Evans, S. M., Jenkins, K. W., Jenkins, W. T., Dilling, T., Judy, K. D., Schrlau, A., Judkins, A., Hahn, S. M. and Koch, C. J. Imaging and Analytical Methods as Applied to the Evaluation of Vasculature and Hypoxia in Human Brain Tumors. Radiat. Res. 170, 677–690 (2008).

Tissue hypoxia results from the interaction of cellular respiration, vascular oxygen carrying capacity, and vessel distribution. We studied the relationship between tumor vasculature and regions of low pO₂ using quantitative analysis of binding of the 2-nitroimidazole EF5 given to patients intravenously (21 mg/kg) approximately 24 h preceding surgery. We describe new computer algorithms for determining EF5 binding as a function of radial distance from individual blood vessels and converting this value to tissue pO₂. Tissues from six human brain tumors were assessed. In a hemangiopericytoma, a WHO Grade 2 and WHO Grade 3 glial brain tumor, all tissue pO₂ values calculated by EF5 binding were >20 mmHg (described as “physiologically oxygenated”). In these three tumors, EF5 binding gradients (measured as a function of distance from each observed vessel) were low, with small positive and negative values averaging close to zero. Much lower tissue oxygen levels were found, including near some vessels, in glioblastomas. Gradients of EF5 binding away from vessels were larger in glioblastomas than in the low-grade tumors, but positive and negative values again averaged to near zero. Based on these preliminary data, we hypothesize a new paradigm for tumor blood flow in human brain tumors whereby in-flowing and out-flowing blood patterns may have contrasting effects on average tissue EF5 (and by inference, oxygen) gradients. Our studies also imply that neither distance to the nearest blood vessel nor distance from each observed blood vessel provide reliable estimates of tissue pO₂.

Romanenko, A., Bebeshko, V., Hatch, M., Bazyka, D., Finch, S., Dyagil, I., Reiss, R., Chumak, V., Bouville, A., Gudzenko, N., Zablotska, L., Pilinskaya, M., Lyubarets, T., Bakhanova, E., Babkina, N., Trotsiuk, N., Ledoschuk, B., Belayev, Y., Dybsky, S. S., Ron, E. and Howe, G. The Ukrainian-American Study of Leukemia and Related Disorders among Chornobyl Cleanup Workers from Ukraine: I. Study Methods. Radiat. Res. 170, 691–697 (2008).

There are relatively few data on the risk of leukemia among those exposed to external radiation during cleanup operations after the Chornobyl nuclear accident, and results have not been consistent. To investigate this further, we assembled a cohort of 110,645 male cleanup workers from Ukraine and identified cases of leukemia occurring during the period 1986 to 2000. Detailed interviews were conducted and individual bone marrow doses estimated using a new time-and-motion method known as RADRUE described in companion paper II. For the initial analyses we used a nested case-control approach with a minimum of five controls per case, matched for year of birth, oblast (region) of registration, and residence. All identified cases were reviewed by an international panel of experts; 87 of 111 were confirmed. The dose–response analysis and results are given in companion paper III. As background, we describe herein the design, procedures, outcome of case finding and confirmation, control selection, dose estimation and interviewing of subjects.

Chumak, V. V., Romanenko, A. Y., Voillequé, P. G., Bakhanova, E. V., Gudzenko, N., Hatch, M., Zablotska, L. B., Golovanov, I. A., Luckyanov, N. K., Sholom, S. V., Kryuchkov, V. P. and Bouville, A. The Ukrainian-American Study of Leukemia and Related Disorders among Chornobyl Cleanup Workers from Ukraine: II. Estimation of Bone Marrow Doses. Radiat. Res. 170, 698–710 (2008).

After the accident that took place on 26 April 1986 at the Chornobyl nuclear power plant, hundreds of thousands of cleanup workers were involved in emergency measures and decontamination activities. In the framework of an epidemiological study of leukemia and other related blood diseases among Ukrainian cleanup workers, individual bone marrow doses have been estimated for 572 cases and controls. Because dose records were available for only about half of the study subjects, a time-and-motion method of dose reconstruction that would be applicable to all study subjects, whether dead or alive, was developed. The doses were calculated in a stochastic mode, thus providing estimates of uncertainties. The arithmetic mean individual bone marrow doses were found to range from 0.00004 to 3,300 mGy, with an average value of 87 mGy over the 572 study subjects. The uncertainties, characterized by the geometric standard deviation of the probability distribution of the individual dose, varied from subject to subject and had a median value of about 2. These results should be treated as preliminary; it is likely that the dose calculations and particularly the uncertainty estimates will be improved in the follow-up of this effort.

