Recent studies have suggested that visualization of gamma-H2AX nuclear foci can be used to estimate exposure to very low doses of ionizing radiation. Although this approach is widely used for various purposes, its suitability for individual human biodosimetry has not yet been assessed. We therefore conducted such an assessment with the help of available software for observing and automatically scoring gamma-H2AX foci. The presence of gamma-H2AX foci was evaluated in human peripheral blood lymphocytes exposed ex vivo to gamma rays in a dose range of 0.02 to 2 Gy. We analyzed the response of gamma-H2AX to ionizing radiation in relation to dose, time after exposure, and individual variability. We constructed dose-effect calibration curves at 0.5, 8 and 16 h after exposure and evaluated the threshold of detection of the technique. The results show the promise of automatic gamma-H2AX scoring for a reliable assessment of radiation doses in a dose range of 0.6 Gy to 2 Gy up to 16 h after exposure. This gamma-H2AX-based assay may be useful for biodosimetry, especially for triage to distinguish promptly among individuals the ones who have received negligible doses from those with significantly exposures who are in need of immediate medical attention. However, additional in vivo experiments are needed for validation.

After large-scale accidental overexposure to ionizing radiation, a rapid triage of the exposed population can be performed by scoring dicentrics and ring chromosomes among 50 metaphases. This is rapid but is not accurate because the sensitivity is around 0.5 Gy. After the triage step, dose can be estimated by scoring 500 metaphases. This is lengthy but very accurate because the sensitivity is between 0.1 and 0.2 Gy. To improve the methodology, we propose the use of software for automatic dicentric scoring that was tested on victims of an accident in Dakar. Manual scoring of 50 metaphases was carried out, then manual scoring of 500 metaphases, and automatic scoring. Comparison between the dose classifications obtained with manual scoring on 50 metaphases and 500 metaphases showed 50% misclassification with the manual scoring on 50 metaphases. Comparison between the dose classifications obtained with the automatic scoring and manual scoring on 500 metaphases showed only 4.35% misclassification with the automatic scoring. The automatic scoring method is more accurate than the manual scoring on 50 metaphases and can therefore be used for triage, and in place of the manual scoring on 500 metaphases method for individual dose estimation, because it is as accurate and much faster.

Most studies that aim to understand the interactions between different types of photon radiation and cellular DNA assume homogeneous cell irradiation, with all cells receiving the same amount of energy. The level of DNA damage is therefore generally determined by averaging it over the entire population of exposed cells. However, evaluating the molecular consequences of a stochastic phenomenon such as energy deposition of ionizing radiation by measuring only an average effect may not be sufficient for understanding some aspects of the cellular response to this radiation. The variance among the cells associated with this average effect may also be important for the behaviour of irradiated tissue. In this study, we accurately estimated the distribution of the number of radiation-induced γH2AX foci (RIF) per cell nucleus in a large population of endothelial cells exposed to 3 macroscopic doses of gamma rays from 60Co. The number of RIF varied significantly and reproducibly from cell to cell, with its relative standard deviation ranging from 36% to 18% depending on the macroscopic dose delivered. Interestingly, this relative cell-to-cell variability increased as the dose decreased, contrary to the mean RIF count per cell. This result shows that the dose effect, in terms of the number of DNA lesions indicated by RIF is not as simple as a purely proportional relation in which relative SD is constant with dose. To analyse the origins of this observed variability, we calculated the spread of the specific energy distribution for the different target volumes and subvolumes in which RIF can be generated. Variances, standard deviations and relative standard deviations all changed similarly from dose to dose for biological and calculated microdosimetric values. This similarity is an important argument that supports the hypothesis of the conservation of the association between the number of RIF per nucleus and the specific energy per DNA molecule. This comparison allowed us to calculate a volume of 1.6 μm3 for which the spread of the specific energy distribution could explain the entire variability of RIF counts per cell in an exposed cell population. The definition of this volume may allow to use a microdosimetric quantity to predict heterogeneity in DNA damage. Moreover, this value is consistent with the order of magnitude of the volume occupied by the hydrated sugar-phosphate backbone of the DNA molecule, which is the part of the DNA molecule responsible for strand breaks.

