Renal adaptive response to exposure to low doses of uranyl nitrate and sodium fluoride in mice.

BACKGROUND: Despite their differences in physicochemical properties, both uranium (U) and fluoride (F) are nephrotoxicants at high doses but their adverse effects at low doses are still the subject of debate. METHODS: This study aims to improve the knowledge of the biological mechanisms involved through an adaptive response model of C57BL/6 J mice chronically exposed to low priming doses of U (0, 10, 20 and 40 mg/L) or F (0, 15, 30 and 50 mg/L) and then challenged with acute exposure of 5 mg/kg U or 7.5 mg/kg NaF. RESULTS: We showed that an adaptive response occurred with priming exposures to 20 mg/L U and 50 mg/L F, with decreased levels of the biomarkers KIM-1 and CLU compared to those in animals that received the challenge dose only (positive control). The adaptive mechanisms involved a decrease in caspase 3/7 activities in animals exposed to 20 mg/L U and a decrease in in situ VCAM expression in mice exposed to 50 mg/L F. However, autophagy and the UPR were induced independently of priming exposure to U or F and could not be identified as adaptive mechanisms to U or F. CONCLUSION: Taken together, these results allow us to identify renal adaptive responses to U and F at doses of 20 and 50 mg/L, probably through decrease apoptosis and inflammatory cell recruitment.

Molecular, cellular, and tissue impact of depleted uranium on xenobiotic-metabolizing enzymes.

Enzymes that metabolize xenobiotics (XME) are well recognized in experimental models as representative indicators of organ detoxification functions and of exposure to toxicants. As several in vivo studies have shown, uranium can alter XME in the rat liver or kidneys after either acute or chronic exposure. To determine how length or level of exposure affects these changes in XME, we continued our investigation of chronic rat exposure to depleted uranium (DU, uranyl nitrate). The first study examined the effect of duration (1-18 months) of chronic exposure to DU, the second evaluated dose dependence, from a level close to that found in the environment near mining sites (0.2 mg/L) to a supra-environmental dose (120 mg/L, 10 times the highest level naturally found in the environment), and the third was an in vitro assessment of whether DU exposure directly affects XME and, in particular, CYP3A. The experimental in vivo models used here demonstrated that CYP3A is the enzyme modified to the greatest extent: high gene expression changed after 6 and 9 months. The most substantial effects were observed in the liver of rats after 9 months of exposure to 120 mg/L of DU: CYP3A gene and protein expression and enzyme activity all decreased by more than 40 %. Nonetheless, no direct effect of DU by itself was observed after in vitro exposure of rat microsomal preparations, HepG2 cells, or human primary hepatocytes. Overall, these results probably indicate the occurrence of regulatory or adaptive mechanisms that could explain the indirect effect observed in vivo after chronic exposure.

Increased urinary excretion of albumin, hemopexin, transferrin and VDBP correlates with chronic sensitization to gentamicin nephrotoxicity in rats.

Drug nephrotoxicity is a serious health and economic problem worldwide. Rats can be acutely sensitized to acute kidney injury (AKI) by subnephrotoxic treatments with potentially nephrotoxic drugs. Acquired sensitization to AKI poses a silent risk impossible to diagnose pre-emptively with the technology available at the clinical level. Herein, we hypothesized whether a chronic, subnephrotoxic insult to the kidneys might result in chronically acquired sensitization to AKI, and whether chronic sensitization might be detected through specific urinary markers. To this end, rats were treated with a subtoxic dosage of the experimental nephrotoxin uranyl nitrate (UN) in the drinking water for 21 weeks, or plain water (as control), and then with low-dose gentamicin for 7 days. Renal function and renal tissue damage were evaluated through the experiment. The mild renal damage caused by gentamicin was markedly magnified in rats having received UN chronically, which was evident both at the functional and histological level. Four proteins, namely albumin, hemopexin, transferrin and vitamin D binding protein were increased in the urine in temporal association with the appearance of chronic predisposition. Although further studies are necessary, our results suggest that these proteins might be potentially used as markers of hidden, chronic predisposition to gentamicin nephrotoxicity, in order to appropriately and pre-emptively stratify and handle individuals according to their specific risk in the long term, and to conveniently optimize their life conditions or additional clinical procedures or treatments that might trigger the disease. This might reduce AKI incidence and severity and the associated costs.

