Lung carcinomas in Sprague-Dawley rats after exposure to low doses of radon daughters, fission neutrons, or gamma rays

The effectiveness of radon-daughter inhalation and irradiation with fission neutrons and gamma rays in the induction of lung carcinomas in Sprague-Dawley rats at low doses is compared. Earlier reports which compared radon-daughter inhalations and neutron irradiations over a wider range of doses were based on dosimetry for the radon-daughter inhalations which has recently been found to be faulty. In the present analysis, low-dose experiments were designed to derive revised equivalence ratios between radon-daughter exposures, and fission neutron or gamma irradiations. The equivalence is approximately 15 working level months (WLM) of radon daughters to 10 mGy of neutrons (the earlier value was 30 WLM to 10 mGy). The relative biological effectiveness (RBE) of neutrons is 50 or more at a gamma-ray dose of 1 Gy. In these experiments with low doses and exposures, the lifetime incidences can be estimated from the raw incidences, while the derivation of the time dependence of the prevalence is essential for the estimation of RBE values and equivalence ratios.     

Dose-response modeling of life shortening in a retrospective analysis of the combined data from the JANUS program at Argonne National Laboratory

Life shortening was investigated in both sexes of the B6CF1 (C57BL/6 x BALB/c) mouse exposed to fission neutrons and 60Co gamma rays. Three basic exposure patterns for both neutrons and gamma rays were compared: single exposures, 24 equal once-weekly exposures, and 60 equal once-weekly exposures. Ten different dose-response models were fitted to the data for animals exposed to neutrons. The response variable used for all dose-response modeling was mean after-survival. A simple linear model adequately described the response to neutrons for females and males at doses less than or equal to 80 cGy. At higher neutron dose levels a linear-quadratic equation was required to describe the life-shortening response. An effect of exposure pattern was observed prior to the detection of curvature in the dose response for neutrons and emerged as a potentially significant factor at neutron doses in the range of 40-60 cGy. Augmentation of neutron injury with dose protraction was observed in both sexes and began at doses as low as 60 cGy. The life-shortening response for all animals exposed to gamma rays (22-1918 cGy) was linear and inversely dependent upon the protraction period (1 day, 24 weeks, 60 weeks). Depending on the exposure pattern used for the gamma-ray baseline, relative biological effectiveness (RBE) values ranged from 6 to 43. Augmentation, because it occurred only at higher levels of neutron exposure, had no influence on the estimation of RBEm.     

Life shortening in mice exposed to fission neutrons and gamma rays. VII. Effects of 60 once-weekly exposures

A total of 6316 B6CF1 mice were exposed to 60 equal once-weekly doses of 0.85-MeV fission neutrons (0.033 to 0.67 cGy per weekly fraction) or 60Co gamma rays (1.67 to 10 cGy per weekly fraction) and were observed until they died. The mean aftersurvival times showed that the dose-response curves for both neutron and gamma-ray exposures were indistinguishable from linear over all doses except the highest neutron dose. The relative biological effectiveness (RBE) for neutrons, calculated as the ratio of the initial slopes of the dose-response curves, was about 20 for both males and females. Essentially the same value was obtained by a number of other analyses of the data. Virtually all of the radiation-specific excess mortality could be attributed to tumors; after decrementation of the population for nontumor deaths, the value of the RBE was not significantly changed.     

Studies of alveolar cell morphometry and mass clearance in the rat lung following inhalation of an enriched uranium dioxide aerosol

Summary Seventy-three rats were exposed to an aerosol of enriched uranium dioxide (UO₂), giving initial lung burdens of 26 to 447 µg at 6 days post-inhalation (PI). At 7 days PI 35 of these rats were further exposed to thermalised neutrons at a fluence of 1 x 10¹² neutrons CM^–2. There was no significant difference between the two groups in the clearance rate of the UO₂ particles from the lung, up to 590 days PI. The particles cleared relatively slowly over this period with a retention halftime in the lung of 160 to 176 days.Transmission electron microscope (TEM) studies of tissue from the alveolar region at 8 days PI showed that inhalation of UO₂ particles significantly increased the sizes of macrophage and type II cells, and the number of macrophage and type I cells. There was also a significant increase in the size of lysosomal granules within the macrophages after exposure to the UO₂ particles. The exposure to UO₂, neutrons and²³⁵U fission fragments had no significant effect on any of the cells above that observed in the animals exposed to UO₂ alone.Additional rats were exposed to the same neutron fluence without prior UO₂ inhalation. The alveolar cells of neutron-only exposed rats were, in size and number, typically no different from those in the completely unexposed control rats.     

