The role of irreparable DNA damage in initiating chromosomal mutagenesis in Chinese hamster cells irradiated with gamma and secondary proton radiation with an energy of 70 GeV

The study on the kinetics of DNA injury repair in Chinese hamster cells exposed to gamma and secondary proton radiation (70 GeV) has demonstrated the absence of distinctions in the kinetics of repair of rapidly repaired damages, a decreased rate of repair of slowly repaired lesions, and an increased residual irreparable damage induced by secondary radiation that reliably correlates with the RBE value estimated by the cytogenetic effect.     

Action of secondary radiation from 70-Gev protons on the cellular DNA of mammals

A study was made of the effect of secondary radiation of 70 GeV protons on DNA of Chinese hamster cells. With a reference to fibroblast DNA, lymphoid cell DNA, and the lethal effect of radiation on the survival of Chinese hamster cells the RBE was 1.6-7.6, 1.1-3.8 and 1.14-1.7, respectively. DNA breaks were repaired to an equal level after exposure to secondary radiation from the accelerator and gamma-radiation from 60Co in equally effective doses.     

Biological effect of 9 GeV protons and the radioprotective effect of ATP and AMP on epithelial cells of the mouse cornea

The cytogenetic and cytological effects induced in mouse cornea epithelium cells by 9 GeV protons and "standard" radiation have been studied. The RBE coefficients are different at different times of observation. ATP and AMP are shown to produce a protective effect on the frequency of formation of aberrant mitoses. DMF values for protons determined 24 and 72 h following irradiation are 1.8 +/- 0.2 and 1.7 +/- 0.2, respectively.     

The characteristics of the realization of cytogenetic damage in mammalian and plant cells exposed to low doses of radiation]

A complicated character of the cytogenetic injury dependence upon radiation dose was revealed after low-level gamma irradiation of Vicia faba seedlings and Chinese hamster fibroblasts. The dependence was linear with low-level secondary exposure to 70 GeV protons. The authors discuss a threshold nature of induction of the cytogenetic damage repair responsible for a high outcome of damages under the effect of low-level gamma radiation.     

Evaluation of the cataractogenic effect of 9 GeV protons

A study was made of the regularities of formation of lenticular opacity in mice exposed to 9 GeV protons and 60Co-gamma-rays. The RBE coefficients, calculated by the nonparametric method, were found to depend upon dose and time after irradiation. It was shown that after small radiation doses (0.25 to 0.50 Gy) the RBE coefficients increased from 1 to 8 with increasing period of observation. With higher doses (up to 5.0 Gy) the RBE coefficient increase in time was less pronounced.     

Effect of secondary radiation from 70-GeV protons and gamma quanta on chromosomes of Chinese hamster cells as dependent on the cell cycle stage

In cultured Chinese hamster cells, no decrease in the number of chromosome aberrations was noted after exposure thereof to 70 GeV protons at the late S-phase as opposed to early one. It is suggested that high biological effectiveness of this type of radiation is associated with its inhibiting effect of cytogenetic damages repair.     

The effect of low-dose rate radiation simulating the high-altitude flight conditions on mice in vivo

In present work, we investigated the peculiarities of the effect of a low-dose rate high-LET radiation that simulates the spectral and component composition of the radiation field formed in the atmosphere at a height of 10 km on mice in vivo. The dose dependence and adaptive response were examined. Irradiation of mice was performed for 24 h a day in the radiation field behind the concrete shield of the Serpukhov accelerator of 70 GeV protons for the time (15-31 days) necessary to accumulate the required doses. The experiments demonstrated that irradiation of mice in vivo in the dose range of 11.5-31.5 cGy leads to an increase in cytogenetic damage to bone marrow cells and induces no adaptive response in bone marrow cells.     