Romanenko, A. Ye., Finch, S. C., Hatch, M., Lubin, J. H., Bebeshko, V. G., Bazyka, D. A., Gudzenko, N., Dyagil, I. S., Reiss, R. F., Bouville, A., Chumak, V. V., Trotsiuk, N. K., Babkina, N. G., Belyayev, Yu., Masnyk, I., Ron, E., Howe, G.;thR. and Zablotska, L. B. The Ukrainian-American Study of Leukemia and Related Disorders among Chornobyl Cleanup Workers from Ukraine: III. Radiation Risks. Radiat. Res. 170, 711–720 (2008).

Leukemia is one of the cancers most susceptible to induction by ionizing radiation, but the effects of lower doses delivered over time have not been quantified adequately. After the Chornobyl (Chernobyl) accident in Ukraine in April 1986, several hundred thousand workers who were involved in cleaning up the site and its surroundings received fractionated exposure, primarily from external γ radiation. To increase our understanding of the role of protracted low-dose radiation exposure in the etiology of leukemia, we conducted a nested case-control study of leukemia in a cohort of cleanup workers identified from the Chornobyl State Registry of Ukraine. The analysis is based on 71 cases of histologically confirmed leukemia diagnosed in 1986–2000 and 501 age- and residence-matched controls selected from the same cohort. Study subjects or their proxies were interviewed about their cleanup activities and other relevant factors. Individual bone marrow radiation doses were estimated by the RADRUE dose reconstruction method (mean dose = 76.4 mGy, SD = 213.4). We used conditional logistic regression to estimate leukemia risks. The excess relative risk (ERR) of total leukemia was 3.44 per Gy [95% confidence interval (CI) 0.47–9.78, P < 0.01]. The dose response was linear and did not differ significantly by calendar period of first work in the 30-km Chornobyl zone, duration or type of work. We found a similar dose–response relationship for chronic and non-chronic lymphocytic leukemia [ERR = 4.09 per Gy (95% CI < 0–14.41) and 2.73 per Gy (95% CI < 0–13.50), respectively]. To further clarify these issues, we are extending the case-control study to ascertain cases for another 6 years (2001–2006).

Kesminiene, A., Evrard, A-S., Ivanov, V. K., Malakhova, I. V., Kurtinaitis, J., Stengrevics, A., Tekkel, M., Anspaugh, L. R., Bouville, A., Chekin, S., Chumak, V. V., Drozdovitch, V., Gapanovich, V., Golovanov, I., Hubert, P., Illichev, S. V., Khait, S. E., Kryuchkov, V. P., Maceika, E., Maksyoutov, M., Mirkhaidarov, A. K., Polyakov, S., Shchukina, N., Tenet, V., Tserakhovich, T. I., Tsykalo, A., Tukov, A. R. and Cardis, E. Risk of Hematological Malignancies among Chernobyl Liquidators. Radiat. Res. 170, 721–735 (2008).

A case-control study of hematological malignancies was conducted among Chernobyl liquidators (accident recovery workers) from Belarus, Russia and Baltic countries to assess the effect of low- to medium-dose protracted radiation exposures on the relative risk of these diseases. The study was nested within cohorts of liquidators who had worked around the Chernobyl plant in 1986–1987. A total of 117 cases [69 leukemia, 34 non-Hodgkin lymphoma (NHL) and 14 other malignancies of lymphoid and hematopoietic tissue] and 481 matched controls were included in the study. Individual dose to the bone marrow and uncertainties were estimated for each subject. The main analyses were restricted to 70 cases (40 leukemia, 20 NHL and 10 other) and their 287 matched controls with reliable information on work in the Chernobyl area. Most subjects received very low doses (median 13 mGy). For all diagnoses combined, a significantly elevated OR was seen at doses of 200 mGy and above. The excess relative risk (ERR) per 100 mGy was 0.60 [90% confidence interval (CI) −0.02, 2.35]. The corresponding estimate for leukemia excluding chronic lymphoid leukemia (CLL) was 0.50 (90% CI −0.38, 5.7). It is slightly higher than but statistically compatible with those estimated from A-bomb survivors and recent low-dose-rate studies. Although sensitivity analyses showed generally similar results, we cannot rule out the possibility that biases and uncertainties could have led to over- or underestimation of the risk in this study.

