultures. In conclusion, the response of thyroid follicular cells to high-LET radiation was not influenced by the presence of gap junctions or the proliferative status of the target cells.

Moore, S. R., Marsden, S., Macdonald, D., Mitchell, S., Folkard, M., Michael, B., Goodhead, D. T., Prise, K. M. and Kadhim, M. A. Genomic Instability in Human Lymphocytes Irradiated with Individual Charged Particles: Involvement of Tumor Necrosis Factor α in Irradiated Cells but not Bystander Cells. Radiat. Res. 163, 183–191 (2005).

Exposure to ionizing radiation can increase the risk of cancer, which is often characterized by genomic instability. In environmental exposures to high-LET radiation (e.g. ²²²Ra), it is unlikely that many cells will be traversed or that any cell will be traversed by more than one α particle, resulting in an in vivo bystander situation, potentially involving inflammation. Here primary human lymphocytes were irradiated with precise numbers of ³He² ions delivered to defined cell population fractions, to as low as a single cell being traversed, resembling in vivo conditions. Also, we assessed the contribution to genomic instability of the pro-inflammatory cytokine tumor necrosis factor α (TNFA). Genomic instability was significantly elevated in irradiated groups (≥twofold over controls) and was comparable whether cells were traversed by one or two ³He² ions. Interestingly, substantial heterogeneity in genomic instability between experiments was observed when only one cell was traversed. Genomic instability was significantly reduced (60%) in cultures in which all cells were irradiated in the presence of TNFA antibody, but not when fractions were irradiated under the same conditions, suggesting that TNFA may have a role in the initiation of genomic instability in irradiated cells but not bystander cells. These results have implications for low-dose exposure risks and cancer.

Canova, S., Fiorasi. F., Mognato. M., Grifalconi, M, Reddi, E., Russo, A. and Celotti, L. “Modeled Microgravity” Affects Cell Response to Ionizing Radiation and Increases Genomic Damage. Radiat. Res. 163, 191–199 (2005).

The aim of this work was to assess whether “modeled microgravity” affects cell response to ionizing radiation, increasing the risk associated with radiation exposure. Lymphoblastoid TK6 cells were irradiated with various doses of γ rays and incubated for 24 h in a modeled microgravity environment obtained by the Rotating Wall Vessel bioreactor. Cell survival, induction of apoptosis and cell cycle alteration were compared in cells irradiated and then incubated in 1g or modeled microgravity conditions. Modulation of genomic damage induced by ionizing radiation was evaluated on the basis of HPRT mutant frequency and the micronucleus assay. A significant reduction in apoptotic cells was observed in cells incubated in modeled microgravity after γ irradiation compared with cells maintained in 1g. Moreover, in irradiated cells, fewer G₂-phase cells were found in modeled microgravity than in 1g, whereas more G₁-phase cells were observed in modeled microgravity than in 1g. Genomic damage induced by ionizing radiation, i.e. frequency of HPRT mutants and micronucleated cells, increased more in cultures incubated in modeled microgravity than in 1g. Our results indicate that modeled microgravity incubation after irradiation affects cell response to ionizing radiation, reducing the level of radiation-induced apoptosis. As a consequence, modeled microgravity increases the frequency of damaged cells that survive after irradiation.

Dubrova, Y. E. Radiation-Induced Mutation at Tandem Repeat DNA Loci in the Mouse Germline: Spectra and Doubling Doses. Radiat. Res. 163, 200–207 (2005).