Uranium in drinking water: effects on mouse oocyte quality.

The aim of this work was to evaluate the reproductive toxicological effects of uranium (U) at 2.5, 5, and 10 mgU/kg/d chronically administered in drinking water for 40 d. Swiss female control mice (n = 28) and mice chronically contaminated with uranyl nitrate in drinking water (n = 36) were tested. The number and quality of ovulated oocytes, chromatin organization, and nuclear integrity were evaluated. No significant differences were obtained in the numbers of ovulated oocytes between the different groups. Nevertheless, in 1,520 of the oocytes examined, dysmorphism increased from 11.99% in the control group to 27.99%, 27.19%, and 27.43% in each of the contaminated groups, respectively, in a dose-independent manner. On the other hand, morphological chromatin organization from 880 oocytes examined showed an increase in metaphase plate abnormalities from 37.20% (+/-7.21) in the control group to 55.13% (+/-21.36), 58.29% (+/-21.72), and 64.10% (+/-12.62) in each of the contaminated groups, respectively. Cumulus cell (CC) micronucleation, a parameter of nuclear integrity, increased from 0.21% (+/-0.31) in the control group to 1.92 (+/-0.95), 2.98 (+/-0.97), and 3.2 (+/-0.98), respectively. Both metaphase plate abnormalities and CC micronucleation showed an increase in a dose-dependent manner (r = 0.9; p < 0.001). The oocyte and its microenvironment showed high sensitivity to uranium contamination by drinking water. The lowest observed adverse effect level for this system is estimated at a level below 2.5 mgU/kg/d for female mice.

Ultrastructural and metabolic changes in osteoblasts exposed to uranyl nitrate.

Exposure to uranium is an occupational hazard to workers who continually handle uranium and an environmental risk to the population at large. Since the cellular and molecular pathways of uranium toxicity in osteoblast cells are still unknown, the aim of the present work was to evaluate the adverse effects of uranyl nitrate (UN) on osteoblasts both in vivo and in vitro. Herein we studied the osteoblastic ultrastructural changes induced by UN in vivo and analyzed cell proliferation, generation of reactive oxygen species (ROS), apoptosis, and alkaline phosphatase (APh) activity in osteoblasts exposed to various UN concentrations (0.1, 1, 10, and 100 microM) in vitro. Cell proliferation was quantified by means of the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, ROS was determined using the nitro blue tetrazolium test, apoptosis was morphologically determined using Hoechst 3332 and APh activity was assayed spectrophotometrically. Electron microscopy revealed that the ultrastructure of active and inactive osteoblasts exposed to uranium presented cytoplasmic and nuclear alterations. In vitro, 1-100 microM UN failed to modify cell proliferation ratio and to induce apoptosis. ROS generation increased in a dose-dependent manner in all tested doses. APh activity was found to decrease in 1-100 microM UN-treated cells vs. controls. Our results show that UN modifies osteoblast cell metabolism by increasing ROS generation and reducing APh activity, suggesting that ROS may play a more complex role in cell physiology than simply causing oxidative damage.

Radiation protection by deferiprone in animal models.

The effectiveness of deferiprone (L1) in removing depleted uranium (DU) and protecting animals from radiation exposure was examined. Rats that had received 2 mg/kg DU via intramuscular injection were orally administered 100, 200 or 400 mg/kg L1 for 3 days. In all of the groups, significant increases in urinary DU excretion and decreases in DU concentration in the injected muscle were observed, indicating that L1 combined with DU and DU was excreted in the urine. No significant increase in the amount of DU in the excreta or decrease in DU concentration in organs other than the muscles was found. As a preliminary test, the effectiveness of L1 in reducing radiation damage was examined in mice injected with 400 mg/kg L1 and rats administered orally with 200 and 400 mg/kg L1 before and just after x-ray exposure. The results were inconclusive.

Renal toxicogenomic response to chronic uranyl nitrate insult in mice.

