An assessment of galactic cosmic radiation quality considering heavy ion track structures within the cellular environment.

Beyond the magnetic influence of the Earth, the flux of galactic cosmic radiation (GCR) represents a radiological concern for long-term manned space missions. Current concepts of radiation quality and equivalent dose are inadequate for accurately specifying the relative biological "efficiency" of low doses of such heavily ionising radiations, based as they are on the single parameter of Linear Energy Transfer (LET). Such methods take no account of the mechanisms, nor of the highly inhomogeneous spatial structure, of energy deposition in radiation tracks. DNA damage in the cell nucleus, which ultimately leads to the death or transformation of the cell, is usually initiated by electrons liberated from surrounding molecules by the incident projectile ion. The characteristics of these emitted "delta-rays", dependent primarily upon the charge and velocity of the ion, are considered in relation to an idealised representation of the cellular environment. Theoretically calculated delta-ray energy spectra are multiplied by a series of weighting algorithms designed to represent the potential for DNA insult in this environment, both in terms of the quantity and quality of damage. By evaluating the resulting curves, and taking into account the energy spectra of heavy ions in space, a relative measure of the biological relevance of the most abundant GCR species is obtained, behind several shielding configurations. It is hoped that this method of assessing the radiation quality of galactic cosmic rays will be of value when considering the safety of long-term manned space missions.     

Possible effects of protracted exposure on the additivity of risks from space radiations.

Conventional radiation risk assessments are presently based on the additivity assumption. This assumption states that risks from individual components of a complex radiation field involving many different types of radiation can be added to yield the total risk of the complex radiation field. If the assumption is not correct, the summations and integrations performed to obtain the presently quoted risk estimates are not appropriate. This problem is particularly important in the area of space radiation risk evaluation because of the many different types of high- and low-LET radiation present in the galactic cosmic ray environment. For both low- and high-LET radiations at low enough dose rates, the present convention is that the addivity assumption holds. Mathematically, the total risk, Rtot is assumed to be Rtot = summation (i) Ri where the summation runs over the different types of radiation present. If the total dose (or fluence) from each component is such that the interaction between biological lesions caused by separate single track traversals is negligible within a given cell, it is presently considered to be reasonable to accept the additivity assumption. However, when the exposure is protracted over many cell doubling times (as will be the case for extended missions to the moon or Mars), the possibility exists that radiation effects that depend on multiple cellular events over a long time period, such as is probably the case in radiation-induced carcinogenesis, may not be additive in the above sense and the exposure interval may have to be included in the evaluation procedure. It is shown, however, that "inverse" dose-rate effects are not expected from intermediate LET radiations arising from the galactic cosmic ray environment due to the "sensitive-window-in-the-cell-cycle" hypothesis.     

Stochastics of HZE-induced microlesions.

Fundamental biological experiments with bacteria, yeast, and mammalian cells irradiated with ions heavier than helium indicate that maximal probability of single-hit inactivation does not occur when the ion has LET below about 100-200 keV/micrometer. Theoretical treatments of cell inactivation data and the radiation chemistry in particle tracks are consistent with this finding. If a "microlesion" is defined as a linear array, within a tissue, of cells inactivated with maximum probability, surrounded by non-lethally damaged cells, then, by this definition, there must be an LET below which "microlesion" damage cannot be expected. In a retrospective survey of experimental literature in which single-particle effects in tissues were sought, it was found that little or no evidence has been reported supporting single-particle effects in tissues when LET was below 200 keV/micrometer, while some experimenters who irradiated tissues with particles having LET greater than 200 keV/micrometer reported effects that could be attributed to single-particle tracks.     

Early and late damages induced by heavy charged particle irradiation in embryonic tissue of Arabidopsis seeds.

