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Purpose: Plotting dose-effect curves for translocations identified using the tricolor FISH method based on the results of cytogenetic analysis of cultures of peripheral blood lymphocytes of healthy donors after in vitro gamma irradiation. Material and methods: Venous blood was obtained from three donors (2 men and 1 woman aged from 28 to 41 years) and subjected to in vitro gamma irradiation from a 60Co source at doses of 0.10; 0.15; 0.25; 0.35; 0.50; 0.75; 1.00; 1.50; 2.00 and 3.00 Gy at 37 ° C (dose rate 0.5 Gy / min). For tricolor FISH staining, two different sets of DNA probes were used for chromosome pairs 1, 4, 12 and 2, 3, 8. Metaphases with a quasi-diploid number of chromosomes (40-46) and a complete set of all FISH-stained chromosomes, taking into account their total length, were selected for analysis. Differentiation of stable and unstable cells was also carried out. In the cytogenetic analysis, traditional terminology was used with the designation of translocations as reciprocal (complete, two-sided), non-reciprocal (terminal, incomplete, or unilateral), or interstitial. Results: The obtained numerical data were used to statistically compare the frequencies of FISH-recorded translocations when using different sets of DNA probes, when calculating of chromosome aberrations were in all (unstable and stable) and stable metaphase cells, when comparing of the frequencies of FISH-recorded translocations and dicentrics, and assessing of the contribution of the level of translocations between FISH-stained chromosome pairs in the total translocation frequency. The plotted dose-effect curves generally corresponded to the linear-quadratic form. Conclusion: Dose dependences obtained for translocations using two different selected tricolor sets of DNA probes did not differ statistically significantly. At the same time, cytogenetic analysis of only stable metaphase cells revealed a tendency to register lower levels of translocations than when analyzing all cells (unstable and stable ), at the highest doses of 2 and 3 Gy. The levels of dicentrics formed with the participation of FISH-stained chromosomes were significantly lower than the number of observed translocations. The quantitative contribution of translocations between FISH-stained pairs of chromosomes turned out to be very low, which clearly does not contribute to an increase in the sensitivity of the FISH method of retrospective dose estimation as compared to its one-color version. At the same time, the three-color FISH-staining makes it possible to identify such variants of chromosomal rearrangements that are not recorded using the one-color FISH method.

Purpose To evaluate radiation induced chromosome aberration frequency in peripheral blood lymphocytes of cosmonauts who participated in flights on Mir Orbital Station and ISS (International Space Station). Materials and methods Cytogenetic examination which has been performed in the period 1992–2008 included the analysis of chromosome aberrations using conventional Giemsa staining method in 202 blood samples from 48 cosmonauts who participated in flights on Mir Orbital Station and ISS. Results Space flights led to an increase of chromosome aberration frequency. Frequency of dicentrics plus centric rings (Dic+Rc) depend on the space flight duration and accumulated dose value. After the change of space stations (from Mir Orbital Station to ISS) the radiation load of cosmonauts based on data of cytogenetic examination decreased. Extravehicular activity also adds to chromosome aberration frequency in cosmonauts’ blood lymphocytes. Average doses after the first flight, estimated by the frequency of Dic+Rc, were 227 and 113 mGy Eq for long-term flights (LTF) and 107 and 53 mGy Eq for short-term flights (STF). Conclusion Cytogenetic examination of cosmonauts can be applied to assess equivalent doses.

To investigate within the framework of a multilaboratory study the suitability of FISH chromosome painting to measure so-called stable translocations in peripheral lymphocytes of Mayak nuclear-industrial workers (from the Southern Urals) and their use for retrospective biodosimetry.Chromosime analyses were carried out from 69 workers who had received protracted occupational radiation exposures (0.012-6.065 Gy) up to approximately 40 years before blood sampling. Twenty-one unexposed people living in the same area were controls. A multicolour FISH-painting protocol with the target chromosomes 1, 4 and 8 simultaneously with a pancentromeric probe was used to score potentially transmissible chromosome-type aberrations (reciprocal translocations 2B and related 'one-way' patterns I-III according to the SS classification).Individual biodosimetry estimates were obtained in terms of these potentially long-term surviving aberration types based on the linear component of a low dose-rate gamma-ray calibration curve produced using identical staining and scoring protocols. For comparison, the workers personal and total background doses were converted to red bone marrow doses. The estimated doses were mainly lower than would be predicted by the calibration curve, particularly at accumulated higher dose levels.Owing to the limited life-time of circulating T-lymphocytes, the long-term persistence of translocations in vivo requires the assumption of a clonal repopulation of these naturally senescing cells from the haemopoietic stem cell compartments. Obviously such a replacement cannot be fully achieved, leading to a temporal decline even of the yield of transmissible aberrations types. Assuming further a highly selective capacity of stem cells against any type of chromosomal damage and the fact that one must rely on partial genome findings, the potential of FISH chromosome painting for retrospective dose reconstruction is probably limited to a decade or so after high-level protracted radiation exposure.

