DNA methylation and potential multigenerational epigenetic effects linked to uranium chronic low-dose exposure in gonads of males and females rats.

INTRODUCTION: An increased health problem in industrialised countries is the contemporary concern of public and scientific community as well. This has been attributed in part to accumulated environmental pollutants especially radioactive substances and the use of nuclear power plants worldwide. However, the outcome of chronic exposure to low doses of a radionuclide such as uranium remains unknown. Recently, a paradigm shift in the perception of risk of radiotoxicology has emerged through investigating the possibility of transmission of biological effects over generations, in particular by epigenetic pathways. These processes are known for their crucial roles associated with the development of several diseases. OBJECTIVE: The current work investigates the epigenetic effect of chronic exposure to low doses of uranium and its inheritance across generations. Materials and Methods To test this proposition, a rodent multigenerational model, males and females, were exposed to a non-toxic concentration of uranium (40mgL-1 drinking water) for nine months. The uranium effects on were evaluated over three generations (F0, F1 and F2) by analysing the DNA methylation profile and DNMT genes expression in ovaries and testes tissues. RESULTS: Here we report a significant hypermethylation of testes DNA (p <0.005) whereas ovaries showed hypomethylated DNA (p <0.005). Interestingly, this DNA methylation profile was significantly maintained across generations F0, F1 and F2. Furthermore, qPCR results of both tissues imply a significant change in the expression of DNA methyltransferase genes (DNMT 1 and DNMT3a/b) as well. CONCLUSION: Altogether, our work demonstrates for the first time a sex-dependance and inheritance of epigenetic marks, DNA methylation, as a biological response to the exposure to low doses of uranium. However, it is not clear which type of reproductive cell type is more responsive in this context.

Assessment of the Central Effects of Natural Uranium via Behavioural Performances and the Cerebrospinal Fluid Metabolome.

Natural uranium (NU), a component of the earth's crust, is not only a heavy metal but also an alpha particle emitter, with chemical and radiological toxicity. Populations may therefore be chronically exposed to NU through drinking water and food. Since the central nervous system is known to be sensitive to pollutants during its development, we assessed the effects on the behaviour and the cerebrospinal fluid (CSF) metabolome of rats exposed for 9 months from birth to NU via lactation and drinking water (1.5, 10, or 40 mg·L(-1) for male rats and 40 mg·L(-1) for female rats). Medium-term memory decreased in comparison to controls in male rats exposed to 1.5, 10, or 40 mg·L(-1) NU. In male rats, spatial working memory and anxiety- and depressive-like behaviour were only altered by exposure to 40 mg·L(-1) NU and any significant effect was observed on locomotor activity. In female rats exposed to NU, only locomotor activity was significantly increased in comparison with controls. LC-MS metabolomics of CSF discriminated the fingerprints of the male and/or female NU-exposed and control groups. This study suggests that exposure to environmental doses of NU from development to adulthood can have an impact on rat brain function.

Influence of depleted uranium on hepatic cholesterol metabolism in apolipoprotein E-deficient mice.

