The Clinical and Pathological Response of Goats to Whole-body Irradiation by Gamma-rays and Fission Neutrons

Summary

Fifty adult male British Saanen goats in equal numbers were given acute whole-body irradiation with either fission neutrons or gamma-rays of 2·5 mev mean energy. The LD 50/30 was about 505 rad for neutrons and 395 r for gamma-rays, giving an RBE of 0·8.

Higher doses resulted in a fatal ‘gastro-intestinal’ syndrome characterized by severe to absolute anorexia, fluid intake greatly reduced to absent, progressive haemoconcentration from 48 hours and diarrhoea from 4 days after exposure. At post-mortem there were mucus casts within the lumen of the small intestine. Deaths in the third and fourth weeks were commonly the result of widespread haemorrhage with severe thrombocytopenia following bone-marrow damage. Severe anaemia was seen only once and that in a survivor. Although few deaths could be confidently attributed to infection and blood-cultures were sterile, bacterial aggregates were nearly always found in the tissues, most commonly in the intestines, liver and lungs, and fever occurred invariably in the few days before death.

Blood urea, proteins, glucose, chloride and electrolyte measurements showed no radiation-induced changes.

No deaths were seen after 30 days in a one-year follow-up. Platelet recovery when it began was usually rapid. The most reliable prognostic sign was a progressive rise in platelet count. Blood lymphocytes began to increase in number at the end of the first week, even in animals which were going to die, but recovery was so slow that even after one year, values were only about half the pre-irradiation mean.

The response to the two kinds of radiation was qualitatively very similar. However, epilation was common after neutrons and rare after gamma-radiation, and in survivors platelet recovery was distinctly faster after neutron-than after gamma-radiation.     

The Relative Biological Efficiency of Single Doses of Fast Neutrons and Gamma-rays on Vicia Faba Roots and the Effect of Oxygen

Summary

The frequency of micronuclei resulting from acentric chromosome and chromatid fragments induced by cobalt-60 gamma radiation and by ～3 mev fast neutrons has been studied in the root-tip meristematic cells of Vicia faba. Roots irradiated with gamma-ray doses of 107 and 188 rads whilst in aerated water showed a greater micronuclear frequency than roots irradiated with similar doses in the absence of oxygen. The ratio of doses given in nitrogen and in air in order to produce the same micronuclear yield was found to be about 2·4. With roots irradiated with fast-neutron doses of 2·02, 4·95 and 9·94 rads the effect of oxygen is less marked, the nitrogen/air dose-ratio being about 1·4.

For a given level of chromosome damage produced under conditions of aeration, neutron radiation was found to be about 10·5 times as effective as gamma-rays. Because of the quantitative difference in the response of the two types of radiation to the presence of oxygen, the relative biological efficiency of neutrons to gamma-rays under anoxic conditions would be about 18 to 1.     

The Effect of Oxygen on the Sensitivity of Mammalian Tumour Cells to Neutrons and X-rays

Summary

Telophase abnormalities in Ehrlich ascites-tumour cells were observed 18 hours after the cells had been irradiated either with 1·5 mev x-rays or fast neutrons. The sensitivity of well-oxygenated tumour was greater than that of anoxic tumour by a factor of 2·5 in the case of x-rays, but of only 1·3 in the case of neutrons. After neutron-irradiation the abnormalities showed an exponential relationship to dose, but after x-irradiation the abnormalities increased more rapidly than the first power of the dose. When the proportion of abnormalities was 50 per cent the RBE in O₂ was 3·2, but in N₂ it was 6·5.     

The Production of Chromosome Aberration in Chinese Hamster Fibroblasts Exposed to 24 keV Neutrons

Summary

A filtered reactor beam, consisting mainly of 24 keV neutrons, was used to study the induction of chromosome aberrations in the V79/4(AH1) Chinese hamster cell line. The yields of both dicentrics and acentrics were linear with dose and the value of relative biological effectiveness (RBE) for dicentrics at low doses was 6·5 ± 1·4. This value was similar to that found previously for a neutron spectrum with mean energy 2·1 MeV, and suggests that the RBE of neutrons does not increase to very high values in the energy region below 100 keV. This result does not support the suggestions of Davy (1969) and Key (1971) that the neutron RBE rises to very high values in the intermediate energy range.     