Effects of heavy ions and energetic protons on normal human fibroblasts

At the low particle fluences of radiation to which astronauts are exposed in space, "non-targeted" effects such as the bystander response may have increased significance. The radiation-induced bystander effect is the occurrence of biological responses in unirradiated cells near to or sharing medium with cells traversed by radiation. The objectives of this study were to establish the responses of AG01522 diploid human fibroblasts after exposure to several heavy ions and energetic protons, as compared to X-rays, and to obtain initial information on the bystander effect in terms of cell clonogenic survival after Fe ion irradiation. Using a clonogenic survival assay, relative biological effectiveness (RBE) values at 10% survival were 2.5, 2.3, 1.0 and 1.2 for 1 GeV/amu Fe, 1 GeV/amu Ti, 290 MeV/amu C and 1 GeV/amu protons, respectively, compared to 250 kVp X-rays. For induction of micronuclei (MN), compared to the low LET protons, Fe and Ti are very effective inducers of damage, although C ions are similar to protons. Using a transwell insert system in which irradiated and unirradiated bystander cells share medium but are not touching each other, it was found that clonogenic survival in unirradiated bystander cells was decreased when irradiated cells were exposed to Fe ions or X-rays. The magnitude of the decrease in bystander survival was similar with both radiation types, reaching a plateau of about 80% survival at doses of about 0.5 Gy or larger.     

DNA damage and repair in Chinese hamster fibroblasts exposed to gamma and secondary radiation generated by 70 GeV protons

The cytochemical study of DNA damage and repair in a Chinese hamster fibroblast culture exposed to gamma-rays and secondary radiation from 70 GeV protons showed no significant differences between the two types of radiation.     

Biological effectiveness of helium ions and relativistic-energy protons

It was shown that RBE coefficients of protons (9 GeV) and accelerated helium ions (4 GeV/nucleon) are within the range from 1.0 to 11.6 and 1.0 to 7.2, respectively, depending on the object under study, the criterium of estimation, the registration time, and the dose value.     

Nonparametric method of determining the RBE coefficients of accelerated charged particles by the incidence of neoplasms in rats

The nonparametric method was used to determine RBE coefficients of accelerated charged particles (helium ions of 4 GeV/nucleon and 645 MeV protons) by the incidence of tumors localized in different rat organs or by the absence of tumors. The nonparametric method permitted to find the dose dependence of the RBE coefficients and to make statistical analysis of the results obtained with due regard for come features of developing damages which were not revealed by conventional methods of determining RBE coefficients.     

Induction of 8-azaguanine resistant mutants in human cultured cells exposed to 31 MeV protons

We report results on the induction of 8-azaguanine (8-AG)-resistant mutants in cultured human cells (EUE) exposed to 31 MeV protons. The spontaneous frequency of mutants was 5.6 +/- 0.7 x 10(-6) per viable cell. Gamma rays were taken as reference radiation. Expression times giving the highest frequency of mutants after 31 MeV protons and gamma irradiation were found to be about 10 days for both radiations. The dose-response relationship for mutant induction by protons, as determined at the optimal expression time, was compared to that obtained after gamma rays. The relative biological effectiveness (RBE) is 2.4 +/- 0.5, this value being higher than the RBE value determined for cell survival.     

Is the risk for secondary cancers after proton therapy enhanced distal to the Planning Target Volume? A two-case report with possible explanations

It is often assumed that radiation-induced secondary cancer after proton therapy forms preferentially close to the distal fall-off of the spread-out Bragg peak because of an increased relative biological effectiveness (RBE) with regard to cancer induction of low-energy protons. In this study we analyze to what extent dose gradients distal to the Planning Target Volume (PTV) may, independently from the RBE, contribute to enhanced radiation carcinogenesis. The study is based on two dogs which, out of 30 dogs treated with proton therapy at the Paul Scherrer Institute (PSI), developed a secondary cancer. Both dogs were originally diagnosed and treated for a fibrosarcoma and developed an osteosarcoma 48 and almost 60 months, respectively, after radiotherapy. From the dose distributions of the initial radiotherapy for both dogs three-dimensional maps of secondary cancer complication probability (SCCP) were computed. The SCCP maps were analyzed in the regions where the dogs developed a secondary cancer. The SCCP maps showed an enhanced risk in the regions of the femur where the secondary cancers were detected, as compared to the SCCP of the total femur. Excess risk of radiation-induced cancer at the distal part of proton radiation fields can thus be explained using SCCP calculations on the basis of the physical dose distributions. Therefore, the occurrence of secondary cancer close to the distal dose gradients of proton therapy is not necessarily due to an increased RBE of low-energy protons. More extensive studies based on more patients will be necessary to further elucidate the factors influencing the development of secondary tumors.     

