Heat-shock protein 90 (Hsp90) is a ubiquitous molecular chaperone protein, which is related to the stabilization and activation of various cell cycle checkpoints and signal transduction proteins. Insulin-like growth factor I receptor (IGF-IR) overexpressed HeLa cells (HeLa-IGF-IR) exhibited a radioresistant phenotype. We have studied the radiosensitization effects of an HSP90 inhibitor 17AAG in HeLa-IGF-IR cells. After pretreatment with 17-AAG for 24 hours, cellular radiosensitivities were studied with colony-forming assay, MTT assay, western blotting, SLGA, and immunostaining. 150nM of 17AAG pretreatment significantly radio-sensitized HeLa-IGF-IR cells, but this sensitization was not observed with non-transfected HeLa cells. Protein expression of PARP after irradiation was reduced with the 17-AAG pretreatment, indicating apoptosis formation. X-irradiation with 17-AAG pretreatment in HeLa-IGF-IR cells led to reduction in cell growth kinetics, spheroid formation prevention, and hyper-sensitivity for gamma-H2AX and p-ATM foci formation when compared to HeLa cells. The downstream pathways of IGF-IR (PI3-K/Akt and Raf/MEK/ERK pathways, proliferation signal pathways) were inhibited by 17AAG treatment, and more cells were led to apoptosis and SLGA. 17AAG treatment would be a novel method for certain radio-resistant tumor cells.

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DNA double strand break (DSB) is the main factor for radiation induced cell death. Cells repair those lethal DSBs with two major repair pathways; homologous recombination repair (HRR) and non-homologous end joining (NHEJ). HRR is the error free repair process and efficient in late S to G2-phases of cell cycle. Rad51 proteins, essential for the HRR, form nucleoprotein filaments on the single strand DNA induced by DSB to search homologous pairing for repair and strand exchange reactions between homologous sequences. HRR deficiency enhances cellular radiosensitivity. The aim of this study is to evaluate the radiosensitization of Hsp90 inhibitor 17-Allylamino-17-demethoxygeldanamycin (17AAG), specifically the effect of 17AAG on the DNA DSB repair machinery. Our constant field gel electrophoresis studies indicated that pretreatment with 17AAG for 24 hours inhibited radiation induced DSB repair in two cancer cell lines (DU145 and SQ-5). The treatment of 17AAG alone leads to the reduction of Rad51 protein expression by western blotting, and the combined treatment with X-irradiation caused a delay in the formation of nuclear Rad51 foci by immuno-staining. These results suggest that 17AAG affects the key protein(s) for HRR, resulting in the radiosensitization of tumor cells. Our data show for the first time that 17AAG is a DNA DSB repair inhibitor, predominantly affecting the HRR pathway. The combination of ionizing radiation and 17AAG would be a useful method for the tumor radio-therapy.

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(Background) FGFs are currently composed of 22 members in human. They have important roles in numerous biological events such as angiogenesis, wound repair and so on. Recently several FGFs have been put to the clinical trials. FGF1 has more potential because it can react on all FGFRs. However, there are a few reports about the efficacy of FGF1. (Aim) An efficacy of FGF1 on radiation-induced intestinal injuries was investigated to realize an appropriate condition of its administration. (Methods) For crypt regeneration assay (CRA), Balb/c mice was exposed to 8 Gy irradiation under several conditions of i.p. administration of FGF1. Crypt cell apoptosis was investigated for the mice 24 hours after irradiation by Tunel assay. (Result) Increase of crypt survival was shown with FGF1 administration even after irradiation by CRA. Especially, maximum experimental dose, 100μg, had most beneficial effect. FGF1 decreased cumulative apoptosis of irradiated crypts in a dose-dependent manner. The LD_50/6 (11.5Gy) was not increased by the pretreatment of FGF1, however the jejunum tissue damage was slighter that control. (Discussion) These findings showed that FGF1 had an effect on the radiation-induced intestinal damage in a dose-dependent manner. The jejunum tissue damage was histopathologically improved by FGF1, although LD_50/6 was not increased. The death of highly irradiated mice might be caused by not only intestine damage, but also multiple organ damage. Therefore it was supposed that FGF1 had the potential to improve the intestine damage.

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Lactoferrin (LF) is an iron-binding glycoprotein with a molecular weight of about 80,000. The radioprotective effect of lactoferrin was studied in C3H/He mice following whole-body X-ray irradiation. We already reported that the survival rates of LF-diet (which contained 0.1 % w/w LF) mice after X-ray irradiation were about 23% higher than those of standard diet-mice. The scavenging abilities were evaluated by the ESR spin-trapping method and we showed that lactoferrin is a radical scavenger of hydroxyl radicals. The observations suggested that radical trapping or scavenging might cause the biological response. In this experiment, 52 of C3H/He mice were exposed 6.8 Gy of X-ray irradiation. After the irradiation, LF (4 mg) was administered 26 of mice intraperitonealy and survival rates of these mice were observed within 30 days. The survival rates of lactoferrin-treated mice and non-treated mice were 93 % and 50 %, respectively within 30 days after X-ray irradiation. Lactoferrin inhibited splenocytes apoptosis and damage on a bone marrow cell after X-ray irradiation.

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Purpose: 5-azacitidine (5-Aza-CR), a DNA methyltransferase inhibitor used in some clinical trials, shows anti-tumor activity by inhibiting methylation of several tumor suppressor genes. On the other hand, 5-Aza-CR shows cytotoxicity. Some report say 5-Aza-CR activates apoptotic pathway via p53. We estimated the effect of 5-Aza-CR on radiosensitivity of cell lines with wild or mutant p53. Materials and methods: human non-small lung cancer cell lines with p53 null type transfected with neomycine resistant vector (H1299/neo) or with wild type p53 (H1299/wp53) or mutant p53 (H1299/mp53) were used in the experiment. Cells were subcultured in D-MEM containing 10% of fetal bovine serum at 37C in 5% CO2 atmosphere. Cells with exponentially growing phase were used in the experiment. Radiation was given by 10MV linac X-ray. Cell viability was estimated by colony forming ability. Results: Plating efficiency of 3 cell lines declined around 30% of control by exposing the cells to 100uM of 5-Aza-CR for 48 hours. No difference in cytotoxicity of 5-Aza-CR was seen between 3 cell lines. 5-Aza-CR slightly enhanced radiosensitivity of H1299/wp53 cell line, while in H1299/neo and H1299/mp53 cell lines, 5-Aza-CR showed no modification effect in radiosensitivity. Conclusion: 5-Aza-CR slightly enhanced radiosensitivity of cell lines with wild type p53.