Although the nephrotoxicity of uranium has been established through numerous animal studies, relatively little is known about the effects of long-term environmental uranium exposure. Using a combination of conventional biochemical studies and serial analysis of gene expression (SAGE), we examined the renal responses to uranyl nitrate (UN) chronic exposure. Renal uranium levels were significantly increased 4 months after ingestion of uranium in drinking water. Creatinine levels in serum were slightly but significantly increased compared with those in controls. Although no further significant differences in other parameters were noted, substantial molecular changes were observed in toxicogenomic profiles. UN induced dramatic alterations in expression levels of more than 200 genes, mainly up-regulated, including oxidative-response-related genes, genes encoding for cellular metabolism, ribosomal proteins, signal transduction, and solute transporters. Seven differentially expressed transcripts were confirmed by real-time quantitative polymerase chain reaction. In addition, significantly increased peroxide levels support the implication of oxidative stress in UN toxicant response. This report highlights the potential of SAGE for the discovery of novel toxicant-induced gene expression alterations. Here, we present, for the first time, a comprehensive view of renal molecular events after uranium long-term exposure.

Renal interstitial fibrosis (RIF): II. Ultrasound follow up study of single uranyl nitrate administration causing renal dysfunction in rats--comparison with histologic and functional renal parameters.

A single administration of uranyl nitrate (UN; 0.5 mg/100 g b. wt. i.p.) to adult female Wistar rats reliably induces renal interstitial fibrosis (RIF) providing an experimental model to investigate therapeutic strategies. It was the aim of this study to further characterise a rat model of UN induced RIF which we have studied previously (Appenroth et al. 2001) by the comparison of functional parameters with ultrasonographic examination over a period of 30 weeks after injury. In the acute phase after UN administration (between days 2 and 17) signs of inflammation (increase in renal blood flow, swelling of renal cortex, enlargement of renal pelvis) could be detected by ultrasound. After four weeks UN led to functional changes (decreased creatinine clearance, increased urinary protein excretion and increased OH-proline concentration in renal tissue). In vitro, the accumulation of p-aminohippurate and the gluconeogenesis were reduced. In accordance with the functional changes, distinct ultrasonographic abnormalities could be seen between weeks 10 and 30 after UN with regard to changes in kidney size and shape, reduced renal perfusion and enlargement of renal pelvis. The sensitivity of ultrasonography in small laboratory animals is limited and most useful for follow-up studies of acute renal changes after administration of nephrotoxins. Ultrasonography can not be recommended for non-invasive screening of the progression of chronic renal failure.

Influence of bisphosphonate on the negative erythropoietic effects of uranyl nitrate.

Uranium salts, such as uranyl nitrate, induce severe renal dysfunction and tubular necrosis and a significant impairment of both oxygen dependent erythropoietin production and response to recombinant human erythropoietin. All effects are transient and reach maximal severity on the 7th day post injection. We investigated the effects of ethane 1-hydroxy-1, 1-bisphosphonate, which counteracts the inhibitory effect of uranyl nitrate on bone formation, on the negative erythropoietic effects of uranyl nitrate. Adult female Wistar rats received 1 mg/kg body weight of uranyl acetate by the i.v. route. Ethane 1-hydroxy-1,1-bisphosphonate was injected simultaneously at a dose of 7.5 mg/kg by the same route. Seven days after drug injections, plasma erythropoietin was estimated after hypobaric hypoxemia or cobalt chloride administration. The response to exogenous erythropoietin was also measured in uranyl nitrate- and/or ethane 1-hydroxy-1,1-bisphosphonate-injected rats made polycythemic by transfusion. The erythroid response was quantitated in terms of red blood cell 59iron uptake. Ethane 1-hydroxy-1, 1-bisphosphonate counteracted the effect of uranyl nitrate on oxygen-dependent and cobalt-dependent erythropoietin production, but did not correct the right shift of the dose-response relationship for exogenous erythropoietin induced by uranyl nitrate in the polycythemic rat.

Hormesis in effect of low uranium doses on organisms.

Uranium nitrate applied daily in small amounts per os (0.5-17 mg/l in drinking water) or per cuttim (0.2-0.8 g/kg) exerts positive effect on metabolism of phosphate ions by red blood cells in vitro. This positive effect changes to the negative one with increasing dose or exposure time. It may be assumed that most other cells in the organism are affected in the same way as erythrocytes and that similar effect might be observed with other harmful agents or substances.

Bone growth is impaired by uranium intoxication.

