Long-term follow-up cytogenetic survey and biological dosimetry in persons evacuated from 30-km Chernobyl NPP zone

The paper presents the results of the follow-up cytogenetic survey and biological dosimetry carried out in inhabitants of Pripiat' town and nearby villages, who were departured from the Chernobyl NPP 30-km exclusive zone during first days after the Chernobyl catastrophe. The unstable chromosome aberration level in inhabitants were significantly increased above control in terms up to 1 year after evacuation and declined gardually during next 14 years. In early period the cytogenetic damage frequency in evacuees showed no dependence on gender. The chromosome type aberration level appeared to be lower in young persons comparing with adults. The dicentrics plus centric rings yield had a positive correlation with duration of staying at Chernobyl zone. The average doses of protracted exposure were calculated from the dicentrics and centric rings yields; the dose estimations appeared to be 1.4 times higher in persons evacuated 3-11 days after the accident than that of in persons with shorter departure time. Uing the Bayesian analysis the probabilistic distribution of biological doses was constructed for the studied evacuees group. This distribution was characterized by a mean dose of 360 mGy, the modal doses of 200-450 mGy and 80% of probability density within the dose range 0-1000 mGy, that seems to be sufficient for considering the increased risk of late somatic radiation effects for this cohort.     

Cytogenetic effect in peripheral blood lymphocytes of children living in villages in the Ovruch District, Zhitomir Region of the Ukraine contaminated by radionuclides]

For the purpose of genetic indication of low level of chronic radiation exposure the cytogenetic monitoring of some critical children groups living in two contaminated Ukrainian villages was carried out. In all the groups the mean frequency of aberrant cells and chromosome type aberrations (including dicentrics, centric rings, chromosome translocations as well as polyploid cells) significantly exceeded control level. During the repeated examination of children from Vistupovichi (in 13 months after the first one) the striking increase of cytogenetic effect was revealed. Dicentrics and rings were registered in 79% of persons with the individual rate 0.5-1.5 and mean group frequency 0.57 per 100 cells. The authors tried to evaluate the average cumulative doses of radiation for examined groups using G. Littlefield equation for dicentric outcome under the low dose rate Cs-137 source and taking into account the assumption about the reducing of 50% dicentrics per year. According to this rough calculation, the revealed cytogenetic effect can correspond to the mean total dose of 33 cSv for Vistupovichi children.     

The role of cytogenetic examination for prognosis of remote consequences of irradiation

Correlation between the level of somatic pathology and cytogenetic characteristics of blood was analyzed in a group of liquidators of the accident at the Chernobyl Nuclear Power Plant (ChNPP). A statistically significant correlation was found between the occurrence of cardiovascular diseases and the level of chromosome aberrations (total frequency of chromosome aberrations, frequency of dicentrics and centric rings, frequency of chromatid aberrations). The results obtained are of great importance for the substantiated prediction of the development of postradiation pathologies.     

The low dose and low dose rate cytogenetic effects induced by gamma-radiation in human blood lymphocytes in vitro. II. the results of cytogenetic study

The yield of chromosome aberrations induced by gamma-radiation of 60Co in human blood lymphocytes in vitro at low doses (30 divided by 600 mGy) and low dose rates (0.70, 5.05, 59.2 mGy/min) was investigated. It was found that the observed level of chromosomal aberrations induced by gamma-irradiation was unaffected by the value of the dose rate when using constant dose rate and obtaining different doses by altering the exposure time. However, a relatively enhanced level of chromatid aberrations was found at 5.05 and 59.2 mGy/min dose rates in the dose range less than 250 mGy. We have found that the observed level of the sum of chromosomal aberrations induced by gamma-irradiation at doses less than 250 mGy and a dose rate of 59.2 mGy/min was essentially larger compared with the level extrapolated from high doses (above 300 mGy) using a linear-quadratic dose curve. This complied with our previous finding in 1976, 1977 when the enhanced level of dicentrics was only found at a high dose rate approximately 500 mGy/min. Such a non-linear cytogenetic effect does not manifest itself statistically significantly at dose rates of 0.70 and 5.05 mGy/min for the sum of chromosomal aberrations and does not manifest itself at all for dicentrics at all the examined dose rates.     