The Relative Biological Effectiveness of Different Kinds of Radiations on Chromosome Aberrations in Nigella Damascena Seed

Summary

In Nigella damascena the relative biological effectiveness (RBE) of different kinds of radiations was measured for chromosome breaks induced in G₁ and scored at first mitosis after germination of seeds. The effects of γ-rays from ¹³⁷Cs were used for reference to compute RBE's for 2·8 GeV protons, mono-energetic neutrons, fission neutrons, and α-particles from the reactions ¹⁰B(n, α)⁷Li and ⁶Li(n, α)³H following irradiation with thermal neutrons. For 2·8 GeV protons the RBE was 2·1 for total aberrations. The RBE's for mono-energetic neutrons ranged from 44 (1·8 MeV) to 84 (0·43 MeV) for one-break, and 79 (1·8 MeV) to 154 (0·43 MeV) for two-break aberrations: for fission neutrons the RBE was 78 for total aberrations. The low RBE for α-particles from thermal neutron irradiation of seeds enriched with ¹⁰B and ⁶Li was attributed in part to the non-homogeneous distribution of these atoms in the seeds.     

Lack of Inverse Dose-rate Effect on Fission Neutron Induced Transformation of C3H/10T1/2 Cells

Summary

Exponential and density-inhibited cultures of C3H/10T1/2 cells were exposed to a single dose of 0·3 Gy of fission neutrons delivered at rates ranging from 0·005 to 0·1 Gy/min. No discernible effect upon cell survival or transformation was observed by a lowering of the fission neutron dose rate in either exponential or plateau cultures. At the level of 2·3 × 10^?4 transformants per surviving cell, the RBE for neoplastic transformation was three at acute dose rates and ten at the lowest dose rate studied (0·005 Gy/min for neutrons and 0·01 Gy/min for X-rays).     

Lung Tumour Induction in Mice after X-rays and Neutrons

Summary

Dose–response curves were determined for pulmonary adenomas and adenocarcinomas in mice after single acute doses of 200 kVp X-rays and cyclotron neutrons (E¯ = 7·5 MeV). A serial-killing experiment established that the radiation induction of chromosome aberrations. The validity of the concept of PLD neous cancers. The dose versus incidence (I) of tumours in male and female mice for X-ray doses between 0·25 and 7·5 Gy is ‘bell-shaped’ and best fitted with a purely quadratic induction and exponential inactivation terms, i.e. I = A + BD²e^?αD. In contrast, the tumour dose–response after 0·1–4·0 Gy of neutrons is best fitted by I = A + BD e^?αD and is steeply linear ≤ 1 Gy, peaks between 1 and 3 Gy and sharply declines at 4·0 Gy. The data for the female mice ≤ 1 Gy neutrons are best fitted to the square root of the dose.

A major objective of the experiments was to derive neutron RBE values. Because of the differences between the X-ray (quadratic) and neutron (linear) curves, the RBE_n will vary inversely with decreasing X-ray dose. The RBE values at 1 Gy of X-rays derived from the B coefficients in the above equations are 7·4 ± 3·2 (male and female); 8·6 ± 3·6 (female) and 4·7 ± 1·8 (male). These are high values and imply even higher values at the doses of interest to radiation protection. If, however, one restricts the analysis to the initial, induction side of the response (≤ 1 Gy neutrons, ≤ 3 Gy X-rays) then good linear fits are obtainable for both radiations and indicate neutron RBE values of 7·4 ± 2·3 for female mice and 4·5 ± 1·8 for males, and these are independent of dose level.     