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Apigenin, a common dietary flavonoid present in many fruits and vegetables, is a nonmutagenic chemopreventive agent. In the present study, we investigated the effect of apigenin on the radiosensitivity of SQ-5 cells, which are derived from a human lung carcinoma. Actively growing cells were incubated for 16 h in medium containing 40 µM apigenin. The cells were then irradiated with X-rays and incubated with apigenin for a further 8 h. Radiosensitivity was assessed using a clonogenic assay. Apoptosis and necrosis were assessed using acridine orange/ethidium bromide double staining. Cells incubated with apigenin exhibited significantly greater radiosensitivity and apoptosis levels than cells not incubated with apigenin. Protein levels were measured by Western blotting. Incubation with apigenin increased protein expression of WAF1/p21 and decreased protein expression of Bcl-2. Furthermore, apigenin sensitized SQ-5 spheroids (cell aggregates growing in a three-dimensional structure that simulate the growth and microenvironmental conditions of in vivo tumors) to radiation. Thus, apigenin appears to be a promising radiosensitizing agent for use against human carcinomas.

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In a previous study, we have demonstrated that in the radio-sensitive megakaryocyte progenitor cells, namely colony-forming unit megakaryocytes (CFU-Meg), the degree of X-ray-induced damages was prevented by the post-treatment with several cytokines. On the other hand, heavy ion particles provide unique properties in radiotherapy. However, they have also been shown to pose high risks for both accident victims and astronauts participating in space missions. We therefore analyzed that the effects of cytokines on heavy ion particles-irradiated CFU-Meg.
The CD34⁺ CFU-Meg were isolated from the human placental/umbilical cord blood using a magnetic isolation kit and then were exposed to a carbon ion beam (LET=50 KeV/micro m). The number of CFU-Meg were calculated by a plasma clot technique. The differentiation to megakaryocytes and the release of platelets and the increase of gamma-H2AX, a marker of DNA double-strand breaks (DSBs), were both analyzed by flow cytometry.
The sensitivity to carbon ion beam was found to be extremely much high and it could not be lowered by any type of cytokine combination unlike X-rays. However, a treatment with thrombopoietin and interleukin-3 potentially induced megakaryocytepoiesis at 14 days after the exposure of a carbon ion beam at 2 Gy. The induction of gamma-H2AX by carbon ion beam irradiation in CD34⁺ cells was not influenced by cytokine treatment. These results showed that the repair of DSBs by cytokine was therefore less strongly promoted in the progenitors to carbon ion beam than in X-rays.

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Since biological effects of radiation are initially induced by formation of free radicals, we have investigated many antioxidants and found that several antioxidants are effective for radioprotector when evaluated by 30 day-survival of mice. In the present study, we examined such antioxidant radioprotectors for X-irradiation-induced thymic lymphoma of mice (C57BL/6NCrj, female, 4-week old). Five reagents (edalavone, HM-PROXYL, POBN, TMG, and cysteamine) were used. The protocol for inducing thymic lymphoma is repeated whole body X-irradiation of 1.6 Gy each, four times with interval of a week. The reagents were administered i.p. before each irradiation. The mice were observed for 200 days and the organ samples were corrected when it died. The incidence of thymic lymphoma for the group of mice with X-irradiation only (control group) was 56%. The incidence for antioxidant-injected groups was 42-75%, which is not significantly different from the control. Only cysteamine prolonged the survival curve. The survival curve of non-thymic lymphoma group showed that cysteamine and TMG were significantly effective for extending the life span. These findings suggest that some antioxidants are effective for reducing some late effects of irradiation other than generation of thymic lymphoma and its related death.

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Both adriamycin (ADM) and hyperthermia show thermal chemo-enhancement. Tolerance induction against ADM in heated cells has been reported resulting in clinical difficulty of cancer therapy. We had reported that thermo-enhancement induced by simultaneous or sequential combination therapy with 42^oC hyperthermia and ADM in Chinese hamster V79 cells in vitro. The simultaneous treatments with 42^oC hyperthermia and ADM resulted in the regression curve of the survival fraction with markedly steeper slope for the initial 1 hour of the treatment, followed by the gentle exponential regression for further 4 hours. In the effect of ADM on thermotolerant V79 cells from step-up hyperthermia (42^oC-44^oC), the thermotolerance in the V79 cells markedly induced by the 42^oC preheating for 2 hours was reduced by ADM, that was examined by the following 44^oC hyperthermia for the graded periods. The enhancement effects of ADM on the sensitivity of a human lung adenocarcinoma A549 cells with wild type p53 and mutant K-ras gene to hyperthermia were investigated in vitro. These results showed an additive thermo-enhancement effect that reduced not only survival, but also the shoulder width. Further thermosensitivity of A549 cells with transformed mutant p53 gene were compared to parent cells with wild type p53 gene. Kinetics of apoptosis induction was investigated after ADM, hyperthermia and the combined treatments.

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Scid cells are sensitive to heat treatment. We analyzed the DNA double-strand breaks induced by heat treatment in scid cells using rH2AX foci formation. Scid cells and Hybrid cells were ideal system to study the DNA double-strand breaks repair mechanisms, because their genetic backgrounds were identical except DNA-PK activity. To clarify the relationship between thermotolerance and heat-induced DSB, scid cells and hybrid cells were conditioned by preheating at 44C and then incubated at 37C for various intervals. Scid cells were heated at 44C for different periods. In the present study, we found that a preheat treatment led to heat-induced thermotoleranace. It was confirmed that the number of rH2AX foci was reduced in preheated cells later treated with a challenge heat. From these results, it is suggested that thermotolerance is related with heat-induced DSB. We considered that the main effect of DNA double-strand break repair of scid cells deduced to the lack of DNA-PK activity.