Acute and chronic uranium intoxication leads to the inhibition of bone formation and impaired bone modeling and remodeling. As these are processes directly involved in bone growth the aim of this paper is to present a biometric study of bone growth--tibiae and mandibles of rats intoxicated with uranium. Wistar ratios weighing 60-80 g were used as follows, a) one intraperitoneal injection (IPI, 2 mg/Kg of body weight)) of uranyl nitrate; b) 30 daily applications on the dorsal skin of aliquots of a mixture of U308, concentrated at 2% and at 4%--percutaneous absorption(PA)-. Tibia and mandible length were smaller in both experimental groups than in their respective controls. Some of the mandibular parameters were lower in intoxicated animals than their controls which in turn results in the alteration of the mandibular shape. We conclude that impairment in bone growth can be achieved by uranium intoxication.

Ethane 1-hydroxy-1, 1-diphosphonate (EHDP) counteracts the inhibitory effect of uranyl nitrate on bone formation.

The beneficial effect of ethane 1-hydroxy-1, 1-diphosphonate (EHDP) in restoring the inhibition of bone formation in cases of acute uranium intoxication is presented. Bone formation was studied histomorphometrically in a model of alveolar bone healing. After tooth extraction, 40 rats were divided into 4 groups that received (1) no further treatment, (2) 10 daily intraperitoneal injections of 7.5 mg/kg of body weight of EHDP, (3) an intraperitoneal injection of 2.0 mg/kg of body weight of uranyl nitrate, and (4) the same treatment as was provided rats in groups 2 and 3. The results showed that the healing of bone did not occur in exposed animals, whereas healing in EHDP-treated exposed animals did not differ from that of nonexposed controls. This effect might result from a blocking and/or competitive action of EHDP and/or the stimulation that EHDP elicits at the doses and in the administration period studied.

Differential effects of the degree of renal damage on p-aminohippuric acid and inulin clearances in rats.

Kidney disease is generally thought to affect all segments of a nephron equally. Bricker and co-workers first proposed this as the Intact Nephron hypothesis in 1971, and evidence to date has usually supported this hypothesis. However, most supporting studies have involved severe renal failure, which may not be suitable to differentiate effects on functional sites or to test the hypothesis. The work included here examines the effects of limited renal failure on two separate functions of the nephron: glomerular filtration, as measured by inulin clearance and proximal tubular organic anion secretory function, as measured by p-aminohippuric acid (PAH) clearance. Renal failure was induced in rats by intravenous administration of uranyl nitrate, a nephrotoxin. Doses used were 0.3, 1.0, and 3.0 mg/kg rat body weight. Five days later, rats were given an intravenous infusion of PAH and inulin. Renal clearance of each compound was calculated. Results obtained in these experiments show that, at the lowest uranyl nitrate dose, PAH clearance was significantly decreased but inulin clearance was not. The ratio of CLPAH/CLIN was decreased from 2.55 in control rats to 1.21 in rats given the low dose of nephrotoxin. At higher uranyl nitrate doses, both clearance rates were significantly decreased and the ratio of CLPAH/CLIN remained close to 1.0. These results indicate that the active transport functions of the nephron can be differentiated from passive transport functions. Caution should be exercised in extrapolating renal disease changes in active renal secretion to changes in passive renal elimination and the reverse.

Absorption and biokinetics of U in rats following an oral administration of uranyl nitrate solution.

The absorption of U within the male Wistar rat was determined following oral gavage with uranyl nitrate solutions at seven different dosages. Gavage levels ranged from 0.003 to 45 mg U per kilogram body weight. Uranium tissue burdens were determined at 0.25, 0.5, 1, 2, 4, 8, 24, 48, 96 and 240 h following gavage. Blood, kidney, liver and bone were analyzed for U content using neutron activation followed by delayed neutron counting. Uranium rapidly localized in the kidneys and bone following ingestion. Bone was found to be the primary tissue of deposition. Skeletal and kidney burdens closely paralleled each other from 15 min to 10 d after oral gavage. Uranium burdens in the blood reached a maximum within 30 min but declined rapidly thereafter. Burdens of all tissues were well correlated with each other and with dosage at all dose levels. Equations relating body burdens with blood levels were developed and found to be useful for predicting body burdens for the initial 8 h following gavage. Gastrointestinal absorption (f1) was 0.6-2.8% over the range of U administered. Movement of U through the GI tract was assessed at two dosages. The transit time of U through the GI tract was approximately 48 h. Uranium loss from the stomach was described as a power function of time. The maximum value in the small intestine was attained within 2 h, and thereafter its rapid loss was linear up to 8 h. A minor residual loss component from the small intestine was evident beyond 8 h post-gavage.

