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To study the environmental radiation effects of wild animals after the Fukushima Dai-ichi nuclear power plant accident, we assessed effects on hematopoietic progenitor cells (HPCs) in large Japanese field mice (A. speciosus were collected from three contaminated sites and control area. The air dose-rates at the control and contaminated areas were 0.96 ± 0.05 μGy/d (Hirosaki), 14.4 ± 2.4 μGy/d (Tanashio), 208.8 ± 31.2 μGy/d (Ide), 470.4 ± 93.6 μGy/d (Omaru), respectively. We investigated possible DNA damage and pro-inflammatory markers in the bone marrow (BM) cells. The colony-forming potential of BM cells was estimated by the number of HPC colony-forming cells. Radiation-induced genomic instability (RIGI) in HPCs was also analyzed by quantifying delayed DNA damage in CFU-GM clones.Although no significant differences in DNA damage and inflammation markers in BM cells from control and contaminated areas, the number of HPC colonies exhibited an inverse correlation with air dose-rate. With regard to RIGI, no significant differences in DNA damage of CFU-GM clones between the mice from the control and the three contaminated areas.Our study suggests that low dose-rate radiation of more than 200 Gy/d reduced HPCs, possibly eliminating genomically unstable HPCs.

ADAR1 is involved in adenosine-to-inosine RNA editing. The cytoplasmic ADAR1p150 edits 3’UTR double-stranded RNAs and thereby suppresses induction of interferons. Loss of this ADAR1p150 function underlies the embryonic lethality of Adar1 null mice, pathogenesis of the severe autoimmune disease Aicardi-Goutières syndrome, and the resistance developed in cancers to immune checkpoint blockade. In contrast, the biological functions of the nuclear-localized ADAR1p110 remain largely unknown. Here, we report that ADAR1p110 regulates R-loop formation and genome stability at telomeres in cancer cells carrying non-canonical variants of telomeric repeats. ADAR1p110 edits the A-C mismatches within RNA:DNA hybrids formed between canonical and non-canonical variant repeats. Editing of A-C mismatches to I:C matched pairs facilitates resolution of telomeric R-loops by RNase H2. This ADAR1p110-dependent control of telomeric R-loops is required for continued proliferation of telomerase-reactivated cancer cells, revealing the pro-oncogenic nature of ADAR1p110 and identifying ADAR1 as a promising therapeutic target of telomerase positive cancers., One type of RNA editing involves ADAR-mediated conversion of adenosine to inosine. Here the authors show that ADAR1 nuclear isoform p110 regulates R loop formation and genome stability at telomeres in cancer cells.

Several past studies using a mouse model of radiation-induced AML (rAML) have shown that hemizygous deletion of the Sfpi1 gene (HDSG) is an initiating event for the development of rAML. In this study, we examined the difference in frequency of HDSG in hematopoietic stem cells (HSCs) Rich hematopoietic Cell population (HRCs) from bone marrow (BM) and spleen of C3H mice irradiated with 3 Gy X-rays.8-weeks old male C3H mice were irradiated 3Gy of whole body X-ray (1 Gy/min) and mice were sacrificed at 1, 4, 8, and 26 weeks. Then, HSPCs were isolated from BM of femur and spleen, the frequency of HRCs with Sfpi1 gene deletion was analyzed by fluorescence in situ hybridization (FISH).The frequency of HRCs with HDSG in both BM and spleen was increased 1 week after X-irradiation. Then, the frequency of HRCs with HDSG in BM showed a gradual decrease from 4 to 26 weeks, whereas HRCs with HDSG in spleen remained high, even at 26 weeks after X-irradiation. HDSG is less likely to be eliminated, particularly in the spleen, after X-irradiation. The spleen as well as BM of the femur may be major sites of rAML development.

After the Fukushima Daiichi Nuclear Power Plant (FDNPP) accident in Japan on March 11 2011, the surroundings became contaminated with radionuclides. To understand the possible biological effects after chronic low dose-rate radiation in contaminated areas of Fukushima, we assessed the effects in large Japanese field mice (We collectedAlthough the mice in the contaminated areas were chronically exposed, there was no radiation-specific chromosome aberrations observed, such as dicentric chromosomes and rings. Some structural aberrations such as gaps and breaks were observed, and these frequencies decreased annually in mice from Namie Town.These findings suggest that chromosome aberration analysis is useful to evaluate and monitor radiation effects in wild animals.

Primary hepatocyte culture is a crucial tool for investigations of liver function and for evaluating the toxic effects of drugs. In addition, chromosomal analysis of hepatocytes could also prove useful for understanding the mechanisms of hepatocarcinogenesis. However, cultivation of primary hepatocytes for chromosome analysis has been hampered by the specific equipment and skill required to perform the in situ perfusion step necessary for isolation of primary hepatocytes. In the present study, we aimed to establish a simple and efficient method of isolating hepatocytes suitable for chromosome analysis. We performed hepatocyte isolation without using collagenase perfusion, instead digesting liver tissues using collagenase in tubes. In addition, we examined hepatocyte and bone marrow cell (BMC) co-culture and cultivation of hepatocytes with medium containing BMC culture medium supernatants. We found that hepatocyte viability and attachment rate were significantly improved, both by co-culture with BMCs and medium containing BMC culture media supernatants, with the latter also significantly increasing the mitotic index. Using this simple method of isolation and cultivation, we could successfully perform chromosomal analysis of mouse primary hepatocytes. This method has the potential to help understand the mechanisms underlying chromosomal instability-mediated hepatocarcinogenesis.