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We investigated the enhancement of hyperthermic effects and immune activity using extracts of mushroom (Iwade strain 101;IH and Iwade Grifola Gargal;IGG).The used animals were C57BL mice bearing EL-4 tumor cells. The heat treatment was given by hot water bath(42 degree, 30min) after administration of IH(150mg/kg) and IGG(150mg/kg) to pelvis region. The spleen was removed after each treatment and single cell suspension with PBS solution. The EL-4 and YAC-1 cells were labeled with Cr-51 and single cell of spleen were mixed with labeled cells for NK cell activity assay. When the control group of NK cells was regard as 1.00, 1.75 in heat alone group, 1.44 in IH alone group, 2.25 in IGG alone group, 1.11 in tumor transplantation group, 3.80 in IH combined with heat group, 4.03 in IGG combined with heat group. Also indicated the anti-tumor effects in IH and IGG combined with heat treatment due to increase of immunological activity.These results suggest that IH and IGG could be increase the immune activity and enhancemnt of hyperthermic effects without side effect, and maintain the QOL of patients for clinically.

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Functional imaging using S-[methyl-11C]methionine (11C-Met) provides earlier estimation of tumor response after therapy compared to anatomical methods. However, cyclotron facilities are required to produce and use 11C-Met, due to its short half-life (20 min). Since intracellular energy-dependent transmethylation would be involved in tumor accumulation of 11C-Met, development of 18F (T1/2: 110 min) or 123I (T1/2: 13 h)-labeled radiopharmaceuticals affording diagnosis information similar to that of 11C-Met has been hampered. In this study, to find out the cellular functions that provide diagnosis information similar to energy-dependent transmethylation of 11C-Met, we evaluated the energy-dependent amino acid transport system A, using its specific substrate, [14C] α-(methylamino) isobutyric acid (14C-MeAIB). The carbon ion beam irradiation (3Gy) reduced the uptake of both 14C-MeAIB and 14C-Met to HSG human salivary gland tumor cells. The decrease in the uptake of both 14C-MeAIB and 14C-Met to the tumor cells proceeded in advance of the decrease in cell number and intracellular net ATP levels. The decreased uptake of 14C-MeAIB to carbon ion beam irradiated-HSG cells was caused by the decrease in amino acid transport system A-mediated uptake. These results indicated that the amino acid transport system A would constitute a sensitive target to predict therapeutic efficacies of tumor therapy.

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Medical Internal Radiation Dosimetry Methods assume that the beta particle energy is absorbed completely in the source organ. However, the organs of mice used in fundamental experiments for internal radiotherapy are relatively small compared to the range of high-energy ⁹⁰Y beta particles. Therefore, it is well known that the high-energy beta particle can escape the source organ, resulting in large cross-organ doses. The more realistic geometric model such as a voxel-phantom based on Computed Tomography scans for the mouse has been needed for accurately estimating internal doses from administered beta particle emitters. Then it becomes important to simulate the beta particle transport precisely using the Monte Carlo code. The purpose of this study is to validate a user code to simulate the beta particle transport in the Monte Carlo code system (EGS5) by comparing with the absorbed dose measured with a number of small Fluorescent Glass Dosimeters and two capsules (9.6MBq, 1.1MBq) of ⁹⁰YCl₃ solution implanted in a small phantom (Tough-Water; 30mm x 30mm x 70mm) simulated mice. It was found that the absorbed dose distribution in this phantom simulated with this user code gives rather good agreement with the measured results.

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<Purpose> In A-bomb survivors, Sawada noted that the number of women experiencing menopause immediately after exposure was significantly higher in the proximally exposed group compared with the distally exposed group, with average menopausal age tending to be significantly younger particularly among those with acute radiation syndrome group. However, he concluded that the effects on lowering the age at menopause from the serious general symptoms of radiation syndrome should not be ignored. In this study, we determined relationship between age at menopause occurring several decades after exposure and radiation exposure in the Life Span Study (LSS) population of the Radiation Effects Research Foundation (RERF). <Methods> RERF has thus far conducted three mail surveys (in 1969, 1978 and 1991) on the female subjects of the LSS population, which it continues to study on a follow-up basis. About 18,000 subjects among those with available information on radiation exposure dose replied to the question about menopausal age in one of three surveys. The study population was divided into two groups, whether exposure occurred before or after menarche, because it was thought that association of radiation and age at menopause would be different between these groups. We also considered several factors considered related to age at menopause. <Result> Age at menopause was significantly younger with increased radiation dose in the group exposed after menarche. In the group exposed before menarche, the same tendency was observed, but the difference was not significant.

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We evaluated the accuracy of information on acute radiation injury in atomic bomb survivors by comparing the information obtained from two surveys: one conducted by US-Japan Joint Commission in 1945-1946 (Survey A) and the other conducted by Nagasaki city office administrating self-reported questionnaire in 1960-1965 (Survey B). Information on name, age and place at the bombing identified 627 individuals who were in both surveys. The frequency of respective acute injuries observed in Survey A and Survey B was as follows: diarrhea—35% vs. 32%; vomiting—28% vs. 20%; fever—22% vs. 30%; stomatitis—20% vs. 13%; epilation—14% vs. 23%; gingival bleeding—12% vs. 18%; subcutaneous bleeding—12% vs. 16%; and nasal bleeding—3% vs. 9%. Epilation was kept best in memory by survivors; 74 % of those who had been identified epilation by Survey A also reported epilation in Survey B.

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A low-dose risk is a controversial issue because there are gaps between epidemiology and biology. Cancer is thought to be induced through multi-mutational steps. Some biologically-based models can well describe the age pattern of the cancer incidence among the atomic bomb survivors. However, the role of radiation in carcinogenesis is unclear since model-fitting to data shows a good result but the meanings of biological parameters are different among the models. Recent progress in leukemia research provides a useful information to mathematical modeling. We will present the model for radiation-induced leukemia in support of our "cell kinetics disturbance hypothesis". Note that previous initiation theory of radiation does not have a supportive biological evidence and the spontaneous translocation specific to ALL occur much more frequency than radiation-induced translocation. Moreover, the first step of carcinogenesis during fetus is likely to occur irrespective of radiation exposure. We suggested that asymmetric division of stem cells supply mature cells and genomic instability induces delayed cell death. The difference in cellular kinetics between mutated and non-mutated stem cells results in clonal expansion of the mutated cells due to their growth advantage. Consequently, risk assessment for carcinogenesis should be based on extrapolation from the dose response relationship of cellular kinetics of stem cell population.