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 retrospective cytogenetic dosimetry using the results of conventional chromosomal analysis in Chernobyl clean-up workers

The paper presents the results of the cohortal biodosimetry carried out in 435 Chernobyl clean-up workers, who were surveyed with the conventional cytogenetic technique in terms from several days to 10 years after the end of their duties in the Chernobyl accident exclusive zone. An empirical model of the aberrant cell dynamics was utilized for the calculation of mean initial yields of dicentrics and centric rings in groups with different terms and duration of staying in the Chernobyl zone. Corresponding protracted irradiation doses estimated from aberration levels ranged from 79 to 670 mGy. The probabilistic distribution of the radiation doses was constructed by the applying the Bayesian analysis to initial individual chromosome exchange yields extrapolated to the exposure termination moment. This distribution was characterized by the mean dose about 460 mGy and maximum of probability density in the interval of 50-300 mGy. For the late somatic risk assessment in clean-up workers the probabilistic distribution of equivalentally-acute radiation doses was proposed; that had the mean value about 270 mGy, modal classes of 250-350 mGy and 99.8% of the probability density concentrated within the dose range from 0 to 1000 mGy.     

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.     

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.     

Results of cytogenetic studies of the consequences of the Chernobyl accident

The selected results of the cytogenetic studies of the Chernobyl accident consequences were summarised. The chromosomal aberrations were used as a method of biodosimetry for a dose assessment for victims during the initial period after the Chernobyl accident. A good correlation between doses calculated based on chromosomal aberrations (dicentrics) and severity of acute radiation syndrome observed in clinic was found. The biodosimetry based on conventional cytogenetic technique (dicentrics) has been unsuccessful for various groups (rehabilitation workers, evacuees, inhabitants of contaminated areas) sampled long time after the Chernobyl accident. The possible reasons of the failure are analysed. The original results of multiaberration cell yield observed in different cohorts of the Chernobyl victims are presented. The problems related to the phenomena are discussed.     

Biological dosimetry in a group of radiologists by the analysis of dicentrics and translocations

The results of a cytogenetic study carried out in a group of nine radiologists are presented. Chromosome aberrations were detected by fluorescence plus Giemsa staining and fluorescence in situ hybridization. Dose estimates were obtained by extrapolating the yield of dicentrics and translocations to their respective dose-effect curves. In seven individuals, the 95% confidence limits of the doses estimated by dicentrics did not include 0 Gy. The 99 dicentrics observed in 17,626 cells gave a collective estimated dose of 115 mGy (95% confidence limits 73-171). For translocations, five individuals had estimated doses that were clearly higher than the total accumulated recorded dose. The 82 total apparently simple translocations observed in 9722 cells gave a collective estimated dose of 275 mGy (132-496). The mean genomic frequencies (x100 +/- SE) of complete and total apparently simple translocations observed in the group of radiologists (1.91 +/- 0.30 and 2.67 +/- 0.34, respectively) were significantly higher than those observed in a matched control group (0.53 +/- 0.10 and 0.87 +/- 0.13, P < 0.01 in both cases) and in another occupationally exposed matched group (0.79 +/- 0.12 and 1.14 +/-0.14, P < 0.03 and P < 0.01, respectively). The discrepancies observed between the physically recorded doses and the biologically estimated doses indicate that the radiologists did not always wear their dosimeters or that the dosimeters were not always in the radiation field.     

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.     

Chromosome aberration frequencies and chromosome instability in mice after long-term exposure to low-dose-rate gamma-irradiation

Chronological changes of chromosome aberration rates related to accumulated doses in chronically exposed humans and animals at a low-dose-rate have not been well studied. C3H female specific pathogen-free mice (8 weeks of age) were chronically irradiated. Chromosome aberration rate in mouse splenocytes after long-term exposure to low-dose-rate (LDR) gamma-rays was serially determined by conventional Giemsa method. Incidence of dicentrics and centric rings increased almost linearly up to 8000 mGy following irradiation for about 400 days at a LDR of 20 mGy/day. Clear dose-rate effects were observed in the chromosome aberration frequencies between dose rates of 20 mGy/day and 200 Gy/day. Furthermore, the frequencies of complex aberrations increased as accumulated doses increased in LDR irradiation. This trend was also observed for the incidences of micronuclei and trisomies of chromosomes 5, 13 and 18 in splenocytes, detected by micronucleus assay and metaphase fluorescence in situ hybridization (FISH) method, respectively. Incidences of 2-4 micronuclei and trisomy increased in mouse splenocytes after irradiation of 8000 mGy at a LDR of 20 mGy/day. These complex chromosome aberrations and numerical chromosome aberrations seem to be induced indirectly after radiation exposure and thus the results indicate that continuous gamma-ray irradiation for 400 days at LDR of 20 mGy/day induced chromosomal instability in mice. These results are important to evaluate the biological effects of long-term exposure to LDR radiation in humans.     

