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Nuclear factor erythroid 2-related factor 2 (Nrf2), which regulates the expression of critical antioxidant proteins, was recently demonstrated to play a key role in cancer progression. Resistance to radiotherapy is a major obstacle in treating oral squamous cell carcinoma (OSCC). However, little is known about the association between Nrf2 and radioresistance in OSCC. Two OSCC cell lines (SAS and HSC-2) and their clinically relevant radioresistant (CRR) clones (SAS-R, HSC-2-R) were used. The effects of Nrf2 downregulation on radiosensitivity and the involvement of glycolysis in Nrf2-mediated radioresistance were evaluated. Immunohistochemistry of phosphorylated Nrf2 (p-Nrf2) was performed in 110 patients with OSCC who underwent preoperative chemoradiotherapy and surgery. Nrf2 was stably upregulated in CRR cells in vitro and in a mouse xenograft model. Moreover, elevated Nrf2 expression was associated with radioresistance. The enhancement of Nrf2-dependent glycolysis and glutathione synthesis was involved in the development of radioresistance. Additionally, p-Nrf2 expression was closely related to the pathological response to chemoradiotherapy, and its expression was predictive of prognosis in patients with advanced OSCC. Our results suggest that Nrf2 plays an important role in the radioresistance of OSCC accompanied by metabolic reprogramming. Targeting Nrf2 antioxidant pathway may represent a promising treatment strategy for highly malignant OSCC.

Radioresistant cancer cells lead to poor prognosis after radiotherapy. However, the mechanisms underlying cancer cell radioresistance have not been fully elucidated. Thus, the DNA damage response of clinically relevant radioresistant oral squamous cell carcinoma HSC2-R cells, established by long-term exposure of parental HSC2 cells to fractionated radiation, was investigated. The DNA double-strand break (DSB) repair protein-specific inhibitor, NU7441, which targets DNA-dependent protein kinase catalytic subunit (DNA-PKcs) phosphorylation, and IBR2, which targets Rad51, were administered to HSC2 and HSC2-R cells. NU7441 administration eliminated colony formation in both cell lines under 6 Gy X-ray irradiation, whereas IBR2 did not affect colony formation. NU7441 and IBR2 significantly enhanced 6 Gy X-ray irradiation-induced apoptosis in HSC2-R cells. In HSC2-R cells, cell cycle arrest released earlier than in HSC2 cells, and phosphorylated-H2A histone family member X (γH2AX) expression rapidly decreased. Following NU7441 administration, γH2AX expression and the cell percentages of the G2/M phase were not decreased at 48 h after treatment in HSC2-R cells. DNA-PKcs has been demonstrated to regulate non-homologous end-joining (NHEJ) and homologous recombination (HR) repair, and the later phase of DSB repair is dominated by HR. Therefore, the results of the present study indicated that the DSB repair mechanism in HSC2-R cells strongly depends on NHEJ and loss of HR repair function. The present study revealed a potential mechanism underlying the acquired radioresistance and therapeutic targets in radioresistant cancer cells.

Rapid analysis of 90Sr in bone and tooth samples of cattle was achieved by inductively coupled plasma mass spectrometry (ICP-MS) coupled with mass-shift and solid-phase extraction techniques. The mass-shift technique with the O2 reaction mode of ICP-MS was applied to the mass shift of isobaric element 90Zr, effectively removing its interference. The solid-phase extraction technique using a Sr resin, which is a polymeric resin supported by 4,4′(5′)-bis-t-butylcyclohexano-18-crown-6, quantitatively separated 90Sr from the interfering matrix elements and isobars in the bone and tooth samples. The activity concentrations determined by ICP-MS were comparable with those determined by the conventional radiometric method. However, the limit of detection in the ICP-MS measurement of 0.1 g samples was lower than that in the radioactivity measurement. The analytical time of the ICP-MS method was shortened from 20 days to 11 h, compared with the radiometric method. Therefore, the ICP-MS method can be a rapid and useful procedure for determination of 90Sr in small bone and tooth samples taken from terrestrial animals.

