Sisko Salomaa

Non-targeted effects of ionising radiation-Implications for low dose risk

Non-DNA targeted effects of ionising radiation, which include genomic instability, and a variety of bystander effects including abscopal effects and bystander mediated adaptive response, have raised concerns about the magnitude of low-dose radiation risk. Genomic instability, bystander effects and adaptive responses are powered by fundamental, but not clearly understood systems that maintain tissue homeostasis. Despite excellent research in this field by various groups, there are still gaps in our understanding of the likely mechanisms associated with non-DNA targeted effects, particularly with respect to systemic (human health) consequences at low and intermediate doses of ionising radiation. Other outstanding questions include links between the different non-targeted responses and the variations in response observed between individuals and cell lines, possibly a function of genetic background. Furthermore, it is still not known what the initial target and early interactions in cells are that give rise to non-targeted responses in neighbouring or descendant cells. This paper provides a commentary on the current state of the field as a result of the non-targeted effects of ionising radiation (NOTE) Integrated Project funded by the European Union. Here we critically examine the evidence for non-targeted effects, discuss apparently contradictory results and consider implications for low-dose radiation health effects.

Persistence of translocations after accidental exposure to ionizing radiation

PURPOSE To assess the validity of translocations for retrospective biodosimetry using FISH chromosome painting of peripheral lymphocytes in repeated samples of people accidentally exposed to radiation. MATERIALS AND METHODS Blood samples from five people exposed to mainly whole-body irradiation of gamma-rays from a radiation accident in Estonia in 1994 were taken at 2-4 month intervals. A total of eight samplings were carried out, including one directly after the accident. Fluorescence in situ hybridization with probes for chromosomes 1, 2 and 4 was performed on metaphase preparations from 48 h whole blood cultures; 1500 cells were scored from each individual per time point. RESULTS Translocations remained relatively stable during 2 years after exposure in all subjects. A noticeable decrease in complete translocation yields and a significant decrease in incomplete translocations were observed in one person. In addition to whole-body exposure, he had also been exposed to partial-body irradiation. Due to the overall persistence of translocations, dose estimates were very similar throughout the 2 year period. A rapid decline in dicentric frequencies was noted during the first year after exposure. CONCLUSIONS The results suggest that during 2 years the yield of translocations in peripheral lymphocytes remained at a constant level after whole-body exposure. This finding supports the use of translocations for retrospective dosimetry, at least within this relatively short period of follow-up. In the case of partial-body irradiation, however, the elimination of co-existing unstable aberrations reduced the translocation yield over time. Follow-up will be continued in order to determine the stability of translocations over longer times.