Early and late effects of accelerated heavy ions (HZE) on the embryonic tissue of Arabidopsis thaliana seeds were investigated seeing that initial cells of the plant eumeristems resemble the original cells of animal and human tissues with continuous cell proliferation. The endpoints measured were lethality and tumorization in the M1-generation for early effects and embryonic lethality in the M2-generation for late effects. The biological endpoints are plotted as functions of the physical parameters of the irradiation i.e. ion fluence (p/cm2), dose (Gray), charge Z and linear energy transfer (LET). The results presented contribute to the estimation of the principles of biological HZE effects and thus may help to develop a unified theory which could explain the whole sequence from physical and chemical reactions to biological responses connected with heavy ion radiation. Additionally, the data of this paper may be used for the discussion of the quality factor for heavy ion irradiation needed for space missions and for HZE-application in radio-therapy by use of accelerators (UNILAC, (SIS/ESR), BEVALAC).     

A new perspective of carcinogenesis from protracted high-LET radiation arises from the two-stage clonal expansion model.

When applied to the Colorado Plateau miner population, the two-stage clonal expansion (TSCE) model of radiation carcinogenesis predicts that radiation-induced promotion dominates radiation-induced initiation. Thus, according to the model, at least for alpha-particle radiation from inhaled radon daughters, lung cancer induction over long periods of protracted irradiation appears to be dominated by radiation-induced modification of the proliferation kinetics of already-initiated cells rather than by direct radiation-induced initiation (i.e., mutation) of normal cells. We explore the possible consequences of this result for radiation exposures to space travelers on long missions. Still unknown is the LET dependence of this effect. Speculations of the cause of this phenomenon include the suggestion that modification of cell kinetics is caused by a "bystander" effect, i.e., the traversal of normal cells by alpha particles, followed by the signaling of these cells to nearby initiated cells which then modify their proliferation kinetics.     

Radiation exposures during space flight and their measurement.

The paper reviews radiation exposures recorded during space flights of the US and USSR. Most of the data are from manned missions and include discussion of absorbed dose and dose rates as a function of parameters such as altitude, inclination, spacecraft type and shielding. Preliminary data exist on the neutron and HZE-particle component, as well as the LET spectra. For low Earth-orbit missions, the dose encountered is strongly altitude-dependent, with a weaker dependence upon inclination. The doses range from about 6 millirad per day for the Space Transportation System No. 3 flight to about 90 mrad per day for Skylab. The effective quality factor (QF) for the near-Earth orbits and free space has been estimated to be about 1.5 and about 5.5 respectively. Complete shielding from the galactic cosmic rays does not appear practical because of spacecraft weight limitations.     

Induction and repair of DNA strand breaks in bovine lens epithelial cells after high LET irradiation.

The lens epithelium is the initiation site for the development of radiation induced cataracts. Radiation in the cortex and nucleus interacts with proteins, while in the epithelium, experimental results reveal mutagenic and cytotoxic effects. It is suggested that incorrectly repaired DNA damage may be lethal in terms of cellular reproduction and also may initiate the development of mutations or transformations in surviving cells. The occurrence of such genetically modified cells may lead to lens opacification. For a quantitative risk estimation for astronauts and space travelers it is necessary to know the relative biological effectiveness (RBE), because the spacial and temporal distribution of initial physical damage induced by cosmic radiation differ significantly from that of X-rays. RBEs for the induction of DNA strand breaks and the efficiency of repair of these breaks were measured in cultured diploid bovine lens epithelial cells exposed to different LET irradiation to either 300 kV X-rays or to heavy ions at the UNILAC accelerator at GSI. Accelerated ions from Z=8 (O) to Z=92 (U) were used. Strand breaks were measured by hydroxyapatite chromatography of alkaline unwound DNA (overall strand breaks). Results showed that DNA damage occurs as a function of dose, of kinetic energy and of LET. For particles having the same LET the severity of the DNA damage increases with dose. For a given particle dose, as the LET rises, the numbers of DNA strand breaks increase to a maximum and then reach a plateau or decrease. Repair kinetics depend on the fluence (irradiation dose). At any LET value, repair is much slower after heavy ion exposure than after X-irradiation. For ions with an LET of less than 10,000 keV micrometers-1 more than 90 percent of the strand breaks induced are repaired within 24 hours. At higher particle fluences, especially for low energetic particles with a very high local density of energy deposition within the particle track, a higher proportion of non-rejoined breaks is found, even after prolonged periods of incubation. At the highest LET value (16,300 keV micrometers-1) no significant repair is observed. These LET-dependencies are consistent with the current mechanistic model for radiation induced cataractogenesis which postulates that genomic damage to the surviving fraction of epithelial cells is responsible for lens opacification.     

