ESLAV/ECLAM/LAVA/EVERI recommendations for the roles, responsibilities and training of the laboratory animal veterinarian and the designated veterinarian under Directive 2010/63/EU.

Directive 2010/63/EU was adopted in September 2010 by the European Parliament and Council, and became effective in January 2013. It replaces Directive 86/609/EEC and introduces new requirements for the protection of animals used for scientific purposes. In particular, it requires that establishments that breed, supply or use laboratory animals have a designated veterinarian (DV) with expertise in laboratory animal medicine, or a suitably qualified expert where more appropriate, charged with advisory duties in relation to the well-being and treatment of the animals. This paper is a report of an ESLAV/ECLAM/LAVA/EVERI working group that provides professional guidance on the role and postgraduate training of laboratory animal veterinarians (LAVs), who may be working as DVs under Directive 2010/63/EU. It is also aimed at advising employers, regulators and other persons working under the Directive on the role of the DV. The role and responsibilities of the DV include the development, implementation and continuing review of an adequate programme for veterinary care at establishments breeding and/or using animals for scientific purposes. The programme should be tailored to the needs of the establishment and based on the Directive's requirements, other legislations, and current guidelines in laboratory animal medicine. Postgraduate laboratory animal veterinary training should include a basic task-specific training module for DVs to complement veterinary competences from graduation, and continuing professional development on the basis of a gap analysis. A tiered approach to further training in laboratory animal veterinary medicine and science offers career development pathways that are mutually beneficial to LAVs and establishments.

Short-term effects of depleted uranium on immune status in rat intestine.

In the event of ingestion, the digestive tract is the first biological system exposed to depleted uranium (DU) intake via the intestinal lumen. However, little research has addressed the biological consequences of a contamination with depleted uranium on intestinal properties such as the barrier function and/or the immune status of this tissue. The aim of this study was to determine if the ingestion of depleted uranium led to changes in the gut immune system of the intestine. The experiments were performed at 1 and 3 d following a per os administration of DU to rats at sublethal dose (204 mg/kg). Several parameters referring to the immune status, such as gene and protein expressions of cytokines and chemokines, and localization and density of immune cell populations, were assessed in the intestine. In addition, the overall toxicity of DU on the small intestine was estimated in this study, with histological appearance, proliferation rate, differentiation pattern, and apoptosis process. Firstly, the results of this study indicated that DU was not toxic for the intestine, as measured by the proliferation, differentiation, and apoptosis processes. Concerning the immune properties of the intestine, the ingestion of depleted uranium induced some changes in the production of chemokines and in the expression of cytokines. A diminished production of monocyte chemoattractant protein-1 (MCP-1) was noted at 1 day post exposure. At 3 d, the increased gene expression of interferon gamma (IFNgamma) was associated with an enhanced mRNA level of Fas ligand, suggesting an activation of the apoptosis pathway. However, no increased apoptotic cells were observed at 3 d in the contaminated animals. There were no changes in the localization and density of neutrophils, helper T lymphocytes, and cytotoxic T lymphocytes after DU administration. In conclusion, these results suggest that depleted uranium is not toxic for the intestine after acute exposure. Nevertheless, DU seems to modulate the expression and/or production of cytokines (IFNgamma) and chemokines (MCP-1) in the intestine. Further experiments need to be performed to determine if a chronic contamination at low dose leads in the long term to modifications of cytokines/chemokines patterns, and to subsequent changes in immune response of the intestine.

Short-term hepatic effects of depleted uranium on xenobiotic and bile acid metabolizing cytochrome P450 enzymes in the rat.

The toxicity of uranium has been demonstrated in different organs, including the kidneys, skeleton, central nervous system, and liver. However, few works have investigated the biological effects of uranium contamination on important metabolic function in the liver. In vivo studies were conducted to evaluate its effects on cytochrome P450 (CYP) enzymes involved in the metabolism of cholesterol and xenobiotics in the rat liver. The effects of depleted uranium (DU) contamination on Sprague-Dawley were measured at 1 and 3 days after exposure. Biochemical indicators characterizing liver and kidney functions were measured in the plasma. The DU affected bile acid CYP activity: 7alpha-hydroxycholesterol plasma level decreased by 52% at day 3 whereas microsomal CYP7A1 activity in the liver did not change significantly and mitochondrial CYP27A1 activity quintupled at day 1. Gene expression of the nuclear receptors related to lipid metabolism (FXR and LXR) also changed, while PPARalpha mRNA levels did not. The increased mRNA levels of the xenobiotic-metabolizing CYP3A enzyme at day 3 may be caused by feedback up-regulation due to the decreased CYP3A activity at day 1. CAR mRNA levels, which tripled on day 1, may be involved in this up-regulation, while mRNA levels of PXR did not change. These results indicate that high levels of depleted uranium, acting through modulation of the CYP enzymes and some of their nuclear receptors, affect the hepatic metabolism of bile acids and xenobiotics.

The European Radiobiology Archives (ERA)--content, structure and use illustrated by an example.