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A total of 1237 Nagasaki A-bomb survivors were interviewed in 1997 and were asked to talk freely about atomic bombing, health concerns, relatives' death, and mental health. Their talk was recorded on tape if allowed. The 30-item version of General Health Questionnaire was also administered to assess their mental health conditions. In the present study, we analyzed the talks of 928 Nagasaki A-bomb survivors who completely responded to all items. One of interview item was "When you are in bad condition, are you concerned about the effects of radiation exposure?" The subjects were classified into two groups of "concerned" and "not concerned" according as their response was "yes" or "no".
We used Wordminer for the analysis: we first decomposed texts into words; then evaluated their frequency in the texts; and finally used them for correspondence analysis.
The words appearing most frequently in the "concerned" group were those about physical conditions, e.g. body, anemia and bone, and those about of diseases, e.g. liver, thyroid and operation. On the other hand, these words appeared rather few in the "not concerned" group. Especially, in those bombed at 19 years of age or less, the words about physical conditions and diseases were frequent in the "concerned" groups.

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Effects of in utero exposure of male fetal Wistar rats on embryonic day 15 to accelerated heavy ions on the fetal gonads, postnatal testicular development and reproductivity were investigated in a dose range from 0.1 Gy to 2.0 Gy. The effects of X-rays at 200 kVp were studied for comparison. A dose of 2.0 Gy caused 100%, 18%, and 12% preweaning mortality by irradiations with neon ions, carbon ions and X-rays, respectively. For neon ions, a dose as low as 0.1 Gy led to significant alterations to such as preweaning mortality, testis weight, and ratio of testis weight to body weight. For carbon ions or X rays, a dose of 0.5 Gy or more was usually needed to induce a marked effect. Irradiation with 0.5 Gy or more, resulted in a dose-dependent induction of apoptosis in gonocytes, increased prenatal death, delayed accomplishment in testis descent, low testis weight, changed ratios of testis weight to body weight, and increased malformed testicular tubules. When the prenatal dose was at 1.0 Gy or 1.5 Gy, marked decrease in successful mating rates with the non-irradiated female was also scored in the prenatally irradiated male. On the other hand, a significant increase in prenatal death and preweaning mortality was observed in animals that were descended from the irradiated male receiving prenatally 0.5 Gy or more and the non-irradiated female. These findings indicated that fetal irradiations on embryonic day 15 generally induced markedly detrimental effect on fetal development, postnatal testicular maturation and reproductivity in male rats, which was a dose and LET-related event.

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Individual radiation dose received by radiation accidents are obtained from the amount of paramagnetic radicals in tooth enamel by ESR (electron spin resonance) measurements. The tooth enamel is a sensitive dosimeter made of hydroxyapatite, while dentin had not been used for dosimetry. Recently, it was reported¹ that neutron doses can be independently obtained when we utilize the difference in the sensitivity of neutron dose relative to that of gamma ray dose between enamel and dentin.

In the present study, tooth enamel and dentin from the victims of JCO accident were measured with ESR. The enamel and dentin were mechanically separated with a high speed diamond saw. They are subsequently crushed to grains of about 1mm in diameter. ESR measurements were done with an ESR spectrometer, JEOL FA-100 at RIRBM, Hiroshima University. The intensities of the dosimetric signal probably due to CO₂^- were obtained by using a computer program named "New ER"². The doses equivalent to gamma ray dose were calculated from the calibration line. The obtained doses from enamel ranged from 11 to 13 Gy which are consistent with the previous work³. Unfortunately, the doses obtained from dentin scattered too much to discuss the neutron doses.

1. F. Trompier et al. (in press) Radiat. Protec. Dosim.
2. A. Ivannikov et al. (2001) Health Phys., 81, 124-137.
3. K. Shiraishi et al. (2002) Jour. Raidat. Res., 43, 331-335.

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Japanese nuclear power plants (NPPs) located in 12 prefectures from Hokkaido to Kyushu islands. Cancer mortality in 1972-97 was analyzed mainly in Ibaraki prefecture where the earliest NPP was located. Standard mortality ratio (SMR) was examined for 85 municipalities which the prefecture consisted of as the end of 1997. The empirical Bayes estimate of SMR, EBSMR was also calculated for the latest 5 years period. EBSMR was less than or about 1.0 except some municipalities where it was more than 1.1 for digestive cancers. Adult T-cell leukemia (ATL) may count for 3% of leukemia deaths in the prefecture. Superficial increase after the commissioning year can be observed for cancers of lymphoid-hemopoietic tissues in females, particularly among the 75 years old or more, commonly through all municipalities with an NPP in Japan compared with selected control areas. Superficial decrease can be observed for solid cancers among the 50 years old or more. The confounding was likely to be caused by temporal changes of age specific mortality and of the number of cancer deaths in the elderly. The proportion of the latter in all cancer deaths became larger in females than in males except non-digestive solid cancers. SMR by large areas such as six blocks through Japan is relatively constant in the long period but not in the ATL endemic area for leukemia and malignant lymphoma. Estimated risk was also influenced by a short term variation in the selected municipalities; for an example, 1983-87 low mortality for non-digestive solid cancer in the area with an NPP in Ibaraki prefecture.

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The treatment of intestinal injury induced by radiation is almost carried out by symptomatic therapy now. We try to develop a treatment method that reproduction of intestines by transplantation of embryonic stem (ES) cells for prevention against the intestinal injury. Intestines of mice (female ICR nu/nu) were irradiated at a single dose of 30 Gy. After irradiation, the mice were transplanted directly with ES cells (male 129/Sv) as a donor to the irradiated intestines. On day 13 to 27 after transplantation, the intestine was removed. To confirm the proliferation and differentiation of ES cells, we determined the Y-chromosome DNA of ES cells by PCR and observed the intestine by histology (H.E.) and immunohistochemistry (SSEA1, bFGF, αSMA, cytokeratin AE1/AE3). The results showed that the cells of donor origin were identified in the intestine of irradiated mice. The transplanted ES cells could proliferate and differentiate in the intestinal tissue that injured by radiation. On day 13 after transplantation, the crypt-like cells were observed, but they are not epithelial cells. Interestingly, we observed that ES cells differentiated into epithelial cells in other area. On day 27, the transplanted ES cells differentiated into various cell types including teratoma. It may be necessary to differentiate the ES cells to epithelial cells in vitro before transplantation, or differentiate transplanted ES cells to crypt cells in vivo.