To better understand the cancer risk posed by radiation and the development of radiation therapy resistant cancer cells, we investigated the involvement of the cancer risk factor, APOBEC3B, in the generation of radiation-induced mutations. Expression of APOBEC3B in response to irradiation was determined in three human cancer cell lines by real-time quantitative PCR. Using the hypoxanthine-guanine phosphoribosyl transferase (HPRT) mutation assay, mutations in the HPRT gene caused by irradiation were compared between APOBEC3B-deficient human hepatocellular carcinoma (HepG2) cells [APOBEC3B knocked out (KO) using CRISPR-Cas9 genome editing] and the parent cell line. Then, HPRT-mutated cells were individually cultured to perform PCR and DNA sequencing of HPRT exons. X-Irradiation induced APOBEC3B expression in HepG2, human cervical cancer epithelial carcinoma (HeLa) and human oral squamous cell carcinoma (SAS) cells. Forced expression of APOBEC3B increased spontaneous mutations. By contrast, APOBEC3B KO not only decreased the spontaneous mutation rate, but also strongly suppressed the increase in mutation frequency after irradiation in the parent cell line. Although forced expression of APOBEC3B in the nucleus caused DNA damage, higher levels of APOBEC3B tended to reduce APOBEC3B-induced γ-H2AX foci formation (a measure of DNA damage repair). Further, the number of γ-H2AX foci in cells stably expressing APOBEC3B was not much higher than that in controls before and after irradiation, suggesting that a DNA repair pathway may be activated. This study demonstrates that irradiation induces sustained expression of APOBEC3B in HepG2, HeLa and SAS cells, and that APOBEC3B enhances radiation-induced partial deletions.

Seasonally, bred wild mice provide a unique bioresource, with high genetic diversity that differs from wild-derived mice and laboratory mice. This study aimed to establish an alternative superovulation method using wild large Japanese field mice (Apodemus speciosus) as the model species. Specifically, we investigated how the application of inhibin antiserum and equine chorionic gonadotropin (IASe) during both the reproductive and non-reproductive seasons impact the ovulation rate and competence of embryo development after in vitro fertilization (IVF) with fresh and cryopreserved sperm. When the wild mice were superovulated by injecting eCG followed by human chorionic gonadotropin (hCG), few oocytes were collected during the reproductive and non-reproductive seasons. In comparison, the number of ovulated oocytes was dramatically enhanced by the administration of IASe, followed by isolation of ovulated oocytes 24 hr after 30 IU hCG administration. The IVF oocytes that were in vitro cultured (IVC) with medium containing serum further developed to the 2- and/or 4-cell stage using both fresh and frozen-thawed sperm. In conclusion, we successfully established an alternative protocol for collecting ovulated oocytes from wild large Japanese field mice by administering IASe and hCG during both the reproductive and non-reproductive seasons. This study is the first to develop IVF-IVC wild large Japanese field mice beyond the 2- and/or 4-cell stage in vitro using fresh and cryopreserved sperm. This approach could be used in other species of wild or endangered mice to reduce the number of animals used for experiments, or in maintaining stocks of germ cells or embryos.

MicroRNA (miRNA) is a non-coding RNA involved in regulating both cancer gene promotion and suppression. We investigated the role of miRNA in inducing radiation resistance in cancer cell lines using clinically relevant radioresistant (CRR) cells. Analysis using miRNA arrays and qPCR revealed that miR-7-5p is highly expressed in all examined CRR cells. Additionally, CRR cells lose their radioresistance when daily irradiation is interrupted for over 6 months. MiR-7-5p expression is reduced in these cells, and treating CRR cells with a miR-7-5p inhibitor leads to a loss of resistance to irradiation. Conversely, overexpression of miR-7-5p in CRR cells using a miR-7-5p mimic shows further resistance to radiation. Overexpression of miR-7-5p in parent cells also results in resistance to radiation. These results indicate that miR-7-5p may control radioresistance in various cancer cells at the clinically relevant dose of irradiation. Furthermore, miR-7-5p downregulates mitoferrin and reduces Fe2+, which influences ferroptosis. Our findings have great potential not only for examining radiation resistance prior to treatment but also for providing new therapeutic agents for treatment-resistant cancers.