Exosome-like vesicles (ELV) are involved in mediating radiation-induced bystander effect (RIBE). Here, we used ELV from control cell conditioned medium (CCCM) and from 4 Gy of X-ray irradiated cell conditioned medium (ICCM), which has been used to culture normal human fibroblast cells to examine the possibility of ELV mediating RIBE signals. We investigated whether ELV from 4 Gy irradiated mouse serum mediate RIBE signals. Induction of DNA damage was observed in cells that were treated with ICCM ELV and ELV from 4 Gy irradiated mouse serum. In addition, we treated CCCM ELV and ICCM ELV with RNases, DNases, and proteinases to determine which component of ELV is responsible for RIBE. Induction of DNA damage by ICCM ELV was not observed after treatment with DNases. After treatment, DNA damages were not induced in CCCM ELV or ICCM ELV from mitochondria depleted (ρ0) normal human fibroblast cells. Further, we found significant increase in mitochondrial DNA (mtDNA) in ICCM ELV and ELV from 4 Gy irradiated mouse serum. ELV carrying amplified mtDNA (ND1, ND5) induced DNA damage in treated cells. These data suggest that the secretion of mtDNA through exosomes is involved in mediating RIBE signals.

Radiation-induced acute myeloid leukemia (rAML) in C3H mice is commonly developed through inactivation of PU.1 transcription factor encoded in Sfpi1 on chromosome 2. PU.1 inactivation involves two steps: hemizygous deletion of the Sfpi1 gene (DSG) and point mutation of the allele Sfpi1 gene (PMASG). In this study, we investigated the dose-rate dependence of the frequency of both DSG and PMASG in hematopoietic stem cells (HSCs) of C3H mice that received a total of 3 Gy gamma-ray exposure at dose rates of 20 mGy/day, 200 mGy/day or 1,000 mGy/min. All mice were followed for 250 days from start of irradiation. Fluorescent in situ hybridization of the Sfpi1 gene site indicated that frequency of HSCs with DSG was proportional to dose rate. In cell surface profiles, PU.1-inactivated HSCs by both DSG and PMASG were still positive for PU.1, but negative for GM-CSF receptor-α (GMCSFRα), which is transcriptionally regulated by PU.1. Immunofluorescent staining analysis of both PU.1 and GM-CSFRα also showed dose-rate-dependent levels of PU.1-inactivated HSCs. This study provides evidence that both DSG and PMASG are dose-rate dependent; these experimental data offer new insights into the dose-rate effects in HSCs that can lead to radiation-induced leukemogenesis.

Chromosome missegregation can lead to a change in chromosome number known as aneuploidy. Although aneuploidy is a known hallmark of cancer cells, the various mechanisms by which altered gene and/or DNA copy number facilitate tumorigenesis remain unclear. To understand the effect of aneuploidy occurring in non-tumorigenic human breast epithelial cells, we generated clones harboring artificial aneuploidy using microcell-mediated chromosome transfer. Our results demonstrate that clones with artificial aneuploidy of chromosome 8 or chromosome 22 both show inhibited proliferation and genomic instability. Also, the increased autophagy was observed in the artificially aneuploidy clones, and inhibition of autophagy resulted in increased genomic instability and DNA damage. In addition, the intracellular levels of reactive oxygen species were up-regulated in the artificially aneuploid clones, and inhibition of autophagy further increased the production of reactive oxygen species. Together, these results suggest that even a single extraneous chromosome can induce genomic instability, and that autophagy triggered by aneuploidy-induced stress is a mechanism to protect cells bearing abnormal chromosome number.

Werner syndrome (WS) is an autosomal recessive disease characterized by premature aging and caused by mutations of the WRN gene mapped at 8p12. To examine functional complementation of WS phenotypes, we introduced a normal human chromosome 8 into a strain of WS fibroblasts (WS3RGB) immortalized by expressing a human telomerase reverse transcriptase subunit (hTERT) gene. Here, we demonstrate that the abnormal WS phenotypes including cellular sensitivities to 4-nitroquinoline-1-oxide (4NQO) and hydroxy urea (HU), and chromosomal radiosensitivity at G2 phase are corrected by expression of the WRN gene mediated by introducing a chromosome 8. This indicates that those multiple abnormal WS phenotypes are derived from a primary, but not secondary, defect in the WRN gene., Journal of radiation research, 50(3), pp.253-259; 2009

Item does not contain fulltext Adenosine deaminases acting on RNA (ADARs) are involved in RNA editing that converts adenosine residues to inosine specifically in double-stranded RNAs. In this study, we investigated the interaction of the RNA editing mechanism with the RNA interference (RNAi) machinery and found that ADAR1 forms a complex with Dicer through direct protein-protein interaction. Most importantly, ADAR1 increases the maximum rate (Vmax) of pre-microRNA (miRNA) cleavage by Dicer and facilitates loading of miRNA onto RNA-induced silencing complexes, identifying a new role of ADAR1 in miRNA processing and RNAi mechanisms. ADAR1 differentiates its functions in RNA editing and RNAi by the formation of either ADAR1/ADAR1 homodimer or Dicer/ADAR1 heterodimer complexes, respectively. As expected, the expression of miRNAs is globally inhibited in ADAR1(-/-) mouse embryos, which, in turn, alters the expression of their target genes and might contribute to their embryonic lethal phenotype.