Estimation of risk based on multiple events in radiation carcinogenesis of rat skin.

In the multistage theory of carcinogenesis, cells progress to cancer through a series of discrete, irreversible, heritable genetic alterations or mutations. However data on radiation-induced cancer incidence in rat skin suggests that some part of an intermediate repairable alteration may occur. Data are presented on cancer induction in rat skin exposed to the following radiations: 1. an electron beam (LET=0.34 keV/um, 2. a neon ion beam (LET=25 keV/um and 3. an argon ion beam (LET=125 keV/um. The latter 2 beams were generated by the Bevalac at the Lawrence Berkeley Laboratory, Berkeley, CA. About 6.0 cm2 of skin was irradiated per rat. The rats were observed every 6 weeks for at least 78 weeks and tumors were scored at first occurrence. Several histological types of cancer, including squamous and basal cell carcinomas, were induced. The cancer yield versus radiation dose was fitted by the quadratic equation (Y(D)=CLD+BD2), and the parameters C and B were estimated for each type of radiation. Analysis of the DNA from the electron-induced carcinomas indicated that K-ras and/or c-myc oncogenes were activated in all tumors tested, although only a small proportion of neon-induced tumors showed similar activation. In situ hybridization indicated that the cancers contain subpopulations of cells with differing amounts of c-myc and H-ras amplification. The results are consistent with the idea that ionizing radiation produces carcinogenically relevant lesions via 2 repairable events at low LET and via a non-repairable, linked event pathway at high LET; either pathway may advance the cell by 1 stage in the multistage model. The model, if validated, permits the direct calculation of cancer risk in rat skin in a way that can be subjected to experimental testing.     

Cell cycle delays induced by heavy ion irradiation of synchronous mammalian cells.

Cell cycle effects of very high LET particles on synchronous V79 Chinese Hamster cells have been studied in a track segment experiment by means of flow cytometric methods. Cells were irradiated with 10 MeV/u Pb-ions (LET = 13500 keV/micrometers) at an average fluence of 2 particles per cell nucleus, corresponding to a survival level of about 25%. Instantaneous drastic reductions of cell proliferation in all cycle phases have been observed, which affect the cell cycle for at least 50 hours after exposure to heavy ions. These findings are in clear contrast to the results from low LET radiation experiments, where significant delays can only be observed in S-phase and G2M-phase and for comparatively short time intervals of a few hours. Additionally, high LET radiation gives rise to prolonged DNA synthesis bypassing cell division, which leads to cells with DNA content greater than that of G2M-cells.     

Biological effects of heavy ions from the standpoint of target theory.

The biological effect of heavy ions is best described through the action cross section, as a function of the end-point of interest and the charge and speed of the ion. In track theory this is called the "ion-kill" cross section, for it is the effect produced by a single heavy ion and its delta rays. As with nuclear emulsions the biological track structure passes from the grain count regime to the track width regime to the thindown region with an increase in LET. With biological cells, as with any detector capable of storing sublethal damage, with low LET irradiation the action cross section (in the ion-kill mode) is increasingly obscured by the effect of "gamma-kill", by the influence of overlapping delta rays from neighboring heavy ions. Thus at low LET response is dominated by the gamma-kill mode, so that the RBE approaches 1. The theory requires 4 radiosensitivity parameters for biological detectors, extracted from survival curves at several high LET bombardments passing through the grain count regime, and at high doses. Once these are known the systematic response of biological detectors to all high LET bombardments can be unfolded separating ion kill from gamma kill, predicting the response to a mixed radiation environment, and predicting low dose response even at the level of a single heavy ion. Cell killing parameters are now available for a variety of cell lines. Newly added is a set of parameters for cell transformation.     