Chromosome alterations in cleanup workers sampled years after the Chernobyl accident

Cytogenetic analysis performed 4-13 years after the Chernobyl accident showed an elevated frequency of acentrics, chromatid exchanges, dicentrics and rings in Chernobyl cleanup workers compared to the control group. Cytogenetic data were analyzed according to the information on exposure to radiation (the year when the cleanup workers worked at the Chernobyl station, doses rates, time elapsed since exposure to radiation, and cytogenetic examination) and some lifestyle factors. The data obtained suggested that some types of chromosome aberrations could be influenced by the action of different environmental factors or lifestyle factors. The frequency of acentrics was correlated with the age of the cleanup workers, and the increased chromatid exchange frequency was attributed to smoking. The numbers of dicentrics and rings suggested a genotoxic effect of ionizing radiation that is still present over 13 years after the exposure.     

Genotype dependence of cytogenetic and epidemiological characteristics in the liquidators of the accident at the ChNPP

The dependence of the level of unstable chromosome aberrations and nononcological diseases on the genotype in 57 liquidators of the ChNPP accident was studied. Candidate genes presumably affecting radiosensitivity were highly polymorphic loci of xenobiotic detoxication genes (glutathione-S-transferases GSTM1, GSTT1, GSTP1) and the 5,10-methylenetetrahydrofolate reductase gene (MTHFR) involved in DNA methylation and synthesis. An increased frequency (0.014 +/- 0.001 per cell) of unstable chromosome aberrations, including radiation-specific dicentrics and centric rings (0.0015 +/- 0.0002 per cell), has been found to be preserved in the group of liquidators examined in 2006-2007. No associations of polymorphism for each of the studied genes with cytogenetic parameters were revealed. Increased frequencies of chromosome aberrations were recorded in homozygous carriers of a deletion at the GSTM1 locus in combination with homozygosity for minor alleles at the MTHFR and GSTP1 loci (p = 0.00002 and p = 0.0233, respectively). The number of homozygous carriers of the minor allele GSTP1 was increased among patients with chronic obstructive pulmonary disease and in liquidators with acute circulation disturbances (p = 0.014 and p = 0.04, respectively). Double homozygotes for GSTM1 and GSTT1 deletions were significantly more frequent among subjects with benign tumors (cysts, polyps, p = 0.015) and with benign thyroid tumors (p = 0.017). This genotype has proved to be protective for patients with severe cardiovascular diseases (acute circulation disturbances, p = 0.027).     

Chromosomal damage induced in human lymphocytes by low doses of D-T neutrons

Unstable chromosome aberrations induced by in vitro irradiation with zero plus seven low doses of 14.8 MeV D-T neutrons in the range 3.55-244 mGy have been analysed in human peripheral blood lymphocytes. In order to obtain the required large numbers of scored cells for such low doses, fourteen laboratories participated in the experiment. The dose responses for dicentrics, excess acentrics and total aberrations, fitted well to the Y = alpha D model. The alpha coefficient of yield for dicentrics, 1.60 +/- 0.07 X 10(-2) Gy-1, compares well with the values obtained in previous studies with D-T neutrons at somewhat higher doses. Results from a previous collaborative study using 250 kVp X-rays over a comparable dose range indicated the possible existence of a threshold below 50 mGy. In the present study there is no clear evidence for neutrons for such a threshold. However, the data were insufficient to permit the rejection of a possible threshold below approximately 10 mGy.     

The dose response of chromosome aberrations in human lymphocytes induced in vitro by very low-dose γ rays

This paper considers the dose-effect relationship for unstable chromosome aberration yields in human lymphocytes in very low-dose range. Data are presented for (60)Co γ-ray doses of 0, 10, 20, 40 and 1000 mGy. More than 5,000 metaphases were scored for each data point at the very low doses, and each cell was double-checked using a semi-automated metaphase finding/relocation system. Aberration yields of dicentrics plus centric rings followed an excellent linear dose response down to zero dose; the yields were significantly above the control frequency from 20 mGy.     

