Estimation of relative biological effectiveness of tritium according to chromosome aberration frequency in human blood lymphocytes

The goal of this work was to determine the relative biological effectiveness (RBE) of tritium β-radiation according to the chromosome aberration frequency in the peripheral blood lymphocytes after in vitro and in vivo radiation exposures. The experimental RBE assessment of tritium β-radiation relative to ⁶⁰Co γ-radiation according to unstable chromosome aberration frequency in the peripheral blood lymphocytes under particular conditions is described. It has been demonstrated that tritium β-radiation is, in general, more effective in the dose range of up to 1 Gy, which is most pronounced at low doses. The RBE value of tritium β-radiation at minimum doses reached 2.2 and decreased at higher doses (1 Gy) to 1.25. The data on comparative analysis of the frequency of stable chromosome aberrations in the blood lymphocytes of professional nuclear workers (Sarov, Russia) after long-term chronic exposure to tritium β-radiation, as compared with γ-irradiation, are reported for the first time. The higher biological effectiveness of tritium β-radiation was demonstrated and was estimated as 2.5.     

Induction of chromosome aberrations and micronuclei in human peripheral blood lymphocytes at low dose of radiation

The chromosome damage induced by the doses of y-irradiation 6)Co in peripheral blood lymphocytes was studied using different cytogenetic assays. Isolated lymphocytes were exposed to 0.01-1.0 Gy, stimulated by PHA, and analysed for chromosome aberrations at 48 h postirradiation by metaphase method, at 49 h--by the anaphase method, at 58 h by micronucleus assay with cytochalasin B and, additionally, micronuclei were counted at 48 h on the slides prepared for the metaphase analysis without cytochalasin B. Despite of the quantitative differences in the amount of chromosome damage revealed by different methods all of them demonstrated complex nonlinear dose dependence of the frequency of aberrant cells and aberrations. At the dose range from 0.01 Gy to 0.05-0.07 Gy the cells had the highest radiosensitivity mainly due to chromatid-type aberration induction. With dose increasing the frequency of the aberrant cells and aberrations decreased significantly (in some cases to the control level). At the doses up to 0.5-0.7 Gy the dose-effect curves have become linear with the decreased slope compare to initial one (by factor of 5 to 10 for different criteria) reflecting the higher radioresistance of cells. These data confirm the idea that the direct linear extrapolation of high dose effect to low dose range--the procedure routinelly used to estimate genetic risk of low dose irradiation--cannot be effective and may lead to underestimation of chromosome damage produced by low radiation doses. Preferences and disadvantages of used cytogenetic assays and possible mechanisms of low ionising radiation doses action were discussed.     

The human lymphocyte micronucleus assay. Response of cord blood lymphocytes to gamma-irradiation and bleomycin

The induction of micronuclei in human cord blood lymphocytes by treatment with gamma-irradiation and bleomycin has been measured. Culture durations which gave peak MN frequencies were determined. The lowest tested doses, 0.1 Gy irradiation and 1.25 micrograms/ml bleomycin, produced significant increases in the frequency of micronuclei. The spontaneous frequency of micronucleated lymphocytes in 28 cord blood samples ranged between 0.5 and 9.5 per thousand lymphocytes, with a modal value of 2.5. The method is evaluated for its potential usefulness in monitoring populations for chromosome breakage.     

Mechanism of the antimutagenic action of interferon: the capacity to protect the human cellular repair system

It has been shown that premutagenic treatment with leukocytic interferon (10, 100 IU/ml) of human peripheral blood lymphocytes cultivated in vitro at the G1-stage of the mitotic cycle results in different cell response to gamma-radiation in doses of 0.5, 1, 2, 4 Gy according to chromosome aberration. The antimutagenic effect failed to be attained with the doses 0.5 and 1 Gy, being maximal at the dose 2 Gy. According to sister chromatid exchanges (SCE) cell pretreatment with interferon leads to a reduction in the effect of gamma-radiation at the dose 2 Gy to the level obtained in the cells after exposure to interferon. In experiments with 4-nitroquinoline-I-oxide, there was a significant decrease in the number of SCE in interferon-treated cells.     