Fluence-related risk coefficients using the Harderian gland data as an example.

The risk of radiation-induced cancer to space travelers outside the earth's magnetosphere will be of concern on missions to the Moon and beyond to Mars. High energy galactic cosmic rays with high charge (HZE particles) will penetrate the spacecraft and the bodies of the astronauts, sometimes fragmenting into nuclear secondary species of lower charge but always ionizing densely, thus causing cellular damage which may lead to malignant transformation. To quantitate this risk, the concept of dose equivalent (in which a quality factor Q as a function of LET is assumed) may not be adequate, since different particles of the same LET may have different efficiencies for tumor induction. Also, RBE values on which quality factors are based depend on response to low-LET radiation at low doses, a very difficult region for which to obtain reliable experimental data. Thus, we introduce a new concept, a fluence-related risk coefficient (F), which is the risk of a cancer per unit particle fluence and which we call the risk cross section. The total risk is the sum of the risk from each particle type: sigma i integral Fi(Li) phi i(Li) dLi, where Li is the LET and phi i(Li) is the fluence-LET spectrum of the ith particle type. As an example, tumor prevalence data in mice are used to estimate the probability of mouse Harderian gland tumor induction per year on an extra-magnetospheric mission inside an idealized shielding configuration of a spherical aluminum shell 1 g/cm2 thick. The combined shielding code BRYNTRN/GCR is used to generate the LET spectra at the center of the sphere. Results indicate a yearly prevalence at solar minimum conditions of 0.06, with 60% of this arising from charge components with Z between 10 and 28, and two-thirds of the contribution arising from LET components between 10 and 200 keV/micrometers.     

The influence of radiation quality on the formation of DNA breaks.

We have aimed to present a comprehensive review of our understanding to date of the formation of DNA strand breaks induced by high LET radiation. We have discussed data obtained from DNA in solution as well as from the formation and "repair" of strand breaks in cell DNA. There is good agreement, qualitatively, between these two systems. Results were evaluated for two parameters: (1) effectivity per particle, the cross section (sigma) in micrometers 2/particle; and (2) the strand break induction frequency as number of breaks per Gy per unit DNA (bp or dalton). A series of biological effects curves (one for each Z-number) is obtained in effectivity versus LET plots. The relationships between induction frequencies of single-strand breaks, or double-strand breaks, or the residual "irrepairable" breaks and LET-values have been evaluated and discussed for a wide spectrum of heavy ions, both for DNA in solution and for DNA in the cell. For radiation induced total breaks in cell DNA, the RBE is less than one, while the RBE for the induction of DSBs can be greater than one in the 100-200 keV/micrometers range. The level of irrepairable strand breaks is highest in this same LET range and may reach 25 percent of the initial break yield. The data presented cover results obtained for helium to uranium particles, covering a particle incident energy range of about 2 to 900 MeV/u with a corresponding LET range of near 16 to 16000 keV/micrometers.     

Microscopic track structure of equal-LET heavy ions.

The spatial distributions of ionization and energy deposition produced by high-velocity heavy ions are crucial to an understanding of their radiation quality as exhibited eg., in track segment experiments of cell survival and chromosome aberrations of mammalian cells. The stopping power (or LET) of a high velocity ion is proportional to the ratio z2/v2, apart from a slowly varying logarithmic factor. The maximum delta-ray energy that an ion can produce is proportional to v2 (non-relativistically). Therefore, two HZE ions having the same LET, but in general differing z and v will have different maximum delta-ray energies and consequently will produce different spatial patterns of energy deposition along their paths. To begin to explore the implications of this fact for the microscopic dosimetry of heavy ions, we have calculated radial distributions in energy imparted and ionization for iron and neon ions of approximately equal LET in order to make a direct comparison of their delta-ray track structure. Monte Carlo techniques are used for the charged particle radiation transport simulation.     