Thyroid nodularity and cancer among Chernobyl cleanup workers from Estonia

Thyroid examinations, including palpation, ultrasound and, selectively, fine-needle aspiration biopsy, were conducted on nearly 2,000 Chernobyl cleanup workers from Estonia to evaluate the occurrence of thyroid cancer and nodular thyroid disease among men with protracted exposure to ionizing radiation. The examinations were conducted in four cities in Estonia during March-April 1995, 9 years after the reactor accident. The study population was selected from a predefined cohort of 4,833 cleanup workers from Estonia under surveillance for cancer incidence. These men had been sent to Chernobyl between 1986 and 1991 to entomb the damaged reactor, remove radioactive debris and perform related cleanup activities. A total of 2,997 men were invited for thyroid screening and 1,984 (66%) were examined. Estimates of radiation dose from external sources were obtained from military or other institutional records, and details about service dates and types of work performed while at Chernobyl were obtained from a self-administered questionnaire. Blood samples were collected for assay of chromosomal translocations in circulating lymphocytes and loss of expression of the glycophorin A (GPA) gene in erythrocytes. The primary outcome measure was the presence or absence of thyroid nodules as determined by the ultrasound examination. Of the screened workers, 1,247 (63%) were sent to Chernobyl in 1986, including 603 (30%) sent in April or May, soon after the accident. Workers served at Chernobyl for an average of 3 months. The average age was 32 years at the time of arrival at Chernobyl and 40 years at the time of thyroid examination. The mean documented radiation dose from external sources was 10.8 cGy. Biological indicators of exposure showed low correlations with documented dose, but did not indicate that the mean dose for the population was higher than the average documented dose. Ultrasound examinations revealed thyroid nodules in 201 individuals (10.2%). The prevalence of nodules increased with age at examination, but no significant associations were observed with recorded dose, date of first duty at Chernobyl, duration of service at Chernobyl, building the sarcophagus or working on the roof of neighboring buildings or close to the damaged reactor. Nodularity showed a nonsignificant (p(1) = 0.10) positive association with the proportion of lymphocytes with chromosome translocations, but associations with the frequency of variant erythrocytes in the GPA assay were weak and unstable (p(1) > or = 0.46). The majority of fine-needle biopsies taken on 77 study participants indicated benign nodular disease. However, two cases of papillary carcinoma and three benign follicular neoplasms were identified and referred for treatment. Both men with thyroid cancer had been sent to Chernobyl in May of 1986, when the potential for exposure to radioactive iodines was greatest. Chernobyl cleanup workers from Estonia did not experience a markedly increased risk of nodular thyroid disease associated with exposure to external radiation. Possible reasons for the apparent absence of effect include low radiation doses, the protracted nature of the exposure, errors in dose measurement, low sensitivity of the adult thyroid gland or the insufficient passage of time for a radiation effect to be expressed.

Do Recorded Doses Overestimate True Doses Received by Chernobyl Cleanup Workers? Results of Cytogenetic Analyses of Estonian Workers by Fluorescence In Situ Hybridization

Studies of workers who were sent to Chernobyl after the 1986 reactor accident are being conducted to provide a better understanding of the effects of chronic low-dose radiation exposures. A crucial component to these investigations is an accurate assessment of the radiation doses received during the cleanup activities. To provide information on biological measurements of dose, fluorescence in situ hybridization (FISH) with whole-chromosome painting probes has been applied to quantify stable chromosome aberrations (translocations and insertions) among a defined cohort of 4,833 cleanup workers from Estonia. Cytogenetic analysis of 48-h lymphocyte cultures from 118 Estonian cleanup workers (10.3 cGy mean recorded dose; 25 cGy maximum), 29 Estonian population controls and 21 American controls was conducted by three laboratories. More than 258,000 painted metaphases were evaluated. Overall, we observed lower translocation frequencies than has been reported in previous studies using FISH among Chernobyl cleanup workers. In our data, a clear association with increased levels of translocations was seen with increasing age at blood drawing. There was no correlation, however, between aberration frequency and recorded measurements of physical dose or any category of potential high-dose and high-dose-rate exposure such as being sent to Chernobyl in 1986, working on the roof near the damaged nuclear reactor, working in special zones or having multiple tours. In fact, the translocation frequency was lower among the exposed workers than the controls, though not significantly so. To estimate the level of effect that would have been expected in a population of men having an average dose of approximately 10 cGy, blood from six donors was exposed to low-LET radiation, and more than 32,000 metaphases were scored to estimate dose-response coefficients for radiation-induced translocations in chromosome pairs 1, 2 and 4. Based on these results, we estimate that had this group of 118 men received an average whole-body dose of 10-11 cGy, as chronic or acute exposures, an increase in the mean frequency of chromosome translocations of more than 40-65% would have been observed in their lymphocytes compared to findings in nonirradiated controls. In spite of evaluating more than a quarter of a million metaphases, we were unable to detect any increase in the mean, median or range in chromosome aberrations in lymphocyte cultures from a group of Estonian men who took part in the cleanup of the Chernobyl nuclear power site and those who did not. We conclude that it is likely that recorded doses for these cleanup workers overestimate their average bone marrow doses, perhaps substantially. These results are consistent with several negative studies of cancer incidence in Chernobyl cleanup workers and, if borne out, suggest that future studies may not be sufficiently powerful to detect increases in leukemia or cancer, much less distinguish differences between the effects of chronic compared to brief radiation exposures.