4‐Hydroxynonenal (HNE) is an important product of plasma membrane lipid peroxidation, which is a cause of cell and tissue injury. Mitochondrial DNA (mtDNA)‐depleted ρ0 cells were established using human cervical cancer and oral squamous cell carcinoma cell lines. We investigated the effect of reactive oxygen species in ρ0 cells, especially the mechanism of hydrogen peroxide (H2O2)‐mediated cell death. These cell were subjected to high oxidative stress and, compared with their parental cells, showed greater sensitivity to H2O2 and high lipid peroxidation. Upregulation of HNE in the plasma membrane was observed prior to the increase in intracellular H2O2. The amount of oxidized lipid present changed H2O2 permeability and administration of oxidized lipid led to further cell death after treatment with H2O2. Expression levels of lipoxygenase ALOX genes (ie ALOX5, ALOX12, and ALOX15) were upregulated in ρ0 cells, as were expression levels of ALOX12 and ALOX15 proteins. ALOX5 protein was mainly distributed in the nucleus, while ALOX12 and ALOX15 proteins were distributed in the nucleus and the cytoplasm. Although expression of COX2 gene was upregulated, its protein expression did not increase. ALOX (especially ALOX15) may be involved in the sensitivity of cancer cells to treatment. These data offer promise for the development of novel anticancer agents by altering the oxidation state of the plasma membrane. Our results showed that lipid peroxidation status is important for H2O2 sensitivity and that ALOX15 is involved in lipid peroxidation status.

Since the Fukushima Daiichi Nuclear Power Plant (FDNPP) accident, we have established an archive system of livestock and wild animals from the surrounding ex-evacuation zone. Wildlife within the alert zone have been exposed to low-dose-rate (LDR) radiation for a long continuous time. In this study, we analysed the morphological characteristics of the testes and in vitro fertilization (IVF) capacity of cryopreserved sperm of racoons from the ex-evacuation zone of the FDNPP accident. The radioactivity of caesium-137 (137 Cs) was measured by gamma-ray spectrometry, and the measured radioactivity concentration was 300-6,630 Bq/kg in the Fukushima raccoons. Notably, normal spermatogenesis was observed in the seminiferous tubules of the testes, with the germinal epithelium composed of a spermatogenic cell lineage with no evident ultrastructural alterations; freeze-thawing sperm penetration ability was confirmed using the interspecific zona pellucida-free mouse oocytes IVF assays. This study revealed that the chronic and LDR radiation exposure associated with the FDNPP accident had no adverse effect on the reproductive characteristics and functions of male raccoons.

Insulin-like growth factor (IGF) binding protein-3 (IGFBP-3) modulates various cell functions through IGF-dependent or independent mechanisms. However, its biological roles in the radiosensitivity of oral squamous cell carcinoma (OSCC) remain largely unknown. The purpose of this study was to determine the clinical significance and molecular mechanisms of the association between IGFBP-3 and OSCC radiosensitivity. We performed an immunohistochemical analysis of IGFBP-3 in 52 OSCC specimens from patients treated with preoperative chemoradiotherapy and surgery (phase II study). Associations between IGFBP-3 expression and clinicopathological features were also evaluated. In addition, we examined the effects of IGFBP-3 on post-X-ray irradiation radiosensitivity and DNA damage in vitro. High IGFBP-3 expression was significantly correlated with poor chemoradiotherapy responses and prognosis. With IGFBP-3 knockdown, irradiated OSCC cells exhibited significantly higher radiosensitivity compared with that of control cells. Moreover, IGFBP-3 depletion in OSCC cells reduced phosphorylation of the DNA-dependent protein kinase catalytic subunit (DNA-PKcs), which is required for DNA double-strand break repair during non-homologous end joining. These findings indicate that IGFBP-3 may have a significant role in regulating DNA repair and is be a potential biomarker for predicting clinical response to radiotherapy and prognosis in OSCC.

Cellular radioresistance is one of the major obstacles to the effectiveness of cancer radiotherapy. In an attempt to elucidate the implication of HIF-1α and miR-17-92 expressions in refractory radioresistant cells and also in order to study the potential applications of these molecules as novel therapeutic modalities to overcome radioresistant cancers, the current study was conducted. Clinically relevant radioresistant (CRR) cells from human cancer cell lines were established by exposing to long-term fractionated radiation of X-rays. Correspondingly, microarray analysis and real time RT-PCR were performed to find miRNA involved in the CRR phenotype. HIF-1α was down-regulated and miR17-92 cluster was overexpressed in CRR cells by transfection. The expression of miR 17-3p was inhibited by specific inhibitors and miR 19a was enforced by mimics, respectively in parental cells. Overexpression of HIF-1α in parental cells or down regulation of HIF-1α in CRR cells were not involved in radioresistance. However, when HIF-1α was genetically modified to constitutively express under normoxia condition, it was rendered for protection to cells. Exogenous overexpression of miR 17-92 cluster in CRR cells resulted in abolition of HIF-1α expression and restored sensitizations to ionizing radiation. Attenuated expression of miR-17-3p in parental cells protected them from irradiation. Overall, fine-tune deregulation of miR 17-92 cluster in CRR cells might account for the accumulation of HIF-1α in the CRR cells following exposure to irradiation. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s10616-019-00364-9) contains supplementary material, which is available to authorized users.