Flow Cytometric Analysis of the Effects of 0·4 MeV Fission Neutrons on Mouse Spermatogenesis

Summary

(C57B1/Cne × C3H/Cne)F₁ male mice were irradiated with single acute doses of 0·4 MeV neutrons ranging from 0·05 to 2 Gy, and testis cell suspensions were prepared for cytometric analysis of the DNA content 2–70 days after irradiation. Various cell subpopulations could be identified in the control histogram including mature and immature spermatids, diploid spermatogonia and spermatocytes, tetraploid cells and cells in the S-phase. Variations in the relative proportions of different cell types were detected at each dose and time, reflecting lethal damage induced on specific spermatogenetic stages. The reduction of the number of elongated spermatids 28 days after irradiation was shown to be a particularly sensitive parameter for the cytometrical assessment of the radiosensitivity of differentiating gonia. A D₀ value of 0·13 Gy was calculated and compared with data obtained after X-irradiation, using the same experimental protocol. In the latter case a biphasic curve was obtained over the dose range from 0·25 to 10 Gy, possibly reflecting the existence of some cell population heterogeneity. RBE values were estimated at different neutron doses relative to the radiosensitive component of the X-ray curve, and ranged from 3·3 to 4, in agreement with data in the literature. Genotoxic effects were monitored 7 days after irradiation by a dose-dependent increase of the coefficient of variation (CV) values of the round spermatid peak, reflecting the induction of numerical and structural chromosome aberrations, and 14 or 21 days after irradiation by the detection of diploid elongated spermatids, probably arising from a radiation-induced complete failure of the first or second meiotic division.     

Interaction between the biological effects of high- and low-LET radiation dose components in a mixed field exposure

Abstract

Purpose: The relative biological effectiveness of two epithermal neutron sources, a reactor based source at Studsvik, Sweden, and a proton accelerator-based source in Birmingham, UK, was studied in relation to the proportional absorbed dose distribution as a function of neutron energy. Evidence for any interactions between the effects of biological damage induced by high- and low-linear energy transfer (LET) dose components, in this ‘mixed field’ irradiation, was also examined

Materials and methods: Clonogenic survival in Chinese Hamster-derived V79 cells was used to assess biological effectiveness in this study. Cells were irradiated in suspension at 4°C at depths of 20, 35, 50 and 65 mm in a water phantom. This prevented the repair of sublethal damage, predominantly that produced by both incident and induced γ-rays in the field, over the variable periods of exposure required to irradiate cells with the same total absorbed dose. Cell survival, as a function of the absorbed radiation dose and depth in the phantom, was compared with Monte Carlo N-Particle (MCNP) calculations of the proportional absorbed dose distribution as a function of neutron energy for the two sources.

Results: In terms of the dose-related reduction in clonogenic cell survival, the epithermal neutron source at Studsvik was more biologically effective than the Birmingham source at all depths considered in the phantom. Although the contribution from the high-LET dose component was greater for the Studsvik source at 20 mm depth in the phantom, at greater depths the dose contribution from the high-LET dose component at Studsvik overlap with those for the Birmingham source. However, the most striking difference is in the fast neutron component to the dose of the two sources, neutron energies > 1 MeV were only associated with the Studsvik source. The relative biological effectiveness (RBE) of both sources declined slightly with depth in the phantom, as the total high-LET dose component declined. The maximum source RBE for Studsvik was 2.70 ± 0.50 at 20 mm; reduced to 2.10 ± 0.35 at depths of 50 and 65 mm. The corresponding values for Birmingham were 1.68 ± 0.25 and 1.31 ± 0.19, all values relate only to the surviving fraction of V79 cells at 37%, since RBE values are only applicable to the selected endpoint. Based on a dose reduction factor (DRF) of 1.0 for the total low-LET component to the absorbed dose, the RBE values for the high-LET dose component (fast neutrons and induced protons from the nitrogen capture reaction) was 14.5 and 7.05 for the Studsvik and Birmingham neutron sources, respectively. This is well outside the range of RBE historically reported values for V79 cells for the same level of cell survival for fast neutrons. The calculation of RBE values, based on the proportional absorbed dose distribution as a function of neutron energy, from historical data, and using a RBE of 1.8 for the dose from the nitrogen capture reaction, suggests RBE values for the total high-LET dose component of 3.1–2.8 and 2.5–2.0 for Studsvik and Birmingham, respectively, values again declining with depth in the phantom.