In vivo and in vitro measurements of complex-type chromosomal exchanges induced by heavy ions.

Heavy ions are more efficient in producing complex-type chromosome exchanges than sparsely ionizing radiation, and this can potentially be used as a biomarker of radiation quality. We measured the induction of complex-type chromosomal aberrations in human peripheral blood lymphocytes exposed in vitro to accelerated H-, He-, C-, Ar-, Fe- and Au-ions in the LET range of approximately 0.4-1400 keV/micrometers. Chromosomes were analyzed either at the first post-irradiation mitosis, or in interphase, following premature condensation by phosphatase inhibitors. Selected chromosomes were then visualized after FISH-painting. The dose-response curve for the induction of complex-type exchanges by heavy ions was linear in the dose-range 0.2-1.5 Gy, while gamma-rays did not produce a significant increase in the yield of complex rearrangements in this dose range. The yield of complex aberrations after 1 Gy of heavy ions increased up to an LET around 100 keV/micrometers, and then declined at higher LET values. When mitotic cells were analyzed, the frequency of complex rearrangements after 1 Gy was about 10 times higher for Ar- or Fe- ions (the most effective ions, with LET around 100 keV/micrometers) than for 250 MeV protons, and values were about 35 times higher in prematurely condensed chromosomes. These results suggest that complex rearrangements may be detected in astronauts' blood lymphocytes after long-term space flight, because crews are exposed to HZE particles from galactic cosmic radiation. However, in a cytogenetic study of ten astronauts after long-term missions on the Mir or International Space Station, we found a very low frequency of complex rearrangements, and a significant post-flight increase was detected in only one out of the ten crewmembers. It appears that the use of complex-type exchanges as biomarker of radiation quality in vivo after low-dose chronic exposure in mixed radiation fields is hampered by statistical uncertainties.     

Animal studies of life shortening and cancer risk from space radiation.

The U.S. Air Force study of the delayed effects of single, total body exposures to simulated space radiation in rhesus monkeys is now in its 21st year. Observations on 301 irradiated and 57 age-matched control animals indicate that life expectancy loss from exposure to protons in the energy range encountered in the Van Allen belts and solar proton events can be expressed as a logarithmic function of the dose. The primary causes of life shortening are cancer and endometriosis (an abnormal proliferation of the lining of the uterus in females). Life shortening estimates permit comparison of the risk associated with space radiation exposures to be compared with that of other occupational and environmental hazards, thereby facilitating risk/benefit decisions in the planning and operational phases of manned space missions. Calculations of the relative risk of fatal cancers in the irradiated subjects reveal that the total body surface dose required to double the risk of death from cancer over a 20-year post exposure period varies with the linear energy transfer (LET) of the radiation. The ability to determine the integrated dose and LET spectrum in space radiation exposures of humans is, therefore, critical to the assessment of lifetime cancer risk.     

Induction of chromosomal damage in CHO-K1 cells and their repair-deficient mutant XRS5 by X-ray and particle irradiation.

The cytogenetic effects of X-rays and Au ions were investigated in repair-proficient CHO-K1 cells and their radiosensitive mutant strain xrs5, which shows a defect in the rejoining of DNA double-strand breaks. Both cell lines were synchronized by mitotic shake off, irradiated in G1-phase with either 250 kV X-rays or 780 MeV/u Au ions (LET: 1150 keV/micrometer) and chromosome aberrations were analyzed in first post-irradiation metaphases. Isoeffective doses of X-rays for the induction of aberrant cells and aberrations per cell were about 14 times lower for xrs5 than for CHO-K1 cells. After high LET radiation the difference in the cytogenetic response of both cell lines was drastically diminished. Furthermore, the analysis of the aberration types induced by sparsely and densely ionizing radiation showed for both cell lines specific changes in the spectrum of aberration types as LET increases. The experimental results are discussed with respect to the different types of lesions induced by sparsely and densely ionizing radiation.     