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While health risk of exposure from radon and its decay products has been estimated based on epidemiological studies for miners, it is required that risk assessment for general public should be based on study conducted in general environment. Recently, the pooled analyses of North American and European epidemiological studies have been conducted and it is pointed out that the lung cancer risk increased significantly even at relatively low levels of radon concentration. Yunnan province, China is the largest tin-producing district in the world. Although the results of epidemiological study of Yunnan tin miners was employed in BEIR-_IV_ report, there are some issues on dose assessment which might lead to lower risk presumption than the other studies. In order to examine the feasibility of epidemiological survey of residential radon in Yunnan, radon, thoron and their decay products measurements were conducted in this study.
Radon and thoron decay products were measured in 100 dwelling of the farm village and 50 of the urban area in Gejiu city, Yunnan in 2004-2006.
The annual dose from indoor radon and thoron decay products for the local populations was estimated based on the several models and assumptions, indicating the dose from thoron decay products could be higher than that from radon decay products. Those results suggest the importance of thoron dose assessment in epidemiological survey of residential radon, though there remain some uncertainties in dose assessment for thoron.

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Recent epidemiological studies on residential radon and lung cancer suggest that lung cancer risks increase significantly with the increase of radon concentration even at relatively low levels, and that the risk estimates per unit radon concentration from residential studies are consistent with those from underground miner studies. In those studies, several adjustments were done for controlling uncertainty in radon measurements to obtain reliable estimates of radon-associated lung cancer risks, either by statistical modeling for spatial and temporal variations in radon concentrations, or by restrictions of subjects for whom radon measurements were done completely.
Existence of thoron in houses can be another main source of uncertainty in radon measurements. Specific types for radon detectors used in case-control studies so far were not able to discriminate thoron from radon, thus, measured radon concentrations were likely to be overestimated in such epidemiological studies in areas with high thoron concentrations such as loess plateau area in China. However, the influence of overestimation of radon due to thoron disturbance on lung cancer risks has not been well addressed.
In this study, we evaluated the magnitude of bias in risk estimates due to radon overestimation, on the basis of simulation using several assumptions on radon/thoron concentration ratio and other factors. The detail results will be presented at the meeting.

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Breathing rate is an important parameter for estimating internal dose due to inhalation of radionuclides. In general, internal dose due to inhalation increases with breathing rate regardless of the kind of radionuclides. ICRP gives a typical breathing rate for each of four activity levels: sleeping, rest/sitting, light work and heavy work. The breathing rates corresponding to these activities range from 0.45 to 3 m³/h. However, these values were determined on the basis of the data on Caucasian. It should be investigated whether these values can be applied to Japanese. There were few studies on the measurement of breathing rate for Japanese. In the present study, a portable (wearable) instrument for measuring breathing rate was developed to estimate typical breathing rates in various activities. An outline of this instrument will be presented including its performance test.

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Last 25 years, intensive research have been conducting to examine whether electric and magnetic fields affect human health or not, especially in extremely low frequency and radio frequency. In contrast, there are few studies have been conducted to examine the effect of intermediate frequency magnetic field (IF-MF) so far. In this study, we examine mutagenicity of IF-MF using mouse lymphoma assay (MLA) that detect wide spectrum of mutagenesis. We made an exposure device with a flat surface coil and CO₂ incubator made with resin. This device can generate 20kHz, 310uT magnetic field that is approximately 50 times stronger than that of the reference level of ICNIRP general exposure guideline at cell position. L5178Y TK+/-3.7.2c cell suspension(1x10⁵ cells/ml) divided three group and exposure group was exposed to IF magnetic field, control group was incubated in a conventional CO₂ incubator and positive control treated with MMS for 48 hours. After exposure period, cells removed and poured into 96-well plates with fresh medium. Half of those were used for colony efficiency and the other was used for mutant selection by TFT. After two weeks incubation, mutation frequency was analyzed. The result shows that no significant difference in mutation frequency between control and exposure. In addition, no significant difference was observed in colony formation such as shape or frequency of large and small colony between control and exposure. These results suggest that IF-MF does not have mutagenic potential in the present experimental condition.

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In recent years, the biological effects of extremely-low-frequency magnetic fields (ELF MFs) on possible health hazards have been reported. In rat brain cells, 0.25-0.5 mT of 60Hz ELF MF introduces DNA double-strand (ds) breaks. On the other, it is known that static high magnetic fields (st MFs) are much more safety and used for MRI. But, biological responses at the molecular level to the st MFs are little understood. Thus, we study the influences of the st MFs on growth, development, reproduction, ds breaks, apoptosis, and alterations of entire gene expression using a model metazoan C. elegans. Under 5T st MF, the growth, development and reproduction normally proceeded. In order to study whether the ds breaks were introduced by the static MFs, we used him-17 mutant, whose activity of meiotic ds breaks for general recombination was reduced, and measured hatching rate of the eggs and incidence of males (XO animals). Both the frequencies were not significantly changed under 2T and 3T st MFs compared with 0T control. In addition, we analyzed alterations of entire gene expression using DNA microarray under 3T and 5T st MFs. The alteration patterns between the cases of exposure of st MFs and gamma rays irradiation were quite different. Those results indicate st MFs did not introduce any DNA damage. We also found that hsp-16 gene expression increased under the st MFs as well as the ELF MFs reported, indicating that hsp-16 gene was response to several MFs. In this presentation, we would like to discuss the biological responses to st MFs including other altered gene expression.

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We measured the amount of AP sites (Apurinic/apyrimidinic sites) in human glioma A172 cells to investigate the effect of extremely low frequency electromagnetic fields (ELF-EMF) at 5 mT. The cells were exposed to ELF-EMF alone, H₂O₂ (hydrogen peroxide) alone, or ELF-EMF with H₂O₂ in medium. The duration of exposure was 2, 4, 8, 16 and 24 hours. After the exposure, the cells were collected by cell-scraper, washed with PBS 3 times, and then we extracted DNA from the cells. After isolating DNA, ARP (Aldehyde Reactive Probe) reagents were added to DNA. Then, AP sites were tagged with biotin residues, which could be quantified using avidin-biotin assay, followed by a colorimetric detection of 450 nm. The number of AP sites were determined by calibration curve of 0∼40ARP/10⁵ bp in standard DNAs. There was no difference in the number of AP sites between ELF-EMF exposed cells and sham control. The level of AP sites was increased H₂O₂ treatment alone. The combined treatment of ELF-EMF and H₂O₂ increased AP site levels on 8, 16 and 24 h exposure compared with H₂O₂ treatment alone. In this study, the exposure of ELF-EMF alone may have no effect to produce AP site. However, AP sites induced by H₂O₂ might be enhanced by ELF-EMF exposure.