Conclusions: The overall biological effectiveness of the mixed field irradiation from an epithermal neutron sources depends on the composition and quality of the different dose components. The experimentally derived RBE values for the total high-LET dose components in these ‘mixed field’ irradiations are well in excess of historical data for fast neutrons. The difference between the historically expected and the observed RBE values is attributed to the interactions between the damage produced by high- and low-LET radiation.     

The Effect of 238Pu α-particles on the Mouse Fibroblast Cell Line C3H 10T1/2: Characterization of Source and RBE for Cell Survival

Summary

Considerable interest has been aroused in recent years by reports that the transforming and carcinogenic effectiveness of low doses of high LET radiations can be increased by reducing the dose rate, especially for transformation of 10T1/2 cells in vitro by fission-spectrum neutrons. We report on conditions which have been established for irradiation of 10T1/2 cells with high LET monoenergetic α-particles (energy of 3·2 MeV, LET of 124 keV μm^?1) from ²³⁸Pu. The α-particle irradiator allows convenient irradiation of multiple dishes of cells at selectable high or low dose rates and temperatures. The survival curves of irradiated cells showed that the mean lethal dose of α-particles was 0·6 Gy and corresponded to an RBE, at high dose rates, of 7·9 at 80 per cent survival and 4·6 at 5 per cent survival, relative to ⁶⁰Co γ-rays. The mean areas of the 10T1/2 nuclei, perpendicular to the incident α-particles, was measured as 201 μm², from which it follows that, on average, only one in six of the α-particle traversals through a cell nucleus is lethal. Under the well-characterized conditions of these experiments the event frequency of α-particle traversals through cell nuclei is 9·8 Gy^?1.     

The Relative Biological Efficiency of 24 MeV and 200 kV X-rays Measured by the Reduction in Growth of Roots of Vicia faba

Summary

The RBE of 24 mev x-rays and 200 kv x-rays has been measured at five dose-levels by a comparison of their effects on the growth of roots of Vicia faba. The results support the expectation that the RBE increases as the dose is increased.     

Absence of A Dose-rate Effect in the Transformation of C3H 10T1/2 Cells by α-particles

Summary

The findings of Hill et al. (1984) on the greatly enhanced transformation frequencies at very low dose rates of fission neutrons induced us to perform an analogous study with α-particles at comparable dose rates. Transformation frequencies were determined with γ-rays at high dose rate (0·5 Gy/min), and with α-particles at high (0·2 Gy/min) and at low dose rates (0·83–2·5 mGy/min) in the C3H 10T1/2 cell system.

α-particles were substantially more effective than γ-rays, both for cell inactivation and for neoplastic transformation at high and low dose rates. The relative biological effectiveness (RBE) for cell inactivation and for neoplastic transformation was of similar magnitude, and ranged from about 3 at an α-particle dose of 2 Gy to values of the order of 10 at 0·25 Gy. In contrast to the experiments of Hill et al. (1984) with fission neutrons, no increased transformation frequencies were observed when the α-particle dose was protracted over several hours.     

Biological Effectiveness of Low Energy Protons. I. Survival of Chinese Hamster Cells

Summary

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 γ-rays were used as reference radiation to assess proton relative biological effectiveness (RBE). Survival curves were obtained for the γ-ray and proton irradiations, and the relation S = exp (?αD ? βD²) was fitted to the data and the parameters α and β 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.     

Effects of 14 MeV Neutrons and 300 kVp X-rays on Geotropic Response and Sedimentation Velocity of Statoliths in Barley Roots

Summary

The effects of 300 kVp x-rays and 14 MeV neutrons on the geotropic response and the sedimentation of statoliths in the statocytes of barley roots have been studied. 24 hours after irradiation with 100 rads of x-rays or 60 rads of 14 MeV neutrons in air, the geotropic response was inhibited by 25 and 15 per cent, respectively. Higher doses up to 800 rads of x-rays or 220 rads of neutrons had no greater effect. Irradiation in the absence of oxygen did not affect the geotropic response, and a slight enhancement was observed. Sedimentation of statoliths was slowed down after irradiation in air and speeded up after irradiation in the absence of oxygen. The hypothesis is put forward that the effects of radiation on geotropic response are due to the changes in the viscosity of the protoplasm; irradiation in air increases the viscosity; whereas irradiation in the absence of oxygen decreases it. Mechanisms for the effects of radiation on the viscosity of the protoplasm are suggested.     

