Radiation-induced risks at low dose: moving beyond controversy towards a new vision.

The paper recently published by Mothersill and Seymour (Radiat Environ Biophys 2013, doi: 10.1007/s00411-013-0472-y ) is commented upon by emphasizing on the recommendation not to confound the fields of radiation protection and radiobiological science as a source of controversy. Instead, these authors are proposing a new vision which suggests novel lines of scientific investigations to be addressed. At the moment, these include moving beyond the conceptual approach of DNA alteration through energy deposition in cells, and exploring the striking parallel currently existing between the ongoing individual/population debate in radioecology and that for cells/tissues in radiobiology. These interesting issues are briefly discussed and supported.

Influence on the mouse immune system of chronic ingestion of 137Cs.

The aim of this work was to determine the possible occurrence of damage to the immune system during the course of chronic ingestion of (137)Cs. BALB/C mice were used, with (137)Cs intake via drinking water at a concentration of 20 kBq l(-1). Adults received (137)Cs before mating and offspring were sacrificed at various ages between birth and 20 weeks. Phenotypic analysis of circulating blood cells and thymocytes did not show any significant modification of immune cell populations in animals ingesting (137)Cs as compared with control animals, with the exception of a slight increase in Treg percentage at the age of 12 weeks. Functional tests, including proliferative response to mitogens such as phytohaemagglutinin, response to alloantigens in mixed lymphocyte reaction and immunoglobulin response to vaccine antigens such as tetanus toxin and keyhole limpet haemocyanin did not show any significant functional modification of the immune system in (137)Cs-ingesting animals as compared with control animals. Overall, our results suggest that chronic ingestion of a low concentration of (137)Cs in drinking water in the long term does not have any biologically relevant effect on the immune system.

Brain accumulation of inhaled uranium in the rat depends on aerosol concentration, exposure repetitions, particle size and solubility.

A rostro-caudal gradient of uranium (U) in the brain has been suggested after its inhalation. To study the factors influencing this mapping, we first used 30-min acute inhalation at 56 mg/m3 of the relatively soluble form UO4 in the rat. These exposure parameters were then used as a reference in comparison with the other experimental conditions. Other groups received acute inhalation at different concentrations, repeated low dose inhalation of UO4 (10 exposures) or acute low dose inhalation of the insoluble form UO2. At 24 h after the last exposure, all rats showed a brain U accumulation with a rostro-caudal gradient as compared to controls. However, the total concentration to the brain was greater after repeated exposure than acute exposure, demonstrating an accumulative effect. In comparison with the low dose soluble U exposure, a higher accumulation in the front of the brain was observed after exposure to higher dose, to insoluble particles and following repetition of exposures, thus demonstrating a dose effect and influences of solubility and repetition of exposures. In the last part, exposure to ultrafine U particles made it possible to show 24 h after exposure the presence of U in the brain according to a rostro-caudal gradient. Finally, the time-course after exposure to micronic or nanometric U particles has revealed greater residence times for nanoparticles.

Biodistribution of (137)Cs in a mouse model of chronic contamination by ingestion and effects on the hematopoietic system.

The aim of this work was to define the possible occurrence of hematological changes during the course of a chronic ingestion of (137)Cs. A mouse model was used, with ingestion through drinking water with a cesium concentration of 20 kBq l(-1). Ingestion started in parent animals before mating, and (137)Cs intake and its effect on the hematopoietic system was studied in offspring at various ages between birth and 20 weeks. (137)Cs content was measured in various organs, indicating that (137)Cs was distributed throughout the organism including lympho-hematopoietic organs, i.e., femurs, spleen and thymus. However, we did not observe any effect on the hematopoietic system, whatever the parameter used. In fact, blood cell counts, mononuclear cell counts and progenitor frequency in bone marrow and spleen, and Flt3-ligand, Erythropoietin, G-CSF and SDF-1 concentration in plasma remained unchanged when compared to control animals. Moreover, phenotypic analysis did not show any change in the proportions of bone marrow cell populations. These results indicate that, although (137)Cs was found in all organs implicated in the hematopoietic system, this did not induce any changes in bone marrow function.

Evolution of the percutaneous penetration and distribution of uranyl nitrate as a function of skin-barrier integrity: an in vitro assessment.