Shielding of relativistic protons

Protons are the most abundant element in the galactic cosmic radiation, and the energy spectrum peaks around 1 GeV. Shielding of relativistic protons is therefore a key problem in the radiation protection strategy of crewmembers involved in long-term missions in deep space. Hydrogen ions were accelerated up to 1 GeV at the NASA Space Radiation Laboratory, Brookhaven National Laboratory, New York. The proton beam was also shielded with thick (about 20 g/cm²) blocks of lucite (PMMA) or aluminium (Al). We found that the dose rate was increased 40–60% by the shielding and decreased as a function of the distance along the axis. Simulations using the General–Purpose Particle and Heavy-Ion Transport code System (PHITS) show that the dose increase is mostly caused by secondary protons emitted by the target. The modified radiation field after the shield has been characterized for its biological effectiveness by measuring chromosomal aberrations in human peripheral blood lymphocytes exposed just behind the shield block, or to the direct beam, in the dose range 0.5–3 Gy. Notwithstanding the increased dose per incident proton, the fraction of aberrant cells at the same dose in the sample position was not significantly modified by the shield. The PHITS code simulations show that, albeit secondary protons are slower than incident nuclei, the LET spectrum is still contained in the low-LET range (<10 keV/μm), which explains the approximately unitary value measured for the relative biological effectiveness.     

The possibility for modification of the cytogenetic action of secondary radiation with 70-GeV protons on Chinese hamster fibroblasts

As estimated by the cytogenetic injury induced in Chinese hamster cells by secondary 70-GeV proton radiation, phenylmethylsulfonyl fluoride, an inhibitor of chromatin proteinases, has a radioprotective effect. The poly(ADP)-ribosylation-independent participation of the inhibitor in radiation cytogenetic mutagenesis has been shown.     

Using electron beam radiation to simulate the dose distribution for whole body solar particle event proton exposure

As a part of the near solar system exploration program, astronauts may receive significant total body proton radiation exposures during a solar particle event (SPE). In the Center for Acute Radiation Research (CARR), symptoms of the acute radiation sickness syndrome induced by conventional radiation are being compared to those induced by SPE-like proton radiation, to determine the relative biological effectiveness (RBE) of SPE protons. In an SPE, the astronaut’s whole body will be exposed to radiation consisting mainly of protons with energies below 50 MeV. In addition to providing for a potentially higher RBE than conventional radiation, the energy distribution for an SPE will produce a relatively inhomogeneous total body dose distribution, with a significantly higher dose delivered to the skin and subcutaneous tissues than to the internal organs. These factors make it difficult to use a ⁶⁰Co standard for RBE comparisons in our experiments. Here, the novel concept of using megavoltage electron beam radiation to more accurately reproduce both the total dose and the dose distribution of SPE protons and make meaningful RBE comparisons between protons and conventional radiation is described. In these studies, Monte Carlo simulation was used to determine the dose distribution of electron beam radiation in small mammals such as mice and ferrets as well as large mammals such as pigs. These studies will help to better define the topography of the time-dose-fractionation versus biological response landscape for astronaut exposure to an SPE.     

Effects of very low fluences of high-energy protons or iron ions on irradiated and bystander cells

In space, astronauts are exposed to radiation fields consisting of energetic protons and high atomic number, high-energy (HZE) particles at very low dose rates or fluences. Under these conditions, it is likely that, in addition to cells in an astronaut's body being traversed by ionizing radiation particles, unirradiated cells can also receive intercellular bystander signals from irradiated cells. Thus this study was designed to determine the dependence of DNA damage induction on dose at very low fluences of charged particles. Novel techniques to quantify particle fluence have been developed at the NASA Space Radiation Biology Laboratory (NSRL) at Brookhaven National Laboratory (BNL). The approach uses a large ionization chamber to visualize the radiation beam coupled with a scintillation counter to measure fluence. This development has allowed us to irradiate cells with 1 GeV/nucleon protons and iron ions at particle fluences as low as 200 particles/cm(2) and quantify biological responses. Our results show an increased fraction of cells with DNA damage in both the irradiated population and bystander cells sharing medium with irradiated cells after low fluences. The fraction of cells with damage, manifest as micronucleus formation and 53BP1 focus induction, is about 2-fold higher than background at doses as low as ～0.47 mGy iron ions (～0.02 iron ions/cell) or ～70 μGy protons (～2 protons/cell). In the irradiated population, irrespective of radiation type, the fraction of damaged cells is constant from the lowest damaging fluence to about 1 cGy, above which the fraction of damaged cells increases with dose. In the bystander population, the level of damage is the same as in the irradiated population up to 1 cGy, but it does not increase above that plateau level with increasing dose. The data suggest that at fluences of high-energy protons or iron ions less than about 5 cGy, the response in irradiated cell populations may be dominated by the bystander response.     