Analysis of chromosome aberrations in lymphocytes of peripheral blood has been long successfully used in radiation researches for biological indication and quantitative estimation of radiation influence on a human organism. “Biological doses” calculated on the basis of cytogenetic research data take into account individual radiosensitivity of the organism and its state at the moment of radiation, and this, in its turn, allows to forecast early and more distant consequences of radiation more precisely. Frequency of chromosome aberrations in lymphocytes of peripheral blood is also one of the most frequently used indices in studying the process of mutagenesis in somatic cells. There are data on the possible connection between the increased level of chromosome aberrations in lymphocytes of peripheral blood and the risk of development of oncological diseases. That’s why the frequency of chromosome aberrations in lymphocytes of peripheral blood may be regarded as an integral biological marker of the risk of development of pathological states, first of all, oncological diseases. On the one hand this marker reflects in the influence of genotoxic factor on a human organism, and, on the other hand, individual diathesis to cancer genesis. That’s why it is considered extremely important to carry out cytogenetic examination of people exposed to radiation as a result of extraordinary and emergency situations. Results of the researches allow to estimate the influence of radiation on the genome of a human being, as well as forecast the risk of development of unfavorable consequences of radiation. Results of long-term cytogenetic monitoring of the Chernobyl clean-up workers (liquidators) and the results of cytogenetic examination of the cosmonauts who took part in long flight at “Mir” and ICS stations will be presented in the report. Summing up the data received one should note the following: - During the whole period of research (for more than 20 years) the average frequency of chromosome aberrations in lymphocytes of peripheral blood of the liquidators was significantly higher than the control level. The frequency of dicentrics and centric rings – markers of radiation was higher than the analogous indices in the control group 3,5 – 7 times depending on the period of work and staying in the zone of Chernobyl accident. - Long space flights lead to increasing chromosome aberrations in lymphocytes of cosmonaut's peripheral blood and the frequency of dicentrics and centric rings depends on the duration of a space flight and the amount of the accumulated dose of radiation.

Background and aims: Cosmic radiation is one of the main hazards for manned space exploration. Uncertainty in radiation risk estimates for crews of long-term missions are very high, and direct biological measurements are necessary. We measured chromosomal aberrations in peripheral blood lymphocytes from 33 cosmonauts involved in space missions during the past 11 years. Methods: Blood lymphocytes from the cosmonauts were stimulated to grow in vitro and were harvested at their first mitosis. Slides were either stained with Giemsa stain for dicentrics analysis, or painted with whole-chromosome DNA probes for translocation analysis (FISH). Results: A statistically significant increase in the yield of chromosomal aberrations was measured following long-term space missions in lymphocytes from cosmonauts at their first flight. No significant changes in aberration frequencies were observed for short-term taxi flights. The increase in long-term missions was consistent with the values calculated from physical dosimetry data. However, for cosmonauts involved in two or more space flights, the yield of interchromosomal exchanges was not related to the total duration of space sojourn or integral absorbed dose. Indeed, the yield of aberrations at the end of the last mission was generally in the range of background frequencies measured before the first mission. Conclusions: Chromosome aberration dosimetry can detect radiation damage during space flight, and biological measurements support the current risk estimates for space radiation exposure. However, for cosmonauts involved in multiple space missions the frequency of chromosomal aberrations is lower than expected, suggesting that the effects of repeated space flights on this particular endpoint are not simply additive. Changes in the immune system in microgravity and/or adaptive response to space radiation may explain the apparent increase in radioresistance after multiple space flights.