Androgens stimulate the proliferation of epithelial cells in the prostate by activating topoisomerase 2 (TOP2) and regulating the transcription of target genes. TOP2 resolves the entanglement of genomic DNA by transiently generating double‐strand breaks (DSBs), where TOP2 homodimers covalently bind to 5′ DSB ends, called TOP2‐DNA cleavage complexes (TOP2ccs). When TOP2 fails to rejoin TOP2ccs generating stalled TOP2ccs, tyrosyl DNA phosphodiesterase‐2 (TDP2) removes 5′ TOP2 adducts from stalled TOP2ccs prior to the ligation of the DSBs by nonhomologous end joining (NHEJ), the dominant DSB repair pathway in G0/G1 phases. We previously showed that estrogens frequently generate stalled TOP2ccs in G0/G1 phases. Here, we show that physiological concentrations of androgens induce several DSBs in individual human prostate cancer cells during G1 phase, and loss of TDP2 causes a five times higher number of androgen‐induced chromosome breaks in mitotic chromosome spreads. Intraperitoneally injected androgens induce several DSBs in individual epithelial cells of the prostate in TDP2‐deficient mice, even at 20 hr postinjection. In conclusion, physiological concentrations of androgens have very strong genotoxicity, most likely by generating stalled TOP2ccs., This work was conducted through the Joint Research Program of the Radiation Biology Center, Kyoto University and supported by the following grants: a Grant‐in‐Aid from the Ministry of Education, Science, Sport and Culture to S.T. (KAKENHI 25650006, 23221005 and 16H06306) and H.S. (KAKENHI 16H02953, 18H04900 and 19H04267), the Takeda Research Foundation, and Mitsubishi Foundation (to H.S.) and JSPS Core‐to‐Core Program, A. Advanced Research Networks (to S.T.), and Andalusian Government grants SAF2010‐21017, SAF2013‐47343‐P, SAF2014‐55532‐R, CVI‐7948 and FEDER funds (to F.C.‐L.).

We present data about mitochondrial DNA (mtDNA) copy number and aquaporin (AQP) gene expression in clinically radioresistant (CRR), ρ0, and their parental cells from human cervical cancer and human tongue squamous cell carcinoma. In both ρ0 and CRR cells, the mtDNA copy number was lower than for the parental strain. In addition, the obtained data suggest an association between the gene expression levels of AQP (1, 3, 8, and 9) and the difference in hydrogen peroxide (H2O2) sensitivity between ρ0 and CRR cells. Here, the composition of cell culture medium differs between CRR and ρ0 cells. To compare the gene expression of AQPs between ρ0 and CRR cells, therefore, we showed the data as the ratio to that in their parental cells. Keywords: Mitochondria, Aquaporin, Hydrogen peroxide, ρ0 cells, Clinically relevant radioresistant cells

Excessive intake of a high-calorie diet has been implicated in the development of non-alcoholic fatty liver disease (NAFLD). Several studies have investigated the effect of NAFLD on liver regeneration, but the effects of simple steatosis have been found to be inconsistent. We aimed to assess whether the initial phase of diet-induced lipid accumulation, induced by a diet containing moderate levels of fat, impairs liver regeneration after partial hepatectomy (PHx) of mice.Male ddY mice are prone to obesity, even when fed a relatively low-fat diet (FD). A model of early simple steatosis was created by feeding a 10% FD for 6 weeks to male ddY mice. Liver regeneration rate, DNA synthesis in hepatocytes, and DNA damage were then assessed.FD-fed mice had a slightly higher body mass (44.5 ± 2.6 grams vs. 48.1 ± 3.6 grams, P 0.05), but not liver mass (2.55 ± 0.37 grams vs. 2.69 ± 0.26 grams). Lipid droplets appeared in FD-fed mouse hepatocytes and Oil Red O staining was five times as intense as in control mice. In FD-fed mice, liver regeneration rate after two-thirds PHx was lower and impaired DNA replication was also observed. FD induced the expression of the cyclin-dependent kinase inhibitor p27KIP, but not p21CIP. Moreover, greater histone 2A.X phosphorylation was observed, indicating that FD caused DNA damage.Even short-term feeding of a moderately high FD to male ddY mice results in lipid accumulation in hepatocytes, DNA damage, and greater expression of p27KIP, implying lower DNA synthesis.

