Joan Francesc Barquinero

Cytogenetic analysis of lymphocytes from hospital workers occupationally exposed to low levels of ionizing radiation

Cytogenetic studies were performed in lymphocytes from hospital workers exposed to low doses of radiation (1.6-42.71 mSv). When compared with controls, exposed workers showed a significant increase in structural chromosome-type aberrations, acentric fragments being the most frequent alteration. Our results suggest that acentric fragments are good indicators of exposure to very low doses of radiation, although no dose-effect correlation was observed. The incidence of numerical abnormalities (hyperdiploidy) was significantly increased.

Translocation yields in peripheral blood lymphocytes from control populations

Purpose: To record the latest information on control levels of translocations in cultured human lymphocytes. Materials and methods: Control-level data from seven European laboratories that are using fluorescence in situ hybridization (FISH) techniques for retrospective biological dosimetry have been combined in a meta-analysis. After correction for the differing probe combinations used, tests of consistency are performed. The combined data have been used to test for individual variation, systematic variation with age, gender and smoking habits. Results: There is a strong variation of translocation yield with age but no variation was detectable with gender or smoking habits. After correction for age, homogeneity tests showed that about 10% of individuals were outside the 95% confidence limits as opposed to 5% expected. From a total of 385, there is an excess of about 20 individuals most of whom have an unexpectedly high yield of translocations. Conclusions: For retrospective biological dosimetry purposes a generic age-dependent control level can be assumed. No other lifestyle factors such as smoking appear to have a significant effect on translocation yield.

Relationship between the DNA content of human chromosomes and their involvement in radiation-induced structural aberrations, analysed by painting.

PURPOSE To study the relationship between the DNA content of human chromosomes and their involvement in radiation-induced structural chromosome aberrations. MATERIAL AND METHODS Human lymphocytes were cultured after exposure to 5 Gy of X-rays. FISH-painting was performed for all human chromosomes. RESULTS The results indicate that with the exception of chromosome 20, there was a good fit between the DNA content and the number of exchange-type aberrations and the number of breaks. However, there was a significant tendency for short chromosomes to be more affected than expected and for long chromosomes to be less affected than expected. A better fit was observed when, instead of DNA content, the surface area of the chromosome territories obtained from a spherical model was taken into consideration. CONCLUSIONS The tendency for short chromosomes to be more involved and long chromosomes to be less involved in exchange-type aberrations could be related not only to their DNA content but also to their interphase territory surface area.

Biological dosimetry in simulated in vitro partial irradiations

The assessment by biological dosimetry of the dose received in cases of partial-body exposure to ionizing radiation can be underestimated because irradiated lymphocytes are mixed with non-irradiated ones. To determine if the exposure affects the whole body or only part of it, it may be useful to know the distribution of cells with more than one dicentric chromosome. We established a dose-effect calibration curve for X-rays by analysis of chromosome aberrations. Moreover, in the present work, 20 partial irradiations for four different doses of X-rays (2, 3, 4 and 5 Gy) have been simulated by mixing irradiated and non-irradiated blood in different proportions. In all cases, the 95% confidence intervals of the estimated dose included the real dose of irradiation. However, some difficulties were found for the estimation of the fraction of irradiated cells. In the present study, D0 = 3.8 allows to obtain the best fit between the estimated and the real fraction of irradiated cells.

Realising the European network of biodosimetry: RENEB—status quo

Creating a sustainable network in biological and retrospective dosimetry that involves a large number of experienced laboratories throughout the European Union (EU) will significantly improve the accident and emergency response capabilities in case of a large-scale radiological emergency. A well-organised cooperative action involving EU laboratories will offer the best chance for fast and trustworthy dose assessments that are urgently needed in an emergency situation. To this end, the EC supports the establishment of a European network in biological dosimetry (RENEB). The RENEB project started in January 2012 involving cooperation of 23 organisations from 16 European countries. The purpose of RENEB is to increase the biodosimetry capacities in case of large-scale radiological emergency scenarios. The progress of the project since its inception is presented, comprising the consolidation process of the network with its operational platform, intercomparison exercises, training activities, proceedings in quality assurance and horizon scanning for new methods and partners. Additionally, the benefit of the network for the radiation research community as a whole is addressed.