Simulation of DNA damage after proton irradiation

The biophysical radiation track simulation model PARTRAC was improved by implementing new interaction cross sections for protons in water. Computer-simulated tracks of energy deposition events from protons and their secondary electrons were superimposed on a higher-order DNA target model describing the spatial coordinates of the whole genome inside a human cell. Induction of DNA double-strand breaks was simulated for proton irradiation with LET values between 1.6 and 70 keV/microm and various reference radiation qualities. The yield of DSBs after proton irradiation was found to rise continuously with increasing LET up to about 20 DSBs per Gbp and Gy, corresponding to an RBE up to 2.2. About half of this increase resulted from a higher yield of DSB clusters associated with small fragments below 10 kbp. Exclusion of experimentally unresolved multiple DSBs reduced the maximum DSB yield by 30% and shifted it to an LET of about 40 keV/microm. Simulated fragment size distributions deviated significantly from random breakage distributions over the whole size range after irradiation with protons with an LET above 10 keV/microm. Determination of DSB yields using equations derived for random breakage resulted in an underestimation by up to 20%. The inclusion of background fragments had only a minor influence on the distribution of the DNA fragments induced by radiation. Despite limited numerical agreement, the simulations reproduced the trends in proton-induced DNA DSBs and fragment induction found in recent experiments.     

Biological effectiveness of low energy protons. I. Survival of Chinese hamster cells

The biological effectiveness of monoenergetic protons was investigated with the track-segment method. Protons were accelerated by a Tandem Van de Graaff accelerator and their final energies were 3.0 and 7.4 MeV. The biological system used was Chinese hamster V-79 cells and their survival ability following proton irradiation was investigated. Cobalt-60 gamma-rays were used as reference radiation to assess proton relative biological effectiveness (RBE). Survival curves were obtained for the gamma-ray and proton irradiations, and the relation S = exp (-alpha D-beta D2) was fitted to the data and the parameters alpha and beta were determined. The RBE values, calculated on the basis of the mean inactivation dose D and other pertinent parameters, were found to be 1.7 +/- 0.1 and 2.8 +/- 0.2 for 7.4 and 3.0 MeV protons, respectively. Comparisons were made with the results published by other investigators and it was concluded that in this low energy range the biological effectiveness increases substantially with decreasing proton energy.     

Proton-HZE-particle sequential dual-beam exposures increase anchorage-independent growth frequencies in primary human fibroblasts

The radiation field in deep space contains high levels of high-energy protons and substantially lower levels of high-atomic-number, high-energy (HZE) particles. Calculations indicate that cellular nuclei of human space travelers will be hit during a 3-year Mars mission by approximately 400 protons and approximately 0.4 HZE particles. Thus most cells in astronauts will be hit by a proton(s) before being hit by an HZE particle. To investigate effects of dual ion irradiations on human cells, we irradiated primary human neonatal fibroblasts with protons (1 GeV/nucleon, 20 cGy) followed from 2.5 min to 48 h later by iron or titanium ions (1 GeV/nucleon, 20 cGy) and then measured clonogenic survival and frequency of anchorage-independent growth. This frequency depends on the interval between hydrogen- and iron-ion irradiation, with a critical window between 2.5 min and 1 h producing about three times more anchorage-independent colonies per survivor than expected from simple addition of the two ions separately. The hydrogen-titanium-ion dual-beam irradiation produced similar increases that persisted to approximately 6 h. At longer intervals, anchorage-independent growth frequencies were similar to those expected for additivity. However, irradiation of cells with either an iron or a titanium particle first followed by protons produced only additive levels.