Recovery of murine lens epithelial cells from single and fractionated doses of X rays and neutrons

Subpopulations of mouse lens epithelial cells, differing in proliferative status, were irradiated with either X rays or fission spectrum neutrons given singly or in four weekly fractions. After various times, epithelia were mitogenically stimulated by wounding and DNA synthesis responses were determined by incorporation of [3H]thymidine. At 1 h following both X and neutron irradiations, significant suppression of the wound response after single doses and a sparing effect of fractionation were evident in both the mitotically quiescent and the slowly proliferating subpopulations. At 1 week following single or fractionated doses of both radiations, recovery was evident in both subpopulations. By 4 weeks, the quiescent subpopulation showed significant recovery after both single and fractionated doses of X rays or neutrons. In contrast, a marked decreased ability to respond after neutron irradiation and, in addition, a significant enhancement effect of neutron fractionation were observed for the slowly proliferating subpopulation. Per gray, neutrons were about 7.5 times more effective than X rays as a single dose and 25 times more effective in four equal fractions. The shift from an initial sparing to a final enhancing effect of neutron fractionation for the slowly proliferating subpopulation has importance for understanding divergent early and late radiation responses following dose fractionation.     

Induction of chromosome aberrations in G0 human lymphocytes by low doses of ionizing radiations of different quality

Human peripheral blood lymphocytes from two donors were exposed to low doses (0.05 to 2.0 Gy) of gamma rays, X rays, or fast neutrons of different energies. Chromosome aberrations were analyzed in metaphase of first-division cells after a culture time of 45-46 hr. At this time, less than 5% of the cells were found in second division. Different dose-response relationships were fitted to the data by using a maximum likelihood method; best fits for radiation-induced dicentric aberrations were obtained with the linear-quadratic law for all radiations. The linear component of this equation predominated, however, for neutrons in the range of doses studied, and the frequency of dicentrics induced by d(16)+Be neutrons up to 1.0 Gy could also be described by a linear relationship. The relative biological efficiency (RBE) of X rays and d(16)+Be, d(33)+Be, and d(50)+Be neutrons compared to 60Co gamma rays in the low dose range was calculated from the dose-effect relationships for the dicentrics produced. The RBE increased with decreasing neutron dose and with decreasing neutron energy from d(50)+Be to d(16)-+Be neutrons. The limiting RBE at low doses (RBEo) was calculated to be about 1.5 for X rays and 14.0, 6.2, and 4.7 for the d(16)+Be, d(33)+Be, and d(50)+Be neutrons, respectively.     

Neutron RBEs and the radiosensitive target for mouse immature oocyte killing

The highly radiosensitive immature oocytes of mice were irradiated in vivo with graded doses of 252Cf fission radiation, 0.43- or 15-MeV neutrons, or 60Co gamma rays. Comparisons of oocyte survival for neutrons and for gamma rays demonstrate that neutron RBEs for the killing of these important cells do not reach the high values (30-50 or more) at low doses observed for several other biological end points. Rather, neutrons differ little in effectiveness from gamma rays in killing these extremely sensitive murine oocytes. For 0.43-MeV neutrons, RBEs obtained from fitted survival curves reach only 1.7 at 0.1 rad. For 15-MeV neutrons, they are not significantly different from 1 at any dose tested (lowest, 4.5 rad). For 252Cf fission neutrons (E = 2.15 MeV), RBEs are intermediate between those for 0.43- and 15-MeV neutrons. For all neutron energies tested, the RBEs are particularly low in the juvenile period, a time when murine immature oocytes are especially radiosensitive. With exposure just prior to birth, however, when these cells are much less easily killed, higher, more usual RBEs are found. The minimum size of the lethality target in mouse immature oocytes, estimated from the inactivation constant for 0.43-MeV neutrons and microdosimetric values, is larger than the nucleus but not larger than the cell. This and related analytical considerations suggest that the hypersensitive target in these particular oocytes is the plasma membrane, a finding which is in excellent accord with results from other experiments using different, contrasting radiations and dose deliveries (accelerated Si14+ ions, gamma rays, and beta rays from 3HOH compared with those from [3H]thymidine).     