The characterization and transmissibility of chromosome aberrations induced in peripheral blood lymphocytes by in vitro alpha-particle radiation

Peripheral blood lymphocytes were irradiated in vitro with (213)Bi alpha particles at doses of 0, 10, 20, 50, 100, 200 and 500 mGy. Chromosome analysis was performed on 47-h cultures using single-color fluorescence in situ hybridization (FISH) to paint chromosomes 1, 3 and 5. The whole genome was analyzed for unstable aberrations to derive aberration frequencies and determine cell stability. The dose response for dicentrics was 33.60 +/- 0.47 x 10(-2) per Gy. A more detailed analysis revealed that the majority of aberrations scored as dicentrics were part of complex/multiple aberrations, with the proportion of cells containing complexes increasing with dose. Cells containing aberrations involving painted chromosomes (FISH aberrations) were further classified according to cell stability and complexity. The majority of cells with FISH aberrations were unstable. The proportion of aberrant FISH cells with complex/multiple aberrations ranged from 56% at 10 mGy to 89% at 500 mGy. A linear dose response for genomic frequencies of translocations in stable cells fitted the data from 0 to 200 mGy with a dose response of 7.90 +/- 0.98 x 10(-2) per Gy, thus indicating that they are likely to be observed in peripheral blood lymphocytes from individuals with past or chronic exposure to high-LET radiation. Comparisons with the dose response for low-LET radiation suggest an RBE of 13.6 for dicentrics in all cells and 3.2 for translocations in stable cells. Since stochastic effects of radiation are attributable to genetic changes in viable cells, translocations in stable cells may be a better measure when considering the comparative risks of different qualities of radiation.     

Chromosome aberration frequencies produced by a 70-MeV proton beam

The effectiveness of a 70-MeV proton beam in the induction of chromosome aberrations was studied. We employed peripheral lymphocytes and analyzed the frequencies of dicentrics and rings after irradiation at doses ranging from 0.1 to 8.0 Gy at various depths within a Lucite phantom. The frequency of chromosome aberrations after irradiation with an unmodulated proton beam at 5 mm showed a dose-response relationship similar to that of 60Co gamma rays. However, irradiation at greater depths with the spread-out Bragg peak induced higher aberration frequencies at doses lower than those with gamma rays. Furthermore, the distribution curve of chromosome aberration frequencies as a function of depth was found to be slightly different from the physically measured depth-dose curve. With the spread-out Bragg peak the biological effects were more marked at greater depths, resulting in a distribution of relative biological effectiveness values. The results obtained from chromosome aberration analysis may not be related directly to those for the relationship between dose and cell killing. Slight differences in values for relative biological effectiveness due to the change of dose and site of proton beam irradiation may not be important for practical proton beam therapy, but may be important in the prevention of late radiation injuries.     

Mathematical analysis of genetic effects of low doses of ionizing radiation

The comparative study of effects of low doses of radiation on peripheral blood lymphocytes of persons occupationally exposed to radiation and non-exposed ones was carried out. The main attention was paid to radio-adaptive response forming under consistent exposure to low (0.05 Gy) and damaging (2 Gy) doses of gamma-irradiation. Noticeable heterogeneity in capacity for adaptive response forming in occupational group was revealed. The mathematical model adequate to experimental material was constructed using Kohonen neuronets.     

Quantitative characteristics of the frequency of chromosomal aberrations in a group of residents of a large western Siberian industrial region

The results of a 15-year study of chromosome aberration frequency in cultured peripheral blood lymphocytes from subjects living in the Kuzbass industrial region are presented. The database for the analysis of the main parameters of chromosome aberrations contains data on 925 subjects, with the total number of cells examined being 92,900. It has been found that the total frequency of aberrant metaphases in the database is 3.73 +/- 0.1%, whereas this frequency for the sample of subjects from industrial areas of this region (the basic control group) is significantly lower (2.86 +/- 0.26%). It has been demonstrated that the sex and age of the subjects do not affect substantially the frequency of any type of chromosome aberrations. Tobacco smoking is associated with a slight increase in chromosome damage frequency; however, the difference between smokers and nonsmokers is insignificant even among subjects exposed to occupational hazards. The possible causes that have determined the increased basic and background chromosome aberration frequencies in the population of the Kemerovo oblast compared to these parameters for the European part of Russia and the CIS are discussed.     