Comparison of dose-response curves for chromosomal aberrations established by chromosome painting and conventional analysis

PURPOSE To establish 60Co gamma-ray dose-response curves for dicentrics and translocations visualized by chromosome painting and for dicentrics analysed after conventional solid staining. MATERIALS AND METHODS Analysis of chromosomal aberrations was performed on peripheral blood lymphocytes obtained from 48 h old cultures of irradiated whole blood. Dicentrics were scored from Giemsa-stained preparations, and bi-coloured dicentrics and translocations after FISH painting of chromosomes 1, 2 and 4. RESULTS Equal frequencies of complete dicentrics and translocations, where both members of the exchanges were seen, were observed in the chromosome painting analysis at all doses, resulting in similar calibration curves. Due to differences in scoring criteria, dicentrics scored in conventionally Giemsa-stained slides agreed better with data for total than for complete exchanges. Donor differences for translocations at the control level and at low doses were seen and large uncertainty surrounds the linear component of the dose-response for total translocations. CONCLUSIONS Dose reconstruction of past exposures in cases of low doses is very dependent on the linear coefficient of the curve. Results indicate that total translocations would give less reliable dose estimates and therefore complete translocations are preferred.

Effects of 50 Hz sinusoidal magnetic fields and spark discharges on human lymphocytes in vitro

Abstract Human peripheral blood lymphocytes were exposed in vitro to 50 Hz sinusoidal magnetic fields of 30 μT, 300 μT, and 1 mT, and were monitored for genotoxic effects. No effects on chromosomal aberrations, micronuclei or proliferation indices were noticed. A weak effect on sister chromatid exchanges, noted in one series of experiments, could not be verified when the series was repeated. Exposure of whole blood to spark discharges of up to 3.65 kVcm−1 and 2.6 μs duration (10 pulses) did not result In chromosomal aberrations. Cell destruction from spark discharges was extensive. The conductivity of blood during a spark discharge pulse was examined and was shown to decrease by a factor of 2–3 at the beginning of the pulse owing to the β-relaxation of cells. After 0.5 μs the conductivity approached a constant level.

Biodosimetry after accidental radiation exposure by conventional chromosome analysis and FISH.

A 137Cs source was stolen from a radioactive waste depository in Estonia on 21 October 1994 and kept in a private house for 4 weeks. This resulted in the death of one person, acute radiation injuries to four people and exposure of several other people to lower doses of radiation. Analysis of chromosomal aberrations in peripheral blood lymphocytes was used in the assessment of radiation exposure of 18 people involved in the accident. Dose estimation assessment based on the frequencies of dicentric chromosomes was performed both by the standard method and by considering possible dose protraction and non-uniform exposure. Considerable differences in dose estimates were obtained depending on the approach used, ranging from about 1 Gy to almost 3 Gy in the patients most heavily exposed. In view of the deterministic health effects observed in some of the subjects, it was concluded that the dose estimates involving information on dose protraction were more realistic than those obtained by comparison with the standard high dose-rate calibration curve. Chromosome painting analyses using fluorescence in situ hybridization, with a probe cocktail for chromosomes 1, 2 and 4 and centromere detection, were performed in parallel. Good agreement on dicentric chromosome frequencies was observed between the conventional and painting analyses. The frequencies of complete translocations were comparable with the frequencies of dicentric chromosomes. In addition to the complete translocations, a pronounced increase in the frequency of incomplete translocations was observed. Dose estimates performed on the basis of FISH translocation frequencies were consistent with the dicentric analysis.