The effect of sequential irradiation with X-rays and fast neutrons on the survival of V79 Chinese hamster cells

V79 Chinese hamster cells have been exposed to X-rays or fast neutrons or to the two radiations given sequentially. Cells exposed to a priming dose of X-rays and then exposed immediately to a series of neutron doses regard the X-ray dose as equivalent to a neutron dose giving the same surviving fraction (iso-effective). If the cells are exposed to a neutron dose followed by X-rays the resulting survival is higher than would be obtained if the primary dose had been an iso-effective X-ray dose. However, it is lower than would be expected if the two radiations acted independently. The results imply that there is interaction between the damage caused by X-rays and fast neutrons. If the two radiations are given 3 hours apart they act independently.     

Age dependence of the radiosensitivity of glial progenitors for In vivo fission-neutron and X irradiation

O-2A progenitor cells are the stem cells of the myelin-forming oligodendrocytes in the central nervous system. In the epithermal reactor beams used for boron neutron capture therapy (BNCT) for treatment of brain tumors, fission neutrons are a contaminating component. To estimate the radiosensitivity of the O-2A progenitors for fission neutrons, an in vivo-in vitro clonogenic assay was used. Radiosensitivity of progenitors obtained from the spinal cord of 1- or 5-day-old rats or the optic nerve of 2- or 12-week-old rats for 1 MeV fission neutrons was compared to that for 300 kVp X rays. Dose-survival curves were fitted according to the linear-quadratic model. The resulting beta component was very small to negligible. Progenitor cells obtained from rats of different ages show differences in radiosensitivity, characterized by different alpha values. RBE values for fission neutrons were 3.5 for 1-day-old spinal cord, 3.2 for 5-day-old spinal cord, 3.0 for 2-week-old optic nerve, and 4.3 for 12-week-old optic nerve. These high RBE values indicate the importance of minimizing the fast-neutron component in the epithermal neutron beams used for BNCT.     

Relative biological effectiveness and tolerance dose of fission neutrons in canine skin for a potential combination of neutron capture therapy and fast-neutron therapy

To investigate the potential efficacy of fission neutrons from a fast-neutron reactor for the treatment of radioresistant tumors, the relative biological effectiveness (RBE) and tolerance dose of fission neutrons in canine skin were determined. The forelimbs of 34 healthy mongrel dogs received a single dose of fission neutrons (5.6, 6.8, 8.2, 9.6 or 11 Gy) or 137Cs gamma rays (10, 15, 20, 25 or 30 Gy). Based on observations of radiodermatitis for each radiation, the single-fraction RBE of fission neutrons in the sixth month was calculated as approximately 3. The tolerance doses of fission neutrons and gamma rays, defined as the highest doses giving no moist desquamation on the irradiated skin in the recovery phase, were estimated as 7.6 Gy and 20 Gy, respectively. The tolerance dose of 7.6 Gy of fission neutrons included 5.0 Gy of fast neutrons possessing high anti-tumor effects and 1.4 x 10(12) n/cm2 of thermal neutrons, which could be applicable to neutron capture therapy (NCT). The combination of fast-neutron therapy and NCT using a fast-neutron reactor might be useful for the treatment of radioresistant tumors.     