Genetic damage induced by in vitro irradiation of human G0 lymphocytes with low-energy protons (28 keV/microm): HPRT mutations and chromosome aberrations

Cell survival, mutations and chromosomal effects were studied in primary human lymphocytes exposed in G0 phase to a proton beam with an incident energy of 0.88 MeV (incident LET of 28 keV/microm) in the dose range 0.125-2 Gy. The curves for survival and mutations at the hypoxanthine-guanine phosphoribosyl transferase locus were obtained by fitting the experimental data to linear and linear-quadratic equations, respectively. In the dose interval 0-1.5 Gy, the alpha parameters of the curves were 0.42/Gy and 3.6 x 10(-6) mutants/Gy, respectively. The mutation types at the HPRT locus were analyzed by multiplex-PCR in 94 irradiated and 41 nonirradiated clones derived from T lymphocytes from five healthy donors. All clones showed a normal multiplex-PCR pattern and were classified as point mutations. Chromosome aberration data were fitted as a linear function of dose (alpha = 0.62 aberrations per cell Gy(-1)). By irradiating G0 lymphocytes from a single subject with 28 keV/microm protons and gamma rays, an RBE of 6.07 was obtained for chromosome aberrations. An overinvolvement of chromosome 9 relative to chromosome 7 was found in chromosome breaks after chromosome painting analysis.     

Quantitative Characteristics of Chromosome Aberration Frequency in the Human Population of a Large Western Siberian Industrial Region

The results of a 15-year study of chromosome aberration frequency in cultured peripheral blood lymphocytes from subjects living in the Kuzbass industrial region are presented. The database for the analysis of the main parameters of chromosome aberrations contains data on 925 subjects, with the total number of cells examined being 92 900. It has been found that the total frequency of aberrant metaphases in the database is 3.73 ± 0.1%, whereas this frequency for the sample of subjects from industrial areas of this region (the basic control group) is significantly lower (2.86 ± 0.26%). It has been demonstrated that the sex and age of the subjects do not affect substantially the frequency of any type of chromosome aberrations. Tobacco smoking is associated with a slight increase in chromosome damage frequency; however, the difference between smokers and nonsmokers is insignificant even among subjects exposed to occupational hazards. The possible causes that have determined the increased basic and background chromosome aberration frequencies in the population of the Kemerovo oblast compared to these parameters for the European part of Russia and the CIS are discussed.     

Reciprocal adaptive response of human peripheral lymphocytes induced by bleomycine or gamma rays.

The adaptive response and reciprocal adaptive response induced in vitro by exposure to low doses of gamma rays (0.05 Gy) or bleomycin (0.05 microg/ml) in human peripheral blood lymphocytes were assessed by the frequency of chromosome aberrations. Gamma rays (1.5 Gy) or bleomycin (1.5 microg/ml) were used as the challenge doses. In the experiments, blood samples from 5 healthy donors were investigated. It has been found that low doses of bleomycin and gamma rays induced a reciprocal adaptive response to high doses of gamma rays or bleomycin. Moreover, the results confirmed that the adaptive response did not correlate with the radiosensitivity of the peripheral blood lymphocytes.     

Biological effectiveness of neutrons from Hiroshima bomb replica: results of a collaborative cytogenetic study

The effectiveness of neutrons from a facsimile of the Hiroshima bomb was determined cytogenetically. The "Little-Boy" replica (LBR), assembled at Los Alamos as a controlled nuclear reactor for detailed physical dosimetry, was used. Of special interest, the neutron energy characteristics (including lineal energy) measured 0.74 m from the LBR were remarkably similar to those calculated for the 1945 Hiroshima bomb at 1 to 2 km from the hypocenter, as shown in a companion dosimetric paper (Straume, et al., Radiat. Res. 128, 133-142 (1991)). Thus we examine here the effectiveness of neutrons closely resembling those that the A-bomb survivors received at Hiroshima. Chromosome aberration frequencies were determined in human blood lymphocytes exposed in vitro to graded doses of LBR radiation (97% neutrons, 3% gamma rays). Vials of blood suspended in air at distances up to 2.10 m from the center of the LBR uranium core received doses ranging from 0.02 to 2.92 Gy. The LBR neutrons (E approximately 0.2 MeV) produced 1.18 dicentrics and rings per cell per Gy. They were more effective than the higher-energy fission neutrons (E approximately 1 MeV) commonly used in radiobiology. The maximum RBE (RBEM) of LBR neutrons at low doses is estimated to be 60 to 80 compared to 60Co gamma rays and 22 to 30 compared to 250-kVp X rays. These results provide a quantitative measurement of the biological effectiveness of Hiroshima-like neutrons.     