Large uncertainties are associated with estimates of equivalent dose and cancer risk for crews of longterm space missions. Biological dosimetry in astronauts is emerging as a useful technique to compare predictions based on quality factors and risk coefficients with actual measurements of biological damage in-flight. In the present study, chromosomal aberrations were analyzed in one Italian and eight Russian cosmonauts following missions of different duration on the MIR and the international space station (ISS). We used the technique of fluorescence in situ hybridization (FISH) to visualize translocations in chromosomes 1 and 2. In some cases, an increase in chromosome damage was observed after flight, but no correlation could be found between chromosome damage and flight history, in terms of number of flights at the time of sampling, duration in space and extra-vehicular activity. Blood samples from one of the cosmonauts were exposed in vitro to 6 MeV X-rays both before and after the flight. An enhancement in radiosensitivity induced by the spaceflight was observed.

Measurements of symmetrical translocations by fluorescence in situ hybridization (FISH) were performed for retrospective biodosimetry in a Techa River population exposed to external (gamma-rays) and internal (90Sr, 137Cs) irradiation.Chromosome analyses were carried out on peripheral lymphocytes from 73 radiation-exposed residents from settlements along the Techa River (Southern Urals, Russia) located 7-148 km downstream from the site of release of liquid radioactive waste from the plutonium production facility Mayak. Thirty-nine unexposed persons from non-contaminated areas were used as controls. Whole-chromosome painting probes for chromosomes 1, 4 and 12 were used simultaneously with a pancentromeric probe.A significantly elevated mean translocation frequency compared with controls was found for the total study group and for either of two subgroups of inhabitants residing in villages of the upper regions of the Techa River (7-60 km) during 1950 to 1951, or in villages of the lower regions (78-148 km) until the time of blood sampling. Within the first subgroup, subjects born between 1937 and 1949 showed higher translocation frequencies than those born between 1914 and 1936. Collective biodosimetry estimates for the various groups were between 0.24 and 0.54 Gy. Individual dose estimates for seven subjects with at least five translocations ranged between 0.77 and 1.80 Gy and compared well with doses reconstructed on the basis of 90Sr whole-body counts (WBC) and electronic paramagnetic resonance (EPR) measurements. Individual translocation frequencies from 40 subjects with existing WBC data and calculated cumulative red bone marrow doses below 0.6 Gy fall within the 95% prediction limits of the calibration curve.FISH-based translocation measurements can provide useful information for a retrospective biodosimetric interpretation. However, with the analysed number of cells, individual estimates required for a reliable evaluation of this highly variable exposure situation can be obtained only for a minority of subjects. In addition, influence of a temporal decline on the yield of stable translocations cannot be fully excluded.

Blood samples of 52 Chernobyl clean-up workers were analysed by fluorescence in situ hybridization (FISH) using whole-chromosome painting probes for chromosomes 1, 4 and 12, simultaneously with a pancentromeric probe and by conventional chromosome analysis, for radiation-induced symmetrical translocations and dicentrics in T-lymphocytes. Based on FISH measurements of translocations, individual biodosimetry estimates between 0.32 and 1.0 Gy were obtained from 18 cases. Pooled data for the total group of 52 workers provided a collective biodosimetry estimate of 0.23 Gy. For a group of 34 workers with documented doses, the mean dose estimate of 0.25 Gy compared well with the mean documented dose of 0.26 Gy. However, no correlation between individual translocation frequencies (FG) and documented doses could be found. A statistical analysis of the expected dose-response suggests exposures to higher doses than documented for a substantial fraction of workers with ascribed doses < 0.2 Sv. For subjects working repeatedly at the reactor site between 1986 and 1995 the mean translocation frequency was significantly higher than for those working only in 1986. A comparison of dicentric frequencies obtained by conventional scoring and by FISH measurements showed no significant difference, although only two of 52 cases revealed significantly higher yields than the mean control level. Based on conventionally scored dicentric frequencies, a collective biodosimetry estimate of 0.23 Gy could be derived only of the group of persons working at Chernobyl exclusively in 1986 for which a documented average dose of 0.19 Gy was reported.