Muggenburg, B. A., Guilmette, R. A., Hahn, F. F., Diel, J. H., Mauderly, J. L., Seilkop, S. K. and Boecker, B. B. Radiotoxicity of Inhaled ²³⁹PuO₂ in Dogs. Radiat. Res. 170, 736–757 (2008).

Beagle dogs inhaled graded exposure levels of insoluble plutonium dioxide (²³⁹PuO₂) aerosols in one of three monodisperse particle sizes at the Lovelace Respiratory Research Institute (LRRI) to study the life-span health effects of different degrees of α-particle dose non-uniformity in the lung. The primary noncarcinogenic effects seen were lymphopenia, atrophy and fibrosis of the thoracic lymph nodes, and radiation pneumonitis and pulmonary fibrosis. Radiation pneumonitis/ pulmonary fibrosis occurred from 105 days to more than 11 years after exposure, with the lowest associated α-particle dose being 5.9 Gy. The primary carcinogenic effects also occurred almost exclusively in the lung because of the short range of the α-particle emissions. The earliest lung cancer was observed at 1086 days after the inhalation exposure. The most common type seen was papillary adenocarcinoma followed by bronchioloalveolar carcinoma. These lung cancer results indicate that a more uniform distribution of α-particle dose within the lung has an equal or possibly greater risk of neoplasia than less uniform distributions of α-particle dose. The results are consistent with a linear relationship between dose and response, but these data do not directly address the response expected at low dose levels. No primary tumors were found in the tracheobronchial and mediastinal lymph nodes despite the high α-particle radiation doses to these lymph nodes, and no cases of leukemia were observed.

Dynlacht, J. R., Valluri, S., Lopez, J., Greer, F., DesRosiers, C., Caperell-Grant, A., Mendonca, M. S. and Bigsby, R. M. Estrogen Protects against Radiation-Induced Cataractogenesis. Radiat. Res. 170, 758–764 (2008).

Cataractogenesis is a complication of radiotherapy when the eye is included in the treatment field. Low doses of densely ionizing space radiation may also result in an increased risk of cataracts in astronauts. We previously reported that estrogen (17-β-estradiol), when administered to ovariectomized rats commencing 1 week before γ irradiation of the eye and continuously thereafter, results in a significant increase in the rate and incidence of cataract formation and a decreased latent period compared to an ovariectomized control group. We therefore concluded that estrogen accelerates progression of radiation-induced opacification. We now show that estrogen, if administered continuously, but commencing after irradiation, protects against radiation cataractogenesis. Both the rate of progression and incidence of cataracts were greatly reduced in ovariectomized rats that received estrogen treatment after irradiation compared to ovariectomized rats. As in our previous study, estradiol administered 1 week prior to irradiation at the time of ovariectomy and throughout the period of observation produced an enhanced rate of cataract progression. Estrogen administered for only 1 week prior to irradiation had no effect on the rate of progression but resulted in a slight reduction in the incidence. We conclude that estrogen may enhance or protect against radiation cataractogenesis, depending on when it is administered relative to the time of irradiation, and may differentially modulate the initiation and progression phases of cataractogenesis. These data have important implications for astronauts and radiotherapy patients.

Mitchel, R. E. J., Burchart, P. and Wyatt, H. A Lower Dose Threshold for the in vivo Protective Adaptive Response to Radiation. Tumorigenesis in Chronically Exposed Normal and Trp53 Heterozygous C57BL/6 Mice. Radiat. Res. 170, 765–775 (2008).

Low doses of ionizing radiation to cells and animals may induce adaptive responses that reduce the risk of cancer. However, there are upper dose thresholds above which these protective adaptive responses do not occur. We have now tested the hypothesis that there are similar lower dose thresholds that must be exceeded to induce protective effects in vivo. We examined the effects of low-dose/low-dose-rate fractionated exposures on cancer formation in Trp53 normal or cancer-prone Trp53 heterozygous female C57BL/6 mice. Beginning at 6 weeks of age, mice were exposed 5 days/week to single daily doses (0.33 mGy, 0.7 mGy/h) totaling 48, 97 or 146 mGy over 30, 60 or 90 weeks. The exposures for shorter times (up to 60 weeks) appeared to be below the level necessary to induce overall protective adaptive responses in Trp53 normal mice, and detrimental effects (shortened life span, increased frequency) evident for only specific tumor types (B- and T-cell lymphomas) were produced. Only when the exposures were continued for 90 weeks did the dose become sufficient to induce protective adaptive responses, balancing the detrimental effects for these specific cancers and reducing the risk level back to that of the unexposed animals. Detrimental effects were not seen for other tumor types, and a protective effect was seen for sarcomas after 60 weeks of exposure, which was then lost when the exposure continued for 90 weeks. As shown previously for the upper dose threshold for protection by low doses, the lower dose boundary between protection and harm was influenced by Trp53 functionality. Neither protection nor harm was observed in exposed Trp53 heterozygous mice, indicating that reduced Trp53 function raises the lower dose/ dose-rate threshold for both detrimental and protective tumorigenic effects.