90Sr specific activity in the teeth of young cattle that were abandoned in Kawauchi village and Okuma town located in the former evacuation areas of the Fukushima-Daiichi Nuclear Power Plant (FNPP) accident were measured. Additionally, specific activity in contaminated surface soils sampled from the same area was measured. (1) All cattle teeth examined were contaminated with 90Sr. The specific activity, however, varied depending on the developmental stage of the teeth during the FNPP accident; teeth that had started development before the accident exhibited comparatively lower values, while teeth developed mainly after the accident showed higher values. (2) Values of 90Sr-specific activity in teeth formed after the FNPP accident were higher than those of the bulk soil but similar to those in the exchangeable fraction (water and CH3COONH4 soluble fractions) of the soil. The findings suggest that 90Sr was incorporated into the teeth during the process of development, and that 90Sr in the soluble and/or leachable fractions of the soil might migrate into teeth and contribute to the amount of 90Sr in the teeth. Thus, the concentration of 90Sr in teeth formed after the FNPP accident might reflect the extent of 90Sr pollution in the environment.

Background After the accident at the Fukushima Daiichi Nuclear Power Plant, radioactive contaminants were released over a widespread area. Monitoring the biological effects of radiation exposure in animals in the ex-evacuation zone should be continued to understand the health effects of radiation exposure in humans. The present study aimed to clarify the effects of radiation by investigating whether there is any alteration in the morphology and gene expressions of immune molecules in the intestine of pigs and inobuta (wild boar and domestic pig hybrid) in the ex-evacuation zone in 2012. Gene expression analysis was performed in small intestine samples from pigs, which were collected from January to February 2012, in the ex-evacuation zone. Pigs lived freely in this zone, and their small intestine was considered to be affected by the dietary intake of radioactive contaminants. Results Several genes were selected by microarray analysis for further investigation using real-time polymerase chain reaction. IFN-γ, which is an important inflammatory cytokine, and TLR3, which is a pattern recognize receptor for innate immune system genes, were highly elevated in these pigs. The expressions of the genes of these proteins were associated with the radiation level in the muscles. We also examined the alteration of gene expressions in wild boars 5 years after the disaster. The expression of IFN-γ and TLR3 remained high, and that of Cyclin G1, which is important in the cell cycle, was elevated. Conclusions We demonstrated that some changes in gene expression occurred in the small intestine of animals in the ex-evacuation zone after radiation. It is difficult to conclude that these alterations are caused by only artificial radionuclides from the Fukushima Daiichi Nuclear Power Plant. However, the animals in the ex-evacuation zone might have experienced some changes owing to radioactive materials, including contaminated soil, small animals, and insects. We need to continue monitoring the effects of long-term radiation exposure in living things.

Radiotherapy (RT) is one of the major modalities for the treatment of human cancers and has been established as an excellent local treatment for malignant tumors. Conventional fractionated RT consists of 2-Gy X-rays, fractionated once a day, 5 days a week for 5-7 weeks in total 60 Gy. The efficacy of RT depends on the existence of radioresistant cells, which remains one of the most critical obstacles in RT and radio-chemotherapy. To improve the efficacy of RT, understanding the characteristics of radioresistant cells is one of the important subjects in radiation biology. Several studies have been reported to find out molecules implicated in radioresistance. However, it is noteworthy that cellular radioresistance has been mainly studied among cells with different genetic backgrounds and different origins. Therefore, making a system to compare between radioresistant and sensitive cells with the isogenic background is required. In this review, some aspects of cellular radioresistance mainly focusing on clinically relevant radioresistant (CRR) cell lines that can continue to proliferate even under exposure to 2-Gy X-rays, once a day, for more than 30 days, which is consistent with the conventional fractionated RT are discussed.

