Dependence of the yields of AP sites and AP clusters produced in plasmid DNA on scavenging capacity and LET

椎名 卓也*; 渡辺 立子; 鈴木 雅雄*; 横谷 明徳

Journal of Radiation Research, 55(Suppl.1), p.i15 - i16, 2014/03

BB2013-0476.pdf:0.21MB

https://doi.org/10.1093/jrr/rrt160

The object of the study is to reveal whether the induction process of an abasic site (AP site) and the clustered DNA damage which contain AP sites (AP cluster) depends on ionizing density of radiation. In order to clarify the relation between track structure of C ions and the induction processes of an AP site or AP cluster, we measure the yield of AP using Nfo protein as an enzymatic probe, which converts an AP site to detectable single strand break. Several scavenging capacities in the samples are tested to estimate the effect of OH radicals. We compare those experimental yields with theoretical ones that are obtained from Monte Carlo track simulation to develop a model of AP site induction by irradiation. The yields of AP sites analyzed by the simulation are in good agreement with experimental ones. As a result, we newly determined the branching ratio of induction of the AP the site in the simulation.

Visualization of cell-cycle modification by ionizing irradiation in single HeLa cells using fluorescent ubiquitination-based cell-cycle indicator

神長 輝一; 坂本 由佳; 嘉成 由紀子; 野口 実穂; 横谷 明徳

Journal of Radiation Research, 55(Suppl.1), p.i127 - i128, 2014/03

https://doi.org/10.1093/jrr/rrt159

Fluorescent ubiquitination-based cell cycle indicator (FUCCI) HeLa cells are one of useful model cell lines to visualize a cell-cycle because their nuclei show different colors; orange indicating G1; green indicating S/G2. In order to establish a novel assay system to study cell-cycle modification by high LET irradiation such as ion beams for cancer therapy we have observed time-lapse images of HeLa-FUCCI cells irradiated as a preliminary experiment using conventional X-rays instead of high LET ion beams. The cell-cycle was strongly arrested by irradiation at S/G2 and never progressed to G1. In contrast, cells irradiated at G1 progress to S/G2 with a similar time course as non-irradiated control cells. These results show that single FUCCI cell exposure and live cell imaging are a powerful method to trace the single cell effect of high LET irradiation on the cell-cycle in future.

Radiation-quality-dependent bystander effects induced by the microbeams with different radiation sources

鈴木 雅雄*; Autsavapromporn, N.*; 宇佐美 徳子*; 舟山 知夫; Plante, I.*; 横田 裕一郎; 武藤 泰子*; 鈴木 芳代; 池田 裕子; 服部 佑哉; et al.

Journal of Radiation Research, 55(Suppl.1), P. i54, 2014/03

https://doi.org/10.1093/jrr/rrt219

It is essentially important for evaluating risk such a low-dose-rate exposure as the accident of Fukushima Daiichi Nuclear Power Plants to examine bystander effects induced by low-LET electromagnetic radiations, such as X or rays. We have been studying the cellular responses in normal human fibroblasts by targeted cell nucleus irradiations with monochromatic X-ray microbeams (5.35 keV) produced by Photon Factory in High Energy Accelerator Research Organization. The results indicated that the bystander effect in cell- killing effect was observed in the targeted cell nucleus irradiation, not in the random irradiation containing both cell nucleus and cytoplasm by Poisson distribution. The results suggest that energy deposition in cytoplasm is an important role of inducing bystander effects in case of low-LET radiations. We have also been investigating high-LET-radiation induced bystander effects using the heavy-ion microbeams at Takasaki Ion Accelerators for Advanced Radiation Application in Japan Atomic Energy Agency. Only 0.04% of the total numbers of normal human fibroblasts were irradiated with C-ion (220 MeV), Ne-ion (260 MeV) and Ar-ion (460 MeV) microbeams collimated at 20 micro meter in diameter. Cell-killing effect and gene mutation at HPRT locus in the cells irradiated with C ions were higher beyond our expectations and returned the estimated values that only 0.04% of the total cells were irradiated when using the specific inhibitor of gap junctions. On the other hand, no induced biological effects were observed in Ne and Ar ions whether the inhibitor was applied or not. The result suggested that the C-ion microbeam was capable of inducing bystander cellular effects via gap junction mediated cell-cell communication. There is clear evidence that bystander cellular effects are dependent on radiation quality.

Astronaut dose estimation using the PHITS code in combination with realistic models of the Kibo module and the MATROSHKA phantom

佐藤 達彦; 永松 愛子*; 武田 和雄*; 仁井田 浩二*; Puchalska, M.*; Sihver, L.*; Reitz, G.*

no journal, , 

宇宙飛行士の被ばく線量評価は、長期宇宙滞在を計画するうえで極めて重要となる。そのため宇宙航空研究開発機構(JAXA)は、欧州宇宙機構(ESA)などと協力して、国際宇宙ステーションきぼうモジュール内に人体ファントムMATROSHKAを設置し、その被ばく線量を計測した。本研究では、原子力機構が中心となって開発している粒子線輸送計算コードPHITSに、JAXAが開発したきぼうモジュールモデル及びChalmers工科大学が開発したMATROSHKAファントム数値モデル(NUNDO)を組合せ、測定結果のシミュレーションによる再現精度を検証した。その結果、臓器線量などの測定値と計算値は比較的よく一致することがわかり、PHITSが宇宙飛行士被ばく線量に適応できることを実証した。

Construction of plasmid containing clustered DNA damage to analyze its in vivo repair

高橋 桃子*; 鹿園 直哉

no journal, , 

We constructed lesion-containing plasmids using a novel approach in order to analyze repair of clustered DNA damage. Single stranded circular DNA (the 1st strand) was synthesized in vitro from DNA oligonucleotide as a primer. The template, which is single-stranded circular DNA containing uracil, was then digested with UDG and nucleases. From the resulting 1st strand, the 2nd strand was again synthesized in vitro from a complementary DNA oligonucleotide. As DNA lesions could be placed at any position in the DNA oligonucleotide on either strand, the method allows us to construct a clustered DNA damage site in a plasmid. We have confirmed that the double-stranded plasmid DNA could be constructed with DNA lesions. The constructed plasmid would be highly useful in analyzing in vivo processing of clustered DNA damage. The level of repair of lesions within clustered DNA damage in E. coli is discussed.