The cytogenetic effects of low doses of accelerated charged particles in human blood lymphocytes in vitro

The aim of the investigation was the study of cytogenetic effects in human blood lymphocytes of low doses of ionizing radiation in vitro. The analysis of unstable chromosome aberrations in human lymphocytes after irradiation by the accelerated ions 12C with the energy 500 MeV/nucleon and LET 10.7 keV/microm was carried out. Blood samples were irradiated on Nuclotron of the High Energy Laboratory of the Joint Institute for Nuclear Research. The doses of irradiation were in the range from 0.05 up to 1.0 Gy. Was shown that the frequency of unstable chromosome aberrations depends from the dose of ionizing radiation and can be described by linear function. At the doses 0.25-0.50 Gy the dose-independent curve was obtained for dicentrics and centric rings. The frequencies of dicentrics and centric rings as markers of the radiation action were slightly different for different donors that could be explained by different radiosensitivity. Using the calibration curve obtained earlier for gamma-rays coefficients of relative biological efficiency of accelerated 12C with the energy 500 MeV/nucleon were defined: they varied from 1.0 at the doses (0.5-1.0 Gy) up to 3.2 at the lower doses (0.05-0.25 Gy).     

The intensity of proteolysis in the peripheral blood lymphocytes of rats in the early period of acute radiation injury

Comparative analysis of biochemical indices that reflect the proteolysis rate in rat peripheral blood lymphocytes in normal conditions and during the first 4-6 h after whole-body single uniform gamma-irradiation with lethal (7.5 Gy) and nonlethal (3 Gy) doses has revealed that the pronounced activation of proteolysis in lymphocytes and the disturbances in the inhibitor/protease balance are characteristic of the severe form of radiation damage with fatal outcome. Simultaneously changes occur in biochemical indices of plasma that reflect a catabolic pathway of metabolism in the whole-body. The absence of similar changes in the blood of the survived animals permits to recommend the biochemical indices under study to be used for early prognosis of the radiation affection outcome.     

Choice of model and uncertainties of the gamma-ray and neutron dosimetry in relation to the chromosome aberrations data in Hiroshima and Nagasaki

Chromosome data pertaining to blood samples from 1,703 survivors of the Hiroshima and Nagasaki A-bombs, were utilized and different models for chromosome aberration dose response investigated. Models applied included those linear or linear-quadratic in equivalent dose. Models in which neutron and gamma doses were treated separately (LQ-L model) were also used, which included either the use of a low-dose limiting value for the relative biological effectiveness (RBE) of neutrons of R(0)=70+/-10 or an RBE value of R(1)=15+/-5 at 1 Gy. The use of R(1) incorporates the assumption that it is much better known than R(0), with much less associated uncertainty. In addition, error-reducing transformations were included which were found to result in a 50% reduction of the standard error associated with one of the model fit parameters which is associated with the proportion of cells with at least one aberration, at 1 Gy gamma dose. Several justifiable modifications to the DS86 doses according to recent nuclear retrospective dosimetry measurements were also investigated. Gamma-dose modifications were based on published thermoluminescence measurements of quartz samples from Hiroshima and on a tentative reduction for Nagasaki factory worker candidates by a factor of 0.6. Neutron doses in Hiroshima were modified to become consistent with recent fast neutron activation data based on copper samples. The applied dose modifications result in an increase in non-linearity of the dose-response curve for Hiroshima, and a corresponding decrease in that for Nagasaki, an effect found to be most pronounced for the LQ-L models investigated. As a result the difference in the dose-response curves observed for both cities based on DS86 doses, is somewhat reduced but cannot be entirely explained by the dose modifications applied. The extent to which the neutrons contribute to chromosome aberration induction in Hiroshima depends significantly on the model used. The LQ-L model including an R(1) value of 15 at 1 Gy which is recommended here, would predict between 10% and 20% of the observed chromosome aberrations to be due to neutrons, at all doses. Because of the good agreement between DS86 predictions and the results of retrospective gamma and neutron dosimetry, the modifications applied here to DS86 doses are relatively small. Consequently, the choices of model and RBE values were found to be the major factors dominating the interpretation of the chromosome data for Hiroshima and Nagasaki, with the dose modifications resulting in a smaller influence.     

Comparative study of chromosome aberration formation in lymphocytes culture under pulsed and continuous neutron irradiation

Frequencies of chromosome aberration induced by prolong (continuous) neutron radiation (dose-rate 0.17 Gy/min) and pulsed neutron radiation with ultra-high dose-rate (1-4) x 10(5) Gy/s have been studied in human blood lymphocytes at G0-stage. It was demonstrated that cytogenetic efficiency of pulsed neutrons (after the substraction of approximately 50% gamma-component from the total dose) was 2 times higher than that of continuous neutron radiation.     