The spectra and dose response for mutations at expanded simple tandem repeat (ESTR) loci in the germline of male mice acutely exposed to low-LET X or γ rays at pre-meiotic stages of spermatogenesis were compared in five strains of laboratory mice. Most mutation events involved the gain or loss of a relatively small number of repeat units, and the distributions of length changes were indistinguishable between the exposed and control males. Overall, a significant bias toward gains of repeats was detected, with approximately 60% of mutants showing gains. The values for ESTR mutation induction did not differ substantially between strains. The highest values of doubling dose were obtained for two genetically related strains, BALB/c and C.B17 (mean value 0.98 Gy). The estimates of doubling dose for three other strains (CBA/H, C57BL/6 × CBA/H F₁ and 129SVJ × C57BL/6) were lower, with a mean value of 0.44 Gy. The dose response for ESTR mutation across all five strains was very close to that for the specific loci (Russell 7-locus test). The mechanisms of ESTR mutation induction and applications of this system for monitoring radiation-induced mutation in the mouse germline are discussed.

Herskind, C., Steil, V., Kraus-Tiefenbacher, U. and Wenz, F. Radiobiological Aspects of Intraoperative Radiotherapy (IORT) with Isotropic Low-Energy X Rays for Early-Stage Breast Cancer. Radiat. Res. 163, 208–215 (2005).

The purpose of this study was to model the distribution of biological effect around a miniature isotropic X-ray source incorporating spherical applicators for single-dose or hypofractionated partial-breast intraoperative radiotherapy. A modification of the linear-quadratic formalism was used to calculate the relative biological effectiveness (RBE) of 50 kV X rays as a function of dose and irradiation time for late-reacting normal tissue and tumor cells. The response was modeled as a function of distance in the tissue based on the distribution of equivalent dose and published dose–response data for pneumonitis and subcutaneous fibrosis after single-dose conventional irradiation. Furthermore, the spatial distribution of tumor cell inactivation was assessed. The RBE for late reactions approached unity at the applicator surface but increased as the absorbed dose decreased with increasing distance from the applicator surface. The ED₅₀ for pneumonitis was estimated to be reached at a depth of 6–11 mm in the tissue and that for subcutaneous fibrosis at 3–6 mm, depending on the applicator diameter and whether the effect of recovery was included. Thus lung tissue would be spared because of the thickness of the thorax wall. The RBE for tumor cells was higher than for late-reacting tissue. The applicator diameter is an important parameter in determining the range of tumor cell control in the irradiated tumor bed.

Howell, R. W. and Neti, P. V. S. V. Modeling Multicellular Response to Nonuniform Distributions of Radioactivity: Differences in Cellular Response to Self-Dose and Cross-Dose. Radiat. Res. 163, 216–221 (2005).

Radiopharmaceuticals are distributed nonuniformly in tissue. While distributions of radioactivity often appear uniform at the organ level, in fact, microscopic examination reveals that only a fraction of the cells in tissue are labeled. Labeled cells and unlabeled cells often receive different absorbed doses depending on the extent of the nonuniformity and the characteristics of the emitted radiations. The labeled cells receive an absorbed dose from radioactivity within the cell (self-dose) as well as an absorbed dose from radioactivity in surrounding labeled cells (cross-dose). Unlabeled cells receive only a cross-dose. In recent communications, a multicellular cluster model was used to investigate the lethality of microscopic nonuniform distributions of ¹³¹I iododeoxyuridine (¹³¹IdU). For a given mean absorbed dose to the tissue, the dose response depended on the percentage of cells that were labeled. Specifically, when 1, 10 and 100% of the cells were labeled, a D₃₇ of 6.4, 5.7 and 4.5 Gy, respectively, was observed. The reason for these differences was recently traced to differences in the cellular response to the self- and cross-doses delivered by ¹³¹IdU. Systematic isolation of the effects of self-dose resulted in a D₃₇ of 1.2 ± 0.3 Gy. The cross-dose component yielded a D₃₇ of 6.4 ± 0.5 Gy. In the present work, the overall survival of multicellular clusters containing 1, 10 and 100% labeled cells is modeled using a semi-empirical approach that uses the mean lethal self- and cross-doses and the fraction of cells labeled. There is excellent agreement between the theoretical model and the experimental data when the surviving fraction is greater than 1%. Therefore, when the distribution of ¹³¹I in tissue is nonuniform at the microscopic level, and the cellular response to self- and cross-doses differs, multicellular dosimetry can be used successfully to predict biological response, whereas the mean absorbed dose fails in this regard.