Stable Chromosome Aberrations in the Lymphocytes of a Population Living in the Vicinity of the Semipalatinsk Nuclear Test Site

Abstract Salomaa, S., Lindholm, C., Tankimanova, M. K., Mamyrbaeva, Z. Z., Koivistoinen, A., Hulten, M., Mustonen, R., Dubrova, Y. E. and Bersimbaev, R. I. Stable Chromosome Aberrations in the Lymphocytes of a Population Living in the Vicinity of the Semipalatinsk Nuclear Test Site. Radiat. Res. 158, 591–596 (2002). Translocation analysis using FISH (fluorescence in situ hybridization) chromosome painting was performed to evaluate the magnitude of exposure to ionizing radiation among the human population living close to the Semipalatinsk nuclear test site in Kazakhstan. We studied two generations of people living in villages that were in the path of the radioactive cloud from the first Soviet surface nuclear test performed in August 1949 and from later tests. The older generation (P0) lived in the area at the time of testing, and the younger generation (F1) was exposed to smaller doses from the residual fallout and later tests. In both P0 and F1 generations, similar translocation frequencies were observed in persons living in either the Semipalatinsk area or a noncontaminated area. Assuming translocation stability in peripheral blood lymphocytes over several decades, these findings suggest that on average, the magnitude of exposure of this cohort in the Semipalatinsk area has been considerably smaller than that reported in the literature. Previously reported doses of the order of 1−4.5 Gy (mean 2.9 Gy in the P0 generation) cannot be confirmed by the present data.

Distribution of radiation-induced exchange aberrations in human chromosomes 1, 2 and 4

PURPOSE To examine the distribution of radiation-induced breakpoints in chromosomes 1, 2 and 4 both in relation to their DNA content and by localization of the breaks along each chromosome. MATERIAL AND METHODS The work consisted of two studies, one with chromosomal aberrations found in persons after accidental exposure in Estonia in 1994 and another involving aberrations seen in in vitro-irradiated lymphocytes. Localization of breakpoints in painted chromosomes involved in complete exchange-type aberrations was conducted by applying a computerized measuring system on stored image-files. RESULTS AND CONCLUSIONS The yield of exchanges in chromosomes 1, 2 and 4 in both studies was equal to that expected from their DNA content. In contrast, the breakpoint location of complete exchanges within these chromosomes was not random. Chromosomes 1 and 4 contained more breaks in the middle parts of the p and q arms, whereas breaks were observed more uniformly along chromosome 2. Complete exchanges, however, were very rare in the terminal regions of all three chromosomes, most probably resulting from limitations in the resolution of small painted pieces.

Domestic radon exposure and the frequency of stable or unstable chromosomal aberrations in lymphocytes

PURPOSE To clarify the relationship between domestic radon exposure and the occurrence of chromosomal aberrations, stable translocations especially, in peripheral blood lymphocytes. MATERIALS AND METHODS The study comprised a total of 84 nonsmoking individuals, divided into three groups according to radon concentration measurements performed in their homes: low radon concentration (<100Bq/m3, mean 67Bq/m3), medium (200-400Bq/m3, mean 293Bq/m3) or high (>800Bq/m3, mean 1737Bq/m3). Minimum residence in the present low-rise house was 10 years. The groups were matched with regard to age, gender and medical exposure. Fluorescence in-situ hybridization (FISH) was performed using chromosome paints for chromosomes 1, 2 and 4; 1500 metaphases were scored from each individual. RESULTS Equal frequencies of translocations and also other aberrations, e.g. dicentrics and complex rearrangements, were obtained in each group. Significant correlation of translocations with age was observed, and due to the high mean age (50 years) the genome-corrected frequency of translocations was high: about one translocation in 100 metaphases. CONCLUSIONS Chronic exposure to high concentrations of domestic radon did not increase the rate of stable or unstable chromosomal aberrations in peripheral blood lymphocytes detected by FISH chromosome painting. A strong age effect was observed.