Oncogenic transformation by fractionated doses of neutrons

Oncogenic transformation was assayed after C3H 10T1/2 cells were irradiated with monoenergetic neutrons; cells were exposed to 0.23-, 0.35-, 0.45-, 5.9-, and 13.7-MeV neutrons given singly or in five equal fractions over 8 h. At the biologically effective neutron energy of 0.45 MeV, enhancement of transformation was evident with some small fractionated doses (below 1 Gy). When transformation was examined as a function of neutron energy at 0.5 Gy, enhancement was seen for cells exposed to three of the five energies (0.35, 0.45, and 5.9 MeV). Enhancement was greatest for cells irradiated with 5.9-MeV neutrons. Of the neutron energies examined, 5.9-MeV neutrons had the lowest dose-averaged lineal energy and linear energy transfer. This suggests that enhancement of transformation by fractionated low doses of neutrons may be radiation-quality dependent.     

Tumor induction and life shortening in BC3F1 female mice at low doses of fast neutrons and X rays

Extension of previous investigations at this laboratory regarding life shortening and tumor induction in the mouse has provided more complete dose-response information in the low dose region of X rays and neutrons. A complete observation of survival and late pathology has been carried out on over 2000 BC3F1 female mice irradiated with single doses of 1.5 MeV neutrons (0.5, 1, 2, 4, 8, 16 cGy) and, for comparison, of X rays (4, 8, 16, 32, 64, 128, 256 cGy). Data analysis has shown that a significant life shortening is observable only for individual neutron doses not lower than 8 cGy. Nevertheless, assuming a linear nonthreshold form for the overall dose-effect relationships of both radiation qualities, an RBE value of 12.3 is obtained for the 1.5 MeV neutrons. The induction of solid tumors by neutrons becomes statistically significant at individual doses from 8 cGy and by X rays for doses larger than 1 Gy. Linear dependence on neutron dose appears adequate to interpret the data at low doses. A separate analysis of ovarian tumor induction substantiates the hypothesis of a threshold dose for the X rays, while this is not strictly needed to interpret the neutron data. A trend analysis conducted on the neoplasm incidence confirms the above findings. Death rates have been analyzed, and a general agreement between the shift to earlier times of these curves and tumor induction was found.     

Life-shortening and disease incidence in C57Bl mice after single and fractionated gamma and high-energy neutron exposure

C57Bl Cnb mice were exposed to single or fractionated d(50)+Be neutrons or 137Cs gamma-ray exposure at 12 weeks of age and were followed for life-shortening and disease incidence. The data were analyzed by the Kaplan-Meier procedure using as criteria cause of death and possible cause of death. Individual groups were compared by a modified Wilcoxon test according to Hoel and Walburg, and entire sets of different doses from one radiation schedule were evaluated by the procedure of Peto and by the Cox proportional hazard model. No significant difference was found in life-shortening of C57Bl mice between a single gamma and neutron exposure. Gamma fractionation was clearly less effective in reducing survival time than a single exposure. On the contrary, fractionation of neutrons was slightly although not significantly more effective in reducing life span than a single exposure. Life-shortening appeared to be a linear function of dose in all groups studied. The data on causes of death show that malignant tumors, particularly leukemias including thymic lymphoma, and noncancerous late degenerative changes in lung were the principal cause of life-shortening after a high single gamma exposure. Exposure delivered in 8 fractions 3 h apart was more effective in causing leukemias and all carcinomas and sarcomas than one delivered in 10 fractions 24 h apart or in a single session. Following a single neutron exposure, leukemias and all carcinomas and sarcomas appeared to increase somewhat more rapidly with dose than after gamma irradiation. No significant difference in the incidence of leukemias and all carcinomas and sarcomas was noted between a single and a fractionated neutron exposure.     

Absence of a dose-fractionation effect on neoplastic transformation induced by fission-spectrum neutrons in C3H 10T1/2 cells

We have investigated the effect of fission-spectrum neutron dose fractionation on neoplastic transformation of exponentially growing C3H 10T1/2 cells. Total doses of 10.8, 27, 54, and 108 cGy were given in single doses or in five equal fractions delivered at 24-h intervals in the biological channel of the RSV-TAPIRO reactor at CRE-Casaccia. Both cell inactivation and neoplastic transformation were more effectively induced by fission neutrons than by 250-kVp X rays. No significant effect on cell survival or neoplastic transformation was observed with split doses compared to single doses of fission-spectrum neutrons. Neutron RBE values relative to X rays determined from data for survival and neoplastic transformation were comparable.     