Champion, C., L'Hoir, A., Fainstein, P. D., Rivarola, R. D., Chetioui, A. A Monte Carlo Code for the Simulation of Heavy-Ion Tracks in Water. Radiat. Res. 163, 222–231 (2005).

TILDA, a new Monte Carlo track structure code for ions in gaseous water that is valid for both high-LET (≈10⁴ keV/ μm) and low-LET ions, is presented. It is specially designed for a comparison of the patterns of energy deposited by a large range of ions. Low-LET ions are described in a perturbative frame, whereas heavy ions with a very high stopping power are treated using the Lindhard local density approximation and the Russek and Meli statistical method. Ionization cross sections singly differential with energy compare well with the experiment. As an illustration of the non-perturbative interaction of high-LET ions, a comparison between the ion tracks of light and heavy ions with the same specific energy is presented (1.4 MeV/nucleon helium and uranium ions). The mean energy for ejected electrons was found to be approximately four times larger for uranium than for helium, leading to a much larger track radius in the first case. For electrons, except for the excitation cross sections that are deduced from experimental fits, cross sections are derived analytically. For any orientation of the target molecule, the code calculates multiple differential cross sections as a function of the ejection and scattering angles and of the energy transfer. The corresponding singly differential and total ionization cross sections are in good agreement with experimental data. The angular distribution of secondary electrons is shown to depend strongly on the orientation of the water molecule.

Wan, X. S., Zhou, Z., Ware, J. H. and Kennedy, A. R. Standardization of a Fluorometric Assay for Measuring Oxidative Stress in Irradiated Cells. Radiat. Res. 163, 232–240 (2005).

The present study was undertaken to standardize a dichlorofluorescein (DCF) assay for measurement of radiation-induced oxidation of dichlorofluorescin (DCFH) substrate in MCF-10 cells. This assay was highly sensitive and capable of detecting increased DCFH oxidation in the cells exposed to γ radiation at doses as low as 1.5 cGy with linear dose–response curves. However, the slope of the dose–response curves varied considerably from one experiment to another and was influenced by the fluorescent substrate concentration and cell density. To make the assay reproducible so that results obtained from different experiments could be compared, a series of conversion factors and equations have been established to normalize the data for these variables. The results demonstrate that the DCF assay, as standardized in the present study, is highly reproducible with acceptable assay precision. The normalized results can be compared from one experiment to another even when the experiments were performed using different fluorescent substrate concentrations and/or cell densities. Since changes in DCFH oxidation may be related to changes that are indicative of oxidative stress in cells, this assay can be useful to quantify radiation-induced oxidative stress and evaluate the efficacy of antioxidant agents in protection against radiation-induced oxidative stress.

Funayama, T., Wada, S., Kobayashi, Y. and Watanabe, H. Irradiation of Mammalian Cultured Cells with a Collimated Heavy-Ion Microbeam. Radiat. Res. 163, 241–246 (2005).

As the first step for the analysis of the biological effect of heavy charged-particle radiation, we established a method for the irradiation of individual cells with a heavy-ion microbeam apparatus at JAERI-Takasaki. CHO-K1 cells attached on a thin film of an ion track detector, CR-39, were automatically detected under a fluorescence microscope and irradiated individually with an ⁴⁰Ar¹³ ion (11.5 MeV/nucleon, LET 1260 keV/μm) microbeam. Without killing the irradiated cells, trajectories of irradiated ions were visualized as etch pits by treatment of the CR-39 with an alkaline-ethanol solution at 37°C. The exact positions of ion hits were determined by overlaying images of both cells and etch pits. The cells that were irradiated with argon ions showed a reduced growth in postirradiation observations. Moreover, a single hit of an argon ion to the cell nucleus resulted in strong growth inhibition. These results tell us that our verified irradiation method enables us to start a precise study of the effects of high-LET radiation on cells.