As recommended by OECD Guidelines, percutaneous penetration studies consider intact skin, but rarely injured skin. Recent years have witnessed a growing concern for these two types of dermal exposure in the industry, particularly in the nuclear industry. The aim of this study was to show that a method based on an in vitro device can be used to realistically assess how skin-barrier alterations caused by occupational accidents can modify the percutaneous penetration and distribution of radionuclides, particularly uranium. Wounds encountered in the nuclear industry (i.e., nitric acid burns and abrasion) were simulated on hairless rat skin. Skin-barrier alterations were characterized by means of a histological study and by measuring transepidermal water loss (TEWL) and skin thickness. The percutaneous penetration of uranyl nitrate through intact or injured skin biopsies was then measured in vitro. The maximum uranium flux values obtained for intact skin, skin abrasion with stratum corneum removal, and skin exposed to 2 N HNO(3), 5 N HNO(3), and 14 N HNO(3) were, respectively, 0.6 +/- 0.02, 1.2 +/- 0.03, 1.2 +/- 0.04, 42.0 +/- 1.0, and 174.0 +/- 8.7 ng.cm(-2).h(-1). These results demonstrated that the percutaneous absorption of uranium increased with the increased impairment of the stratum corneum. TEWL, combined with maximum uranium flux values measured in vitro, yielded a good prediction of the percutaneous penetration of uranium through injured skin, previously observed in vivo. To conclude, this in vitro assay provides a conservative estimate of the percutaneous diffusion of uranium through intact or injured skin, making it a good alternative method for toxicological studies and risk assessments.

Neuro-inflammatory response in rats chronically exposed to (137)Cesium.

After the Chernobyl nuclear accident, behavioural disorders and central nervous system diseases were frequently observed in populations living in the areas contaminated by (137)Cs. Until now, these neurological disturbances were not elucidated, but the presence of a neuro-inflammatory response could be one explanation. Rats were exposed for 3 months to drinking water contaminated with (137)Cs at a dose of 400Bqkg(-1), which is similar to that ingested by the population living in contaminated areas in the former USSR countries. Pro-inflammatory and anti-inflammatory cytokine genes were assessed by real-time PCR in the frontal cortex and the hippocampus. At this level of exposure, gene expression of TNF-alpha and IL-6 increased in the hippocampus and gene expression of IL-10 increased in the frontal cortex. Concentration of TNF-alpha, measured by ELISA assays, was also increased in the hippocampus. The central NO-ergic pathway was also studied: iNOS gene expression and cNOS activity were significantly increased in the hippocampus. In conclusion, this study showed for the first time that sub-chronic exposure with post-accidental doses of (137)Cs leads to molecular modifications of pro- and anti-inflammatory cytokines and NO-ergic pathway in the brain. This neuro-inflammatory response could contribute to the electrophysiological and biochemical alterations observed after chronic exposure to (137)Cs.

Bone marrow stromal cells spontaneously produce Flt3-ligand: influence of ionizing radiations and cytokine stimulation.

PURPOSE: To define the ability of human bone marrow (BM) stromal cells to produce fms-like tyrosine kinase 3 (Flt3)-ligand (FL), and the effect of irradiation, tumour necrosis factor-alpha (TNFalpha) or tumour growth factor beta (TGFbeta) on FL production. MATERIAL AND METHODS: Primary BM stromal cell cultures were irradiated at 2-10 Gy or were stimulated with TNFalpha or TGFbeta1. The presence of FL was tested in culture supernatants and in cell lysate. The presence of a membrane-bound form of FL and the level of gene expression were also tested. RESULTS: Primary BM stromal cells spontaneously released FL. This production was increased by TNFalpha but not by TGFbeta1 or by irradiation. Chemical induction of osteoblastic differentiation from BM stromal cells also induced an increase in FL release. CONCLUSIONS: Our results suggest that the observed increase in FL concentration after in vivo irradiation is an indirect effect. The possible implication of BM stromal cells in these mechanisms is discussed.

Chronic exposure to uranium leads to iron accumulation in rat kidney cells.

After it is incorporated into the body, uranium accumulates in bone and kidney and is a nephrotoxin. Although acute or short-term uranium exposures are well documented, there is a lack of information about the effects of chronic exposure to low levels of uranium on both occupationally exposed people and the general public. The objective of this study was to identify the distribution and chemical form of uranium in kidneys of rats chronically exposed to uranium in drinking water (40 mg uranium liter(-1)). Rats were killed humanely 6, 9, 12 and 18 months after the beginning of exposure. Kidneys were dissected out and prepared for optical and electron microscope analysis and energy dispersive X-ray (XEDS) or electron energy loss spectrometry (EELS). Microscopic analysis showed that proximal tubule cells from contaminated rats had increased numbers of vesicles containing dense granular inclusions. These inclusions were composed of clusters of small granules and increased in number with the exposure duration. Using XEDS and EELS, these characteristic granules were identified as iron oxides. Uranium was found to be present as a trace element but was never associated with the iron granules. These results suggested that the mechanisms of iron homeostasis in kidney could be affected by chronic uranium exposure.