The European Radiobiology Archives (ERA), supported by the European Commission and the European Late Effect Project Group (EULEP), together with the US National Radiobiology Archives (NRA) and the Japanese Radiobiology Archives (JRA) have collected all information still available on long-term animal experiments, including some selected human studies. The archives consist of a database in Microsoft Access, a website, databases of references and information on the use of the database. At present, the archives contain a description of the exposure conditions, animal strains, etc. from approximately 350,000 individuals; data on survival and pathology are available from approximately 200,000 individuals. Care has been taken to render pathological diagnoses compatible among different studies and to allow the lumping of pathological diagnoses into more general classes. 'Forms' in Access with an underlying computer code facilitate the use of the database. This paper describes the structure and content of the archives and illustrates an example for a possible analysis of such data.

Absorption of uranium through the entire gastrointestinal tract of the rat.

The aim was to determine the gastrointestinal segments preferentially implicated in the absorption of uranium. The apparent permeability to uranium (233U) was measured ex vivo in Ussing chambers to assess uranium passage in the various parts of the small and large intestines. The transepithelial electrical parameters (potential difference, short-circuit current, transepithelial resistance and tissue conductance) were also recorded for each segment. Determination of in vivo uranium absorption after in-situ deposition of 233U in digestive segments (buccal cavity, ileum and proximal colon) and measurements of uranium in peripheral blood were then made to validate the ex vivo results. In addition, autoradiography was performed to localize the presence of uranium in the digestive segments. The in vivo experiments indicated that uranium absorption from the digestive tract was restricted to the small intestine (with no absorption from the buccal cavity, stomach or large intestine). The apparent permeability to uranium measured with ex vivo techniques was similar in the various parts of small intestine. In addition, the experiments demonstrated the existence of a transcellular pathway for uranium in the small intestine. The study indicates that uranium absorption from the gastrointestinal tract takes place exclusively in the small intestine, probably via a transcellular pathway.

In vivo effects of chronic contamination with depleted uranium on CYP3A and associated nuclear receptors PXR and CAR in the rat.

In addition to its natural presence at high concentrations in some areas, uranium has several civilian and military applications that could cause contamination of human populations, mainly through chronic ingestion. Reports describe the accumulation of this radionuclide in some organs (including the bone, kidney, and liver) after acute or chronic contamination and show that it produces chemical or radiological toxicity or both. The literature is essentially devoid of information about uranium-related cellular and molecular effects on metabolic functions such as xenobiotic detoxification. The present study thus evaluated rats chronically exposed to depleted uranium in their drinking water (1mg/(ratday)) for 9 months. Our specific aim was to evaluate the hepatic and extrahepatic mRNA expression of CYP3A1/A2, CYP2B1, and CYP1A1 as well as of the nuclear receptors PXR, CAR, and RXR in these rats. CYP3A1 mRNA expression was significantly higher in the brain (200%), liver (300%), and kidneys (900%) of exposed rats compared with control rats, while CYP3A2 mRNA levels were higher in the lungs (300%) and liver (200%), and CYP2B1 mRNA expression in the kidneys (300%). Expression of CYP1A1 mRNA did not change significantly during this study. PXR mRNA levels increased in the brain (200%), liver (150%), and kidneys (200%). Uranium caused CAR mRNA expression in the lungs to double. Expression of RXR mRNA did not change significantly in the course of this study, nor did the hepatic activity of CYP2C, CYP3A, CYP2A, or CYP2B. Uranium probably affects the expression of drug-metabolizing CYP enzymes through the PXR and CAR nuclear receptors. These results suggest that the stimulating effect of uranium on these enzymes might lead to hepatic or extrahepatic toxicity (or both) during drug treatment and then affect the entire organism.

Screening of a large panel of gastrointestinal peptide plasma levels is not adapted for the evaluation of digestive damage following irradiation.

The aim of this study was to assess the potential of gastrointestinal peptide plasma levels as biomarkers of radiation-induced digestive tract damage. To this end, plasma levels of substance P, GRP, motilin, PYY, somatostatin-28, gastrin, and neurotensin were followed for up to 5 days in pigs after a 16-Gy whole-body X-irradiation, completed by a histopathological study performed at 5 days. Each peptide gave a specific response to irradiation. The plasma levels of GRP and substance P were not modified by irradiation exposure; neither were those of motilin and PYY. Concerning gastrin, a 2-3-fold increase of plasma concentration was observed in pig, which presented the most important histological alterations of the stomach. The plasma levels of somatostatin, unchanged from 1 to 4 days after irradiation, was also increased by 130% at 5 days. In contrast, a diminution of neurotensin plasma levels was noted, firstly at 1 day (-88%), and from 3 days after exposure (-50%). The present study suggested that changes in gastrin and neurotensin plasma levels were associated with structural alterations of the stomach and ileum, respectively, indicating that they may be relevant biological indicators of radiation-induced digestive damage to these segments.

Does mean lung dose calculated after inhalation of alpha emitters actually reflect the risk of induction of malignant lung tumours?