Depleted uranium (DU) is uranium with a lower content of the fissile isotope U-235 than natural uranium. It is a radioelement and a waste product from the enrichment process of natural uranium. Because of its very high density, it is used in the civil industry and for military purposes. DU exposure can affect many vital systems in the human body, because in addition to being weakly radioactive, uranium is a toxic metal. It should be emphasized that, to be exposed to radiation from DU, you have to eat, drink, or breathe it, or get it on your skin. This particular study is focusing on the health effects of DU for the cholesterol metabolism. Previous studies on the same issue have shown that the cholesterol metabolism was modulated at molecular level in the liver of laboratory rodents contaminated for nine months with DU. However, this modulation was not correlated with some effects at organs or body levels. It was therefore decided to use a "pathological model" such as hypercholesterolemic apolipoprotein E-deficient laboratory mice in order to try to clarify the situation. The purpose of the present study is to assess the effects of a chronic ingestion (during 3 months) of a low level DU-supplemented water (20 mg L(-1)) on the above mentioned mice in order to determine a possible contamination effect. Afterwards the cholesterol metabolism was studied in the liver especially focused on the gene expressions of cholesterol-catabolising enzymes (CYP7A1, CYP27A1 and CYP7B1), as well as those of associated nuclear receptors (LXRα, FXR, PPARα, and SREBP 2). In addition, mRNA levels of other enzymes of interest were measured (ACAT 2, as well as HMGCoA Reductase and HMGCoA Synthase). The gene expression study was completed with SRB1 and LDLr, apolipoproteins A1 and B and membrane transporters ABC A1, ABC G5. The major effect induced by a low level of DU contamination in apo-E deficient mice was a decrease in hepatic gene expression of the enzyme CYP7B1 (-23%) and nuclear receptors LXRα (-24%), RXR (-32%), HNF4α (-21%) when compared to unexposed ones. These modifications on cholesterol metabolism did not lead to increased disturbances that are specific for apolipoprotein E-deficient mice, suggesting that chronic DU exposure did not worsen the pathology in this experimental model. In conclusion, the results of this study indicate that even for a sensitive pathologic model the exposure to a low dose of DU has no relevant impact. The results confirm the results of our first study carried out on healthy laboratory rodents where a sub-chronic contamination with low dose DU did not affect in vivo the metabolism of cholesterol.

Cholesterol 7alpha-hydroxylase (CYP7A1) activity is modified after chronic ingestion of depleted uranium in the rat.

Depleted uranium (DU) is a radioactive heavy metal derived from the nuclear energy production. Its wide use in civilian and military items increases the risk of its environmental dissemination, and thus the risk of internal contamination of populations living in such contaminated territories. Previous studies have shown that vitamin D and cerebral cholesterol metabolisms were affected following chronic ingestion of DU. Even more than the brain, the liver is a crucial organ in cholesterol homeostasis since it regulates cholesterol distribution and elimination at body level. The aim of this work was to assess the impact of a low-level chronic ingestion of DU on hepatic cholesterol metabolism. Rats were contaminated with DU in their drinking water at a concentration of 40mg/l for 9 months. The major effect induced by DU was a decrease of CYP7A1 specific activity (-60%) correlated with a matching decrease of its product 7alpha-hydroxycholesterol in the plasma. Hepatic gene expression of transporters ABC A1, ABC G5, ABC G8 and of nuclear receptor RXR was increased, whereas that of catabolism enzyme CYP7B1 was decreased. Thus, after a chronic ingestion of DU, rats experience a modulation of cholesterol catabolism but overcome it, since their cholesterolemia is preserved and no pathology is declared.

[Uranium: properties and biological effects after internal contamination]. / Uranium: propriétés et effets biologiques après contamination interne.

Uranium is a radionuclide present in the environment since the origin of the Earth. In addition to natural uranium, recent deposits from industrial or military activities are acknowledged. Uranium's toxicity is due to a combination of its chemical (heavy metal) and radiological properties (emission of ionizing radiations). Acute toxicity induces an important weight loss and signs of renal and cerebral impairment. Alterations of bone growth, modifications of the reproductive system and carcinogenic effects are also often seen. On the contrary, the biological effects of a chronic exposure to low doses are unwell known. However, results from different recent studies suggest that a chronic contamination with low levels of uranium induces subtle but significant levels. Indeed, an internal contamination of rats for several weeks leads to detection of uranium in many cerebral structures, in association with an alteration of short-term memory and an increase of anxiety level. Biological effects of uranium on the metabolisms of xenobiotics, steroid hormones and vitamin D were described in the liver, testis and kidneys. These recent scientific data suggest that uranium could participate to increase of health risks linked to environmental pollution.

Enriched uranium affects the expression of vitamin D receptor and retinoid X receptor in rat kidney.