Modifications of inflammatory pathways in rat intestine following chronic ingestion of depleted uranium.

The environmental contamination by dispersion of depleted uranium (DU) might result in its chronic ingestion of DU by local populations. The aim of this study was to determine if chronic ingestion of DU at low doses induces inflammatory reactions in intestine, first biological system exposed to uranium after ingestion. Experiments were performed with rats receiving uranium in drinking water (40 mg/l) during 3, 6, or 9 months. Several parameters referring to prostaglandin, histamine, cytokine, and nitric oxide (NO) pathways were assessed in ileum. Concerning the prostaglandin pathway, a twofold increase in gene expression of cyclooxygenase of type 2 was noted after 6 months, with no changes in prostaglandins levels. At the same time, a decrease in mast cell number was observed without any changes in histamine levels. Experiments on cytokines showed increased gene expression of interleukin (IL)-1beta and IL-10 at 6 months, and decreased messenger RNA level of CCL-2. This change was associated with decreased macrophage density. An opposite effect of DU was induced on neutrophils, since increased number was observed at 3 (x1.7) and 9 months (x3). The results obtained on NO pathway seemed to indicate that DU exposure inhibited this pathway (decreased endothelial NO synthase messenger RNA, inductive NO synthase activity and NO(2)(-)/NO(3)(-) levels) at 6 months. In conclusion, this study demonstrated that chronic ingestion of DU-induced time-dependent modifications of inflammatory pathways, notably in terms of immune cell content. The ultimate effects of DU contamination might be pathogenic by suppressing defense mechanisms or inducing hypersensitivity. Further experiments should be thus performed to determine real consequences on intestinal response to oral antigens.

Heterogeneous accumulation of uranium in the brain of rats.

Recent reports suggest that uranium can accumulate not only in known target organs, that is, kidneys or bones, but also in others such as central nervous system. In the present work, the accumulation of uranium in the brain of rats was studied after repeated exposure by inhalation, chronic exposure by ingestion and acute exposure by injection. For each route of administration, the amount of uranium entering the brain was low. The results showed different accumulation in the brain areas according to the route of intake. Injection gave a rather homogeneous distribution in the different brain areas, whereas both inhalation and ingestion yielded heterogeneous but specific accumulation. These differences in distribution suggest the operation of different mechanisms of delivery of uranium to the brain tissues.

Risk assessment after internal exposure to black sand from Camargue: uptake and prospective dose calculation.

Some beaches in the south of France present high levels of natural radioactivity mainly due to thorium (Th) and uranium (U) present in the sand. Risk assessment after internal exposure of members of the public by either inhalation or ingestion of black sand of Camargue was performed. This evaluation required some information on the human bioavailability of U and Th from this sand. In vitro assays to determine the solubility of U, Th and their progeny were performed either in simulated lung fluid, with the inhalable fraction of sand, or in both simulated gastric and intestinal fluids with a sample of the whole sand. The experimental data show that the bioavailability of these radionuclides from Camargue sand is low in the conditions of the study. Prospective dose assessment for both routes of intake show low risk after internal exposure to this sand.

GHSI EMERGENCY RADIONUCLIDE BIOASSAY LABORATORY NETWORK - SUMMARY OF THE SECOND EXERCISE.

The Global Health Security Initiative (GHSI) established a laboratory network within the GHSI community to develop collective surge capacity for radionuclide bioassay in response to a radiological or nuclear emergency as a means of enhancing response capability, health outcomes and community resilience. GHSI partners conducted an exercise in collaboration with the WHO Radiation Emergency Medical Preparedness and Assistance Network and the IAEA Response and Assistance Network, to test the participating laboratories (18) for their capabilities in in vitro assay of biological samples, using a urine sample spiked with multiple high-risk radionuclides (90Sr, 106Ru, 137Cs, and 239Pu). Laboratories were required to submit their reports within 72 h following receipt of the sample, using a pre-formatted template, on the procedures, methods and techniques used to identify and quantify the radionuclides in the sample, as well as the bioassay results with a 95% confidence interval. All of the participating laboratories identified and measured all or some of the radionuclides in the sample. However, gaps were identified in both the procedures used to assay multiple radionuclides in one sample, as well as in the methods or techniques used to assay specific radionuclides in urine. Two-third of the participating laboratories had difficulties in determining all the radionuclides in the sample. Results from this exercise indicate that challenges remain with respect to ensuring that results are delivered in a timely, consistent and reliable manner to support medical interventions. Laboratories within the networks are encouraged to work together to develop and maintain collective capabilities and capacity for emergency bioassay, which is an important component of radiation emergency response.