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[Purpose] In recent works, sonotherapy have been expected to apply extensively to dermatological disorder and skin homeostasis. Tyrosinase is one of target enzymes of sonotherapy for melanin deposition. In this report, we tried to inactivate tyrosinase in vitro by 1 MHZ ultrasound and to elucidate the basic mechanism of sonotherapy of skin. [Method] As an enzymatic assay of tyrosinase, continuous spectrophotometric rate determination was employed. Reaction mixture (50 nM L-dopa, 2 % DMF, 6 nM MBTH in 1 mM phosphate buffer pH 7.1) containing Mushroom tyrosinase (1 kunits/mg) was prepared and used for the assay. Reactive oxygen species (ROS) were detected by the method combining ESR and spin-trapping. Released Cu²⁺ was quantified with kinetic-catalytic spectrophotometric method. Sonication was performed with 1 MHz Ultax-UX601 (-1.4 W/cm²). [Results] Tyrosinase activity was reduced as increasing power of 1 MHz ultrasound. When the reaction mixture containing spin trap was irradiated, OH-adduct was detected by ESR. However, there was no effect of mannitol (OH scavenger) on the reduction of tyrosinase activity after irradiation. When excess Cu²⁺ was added to the solution, tyrosinase activity was not reduced after irradiation. The analysis of free Cu²⁺ showed that ultrasound mediated Cu²⁺ release from tyrosinase. These results indicate that ultrasound-induced inactivation of tyrosinase is caused by not ROS generated from the cavitaion but the Cu²⁺ release from activation center of the enzyme.

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Mouse lymph node cell (M10) is highly sensitive to X-ray irradiation. This cell is also sensitive against UV-ray irradiation. Growth of this cell depended on the UV-rays (310 nm, band B) irradiation, and the cell growth was suppressed in proportion to the UV irradiation time. UV irradiation doses (10, 20, 30 and 60sec) showed the induction of cell apoptosis and necrosis. Cell sorting analysis distinguished the difference of the apoptosis and the necrosis of M10 cells. Apoptotic cells increased in proportion to UV-irradiation. We also analyzed the gene expression in the UV-irradiated cells using DNA array technology. Dynamic changes of 597 genes after the UV-B (310 nm) irradiation of three doses (10, 20 and 30sec) were investigated on DNA array membranes. Radioactivities obtained from imaging analyzer were analyzed using software of ArrayGauge. The expression of mRNA was further analyzed by EX-ARRAY program. Time course of the gene expression was investigated at 2h and 5h after the irradiation. The expression of almost genes reduced in proportion to the doses. The comparison of the gene expression with the parental cells (L5178Y) by northern blot hybridization demonstrated the activation of several genes (MCP-3, Cdx-2, HMG-14, glucose-regulated protein gene, serotonin receptor gene and Hox-2.4).

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Microgravity and exposure to radiation are major differences between the earth and the space and may be a big stress. Although the liver is the biggest among the intraperitoneal organs and crucial for systemic metabolism, the effect of space environment has not been determined. This study aimed to analyze metabolic changes by measuring expressions of Cytochrome P450 (CYP) and stress-associated molecules in the rat liver after space flight. Male Sprague-Dawley rats after a 9-day space flight (FL) were used. During the mission, rats were singly housed in the research animal holding facility (RAHF) or in the Animal Enclosure Module (AEM), and compared with ground-control (GC). The gene expression levels were determined by Taqman method on total liver RNA. Immunohistochemistry was performed against individual CYP. Within FL animals, CYP2A1, 2C11, 4A1 gene expressions were significantly higher in AEM than in RAHF. CYP3A2 gene expression within GC was significantly higher in the AEM. CYP 1A1, 4A1, Cirbp and Hsp90 gene expressions were significantly higher in FL-AEM than in GC-AEM. The similar tendency was observed in CYP1A1 and 4A1 gene expressions from RAHF. These suggested that CYP1A1 and 4A1 gene expressions were specifically increased by space flight. Immunohistchemistry revealed disarrangement of hepatic architecture after space flight. We concluded that CYP1A1 and 1A4 gene inductions may be sensitive markers for space flight and these induction may be partly caused by architectural changes of the liver tissues.

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Tardigrades are invertebrates whose body length ranges from about 0.1 to 1.0 mm. Terrestrial tardigrades lose their body water almost completely and enter an ametabolic state, called as "anhydrobiosis". Anhydrobiotic tardigrades are known to tolerate extreme environmental conditions, such as high and low temperatures, high pressure and chemicals. In the present study, in order to know radio-tolerance in tardigrades, we examined the survival ability after exposure of heavy ions (⁴He²⁺; 12.5 MeV/amu, LET 16.2KeV/um) and gamma-rays in a tardigrade Milnesium tardigrdum in hydrated and anhydrobiotic states. Hydrated animals showed significantly higher LD₅₀ values (6200 Gy for heavy ions, 5000 Gy for gamma-rays) than anhydrobiotic ones (5200 Gy for heavy ions, 4400 Gy for gamma-rays) 48 hours after irradiation. It is difficult to explain why anhydrobiotic individuals are less radio-tolerant than hydrated ones, because anhydrobiotic animals with extremely low contents of water compared to hydrated ones had been expected to be less damaged by indirect action of radiation compared to hydrated ones. This finding suggests that M. tardigrdum has high ability to repair biological damages after irradiation. And, M. tardigrdum showed higher LD₅₀ values for heavy ions than those for gamma-rays. This result is interesting because high-LET heavy ions generally thought to be more detrimental to organisms than low-LET gamma-rays.

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It is well known that a half of the annual effective dose from natural radiation sources consists of that from radon. The dose is actually derived not from radon gas itself but from its decay products. In actual surveys, however, radon is subject to measure and the dose can be estimated after obtaining its decay products concentration by multiplying an equilibrium factor. In general, the equilibrium factor of radon is assigned to be 0.4 indoors and 0.6 outdoors as the typical value, respectively. Thoron, formed in Th-232 decay series, is present as well as radon in the environment. Previous nationwide indoor radon surveys and many other studies have shown thoron has been detected in the indoor environment. We have measured indoor radon, thoron, thoron decay products concentrations in various environments. The present study describes a summary of the survey results on the equilibrium factor for thoron.