Experimental derivation of relative biological effectiveness of A-bomb neutrons in Hiroshima and Nagasaki and implications for risk assessment

Epidemiological data on the health effects of A-bomb radiation in Hiroshima and Nagasaki provide the framework for setting limits for radiation risk and radiological protection. However, uncertainty remains in the equivalent dose, because it is generally believed that direct derivation of the relative biological effectiveness (RBE) of neutrons from the epidemiological data on the survivors is difficult. To solve this problem, an alternative approach has been taken. The RBE of polyenergetic neutrons was determined for chromosome aberration formation in human lymphocytes irradiated in vitro, compared with published data for tumor induction in experimental animals, and validated using epidemiological data from A-bomb survivors. The RBE of fission neutrons was dependent on dose but was independent of the energy spectrum. The same RBE regimen was observed for lymphocyte chromosome aberrations and tumors in mice and rats. Used as a weighting factor for A-bomb survivors, this RBE system was superior in eliminating the city difference in chromosome aberration frequencies and cancer mortality. The revision of the equivalent dose of A-bomb radiation using DS02 weighted by this RBE system reduces the cancer risk by a factor of 0.7 compared with the current estimates using DS86, with neutrons weighted by a constant RBE of 10.     

Quantitative and qualitative changes in the peripheral blood cells of rats in chronic irradiation of the animals

A study was made of qualitative and quantitative changes in peripheral blood lymphocytes of rats subjected to continuous totalbody gamma-irradiation at dose--rates from 0.002 to 0.35 Gy/day and cumulative doses from 0.7 to 20.0 Gy. After termination of irradiation (on the 400th-500th day), the number of lymphocytes changed in an undulatory manner, i. e., increased at a dose--rate of 0.008 Gy/day, and decreased down to minimum control values at a dose--rate of 0.35 Gy/day, and the fraction of young macrocellular forms increased. The effect increased with increase in dose--rate and cumulative dose.     

The Chernobyl fallout in Salzburg/Austria and its effect on blood chromosomes

The radioactive fallout of the Chernobyl accident caused an increase in radiation dose of 20 to 110 per cent over the normal environmental burden to the inhabitans of Salzburg City in Austria (in a distance of about 1300 km from the accident). The structural chromosome aberration in the lymphocytes of the peripheral blood of 15 test-persons have been investigated one year after the accident. From two of these we know also the aberration frequencies before the accident which were significantly lower. The results from all test persons were pooled according to their Cs137 and Cs134 content, measured by whole body counter. Their mean additional blood doses from the incorporated caesium plus the external fallout radiation were 0.23, 0.36 and 0.55 mGy/yr. The aberration frequencies increased with dose. The slope of the best straight-line through the points was 2.0 +/- 0.7 total chromosome type aberrations in 100 metaphases per mGy/yr. This result fits in well with former investigations of persons with individually calculated radiation burden from the environment. The sharp increase with dose at this low level is not compatible with values extrapolated from high doses. The usual dose assessment based on chromosome aberrations extrapolated from high to low doses is therefore not possible in the range considered in this investigation.     

Retrospective dose estimation in remote period after exposure using different biological methods

Investigation of application of chromosome aberrations of lymphocytes in peripheral blood for biological dosimetry purposes in remote (up to 40 years) period after acute exposure to doses of 1 Gy and more was carried out. The comparative analysis of frequency of unstable and stable (using FISH and G-banding methods) aberrations was performed for 24 subjects accidentally exposed to radiation on nuclear submarines during 1961-1985. Statistically significant increasing of frequency of dicentrics and centric rings was determined in the exposed subjects in remote period after exposure to compare with controls. Their sum frequency in the exposed group varied depending on ARS heaviness from 0.1 to 1.0 aberrations per 100 cells. In control group it was from 0 to 0.2 correspondingly. Translocation frequency (complete + incomplete) fixed by FISH method (2, 4, and 12 chromosomes) varied within the limits of 0.2-16.0 for exposed subjects and 0.3-1.26 translocations per genome per 100 cells for controls. Some examined persons (5 subjects) exposed to accident in 1985 had results of analysis of unstable chromosome aberration in acute period after exposure that allow to estimate obtained doses by dicentrics frequency which having good correlation with ARS heaviness. Individual dosed using traslocation frequency were defined retrospectively in 11 from 21 exposed persons. They correlate with calculated physics doses and doses estimated by haematolotical parameters in acute period and also doses obtained by ESR spectroscopy of tooth enamel in remote period.