Characterization of gene expression profiles at low and very low doses of ionizing radiation

The aim of the present study is to analyse by microarray techniques how gene expression is modulated after exposure to low and very low doses of ionizing radiation, to evaluate if the pattern of gene expression shows dose dependence, and to search for putative regulatory mechanisms behind the observed gene-expression modifications. For this, whole blood samples from five healthy donors were exposed in six doses between 5 and 500mGy. Total RNA extraction from CD4(+) lymphocytes was done at four different post-irradiation times. After mRNA amplification, aRNAs were hybridized on DNA microarrays. The results indicated that up-regulation was twice than down-regulation. Surprisingly, the number of modulated genes does not seem to change drastically with dose, even at the lowest dose of 5mGy. Clustering analysis revealed seven gene expression clusters with different dose dependence profiles. The functional analysis showed that the genes which increased their expression with the dose were related to p53 pathway and DNA damage response. This could be observed from 25mGy, but became very clear at doses equal or greater than 100mGy. On the other hand, genes with a constant modulation of their expression in all the tested doses were related to cellular respiration, ATP metabolic process and chromatin organization. These latter molecular mechanisms seem to be triggered at very low doses (5-25mGy). In silico promoter analysis seems to confirm the implication of transcription factors related to the pathways mentioned above.

Effect of americium‐241 α‐particles on the dose–response of chromosome aberrations in human lymphocytes analysed by fluorescence in situ hybridization

Purpose: To evaluate by the fluorescent in‐situ hybridization (FISH) technique the dose–response and intercellular distribution of α‐particle‐induced chromosome aberrations. In particular, the validity of using the yield of characteristic types of chromosome abnormalities in stable cells as quantitative indicators for retrospective dose reconstruction has been evaluated. Material and methods: Monolayers of human peripheral lymphocytes were exposed at doses from 0.02 to 1 Gy to α‐particles emitted from a source of americium‐241. The most probable energy of the α‐particles entering the cells was 2.7 MeV. FISH painting was performed using DNA probes for chromosomes 2, 4 and 8 in combination with a pan‐centromeric probe. In complete first‐division cells, identified by harlequin staining, aberrations involving painted target chromosomal material were recorded as well as aberrations involving only unpainted chromosomal material. Results: In total, the percentage of complex aberrations was about 35% and no dose dependence was observed. When complex‐type exchanges were reduced to simple base types, the different cell distributions were clearly over‐dispersed, and the linear coefficients of the dose–effect curves for translocations were significantly higher than for dicentrics. For past dose reconstruction, only a few complex aberrations were in stable cells. The linear coefficient obtained for transmissible aberrations in stable cells was more than seven times lower than that obtained in all analysed cells, i.e. including unstable cells. Conclusion: FISH‐based analysis of complex rearrangements allows discrimination between partial‐body exposures to low‐linear energy transfer radiation and high‐linear energy transfer exposures. In assessing past or chronic exposure to α‐particles, the use of a dose–effect curve obtained by FISH‐based translocation data, which had not excluded data determined in unstable cells, would underestimate the dose. Insertions are ineffective biomarkers because their frequency is too low.

Decreased sensitivity to the cytogenetic effects of bleomycin in individuals occupationally exposed to ionizing radiation

In the present work, 12 individuals occupationally exposed to ionizing radiation and 11 unexposed ones were studied to determine the cytogenetic effect of a challenge dose of bleomycin on their phytohemagglutinin stimulated lymphocytes. After bleomycin treatment, the frequencies of chromatid breaks and gaps were significantly lower in the exposed population (p < 0.025 for both types of chromatid alterations). These results could indicate that occupational exposure to ionizing radiation can induce an adaptive response that can be detected by a subsequent treatment with bleomycin.