The effect of graded doses of fission neutrons or X rays on the lymphoid compartment of the thymus in mice

Young adult CBA/H mice were exposed to graded doses of whole-body irradiation with either fast fission neutrons or 300 kVp X rays at center-line dose rates of 0.1 and 0.3 Gy/min, respectively. Dose-response curves were determined at Days 2 and 5 after irradiation for the total thymic cell survival and for the survival of thymocytes defined by monoclonal anti-Thy-1, -Lyt-1, -Lyt-2, and -T-200 antibodies as measured by flow cytofluorometric analysis. Cell dose-response curves of thymocytes show, 2 days after irradiation, a two-component curve with a radiosensitive part and a part refractory to irradiation. The radiosensitive part of the dose survival curve of the Lyt-2+ cells, i.e., mainly cortical cells, has a D0 value of about 0.26 and 0.60 Gy for neutrons and X rays, respectively, whereas that of the other cell types has corresponding D0 values of about 0.30 and 0.70 Gy. The radiorefractory part of the dose-response curves cannot be detected beyond 5 days after irradiation. At that time, the Lyt-2+ cells are again most radiosensitive with a D0 value of 0.37 and 0.99 Gy for neutrons and X rays, respectively. The other measured cell types have corresponding D0 values of about 0.47 Gy. The fission neutron RBE values for the reduction in the thymocyte populations defined by either monoclonal anti-Thy-1, -Lyt-1, -Lyt-2, or -T-200 antibodies to 1.0% vary from 2.6 to 2.8. Furthermore, the estimated D0 values of the Thy-1-, T-200- intrathymic precursor cells which repopulate the thymus during the bone marrow independent phase of the biphasic thymus regeneration after whole-body irradiation are 0.64-0.79 Gy for fission neutrons and 1.32-1.55 Gy for X rays.     

Chromosome aberrations induced in human lymphocytes by neutron irradiation.

In vitro dose--response curves of unstable chromosome aberrations in human lymphocytes have been obtained for neutron spectra of mean energies 0-7, 0-9, 7-6 and 14-7 MeV. The aberration yields have been fitted to the quadratic function Y = alphaD + betaD2, which is consistent with the single-track and two-track model of aberration formation. However with high-LET radiation, the linear component of yield, corresponding to damage caused by single tracks, predominants, and this term becomes more dominant with increasing LET, so that for fission spectrum neutrons the relationship is linear, Y = alphaD. At low doses, such as those recieved by radiation workers, limiting r.b.e. values between 13 and 47 are obtained relative to 60Co gamma-radiation. At higher doses, as used in radiotherapy, the values are much lower; ranging from 2-7 to 8 at 200 rad of equivalent gamma-radiation. Both sets of r.b.e. values correlate well with track-averaged LET but not with dose-averaged LET. When the numbers of cells without aberrations are plotted against radiation dose, curves are obtained which are similar in shape to those for conventional cell-survival experiments with comparable neutron spectra. The Do values obtained in the present study are close to those from other cell system.     

Carcinogenesis in laboratory mice after low doses of ionizing radiation

Experimental data on the incidence of solid tumors from various long-term mouse studies performed at the Casaccia laboratories over several years were reconsidered, limiting the analysis to the results available for doses equal to or less than 17 cGy of neutrons and 32 cGy of X rays since these dose limits are reasonably close to the generally accepted low-dose levels for high- and low-LET radiation (i.e. D(high-LET) < 5 cGy and D(low-LET) < 20 cGy, respectively). The following long-term experiments with BC3F1 mice were reviewed: (a) females treated with single doses of 1.5 MeV neutrons or 250 kVp X rays, (b) males treated with fractionated doses of fission neutrons, and (c) mice of both sexes irradiated in utero 17.5 days post coitus with single doses of fission neutrons or X rays. An experiment with CBA mice of both sexes treated with single doses of fission neutrons was also included in this study. Analysis was done on animals at risk; thus all incidences of tumor-bearing animals were expressed as the percentage excess incidence with respect to the controls. Ovarian tumors and other solid neoplasms were considered. The percentage frequencies and mean survival times of tumor-free mice were also recalculated. The results indicate the existence of a region at low doses where the final incidence of solid neoplasms is indistinguishable from the background incidence. These data reinforce the idea that at low doses the effectiveness of ionizing radiation in inducing solid neoplasms in laboratory mice is very low.     