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The active test using actual spent nuclear fuels was carried out in the Japan's first commercial nuclear fuel reprocessing plant in Rokkasho, Japan from March 2006. Since vertical migration rates of ^239+240Pu or ¹³⁷Cs in soil are important factors for assessing behavior of those nuclides in the environment around the plant, we investigated their vertical profiles in soil in Rokkasho, and estimated the migration rates. The vertical distribution of ^239+240Pu and ¹³⁷Cs were obtained for the forest soil in several sampling sites. Their inventories were larger than the values obtained in un-cultivated fields in Rokkasho by our previously work. Mean ²⁴⁰Pu/²³⁹Pu atom ratio of 0.178±0.014 for all samples indicated that the Pu in the samples was derived from global fallout. The activity ratio of ^239+240Pu/¹³⁷Cs was 0.030, which is also typical for weapon testing fallout. Vertical migration velocities of the nuclides in soil up to 15 cm deep were calculated from their historical fallout patterns and current vertical distributions. Their migration rates ranged from 0.1 to 0.6 cm y^-1 and within the range of reported values in European forest soils. There are negative correlation between the migration rate of Pu or ¹³⁷Cs and ignition losses of the soils, and positive one between the rates and mean diameter of soil particles. This work was supported by a grant from Aomori prefecture, Japan.

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The largest fraction of the short-term and long-term dose from accidental releases and fallout from atomic bomb tests was from iodine isotopes. Among the radioisotopes of iodine, I-129 is of most concern due to its long half-live. It has been expected that the environmental behavior of radioactive I-129 should be basically similar to that of natural I-127. Thus, the biogeochemical cycling of stable iodine in the environment can be used for the safety assessment of I-129. We here report a hyphenation technical between HPLC and SF-ICP-MS for the speciation of stable iodine in seawater because the chemical form is one of the most important factors controlling iodine environmental behavior in the ocean. Several HPLC separation columns, including anion-exchange (ExcelPak ICS-A23), cation-exchange (IonPac CG5A) and size-exclusion (AsahiPak GS-220HQ) columns were investigated for their performance for separation of iodate and iodide. Surprisingly, it was found that the best separation was obtained with the short CG5A cation-exchange column with a TCC2 cation concentrator as the guard column. With a mobile phase of 30 mM ammonium carbonate at a flow rate of 1 ml/min, the analysis of iodate and iodide can be completed within 5 min. Seawater can be directed introduced into the HPLC column for analysis after a 5-fold dilution of seawater. The developed analytical method was successfully applied to the study on the vertical distribution of iodine species in coastal water off Aomori, Japan.

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In the Pacific Ocean there are two principal sources of ²³⁹Pu and ²⁴⁰Pu: global stratospheric fallout and close-in fallout from the North Pacific Proving Grounds in the Marshall Islands. The ²⁴⁰Pu/²³⁹Pu atom ratio is an excellent tracer to characterize the sources of Pu and to investigate the transport and scavenging processes in the oceans. Large-volume seawater samples (250 L each) were collected from the Sagami Bay and their ^239+240Pu activities and ²⁴⁰Pu/²³⁹Pu atom ratios were determined by alpha-spectrometry and SF-ICP-MS, respectively. The ^239+240Pu activity was 13 mBq/m³ in the surface water and increased with depth to a maximum value of 39 mBq/m³. The ^239+240Pu inventory in the water column was 41 Bq/m². The ^239+240Pu inventory in the water column in the western Northwest Pacific Ocean was equivalent to that of the integrated deposition density of atmospheric global fallout at the same latitude of 30-40 N. The ²⁴⁰Pu/²³⁹Pu atom ratios showed no notable variation from the surface to the bottom layer and were significantly higher than the global stratospheric fallout ratio. These high ratios could be attributed to close-in fallout Pu delivered from the North Pacific Proving Grounds.

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In the Central Asian countries, scarcity of clean water for human consumption is one of the most important issues, while the impact of uranium producing industry on the environment have been recognized as a threatening problem due to their large amount of uranium production. In order to cope with those problems, monitoring for radionuclides and metals in the environment has been started recently. In relation to the study of behavior of radionuclides in the environment, we have tried to figure out the dietary intakes of radionuclides and trace elements in the region. Diet samples collected in Tashkent in Uzbekistan with duplicate portion method were chemically processed into acid solution, followed by ICP-MS analysis. It was found that dietary intakes of U and Th were 2.4∼8.4 and 0.4∼1.4 μg/day/person, respectively. More detailed discussion, including comparison with our results from the study of Ukrainian diet will be presented at the conference.

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The behavior of plutonium isotopes was studied in the terrestrial and aquatic environment in the area where the Chernobyl accident took place. Radioactive concentrations and physical and chemical forms of the accident-derived Pu were investigated for soil, surface water, suspended solid, and bottom sediment samples in the vicinity of the Chernobyl nuclear power plant. The migration rate from the contaminated ground of the area by fluvial transport was estimated as about 0.1 % maximally in 1986, the year of the accident. Then it was decreased to as low as 0.01% in 2000. Chemically, most of the plutonium isotopes were found in so-called organic phase and least-soluble phase in soil and suspended solid samples. The same was for the bottom sediment collected at the Pripyat River near the power plant. This chemical form suggests limited mobility and solubility of Pu in the studied environment. This is in contrast to strontium-90 that has been found most soluble among Cs, Sr and Pu radioisotopes in the area and broadly influences on the aquatic environment. This study has also revealed that physical form of transport of Pu in river water was as follows: 60-80% of the total Pu in particulate form in suspended solid ; colloidal form 10-20%; truly dissolved form 10-20%. This phase partitioning is similar to that found for Pu derived from a reprocessing plant effluent in the Rhone River and other instances.

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Determination of the concentration and isotope ratios of uranium is required in environmental monitoring of nuclear contamination in nuclear safeguards. ²³⁸U, ²³⁵U and ²³⁴U are naturally occurring alpha-emitting long- lived radionuclides which are taken up daily at low levels with food and drinking. Natural isotopic composition of ²³⁵U/²³⁸U =0.00725. Therefore, isotope ratio measurements are important because they can provide information on the origin of uranium. Uranium, the heaviest naturally occurring element on earth, plays an important role in daily life because of its use in nuclear power plants. The isotope ratios of uranium, ²³⁴U/²³⁸U, ²³⁵U/²³⁸U and ²³⁶U/²³⁸U were measured by using a VG Sector 54 thermal ionization mass spectrometer (TIMS). Uranium isotopic compositions have been determined in soil samples as well as in some ground water samples in the exclusion zone of Chernobyl nuclear power plant. The isotopic composition of Chernobyl soil samples showed significant deviation from the natural uranium and presence of ²³⁶U is quite noticeable. The ²³⁴U/²³⁸U activity ratio varies in the range 1.06 – 2.1 in case of soil samples. Similar variation was also observed for ground water samples. Enrichment of ²³⁵U was also noticeable for soil as well as ground water samples.