Comprehensive analysis of the renal transcriptional response to acute uranyl nitrate exposure.

BACKGROUND: Chemical and radiological toxicities related to uranium acute exposure have been widely studied in nuclear fuel workers and military personnel. It is well known that uranyl nitrate induces acute renal failure (ARF). However, the mechanisms of this metal-induced injury are not well defined at the molecular level. RESULTS: Renal function and histology were assessed in mice receiving uranyl nitrate (UN(+)) and controls (UN(-)). To identify the genomic response to uranium exposure, serial analysis gene expression (SAGE) of the kidney was performed in both groups. Over 43,000 mRNA SAGE tags were sequenced. A selection of the differentially expressed transcripts was confirmed by real-time quantitative PCR and Western blotting. UN(+) animals developed renal failure and displayed the characteristic histological lesions of UN nephropathy. Of the >14,500 unique tags identified in both libraries, 224 had a modified expression level; they are known to participate in inflammation, ion transport, signal transduction, oxidative stress, apoptosis, metabolism, and catabolism. Several genes that were identified had not previously been evaluated within the context of toxic ARF such as translationally controlled tumor protein, insulin like growth factor binding protein 7 and ribosomal protein S29, all apoptosis related genes. CONCLUSION: We report a comprehensive description of the UN induced modifications in gene expression levels, including the identification of genes previously unrelated to ARF. The study of these genes and the metabolisms they control should improve our understanding of toxic ARF and enlighten on the molecular targets for potential therapeutic interventions.

Genotoxic and inflammatory effects of depleted uranium particles inhaled by rats.

Depleted uranium (DU) is a radioactive heavy metal coming from the nuclear industry and used in numerous military applications. Uranium inhalation can lead to the development of fibrosis and neoplasia in the lungs. As little is known concerning the molecular processes leading to these pathological effects, some of the events in terms of genotoxicity and inflammation were investigated in rats exposed to DU by inhalation. Our results show that exposure to DU by inhalation resulted in DNA strand breaks in broncho-alveolar lavage (BAL) cells and in increase of inflammatory cytokine expression and production of hydroperoxides in lung tissue suggesting that the DNA damage was in part a consequence of the inflammatory processes and oxidative stress. The effects seemed to be linked to the doses, were independent of the solubility of uranium compounds and correlating with the type of inhalation. Repeated inhalations seemed to induce an effect of potentiation in BAL cells and also in kidney cells. Comet assay in neutral conditions revealed that DNA damage in BAL cells was composed partly by double strands breaks suggesting that radiation could contribute to DU genotoxic effects in vivo. All these in vivo results contribute to a better understanding of the pathological effect of DU inhalation.

Evaluation of the effect of chronic exposure to 137Cesium on sleep-wake cycle in rats.

Since the Chernobyl accident, the most significant problem for the population living in the contaminated areas is chronic exposure by ingestion of radionuclides, notably (137)Cs, a radioactive isotope of cesium. It can be found in the whole body, including the central nervous system. The present study aimed to assess the effect of (137)Cs on the central nervous system and notably on open-field activity and the electroencephalographic pattern. Rats were exposed up to 90 days to drinking water contaminated with (137)Cs at a dosage of 400 Bq kg(-1), which is similar to that ingested by the population living in contaminated territories. At this level of exposure, no significant effect was observed on open-field activity. On the other hand, at 30 days exposure, (137)Cs decreased the number of episodes of wakefulness and slow wave sleep and increased the mean duration of these stages. At 90 days exposure, the power of 0.5-4 Hz band of (137)Cs-exposed rats was increased in comparison with controls. These electrophysiological changes may be due to a regional (137)Cs accumulation in the brain stem. In conclusion, the neurocognitive effects of (137)Cs need further evaluation and central disorders of population living in contaminated territories must be considered.