An increasing awareness of the radiological impact of the nuclear power industry and other nuclear technologies is observed nowadays on general population. This led to renew interest to assess the health impact of the use of enriched uranium (EU). The aim of this work was to investigate in vivo the effects of a chronic exposure to EU on vitamin D(3) metabolism, a hormone essential in mineral and bone homeostasis. Rats were exposed to EU in their drinking water for 9 months at a concentration of 40 mg l(-1) (1mg/rat day). The contamination did not change vitamin D plasma level. Vitamin D receptor (vdr) and retinoid X receptor alpha (rxralpha), encoding nuclear receptors involved in the biological activities of vitamin D, showed a lower expression in kidney, while their protein levels were paradoxically increased. Gene expression of vitamin D target genes, epithelial Ca(2+) channel 1 (ecac1) and Calbindin-D28k (cabp-d28k), involved in renal calcium transport were decreased. Among the vitamin D target organs examined, these molecular modifications occurred exclusively in the kidney, which confirms that this organ is highly sensitive to uranium exposure. In conclusion, this study showed that a chronic exposure to EU affects both mRNA and protein expressions of renal nuclear receptors involved in vitamin D metabolism, without any modification of the circulating vitamin D.

In vivo effects of chronic contamination with depleted uranium on vitamin D3 metabolism in rat.

The extensive use of depleted uranium (DU) in today's society results in the increase of the number of human population exposed to this radionuclide. The aim of this work was to investigate in vivo the effects of a chronic exposure to DU on vitamin D(3) metabolism, a hormone essential in mineral and bone homeostasis. The experiments were carried out in rats after a chronic contamination for 9 months by DU through drinking water at 40 mg/L (1 mg/rat/day). This dose corresponds to the double of highest concentration found naturally in Finland. In DU-exposed rats, the active vitamin D (1,25(OH)(2)D(3)) plasma level was significantly decreased. In kidney, a decreased gene expression was observed for cyp24a1, as well as for vdr and rxralpha, the principal regulators of CYP24A1. Similarly, mRNA levels of vitamin D target genes ecac1, cabp-d28k and ncx-1, involved in renal calcium transport were decreased in kidney. In the brain lower levels of messengers were observed for cyp27a1 as well as for lxrbeta, involved in its regulation. In conclusion, this study showed for the first time that DU affects both the vitamin D active form (1,25(OH)(2)D(3)) level and the vitamin D receptor expression, and consequently could modulate the expression of cyp24a1 and vitamin D target genes involved in calcium homeostasis.

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.

Effects of depleted uranium after short-term exposure on vitamin D metabolism in rat.

Uranium is a natural radioactive heavy metal. Its toxicity has been demonstrated for different organs, including bone, kidney, liver and brain. Effects of an acute contamination by depleted uranium (DU) were investigated in vivo on vitamin D(3) biosynthetic pathway. Rats received an intragastric administration of DU (204 mg/kg) and various parameters were studied either on day 1 or day 3 after contamination. Cytochrome P450 (CYP27A1, CYP2R1, CYP27B1, CYP24A1) enzymes involved in vitamin D metabolism and two vitamin D(3)-target genes (ECaC1, CaBP-D9K) were assessed by real time RT-PCR in liver and kidneys. CYP27A1 activity was measured in liver and vitamin D and parathyroid hormone (PTH) level were measured in plasma. In acute treated-rats, vitamin D level was increased by 62% and decreased by 68% in plasma, respectively at day 1 and at day 3, which paralleled with a concomitant decrease of PTH level (90%) at day 3. In liver, cyp2r1 mRNA level was increased at day 3. Cyp27a1 activity decreased at day 1 and increased markedly at day 3. In kidney, cyp27b1 mRNA was increased at days 1 and 3 (11- and 4-fold respectively). Moreover, ecac1 and cabp-d9k mRNA levels were increased at day 1 and decreased at day 3. This work shows for the first time that DU acute contamination modulates both activity and expression of CYP enzymes involved in vitamin D metabolism in liver and kidney, and consequently affects vitamin D target genes levels.

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.

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.