Deposition and early disposition of inhaled 233UO2(NO3)2 and 232UO2(NO3)2 in the rat.

Little information exists on the metabolism and potential health effects of 233U and 232U, high-specific-activity U isotopes associated with Th breeder systems. This paper describes the distribution and retention of the two isotopes following inhalation of uranyl nitrate, a simulated process solution. The lungs of rats exposed to 233UO2(NO3)2 and 232UO2(NO3)2 aerosols contained from 7 to 23% of the total amount of U retained in the rat after a 30-min inhalation exposure. Uranium was translocated rapidly from the lung and was retained mainly in skeleton, kidney and liver. Amounts equivalent to from one-quarter to one-half the initial lung burden (ILB) of U were excreted in urine the first day after inhalation. Radiation dose estimates based on 233U and 232U retention kinetics indicate that lung and skeleton would be the target organs for delayed radiation effects.

Morphologic changes in uranyl nitrate-induced acute renal failure in saline- and water-drinking rats.

The sequential changes in renal morphology that occurred for 5 subsequent days after a subcutaneous injection of uranyl nitrate (10 mg. per kg.) were examined in saline- and water-drinking rats using light microscopy, transmission electron microscopy, and scanning electron microscopy. The cortical proximal tubule exhibited diffuse focal brush border loss and increased vacuolization by 1 hour after administration of the nephrotoxin. By 5 days, the P2 and P3 segments were completely necrotic. Cells of P1 segments accumulated large vacuoles throughout their cytoplasm, and distal nephron segments exhibited considerable cellular swelling and vacuolization. Scanning electron microscopy revealed abnormalities in glomerular epithelial cells similar to those seen in humans with chronic renal disease and in experimental animal models characterized by proteinuria. There was essentially no difference in the morphologic response of saline- and water-drinking rats. Although uranyl nitrate administered at this dosage resulted in the relatively slow development of tubular necrosis, changes in renal morphology could be seen within an hour and progressed insidiously throughout the study with little evidence of regeneration.

[Protective action of contrical in rats with experimental nephritis]. / Zashchitnoe deistvie kontrikala pri éksperimental'nom nefrite u krys.

It has been shown in rats with experimental uranyl-glycerin nephritis that contrykal relieves the severity of the disease acute phase. As compared with untreated animals, this fact is confirmed by the reduced albuminuria and blood urea, as well as by the absence of decreased filtration characteristic of glomerulonephritis. It has been also discovered that contrykal normalizes the urinary sediment, the blood urea and creatinine levels, and eliminates albuminuria and natriuresis.

Chronic uremia syndrome in dogs induced with uranyl nitrate.

11 adult female dogs were given periodic intravenous injections of uranyl nitrate [UO2(NO3)2 . 6H2O] to create a syndrome of chronic uremia. Initially, dogs usually received 2.0 mg/kg of uranyl nitrate; subsequent doses were generally less. After the initial injection, there was an abrupt fall in creatinine clearance and rise in plasma urea nitrogen. Low and relatively constant creatinine clearances (10.2 +/- SD 2.7 ml/min) were easily maintained with further injections. Dogs developed proteinuria, aminoaciduria, weight loss, and plasma amino acid levels similar to those of chronically uremic humans and rats. With creatinine clearances of 4 ml/min or less, dogs became listless and lethargic, and daily activity and food intake decreased. Repeated injections of uranyl nitrate appear to be an easy and reliable method for creating a model of chronic uremia in dogs.

Serum binding of quinidine in experimental acute renal failure.

Quinidine serum binding is increased in some patients with acute renal failure, but in animals with renal failure conflicting results were published. Therefore, serum binding of quinidine, papaverine and phenylbutazone was studied by equilibrium dialysis in rabbits and rats with acute renal failure induced either by injection of uranyl nitrate or ligation of the ureters. From the results it appears that, in animals, quinidine binding changes are different according to the model used for induction of the renal failure, regardless of the species studied. After ligation of the ureters, lipoprotein concentration increases, but the meaning of this increase for increased serum quinidine binding is not clear.

Renal glomerular and tubular lesions in rabbits treated with uranyl nitrate.

Renal injury and regeneration was followed light--and electron microscopically in rabbits treated with uranyl nitrate. The presence and importance of glomerular lesions is noteworthy.