Haemopoietic cell renewal in radiation fields.

Space flight activities are inevitably associated with a chronic exposure of astronauts to a complex mixture of ionising radiation. Although no acute radiation consequences are to be expected as a rule, the possibility of Solar Particle Events (SPE) associated with relatively high doses of radiation (1 or more Gray) cannot be excluded. It is the responsibility of physicians in charge of the health of astronauts to evaluate before, during and after space flight activities the functional status of haemopoietic cell renewal. Chronic low level exposure of dogs indicate that daily gamma-exposure doses below about 2 cGy are tolerated for several years as far as blood cell concentrations are concerned. However, the stem cell pool may be severely affected. The maintenance of sufficient blood cell counts is possible only through increased cell production to compensate for the radiation inflicted excess cell loss. This behaviour of haemopoietic cell renewal during chronic low level exposure can be simulated by bioengineering models of granulocytopoiesis. It is possible to define a "turbulence region" for cell loss rates, below which an prolonged adaptation to increased radiation fields can be expected to be tolerated. On the basis of these experimental results, it is recommended to develop new biological indicators to monitor haemopoietic cell renewal at the level of the stem cell pool using blood stem cells in addition to the determination of cytokine concentrations in the serum (and other novel approaches). To prepare for unexpected haemopoietic effects during prolonged space missions, research should be increased to modify the radiation sensitivity of haemopoietic stem cells (for instance by the application of certain regulatory molecules). In addition, a "blood stem cell bank" might be established for the autologous storage of stem cells and for use in space activities keeping them in a radiation protected container.     

Modifying factors on repair phenomena.

Every step of radiation damage repair can be modified by several factors like temporal and local energy distribution (LET), physiological conditions, biochemical and chemical composition of the target environment. Most interesting in the case of space radiation is the possible influence of high LET and of microgravity which could change the repair i.e. fusion capacity as well as the phenomenon of self assembly.     

Cataractogenesis from high-LET radiation and the Casarett model.

Space radiations, especially heavy ions, constitute significant hazards to astronauts. These hazards will increase as space missions lengthen. Moreover, the dangers to astronauts will be enhanced by the persistence, or even the progression, of biological damage throughout their subsequent life spans. To assist in the assessment of risks to astronauts, we are investigating the long-term effects of heavy ions on specific animal tissues. In one study, the eyes of rabbits of various ages were exposed to a single dose of Bragg plateau 20Ne ions (LET infinity approximately equals 30 keV/micrometer). The development of cataracts has shown a pronounced age-related response during the first year after irradiation, and will be followed for two more years. In other studies, mice were exposed to single or fractionated doses of 12C ions (4-cm spread-out Bragg peak; dose-averaged LET infinity = 70-80 keV/micrometer) or 60Co gamma-photons (LET infinity = 0.3 keV/micrometer). Measurements of the frequency of posterior lens opacification have shown that the tissue sparing observed with dose fractionation of gamma-photons was absent when 12C-ion doses were fractionated. Development of posterior lens cataracts was also followed for long periods (up to 21 months) in mice exposed to single doses of Bragg plateau HZE particles (40Ar, 20Ne and 12C ions: LET infinity approximately equals 100, 30 and 10 keV/micrometer, respectively) or 225 kVp X-rays. Based on average cataract levels at the different observation times, the RBE's (RBE = relative biological effectiveness) for the ions were circa 5, 3 and 1-2, respectively, over the range of doses used (0.05-0.9 Gy). Investigations of cataractogenesis are useful for exploring the model of radiation damage proposed by Casarett and by Rubin and Casarett with a tissue not connected directly to the vasculature.