Dose-response and relative biological effectiveness of fast neutrons: Induction of apoptosis and inhibition of neurogenesis in the hippocampus of adult mice

Purpose:?Our study compared the effects of high linear energy transfer (LET) fast neutrons on the induction of apoptosis and reduction of neurogenesis in the hippocampus of adult ICR mice with those of low-LET ⁶⁰Co γ-rays, to evaluate the relative biological effectiveness (RBE) of fast neutrons in the adult hippocampal dentate gyrus (DG).

Materials and Methods:?The mice were exposed to 35 MeV fast neutrons or ⁶⁰Co γ-rays. We evaluated acutely the incidence of apoptosis and expression of Ki-67 (a protein marker for cell proliferation originally defined by the monoclonal antibody Kiel-67) and doublecortin (DCX: an immature progenitor neuron marker) in the hippocampus after a single whole-body irradiation.

Results:?The number of terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labelling (TUNEL)-positive apoptotic nuclei in the DG increased and both Ki-67- and DCX-positive cells declined in a dose-dependent pattern, with fast neutrons or γ-rays. In the hippocampus, which showed an apoptosis frequency between 2 and 8 per DG, the RBE of fast neutrons was approximately 1.9. Additionally, the inhibitory effects of fast neutrons on the expression frequencies of Ki-67 (4–8) and DCX (8–32) were approximately 3.2 and 2.5 times, respectively, the effects of γ-rays at the same dose.

Conclusions:?Increased apoptotic cell death and decreased neurogenesis in the hippocampal DG were seen in a dose-dependent pattern after exposure to fast neutrons and γ-rays. In addition, the different rate of hippocampal neurogenesis between different radiation qualities may be an index of RBE.     

A novel vertebrate system for the examination and direct comparison of the relative biological effectiveness for different radiation qualities and sources

Purpose: The recent rapid increase of hadron therapy applications requires the development of high performance, reliable in vivo models for preclinical research on the biological effects of high linear energy transfer (LET) particle radiation.

Aim: The aim of this paper was to test the relative biological effectiveness (RBE) of the zebrafish embryo system at two neutron facilities.

Material and Methods: Series of viable zebrafish embryos at 24-hour post-fertilization (hpf) were exposed to single fraction, whole-body, photon and neutron (reactor fission neutrons (<En?=?1?MeV>) and (p (18?MeV)+Be, <En>?=?3.5?MeV) fast neutron) irradiation. The survival and morphologic abnormalities of each embryo were assessed at 24-hour intervals from the point of fertilization up to 192 hpf and then compared to conventional 6?MV photon beam irradiation results.

Results: The higher energy of the fast neutron beams represents lower RBE (ref. source LINAC 6?MV photon). The lethality rate in the zebrafish embryo model was 10 times higher for 1?MeV fission neutrons and 2.5 times greater for p (18?MeV)+Be cyclotron generated fast neutron beam when compared to photon irradiation results. Dose-dependent organ perturbations (shortening of the body length, spine curvature, microcephaly, micro-ophthalmia, pericardial edema and inhibition of yolk sac resorption) and microscopic (marked cellular changes in eyes, brain, liver, muscle and the gastrointestinal system) changes scale together with the dose response.

Conclusion: The zebrafish embryo system is a powerful and versatile model for assessing the effect of ionizing radiation with different LET values on viability, organ and tissue development.     

Mutagenic effect of low energy neutrons on human chromosome 11

Purpose: The shape of the dose–effect curve for neutrons, i.e. the question as to whether the curve is linear or supralinear in the low‐dose region, is still not clear. Therefore, the mutagenic effect of very low doses of low‐energy neutrons was determined.