Here, we present an update on the next level of experiments studying the impact of the gamma radiation environment, created post-March, 2011 nuclear accident at Fukushima Daiichi nuclear power plant, on rice plant and its next generation-the seed. Japonica-type rice (Oryza sativa L. cv. Koshihikari) plant was exposed to low-level gamma radiation (~4 μSv/h) in the contaminated Iitate Farm field in Iitate village (Fukushima). Seeds were harvested from these plants at maturity, and serve as the treated group. For control group, seeds (cv. Koshihikari) were harvested from rice grown in clean soil in Soma city, adjacent to Iitate village, in Fukushima. Focusing on the multi-omics approach, we have investigated the dry mature rice seed transcriptome, proteome, and metabolome following cultivation of rice in the radionuclide contaminated soil and compared it with the control group seed (non-radioactive field-soil environment). This update article presents an overview of both the multi-omics approach/technologies and the first findings on how rice seed has changed or adapted its biology to the low-level radioactive environment.

To clarify the relationship between mitochondrial DNA (mtDNA)-depleted ρ0 cells and the cellular sensitivity to hydrogen peroxide (H2O2), we established HeLa and SAS ρ0 cell lines and investigated their survival rate in H2O2, radical scavenging enzymes, plasma membrane potential status, and chronological change in intracellular H2O2 amount under the existence of extracellular hydrogen peroxide compared with the parental cells. The results revealed that ρ0 cells had higher sensitivity to H2O2 than their parental cells, even though the catalase activity of ρ0 cells was up-regulated, and the membrane potential of the ρ0 cells was lower than their parental cells. Furthermore, the internal H2O2 amount significantly increased only in ρ0 cells after 50 μM H2O2 treatment for 1 h. These results suggest that plasma membrane status of ρ0 cells may cause degradation, and the change could lead to enhanced membrane permeability to H2O2. As a consequence, ρ0 cells have a higher H2O2 sensitivity than the parental cells.

The Fukushima Daiichi Nuclear Power Plant (FNPP) accident, the largest nuclear incident since the 1986 Chernobyl disaster, occurred when the plant was hit by a tsunami triggered by the Great East Japan Earthquake on March 11, 2011. The subsequent uncontrolled release of radioactive substances resulted in massive evacuations in a 20-km zone. To better understand the biological consequences of the FNPP accident, we have been measuring DNA damage levels in cattle in the evacuation zone. DNA damage was evaluated by assessing the levels of DNA double-strand breaks in peripheral blood lymphocytes by immunocyto-fluorescence-based quantification of γ-H2AX foci. A greater than two-fold increase in the fraction of damaged lymphocytes was observed in all animal cohorts within the evacuation zone, and the levels of DNA damage decreased slightly over the 700-day sample collection period. While the extent of damage appeared to be independent of the distance from the accident site and the estimated radiation dose from radiocesium, we observed age-dependent accumulation of DNA damage. Thus, this study, which was the first to evaluate the biological impact of the FNPP accident utilizing the γ-H2AX assays, indicated the causal relation between high levels of DNA damage in animals living in the evacuation zone and the FNPP accident.

To elucidate the molecular mechanisms underlying cellular radioresistance, clinically relevant radioresistant cell lines were established via long-term exposure to X-rays with stepwise dose escalation. Established cells continue to proliferate despite exposure to 2 Gy X-rays/day for more than 30 days, a standard protocol in cancer radiotherapy. DNA repair fidelity in radioresistant and the parental cells by evaluating the mutation frequency at the hypoxanthine phosphoribosyltransferase (HPRT) locus after exposure to X-rays was determined. Mutation spectrum at the HPRT locus was examined by multiplex polymerase chain reaction. Rejoining kinetics of X-ray-induced DNA double strand breaks (dsbs) was evaluated by the detection of phosphorylated histone H2AX (γH2AX) after X-irradiation. The fold increase in the HPRT mutation frequency due to acute radiation was similar between radioresistant and the parental cell lines. However, fractionated radiation (FR) consisting of 2 Gy X-rays/day increased the mutation frequency at the HPRT locus in parental but not in radioresistant cells. Analysis of the FR-induced mutations at the HPRT locus revealed a high frequency of deletion mutations (>70%) in parental but not in radioresistant cells. As assessed by γH2AX immunostaining, DNA dsbs induced by acute exposure to 10 Gy of X-rays were repaired to the control level within 7 days in radioresistant but not in the parental cells. Moreover, 2 Gy × 5 FR increased the number of γH2AX-positive cells in parental cultures but not in radioresistant cultures. DNA dsbs induced by 2 Gy/day FR are repaired with fidelity in radioresistant but not in parental cells.