Sangsuwan, T. and Haghdoost, S. The Nucleotide Pool, a Target for Low-Dose γ-Ray-Induced Oxidative Stress. Radiat. Res. 170, 776–783 (2008).

Oxidative stress occurs when the generation of reactive oxygen species (ROS) exceeds the cellular antioxidant capacity. The excess ROS react with and modify cellular components. Nucleic acid modifications are of principal interest because they may cause mutations. 8-Oxo-7,8-dihydro-2´-deoxyguanosine (8-oxo-dG) is a mutagenic lesion that can be formed by ROS in DNA as well as in the nucleotide pool. 8-Oxo-dG is removed from the DNA by base excision repair and from the nucleotide pool by the nucleotide sanitization enzyme hMTH1. hMTH1 hydrolyzes 8-oxo-dGTP to 8-oxo-dGMP, which is released to the extracellular environment and can serve as a marker of oxidative stress. The aim of this work was to establish the dose–response relationship for radiation-induced extracellular 8-oxo-dG and hMTH1 in the mGy range of γ rays in three cellular model systems: human whole blood, human fibroblasts and stimulated lymphocytes. Extracellular 8-oxo-dG was analyzed with the use of an ELISA and hMTH1 by Western blotting. Our results demonstrate that low-dose ionizing radiation induces a stress response that leads to the formation of extracellular 8-oxo-dG and induction of hMTH1 in all three cellular model systems tested. This suggests that the nucleotide pool is an important target for radiation-induced stress response.

Frankenberg-Schwager, M., Becker, M., Garg, I., Pralle, E., Wolf, H. and Frankenberg, D., The Role of Nonhomologous DNA End Joining, Conservative Homologous Recombination, and Single-Strand Annealing in the Cell Cycle-Dependent Repair of DNA Double-Strand Breaks Induced by H₂O₂ in Mammalian Cells. Radiat. Res. 170, 784–793 (2008).

The purpose of this study was to investigate the cell cycle-dependent role of nonhomologous DNA end joining (NHEJ), conservative homologous recombination (HR), and single-strand annealing (SSA) for the repair of simple DNA double-strand breaks (DSBs) induced by H₂O₂-mediated OH radicals in CHO cells. Cells of the cell lines V3 (NHEJ-deficient), irs1SF (HR-deficient) and UV41 (SSA-deficient) and their parental cell line AA8 were exposed to various concentrations of H₂O₂ in G₁ or S phase of the cell cycle and their colony-forming ability was assayed. In G₁ phase, NHEJ was the most important—if not the only—mechanism to repair H₂O₂-mediated DSBs; this was similar to results obtained in a parallel study of more complex DSBs induced by sparsely or densely ionizing radiation. Unlike HR (irs1SF)- and SSA (UV41)-deficient cells, the sensitivity of NHEJ-deficient V3 cells to H₂O₂ relative to parental AA8 cells in G₁ phase is about 50 times higher compared to 200 kV X rays. This points to a specific role of the catalytic subunit of DNA-PK for efficient NHEJ of H₂O₂-mediated DSBs that are located at sites critical for the maintenance of the higher-order structure of cellular DNA, whereas X-ray-induced DSBs are distributed stochastically. Surprisingly, SSA-deficient cells in G₁ phase showed an increased sensitivity to high concentrations of H₂O₂ relative to the parental wild-type cells and to HR-deficient cells, which may be interpreted in terms of a specific type of H₂O₂-induced damage requiring SSA for repair after its transfer into S phase. In S phase, HR is the most important mechanism to repair H₂O₂-mediated DSBs, followed by NHEJ. In contrast, the action of error-prone SSA may not be beneficial, since SSA-deficient cells are three times more resistant to H₂O₂ than wild-type AA8 cells. This is likely due to channeling of DSBs into the error-free HR repair pathway or into the potentially error-prone NHEJ pathway. Cells with or without a defect in DSB repair are considerably more sensitive to H₂O₂ in S phase compared to G₁ phase. This effect is likely due to the fact that topoisomerase II, which is expressed only in proliferating cells, is a target of H₂O₂, resulting in enhanced accumulation of DSBs and killing of cells treated in S phase with H₂O₂. The relative sensitivities to H₂O₂ differ by orders of magnitude for the four cell lines. This seems to be caused mainly by H₂O₂-mediated poisoning of topoisomerase IIα rather than by a defect in DSB repair.