Materials and methods: Human–hamster hybrid A_L cells contain human chromosome 11, which expresses the membrane protein CD59. This membrane protein can be detected immunologically and quantified by flow cytometry. The A_L cells were irradiated with neutrons of 0.565, 2.5 or 14.8?MeV and the results were compared with those after 200?kVp X‐rays. Before irradiation, cells spontaneously mutated in the CD59 gene were removed by magnetic cell sorting (MACS).

Results: The relative biological effectiveness (RBE) for CD59 mutation induction was 19.8 (±2.7) for 0.565?MeV, 10.2 (±1.9) for 2.5?MeV, and 10.2 (±1.6) for 14.8?MeV neutrons. Linear mutation responses were obtained with all radiations except for 14.8?MeV neutrons where a supralinear curve may be a better fit. The deletion spectrum of mutated cell clones showed 29?Mbp deletions on average after irradiation with 0.069?Gy of 0.565?MeV neutrons. This scale of deletions is similar to that after 3?Gy 100?kV X‐rays (=34?Mbp). For 50% cell survival, the RBE of the neutrons was 11 compared with 200?kV X‐rays.

Conclusion: Neutrons of low energies (0.565 or 2.5?MeV) produce a linear dose–response for mutation in the tested dose range of 0.015–0.15?Gy. The neutron curve of 14.8?MeV can be approximated by a curvilinear or linear function.     

The Radiosensitivity of Embryos of Domestic Chickens and Black-headed Gulls

Summary

Eggs of domestic chickens and black-headed gulls were continuously exposed to gamma-rays during incubation, using dose rates ranging from 0·004 to 0·08 Gy h^?1 for 20 days. Acute-dose experiments were also conducted, and eggs were irradiated on day 10 of incubation with doses of between 1·92 and 28·8 Gy. Hatchability and numbers reaching full-term developed were affected only after chronic doses of 9·6 Gy and acute doses of 4·8 Gy or higher. Maximum embryo mortality occurred around days 10–11 of incubation and just before hatching, in all experiments. An increase in foot and limb deformities was observed above acute and chronic doses of 9·6 Gy.     

A Comparison of the Response to Injected Radioactive Phosphorus and to X-rays in the Haematopoietic Tissue of Rats

The radiation response of haematopoietic tissue in rats following the administration of ³²P is compared with the response to acute whole-body x-irradiation in terms of the radiation dose to different tissues, using thick-section autoradiography for measuring the radiation dose to the bone-marrow in the animals treated with ³²P. Changes in the peripheral blood picture and results of ⁵⁹Fe tracer studies in splenectomized and intact rats are presented.

In spite of considerable differences in the conditions of irradiation, a number of similarities in the response of haematopoietic tissue are obtained with these two methods of irradiation. It is suggested that there is little or no dose-rate effect over the range ～ 30 r/min to ～ 0·2 rads/min for this tissue. A considerably earlier recovery of erythropoiesis in the spleen after ³²P than after x-irradiation is demonstrated.     

Mammary Carcinogenesis in Different Rat Strains after Irradiation and Hormone Administration

Summary

Radiation carcinogenesis of the rat mammary gland was investigated with the objective of investigating the combined effect of oestrogen administration and irradiation. Three rat strains, Sprague—Dawley, Wistar WAG/Rij and Brown Norway, with different susceptibilities to the induction of mammary cancer, have been irradiated with X-rays and mono-energetic neutrons. Increased hormone levels were obtained by subcutaneous implantation of pellets with oestradiol-17β (E₂). The tumour incidence results were corrected for competing risks and were analysed with a continuous failure time distribution. The latency period for the hormone-treated animals is considerably shorter than for animals with normal endocrinological levels. Administration of the hormone results in an appreciable increase in the proportion of rats with malignant tumours. At the level of hormone administration applied in this study, radiation and hormones appear to produce an additive effect. The effect of hormone administration and irradiation for mammary tumourigenesis is equal for hormone administration one week prior to, or 12 weeks after irradiation. The RBE values for induction of mammary carcinomas after irradiation with 0·5 MeV neutrons have a maximum value of 20 and are not strongly dependent on the hormone levels.