A comparison of incidence of lung tumours in rats after inhalation exposure to aerosols containing alpha emitters which have different physico-chemical properties has been performed. Aerosols of radon and progeny, uranium ore dust, NpO2, PuO2 or Cm2O3 were considered for intercomparison with similar or different particle sizes. Dose-effect relationships for the frequency of malignant lung tumours appear linear up to a few Gy and then become infralinear at higher doses delivered to the lungs. The initial slope of the curves reflects the risk of induction of a lung tumour. The highest slopes of incidence were observed for radon and uranium ore dust (about 70 and 20% Gy(-1) respectively) for which the most homogeneous alpha dose distribution to the lungs is expected. In a general trend, increasing the alpha-activity of deposited particles (higher specific activity of constituent radioisotopes or larger particle size) decreases the risk. The comparison of the reported data shows that the risk per Gy at 'low doses' could vary over more than one order of magnitude depending on the physico-chemical properties of the aerosols.

Evaluation of risk factors for lung tumour induction in rats exposed to either NpO(2) or PuO(2) aerosols.

PURPOSE: To compare the incidence of each lung tumour type after inhalation exposure of rats to either NpO(2) or industrial PuO(2) aerosols, which have a similar size. MATERIALS AND METHODS: Male Sprague-Dawley rats were exposed once and followed during their whole life span. At the end of their life, the whole lungs were fixed, embedded and cut into thin sections for histological analysis. The presence of tumours was evaluated on three distinct levels of the lobes for phenotype determination to establish dose-effect relationships. RESULTS: In the range of lung doses studied (0.05 to more than 50 Gy), the general trend was an increased frequency of all types of tumours after inhalation exposure to neptunium compared with plutonium. The linearity of the lower part of the dose-effect relationships for all malignant lung tumours leads to the conclusion that NpO(2) is 3.3-fold more carcinogenic than PuO(2). CONCLUSIONS: According to a linear extrapolation of the data on malignant lung tumour incidence collected among all studies reported on actinide oxide carcinogenesis, the risk of lung tumour appears to vary over a factor of about 10 depending on the nature and/or size of the aerosol. This variation has to be taken into account for a realistic assessment of tumour risk.

[The effect of dose distribution on risk of lung cancer after inhalation of actinide oxides]. / Influence de la distribution de dose sur le risque d'apparition de cancers pulmonaires après inhalation d'oxydes d'actinides.

The aim of this work was to estimate risk of lung tumour occurrence after inhalation of actinide oxides from published studies and rat studies in progress. For the same delivered dose, the risk increases when homogeneity of irradiation increases, i.e., the number of particles deposited after inhalation increases (small particles and (or) low specific alpha activity). The dose-effect relationships reported appear linear up to a few gray, depending on the aerosol considered, and then the slope decreases. This slope, which corresponds with the risk, can vary over one order of magnitude depending on the aerosol used. An effective threshold at about 1 Gy was not observed for the most homogeneous dose distributions. A dosimetric and biological approach is proposed to provide a more realistic risk estimate.

Survival, lung clearance, dosimetry and gross pathology of rats exposed to either NpO2 or PuO2 aerosols.

PURPOSE: To compare survival, lung dosimetry and gross pathology after inhalation exposure of rats to either NpO2 or industrial PuO2 aerosols with similar granulometric parameters. Because the specific alpha activity ratio Pu/Np is about 600, a much more homogeneous lung irradiation was expected for NpO2. MATERIALS AND METHODS: Male Sprague Dawley rats were exposed once and their lung burdens were measured by X-ray spectrometry at different times post-exposure up to death. The time-course of doses delivered to the lungs were estimated, taking into account individual lung clearance parameters and body and lung weights. Gross lung pathologies were scored at autopsy. RESULTS: In the range of initial lung deposits (ILD) studied (0.1-4 kBq), lung clearance impairment and reduced lifespan were only observed after exposure to NpO2. For similar ILD or doses, the highest incidences of lung lesions assumed to be tumours were observed for NpO2 with a saturation of lung tumour induction for doses larger than 8 Gy (ILD: 1.5kBq). Up to 22Gy (ILD: 3.5kBq), such saturation was not observed for PuO2. CONCLUSIONS: NpO2 appears much more toxic than PuO2. Before saturation, lung tumour incidence increased nearly linearly with dose, the slope of the curve for NpO2 being about twice as steep as that for PuO2.

Lung carcinogenesis in rats after inhalation exposure to (237)NpO2.

The results of several studies of experimental carcinogenesis suggest that, after inhalation of alpha-particle emitters, lung tumor incidence varies depending on the exposure rate and dose distribution in the tissue. In the case of transuranics, the main influencing factor would be the specific alpha-particle activity of the inhaled actinide. To confirm these results, long-term studies were performed using male Sprague-Dawley rats exposed to (237)NpO(2) by inhalation. The initial lung burdens of the animals ranged from 0. 1 to about 7 kBq. The rats were followed during their life span and weighed regularly, and their lung burdens were determined in vivo and at death to estimate the lung dose. At death, the incidence of lung tumors and their malignancy and histological types were analyzed. The analysis revealed a typically linear-quadratic dose response for incidence of malignant lung neoplasm and a differential dose response for various types of tumors. Although these results confirm the influence of the activity of the inhaled actinide oxide, further experiments are needed to be able to compare a more homogeneous population of animals.