Groesser, T., Cooper, B. and Rydberg, B. Lack of Bystander Effects from High-LET Radiation for Early Cytogenetic End Points. Radiat. Res. 170, 794–802 (2008).

The aim of this work was to study radiation-induced bystander effects for early cytogenetic end points in various cell lines using the medium transfer technique after exposure to high- and low-LET radiation. Cells were exposed to 20 MeV/ nucleon nitrogen ions, 968 MeV/nucleon iron ions, or 575 MeV/nucleon iron ions followed by transfer of the conditioned medium from the irradiated cells to unirradiated test cells. The effects studied included DNA double-strand break induction, γ-H2AX focus formation, induction of chromatid breaks in prematurely condensed chromosomes, and micronucleus formation using DNA repair-proficient and -deficient hamster and human cell lines (xrs6, V79, SW48, MO59K and MO59J). Cell survival was also measured in SW48 bystander cells using X rays. Although it was occasionally possible to detect an increase in chromatid break levels using nitrogen ions and to see a higher number of γ-H2AX foci using nitrogen and iron ions in xrs6 bystander cells in single experiments, the results were not reproducible. After we pooled all the data, we could not verify a significant bystander effect for any of these end points. Also, we did not detect a significant bystander effect for DSB induction or micronucleus formation in these cell lines or for clonogenic survival in SW48 cells. The data suggest that DNA damage and cytogenetic changes are not induced in bystander cells. In contrast, data in the literature show pronounced bystander effects in a variety of cell lines, including clonogenic survival in SW48 cells and induction of chromatid breaks and micronuclei in hamster cells. To reconcile these conflicting data, it is possible that the epigenetic status of the specific cell line or the precise culture conditions and medium supplements, such as serum, may be critical for inducing bystander effects.

Prisco, M. G., Nasta, F., Rosado, M. M., Lovisolo, G. A., Marino, C. and Pioli, C. Effects of GSM-Modulated Radiofrequency Electromagnetic Fields on Mouse Bone Marrow Cells. Radiat. Res. 170, 803–810 (2008).

We examined the effects of in vivo exposure to a GSM-modulated 900 MHz RF field on the ability of bone marrow cells to differentiate, colonize lymphatic organs, and rescue lethally X-irradiated mice from death. X-irradiated mice were injected with medium alone or containing bone marrow cells from either RF-field-exposed (SAR 2 W/kg, 2 h/day, 5 days/ week, 4 weeks) or sham-exposed or cage control donor mice. Whereas all mice injected with medium alone died, mice that received bone marrow cells survived. Three and 6 weeks after bone marrow cell transplantation, no differences in thymus cellularity and in the frequencies of differentiating cell subpopulations (identified by CD4/CD8 expression) were observed among the three transplanted groups. Mitogen-induced thymocyte proliferation yielded comparable levels in all transplanted groups. As to the spleen, no effects of the RF-field exposure on cell number, percentages of B and T (CD4 and CD8) cells, B- and T-cell proliferation, and IFN-γ production were found in transplanted mice. In conclusion, our results show no effect of in vivo exposure to GSM-modulated RF fields on the ability of bone marrow precursor cells to home and colonize lymphoid organs and differentiate in phenotypically and functionally mature T and B lymphocytes.

Nakamura, H., Ejiri, H. and Kitamura, H. A New Method for Calibration and Response Measurement of a Scintillation Detector Using Radioisotope Source. Radiat. Res. 170, 811–814 (2008).

A new method for calibration was developed that takes the energy deposited by radioisotope sources into account; it was validated with a plastic scintillation plate using a ²⁰⁷Bi source. The energy deposits of the conversion electrons in the ²⁰⁷Bi source led to an energy difference of a few keV for the 976 keV monoenergetic peak. Calibration with high accuracy is essential to estimate radiation doses as well as to evaluate detector performance. The method will be available not only for use in scintillation detectors but also for other detectors.