Suitability of FISH painting techniques for the detection of partial-body irradiations for biological dosimetry.

Abstract Duran, A., Barquinero, J. F., Caballín, M. R., Ribas, M., Puig, P., Egozcue, J. and Barrios, L. Suitability of FISH Painting Techniques for the Detection of Partial-Body Irradiations for Biological Dosimetry. Radiat. Res. 157, 461–468 (2002). Peripheral blood was irradiated with 2, 3, 4 or 5 Gy of X rays and was mixed with nonirradiated blood at five different dilutions to simulate partial-body irradiations. Analysis by FISH was performed using whole-chromosome painting probes for chromosomes 1, 4 and 11 in combination with a pancentromeric probe. Chromosome aberrations affecting the painted fraction were classified according to the PAINT nomenclature; other unstable aberrations affecting the unpainted material were also recorded. To evaluate the suitability of painting for dose assessment in partial-body irradiations, the ability of the u test and a proposed s test to detect the expected overdispersion and the similarity between the real doses and the doses estimated using Dolphins approach were considered. For short-term biodosimetry, compared with solid-stained dicentric analyses, the suitability of FISH painting techniques for the detection of partial-body exposures is reduced, because of the decrease in the frequency of aberrations detected by FISH and in the number of cells with two or more aberrations. For reconstruction of past doses, when only complete apparently simple translocations in cells free of unstable aberrations were considered, the detection of the overdispersion and the accuracy of dose estimations were dramatically reduced. In a partial-body exposure, as the original dose increased, the whole-body dose estimated a long time after irradiation would tend to be lower, and the difference from the original dose would tend to be greater.

Biological Dosimetry Intercomparison Exercise: An Evaluation of Triage and Routine Mode Results by Robust Methods

Abstract Well-defined protocols and quality management standards are indispensable for biological dosimetry laboratories. Participation in periodic proficiency testing by interlaboratory comparisons is also required. This harmonization is essential if a cooperative network is used to respond to a mass casualty event. Here we present an international intercomparison based on dicentric chromosome analysis for dose assessment performed in the framework of the IAEA Regional Latin American RLA/9/054 Project. The exercise involved 14 laboratories, 8 from Latin America and 6 from Europe. The performance of each laboratory and the reproducibility of the exercise were evaluated using robust methods described in ISO standards. The study was based on the analysis of slides from samples irradiated with 0.75 (DI) and 2.5 Gy (DII). Laboratories were required to score the frequency of dicentrics and convert them to estimated doses, using their own dose–effect curves, after the analysis of 50 or 100 cells (triage mode) and after conventional scoring of 500 cells or 100 dicentrics. In the conntional scoring, at both doses, all reported frequencies were considered as satisfactory, and two reported doses were considered as questionable. The analysis of the data dispersion among the dicentric frequencies and among doses indicated a better reproducibility for estimated doses (15.6% for DI and 8.8% for DII) than for frequencies (24.4% for DI and 11.4% for DII), expressed by the coefficient of variation. In the two triage modes, although robust analysis classified some reported frequencies or doses as unsatisfactory or questionable, all estimated doses were in agreement with the accepted error of ±0.5 Gy. However, at the DI dose and for 50 scored cells, 5 out of the 14 reported confidence intervals that included zero dose and could be interpreted as false negatives. This improved with 100 cells, where only one confidence interval included zero dose. At the DII dose, all estimations fell within ±0.5 Gy of the reference dose interval. The results obtained in this triage exercise indicated that it is better to report doses than frequencies. Overall, in both triage and conventional scoring modes, the laboratory performances were satisfactory for mutual cooperation purposes. These data reinforce the view that collaborative networking in the case of a mass casualty event can be successful.