Nonlinear survival curves for cells of solid tumors after large doses of fast neutrons and gamma rays.

We have previously proposed that survival curves for cells of murine NFSa fibrosarcomas after exposure to fast neutrons might demonstrate curvature when the neutron doses reach a level high enough to cure the fibrosarcomas. We report here that this is the case. Murine NFSa fibrosarcomas growing in the hind legs of syngeneic mice were exposed to either gamma rays or fast neutrons. The tumors were removed and retransplanted into fresh recipients to obtain 50% tumor cell doses, from which the dose-cell survival relationship was constructed. Survival curves showed continuous bending down to 10(-7), and were well fitted using the linear-quadratic model. The alpha and beta values for neutrons were larger than those for gamma rays. When the surviving fractions at experimental TCD50 doses were calculated using these values, comparable figures were obtained for neutrons and gamma rays. The RBEs for neutrons were comparable for the TCD50 and TD50 assays. Neutron RBE was independent of dose within a range of 5-28 Gy. The capacity of the tumors to repair the damage caused by large doses of neutrons was identical to that for small doses of neutrons, indicating that cells retained the capacity to repair neutron damage irrespective of the size of the dose.     

Neutron versus Á-ray risk estimates

While it is recognized that neutrons contributed to the excess cancer incidence and mortality among the atomic bomb survivors in Hiroshima, there is no possibility to deduce the magnitude of this contribution from the data. This remains true even if the neutron doses in the dosimetry system DS86 are corrected upwards in line with recent neutron activation measurements. In spite of this fact, important information can be obtained in the form of an inverse relation of the risk coefficients for γ-rays and neutrons. Such an interrelation must apply because the observed excess incidence or mortality is made up of a γ-ray and a neutron component; increased attribution to neutrons decreases the attribution to photons. Computations with the uncorrected and the corrected DS86 are performed for the mortality and the incidence of solid tumors combined. They refer to doses up to 2 Gy and employ the constant relative risk model and a linear-quadratic dose dependence with variable ratio – the neutron relative biological effectiveness (RBE) at low doses – of the linear component for neutrons and γ-rays. In line with past analyses, no quadratic component is obtained with the uncorrected DS86, but it is seen, even in these calculations, that the assumption of increased neutron RBEs does not translate into proportional increases of the risk coefficients of neutrons, because it leads to substantially reduced risk estimates for γ-rays. Calculations with the corrected dosimetry bring out this reciprocity even more clearly. High values of the neutron RBE reduce – in line with recent suggestions by Rossi and Zaider – the risk estimates for γ-rays substantially. Even a purely quadratic dose relation for γ-rays is consistent with the data; it requires no major increase of the nominal risk coefficients for neutrons over the currently assumed values. The cancer data from Hiroshima can still provide `prudent' risk estimates for photons, but with the corrected DS86, they do not prove that there is a linear component in the dose dependence for photons. Received: 20 January 1997 / Accepted in revised form: 14 March 1997     

Neutron RBE for induction of tumors with high lethality in Sprague-Dawley rats

The effectiveness of fission neutrons is compared to that of gamma rays and X rays with regard to the induction of malignancies in male Sprague-Dawley rats. The analysis is based on autopsy results. It is focused on tumors that tend to be present in animals dying early, which is indicative of a high degree of lethality. The relative biological effectiveness (RBE) is deduced from a comparison of the cumulative hazard functions. Different nonparametric models-the constant relative risk model, a time shift model, and an acceleration model-are employed in the comparison, and the resulting values of RBE are seen to be substantially independent of the choice of model. The results are in good agreement with earlier studies of nonlethal lung tumors in the same series of experiments. At neutron doses of 20 to 60 mGy, the RBE of fission neutrons is about 50.