The effect of repeated inhalation on the distribution of uranium in rats.

For the assessment of doses after inhalation of airborne uranium compounds by workers, the International Commission on Radiological Protection (ICRP) developed compartmental models that are used to calculate reference dose coefficients and retention and excretion functions. It is assumed that each acute intake has no effect on the biokinetics of later intakes. Consequently, retention and excretion after multiple or chronic exposure are predicted using the same models as after acute exposure. This assumption was tested here on rats exposed to repeated inhalation of uranium dioxide (UO2). First, excretion and organ retention were determined after a single inhalation of UO2. The follow-up of incorporated activity was used to design a biokinetic model for uranium inhaled by rats. Second, the biokinetics of uranium were monitored in two experiments of repeated inhalations of uranium dioxide under different intake patterns. For these two experiments, the organs' retention and excretion after repeated UO2 inhalation were predicted using the biokinetic model and compared to the experimental measurement. Under the two sets of experimental conditions considered, the prediction of the biokinetic model based on acute exposure data was consistent with the biokinetics observed after repeated UO2 inhalations, with the possible exception of retention in the skeleton.

Distribution and genotoxic effects after successive exposure to different uranium oxide particles inhaled by rats.

In nuclear fuel cycle facilities, workers may inhale airborne uranium compounds that lead to internal contamination, with various exposure scenarios depending on the workplace. These exposures can be chronic, repeated, or acute, and can involve many different compounds. The effect of uranium after multiple scenarios of exposure is unknown. The aim of this study, therefore, was to investigate the genotoxic and biokinetics consequences of exposure to depleted insoluble uranium dioxide (UO2) by repeated or acute inhalation on subsequent acute inhalation of moderately soluble uranium peroxide (UO4) in rats. The results show that UO2 repeated preexposure by inhalation increases the genotoxic effects of UO4 inhalation, assessed by comet assay, in different cell types, when UO4 exposure alone has no effect. At the same time, the study of UO4 bioaccumulation showed that the UO4 biokinetics in the kidneys, gastrointestinal tract, and excreta, but not in the lungs, were slightly modified by previous UO2 exposures. All these results show that both genotoxic and biokinetics effects of uranium may depend on preexposure and that repeated exposure induces a potentiation effect compared with acute exposure.

GHSI EMERGENCY RADIONUCLIDE BIOASSAY LABORATORY NETWORK: SUMMARY OF A RECENT EXERCISE.

The Global Health Security Initiative (GHSI) established a laboratory network within the GHSI community to develop their collective surge capacity for radionuclide bioassay in response to a radiological or nuclear emergency. A recent exercise was conducted to test the participating laboratories for their capabilities in screening and in vitro assay of biological samples, performing internal dose assessment and providing advice on medical intervention, if necessary, using a urine sample spiked with a single radionuclide, 241Am. The laboratories were required to submit their reports according to the exercise schedule and using pre-formatted templates. Generally, the participating laboratories were found to be capable with respect to rapidly screening samples for radionuclide contamination, measuring the radionuclide in the samples, assessing the intake and radiation dose, and providing advice on medical intervention. However, gaps in bioassay measurement and dose assessment have been identified. The network may take steps to ensure that procedures and practices within this network be harmonised and a follow-up exercise be organised on a larger scale, with potential participation of laboratories from the networks coordinated by the International Atomic Energy Agency and the World Health Organization.

Bioaccumulation and behavioural effects of depleted uranium in rats exposed to repeated inhalations.

Depleted uranium has numerous industrial and military uses. Contamination by inhalation of airborne compounds is probably the most important route of exposure. In humans, there are no data clearly demonstrating neurotoxicity of uranium, yet some experimental studies suggest a link between neurological toxicity and uranium exposure. In this work, the bioaccumulation of uranium in male rats after exposure to repeated depleted uranium dioxide inhalation (30 min inhalation at 197 mgm(-3), 4 days a week for 3 weeks) has been studied, together with the behavioural effects. The uranium concentrations in the brain 1 day after the end of the exposure period varied as follows: olfactory bulb>hippocampus>frontal cortex>cerebellum, subsequently decreasing rapidly. The spontaneous locomotion activity of exposed rats was increased 1 day post exposure and the spatial working memory was less efficient 6 days post exposure, compared with control rats. These data suggest that depleted uranium is able to enter the brain after exposure to repeated inhalation, producing behavioural changes.