The significance of cytogenetic investigation for the estimation of Chernobyl accident consequences

The results of the cytogenetic investigation of people, which were exposed to radiation in the result of the Chernobyl NPP accident, were presented. Also the possibilities of the application of cytogenetic findings for dose estimations and for the prediction of the radiation influence consequences were examined. During the period of time since 1986 till 2004 the cytogenetic investigations of 1724 liquidators participating in the liquidation works after the Chernobyl accident were carried out. The radiation dose estimated by the frequency of dicentrics in 1986 was about 0.16 Gy. The doses for liquidators were determined by the frequency of translocations (FISH method) during the period from 1992 till 1995. For liquidators who worked in Chernobyl only in 1986 the average dose of radiation was about 0.19 Gy and for liquidators who worked repeatedly during the period from 1986 till 1995 - 0.39 Gy. There was shown that during the whole period of investigation (1986-2004) the frequency of dicentrics in peripheral blood lymphocytes was significantly higher than the control level. The cytogenetic investigation of Bryansk region inhabitants which was carried out in 1992-1994 discovered heightened value in 5 times than the control one. Findings are of great importance for the prediction of ill effects of radiation and for the development of sensitive criterions for early exposure disturbances in state of health.     

Results of a cytogenetic study of populations with different radiation risks in the Semipalatinsk region

A cytogenetic study was conducted for the first time on human populations neighboring the Semipalatinsk nuclear test site (STS) and exposed to ionizing radiation for a long period of time. In populations with the extreme and maximum radiation risks, high frequencies of radiation-induced chromosomal markers, including acentric fragments (1.99 +/- 0.10 per 100 cells), dicentrics (0.23 +/- 0.01), ring chromosomes (0.38 +/- 0.14), and stable chromosomal aberrations (1.17 +/- 0.02), were found. These frequencies significantly exceeded those in control populations. The spectrum of chromosomal aberrations and the frequencies of the aberrations of different types in persons living in the areas with the highest radionuclide contamination confirmed the mutagenic effect of radiation on chromosomes in the human populations studied.     

The ratio of dicentrics to centric rings produced in human lymphocytes by acute low-LET radiation.

Chromosome aberrations produced by ionizing radiation are assumed to develop from DNA double-strand breaks (DSBs) which interact pairwise, in an exchange event. Dicentrics and centric rings are aberrations that exemplify inter- and intrachromosomal exchanges, respectively. We show from a survey of published data that for acute low-LET irradiation of resting human lymphocytes the observed ratio of dicentrics to centric rings is approximately five times smaller than predicted by a pairwise interaction model which assumes complete randomness. Such a low ratio can be interpreted as evidence for a proximity effect, favoring exchanges of an intrachromosomal type. That is, since DSBs induced close together have an above-average chance of pairwise interaction, the observed excess of centric rings indicates that at the time of irradiation there is some degree of spatial confinement for the two arms of a single chromosome. Assuming the excess of centric rings is indeed due to proximity effects, the data are used to estimate that the volume of a domain, within which any one lymphocyte chromosome is localized at one instant during the G0/G1 phase, is at most approximately 20% of the nuclear volume.     

Possibilities and limitations of fluorescence in situ hybridization technique in retrospective detection of low dose radiation exposure in post-chernobyl human cohorts

Cytogenetic analysis using the fluorescence in situ hybridisation (FISH) technique was performed late time after the Chernobyl accident in groups of liquidators, evacuees from 30 km exclusive zone, residents of radioactively contaminated areas and control donors age-matched to exposed persons. Stable and unstable chromosome type exchanges were recorded using a hybrid conventional-PAINT nomenclature. The mean yield of stable chromosome exchanges in liquidators did not correlate with registered radiation doses but had a clear negative dependence on the duration of liquidators' staying in Chernobyl zone, that was in a good agreement with early data based on conventional dicentrics plus rings analysis. The overspontaneous excess for stable chromosome exchange level appeared to be higher in evacuees 16-40 years old than that of senior persons, whereas no age-dependent difference occurred for initially induced dicentrics plus rings yields in this cohort. The stable chromosome exchange yield, as well as combined yield of dicentrics plus rings and potentially unstable incomplete translocations in residents of radioactively contaminated areas showed a reasonable positive correlation with levels of 137Cs contamination. The observed yields of stable chromosome exchanges in all three exposed groups appeared to be somewhat lower than those of expected from unstable exchange-based doses which were referred to an in vitro dose response of stable exchanges outcome in human lymphocytes. Thus, FISH analysis can be successfully applied for qualitative cytogenetic indication of past and chronic radiation exposure to low doses but further refinement of FISH-based system for quantitative dose assessment is still required. Some practical approaches of solving this task are discussed.