In order to understand the effect of chronic exposure to low-dose-rate radiation on female fertilities and the risk of heritable genetic effects on domestic animals after the Fukushima Daiichi Nuclear Power Plant (FNPP) accident, we assessed the developmental ability of oocytes, and examined the histological characters and structure of ovaries. We showed that potentially viable oocytes could be collected from the ovary of abandoned cattle in the ex-evacuation zone set within a 20-km radius from FNPP, resulting in production of the morphologically normal calves following in vitro culture and embryo transfer. The proliferation of cattle ovarian granulosa cells was confirmed by expression of Ki-67. Apoptosis of oocytes and granulosa cells was few determined by the TUNEL assay. In addition, porcine and inobuta oocytes had the abilities of in vitro maturation. These results suggest that chronic radiation exposure associated with the FNPP accident may have little effect, if any, on the female fertilities of domestic animals.

Substantial amounts of radionuclides including strontium-90 (90Sr) were released by the Fukushima Daiichi Nuclear Power Plant (FNPP) accident. In the present study, we describe and discuss the presence of 90Sr in the ex-evacuation zone of the FNPP accident and its relationship with 90Sr activity concentration in the hard tissue of animals. We found that the activity concentration of 90Sr in the hard tissue exhibited a positive correlation with 90Sr pollution in their corresponding terrestrial and marine environments. Hard tissues, such as the teeth, bones, and otoliths, of animals and fishes could serve as useful tools in assessing 90Sr pollution in the environment during the period of the formation of those tissues.

It is extremely important to understand the mechanisms underlying the effect of radiation on animals including humans. Radiation potentially induces alterations of the immune system, which may cause serious diseases. The Fukushima Daiichi Nuclear Power Plant (FNPP) accident occurred in 2011, in which radioactive contaminants were released over a wide area. After farmers evacuated from the ex-evacuation zone set within 20-km radius from FNPP, pigs were left behind and unleashed in the zone (abandoned pigs). Their small intestine was considered to have been affected by the dietary intake of radioactive contaminants. The present study thus aimed to clarify the effect of radiation by investigating whether there is any alteration in the expression of genes encoding immune-related molecules and in morphology of the intestine of abandoned pigs. Microarray analysis revealed changes of the expression of various genes involved in inflammation and oxidative stress including IFN-γ, which is an important inflammatory cytokine, was highly elevated in abandoned pigs, and its expression level was correlated with radioactive cesium concentration in skeletal muscle. On the other hand, there were no morphological changes in the intestine. At the sampling stage of this study, only 1 year passed since the FNPP accident occurred, it would be premature to conclude that the identified alteration of gene expression was caused only by artificial radionuclides attributed to the FNPP accident. We need to continue monitoring the effect of long-term radiation exposure in living organisms in the ex-evacuation zone.

The Fukushima nuclear power plant (FNPP) accident raised worldwide attention to the health risk of radiation exposure and to its potential transgenerational effects. Here, we analysed transgenerational effects on calf spermatogenesis and on blood plasma metabolome in order to detect alterations associated with paternal exposure to low-dose-rate (LDR) radiation. Sperm was collected from a bull exposed to radiation for 2 years abandoned in the ex-evacuation zone of the FNPP accident (the abandoned bull) and was used for artificial insemination (AI) into a non-radiocontaminated cow. Haematoxylin and eosin stained sections of the testis of a 13-month-old calf revealed spermatogonia, spermatocytes, spermatids, and sperm in normal morphology. Nuclear and acrosomal morphology of sperm was generally normal. Metabolomic profiling of plasma using capillary electrophoresis–mass spectrometry resulted in 104 peaks of candidate compounds suggestive of paternal exposure. A calf was delivered by AI using sperm from the abandoned bull. Regarding glycolysis, the contents of nucleotide sugars tended to be lower in the delivered calf than in the control calf. Among energy carries, AMP and ATP showed different tendency between non-radiocontaminated and delivered calf. In conclusion, there were no apparent transgenerational effects on both spermatogenesis and blood plasma metabolome in a calf obtained by AI using sperm from the abandoned bull exposed to LDR in the ex-evacuation zone of the FNPP accident for about 2 years.