Dicentric chromosome analysis remains the most widely used method in biodosimetry. It has a lower detection limit of about 0.1 Gy, and allows one to distinguish between whole- and partial-body exposures. A drawback of the dicentric analysis is that it is a time consuming method and maybe difficult to implement in a mass casualty event. To try to increase the analysis capacity, automatic dicentric scoring (ADS) using image analysis software is being incorporated in several laboratories. Here we present the results obtained in an emergency exercise simulating 50 victims. The ability to distinguish different radiations scenarios is evaluated. To simulate whole-body exposures peripheral blood samples were irradiated at doses between 0-4.7 Gy, and to simulate partial-body exposures irradiated and nonirradiated blood were mixed in different proportions. With the data obtained from the first slide analyzed (with about 300-400 cells), 32 of 34 simulated whole-body exposures were correctly classified according to radiation exposure levels. For simulated partial-body irradiations, it was possible to detect them as partial exposures at the end of the first slide analyzed but only at the highest doses. In all cases the classification was updated every time the analysis of one additional slide was finished. The comparison between our present results and those reported in the literature for manual scoring shows that for triage purposes the ADS based on 300-400 cells is similar in efficiency to classifying the cases based on manual scoring of 50 cells. However, if one accounts for the associated uncertainties and the time needed for ADS, we suggest that ADS triage scoring should be based on about 1,000 cells. For final dose estimations the number of cells to score will depend on the initial estimated dose, and on the information contributed from physical dose-reconstruction or clinical symptoms. At doses higher than 1 Gy, we propose analysis of 1,500 and for lower doses or suspected partial-body exposures, the number of cells to score should be 3,000.

This technique is appropriate for corroborating a former IR exposure of individuals. When the radiation dose is greater than 1 Gy, the translocations in complex exchanges must be considered. Another relevant point is the use of an appropriate background yield of translocations. The dose assessment, however, also depends on exposure to various genotoxic agents besides IR. If no evidence about the existence of radiation exposure is available, dose assessment is not useful. For this reason, report only the translocation frequency and its comparison with the background yield by age class is preferable.

To compare the responses of the different lymphocyte subtypes after an exposure of whole blood to low doses of ionizing radiation, we examined variations in gene expression in different lymphocyte subpopulations using microarray technology. Blood samples from five healthy donors were independently exposed to 0 (sham irradiation), 0.05 and 0.5 Gy of ionizing radiation. Three and 24 h after exposure, CD56+, CD4+ and CD8+ cells were negatively isolated. RNA from each set of experimental conditions was competitively hybridized on 25k oligonucleotide microarrays. Modifications of gene expression were measured after both intervals and in all cell types. Twenty-four hours after exposure to 0.5 Gy, we observed an induction of the expression of BAX, PCNA, GADD45, DDB2 and CDKN1A. However, the numbers of modulated genes greatly differed between cell types. In particular, 3 h after exposure to doses as low as 0.05 Gy, the number of down-modulated genes was 10 times greater for CD4+ cells than for all other cell types. Moreover, most of these repressed genes were taking part in the cell processes of protein biosynthesis and oxidative phosphorylation. The results suggest that several biological pathways in CD4+ cells could be sensitive to low doses of radiation. Therefore, specifically studying CD4+ cells could help to understand the mechanisms involved in low-dose response and allow their detection.

HAL is a multidisciplinary open access archive for the deposit and dissemination of scientific research documents, whether they are published or not. The documents may come from teaching and research institutions in France or abroad, or from public or private research centers. L'archive ouverte pluridisciplinaire HAL, est destinée au dépôt et à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Disturbances in hemostatic and hemorheological parameters have been investigated in a group of 29 children with nephrotic syndrome: 23 children classified as steroid-sensitive and 6 as steroid-resistant. Studies were performed before prednisone treatment and 3 weeks later, after initiation of steroid therapy. Before treatment, the alterations in hemostatic system involved moderate thrombocytosis with spontaneous aggregation in 19 patients. High levels of fibrinogen, factor VIII, Willebrand factor, protein C, protein S and α₂-macroglobulin (α₂M) were observed. Factor XII and α₁-antitrypsin (α₁AT) were lower than normal. Antithrombin III (ATIII) level was normal in the majority of patients. A plasma and blood hyperviscosity syndrome was also observed as well as an increase in erythrocyte aggregation. During treatment, an improvement in the hemostatic parameters was observed in the patients who responded to prednisone. The expected increase in factor VIII (frequently described in the literature) was not observed, while there was a significant increase in protein C. In the steroid-resistant patients, the only significant changes observed were decreased fibrinogen and increased protein C. The hemorheological parameters showed a tendency towards normality regardless of whether or not the treatment provided remission of NS. The relationship between hemorheological and hemostatic factors changes are discussed.