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At present, 20 years after the Chernobyl nuclear power plant (CNNP) accident, contamination of radioactive cesium still remains in soil. In this report, we have selected mushrooms and potatoes since both are staple food of Ukraine population. Samples were collected in 2003 from Zhytomir district near Chernobyl, which is north part of Ukraine and have measured Cs-137 concentration. Samples were ashed and then put in a U-8 container up to a height of 10mm and radioactive cesium was measured using a germanium semiconductor detector. Furthermore, soil samples were collected from the same area of food samples. The sampling depth was 20 cm with layers:0-2, 2-4, 4-6, 6-8, 8-10, 10-15, and 15-20. We have measured radioactive Cs concentration of seven layers for depth profile and separately from 0-6 cm soil layer that is considered for transfer factor. Concentration of Cs-137 decreases with deeper layer and there is transfer of radioactive Cs from soil to food even now. In case of Mushrooms, we have noticed Cs-137 concentration was more than K-40 concentration as well as Cs-134. In case of potatoes, concentration of Cs-137 was less than mushrooms. The committed effective dose was calculated using the dose conversion coefficient of ICRP Publ.67, assuming that such food is taken for one year. The committed effective dose for one adult is 0.9mSv due to Cs-137 in mushrooms. However, the amount of potatoes consumed in the Ukraine is about 5 times more while comparing with Japan, and the committed effective dose for an adult is about 1.5mSv.

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Belarus is one of the countries most seriously affected by the Chernobyl nuclear power plant accident. Although radionuclide contaminations in foodstuff have been well studied after the accident, intakes of radionuclides through whole diet samples were very rare in the literature. Therefore, we researched radionuclide intakes using total diet samples with gamma-spectrometry. About 30 whole diet samples were collected by a duplicate portion method from Belarus and incinerated in a furnace. Radionuclides in ash samples were measured with Ge-detector. In all diet samples ¹³⁷Cs was found (detected). Daily ¹³⁷Cs intakes were in the range of 2.4 - 90 Bq and geometric mean value is 8.9 Bq. Annual effective dose (0.42 mSv year^-1) for Belarusian was calculated by using geometric mean value of the ¹³⁷Cs intake. This annual effective dose was much higher than those in Japanese whole diets. Cesium-134 was found in only one sample, which showed highest ¹³⁷Cs concentration in all samples analyzed. Daily intakes of ⁴⁰K in Belarusian were almost the same as reported values in Japanese.

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The dietary intakes of radioactivity in 10 regions (13 cities) in Japan were investigated using the market basket method, and the annual effective doses were estimated. This study is one of "Study on the intakes of poisonous substances in foods and evaluation" (The Ministry of Health, Labor and Welfare). The foodstuffs categorized to 14 groups were obtained from 2003 to 2005. The gamma-ray emitting radionuclides in the samples were determined by gamma spectrometry, ⁹⁰Sr and ²³⁸U were analyzed with the low background beta counter and ICP-MS, respectively. The daily intakes of ¹³⁷Cs in each region ranged from 12.5 to <76.6 mBq/day. And the daily intakes of ⁹⁰Sr and ²³⁸U were in the range of 20.8-53.6 mBq/day, 5.9-31.1 mBq/day, respectively. Any clear difference of the radionuclides intakes among these regions was not found in this study. The annual effective doses for the adult population, which were calculated from the above results, were small enough compared with the dose limit for the public (1 mSv: ICRP 1990 Recommendation).

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There is the increasing need for radiological protection of the environment based on scientific principles with increasing recognition of importance of the environment for human species. The soil has a fundamental role in the ecosystem function such as decomposition of organic matters and production of plants. Soil invertebrates play an important role in the ecosystem function of the soil. They catalyze the cycling of elements, become food for above ground fauna, and influence vegetation biodiversity. Because of their ecological importance, the soil invertebrates, especially earthworms and collembiolans, are increasingly used for ecological impact assessment of chemical substances for the terrestrial ecosystem. Earthworm toxicity test guideline was adopted in OECD, and earthworms are considered to be reference organisms by ICRP. For collembolans, toxicicity test guideline is available in ISO, and an international ring-test is in progress to submit the draft of test guideline to OECD.
We aim to provide fundamental data for ecological impact assessment of radiation for the terrestrial ecosystem. In the present paper, we report the dose-effect relationship of (acute or chronic) gamma-irradiation on reproduction or survival of the earthworm, Eisenia fetida, and the collembolan, Folsomia candida, in laboratory tests.

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A strange phenomena that gamma rays from radon daughter Pb-214 and Bi-214 become significantly low when uranium in radioactive equilibrium state and Ammonium Phosphomolybdate (AMP) was mixed. Extent of the decrease was evaluated to be 1/2 at room temperature and 1/4 at 60 degree. When the sample was stored at liquid nitrogen temperature, this phenomena disappeared but was observed again when the sample was returned to room temperature indicating that this phenomena is temperature-reversible. These observations suggest the importance of physico-chemical state of gaseous radon. Sodium salt showed much higher decrease, i.e,1/10 at room temperature and 1/30 at 60 degree. Furthermore strange, the decrease occurs independent of energy from KX-ray to 2 MeV. These phenomena can not be understood without considering suppression of radioactive decay or photon emission by chemical material due to unknown mechanism. Another possible explanation is the conversion of photon to invisible material . The decrease is caused not by the ordinary shielding effect by lead or iron and requires construction of new physics. .If this phenomena was explained theoretically, dreamy application field such as confinement of nuclear waste will be .developed

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Neutron activatoins were examined for commercial dosimeters and dosimeter candidates; these materials were exposed to d(4)+Be neutrons (major energy range: 1-2 MeV) produced by the tandem electrostatic accelerator NASBEE at NIRS. The dose of 2Gy (air kerma) was given at the dose rate of 0.027Gy/min. After irradiation, Ag-110 was observed in Ag-doped phosphate glass dosimeter; Ta-182 was observed in a tranparent ceramic. Applicability of these materials as neutron personal dosimeter will be investigated.

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