Effects of oral exposure to silver nanoparticles on the sperm of rats.

It has been demonstrated that exposure to silver nanoparticles (AgNPs) can induce toxicological effects in rodents. In this study, we investigated whether sub-chronic oral exposure to different doses of polyvinil pyrrolidone (PVP)-coated AgNPs (PVP-AgNPs) (50, 100 and 200mg/kg/day) could induce harmful effects on epididymal sperm rat parameters. Sperm motility, viability and morphology were examined. Moreover, a histological evaluation of testis and epididymis was also performed. High doses of PVP-AgNPs showed higher sperm morphology abnormalities, while a progressive, but not significant effect, was observed in other sperm parameters. The current results suggest that oral sub-chronic exposure to PVP-AgNPs induces slight toxicological effects in sperm rat parameters.

Comparative analysis of laparoscopic and ultrasound-guided biopsy methods for gene expression analysis in transgenic goats.

The present study aimed to compare laparoscopic (LP) and ultrasound-guided (US) biopsy methods to obtain either liver or splenic tissue samples for ectopic gene expression analysis in transgenic goats. Tissue samples were collected from human granulocyte colony stimulating factor (hG-CSF)-transgenic bucks and submitted to real-time PCR for the endogenous genes (Sp1, Baff, and Gapdh) and the transgene (hG-CSF). Both LP and US biopsy methods were successful in obtaining liver and splenic samples that could be analyzed by PCR (i.e., sufficient sample sizes and RNA yield were obtained). Although the number of attempts made to obtain the tissue samples was similar (P > 0.05), LP procedures took considerably longer than the US method (P = 0.03). Finally, transgene transcripts were not detected in spleen or liver samples. Thus, for the phenotypic characterization of a transgenic goat line, investigation of ectopic gene expression can be made successfully by LP or US biopsy, avoiding the traditional approach of euthanasia.

Aluminum-induced pro-oxidant effects in rats: protective role of exogenous melatonin.

In recent years, it has been suggested that oxidative stress is a feature of Alzheimer's disease in which aluminum (Al) could exacerbate oxidative events. The goal of the present study was to assess in rats the pro-oxidant effects induced by Al exposure, as well as the protective role of exogenous melatonin. Two groups of male rats were intraperitoneally injected with Al only or melatonin only, at doses of 5 and 10 mg/kg/day, respectively for 8 wk. During this period, a third group of animals received Al (5 mg/kg/day) and melatonin (10 mg/kg/day). At the end of the treatment period, rats were anesthesized and arterial blood was obtained. Thereafter, animals were killed and liver and brain (cortex, hippocampus and cerebellum) were removed. These tissues were processed to examine oxidative stress markers: glutathione transferase (GST), reduced glutathione (GSH), oxidized glutathione (GSSG), superoxide dismutase (SOD), glutathione reductase (GR), glutathione peroxidase (GPx), catalase (CAT), thiobarbituric acid reactive substances (TBARS), as well as protein content. Samples of these tissues were also used to determine Al, Fe, Mn, Cu and Zn concentrations. The results show that Al exposure promotes oxidative stress in different neural areas, including those in which Al concentrations were not significantly increased. The biochemical changes observed in neural tissues show that Al acts as pro-oxidant, while melatonin exerts an antioxidant action in Al-treated animals. The protective effects of melatonin against cellular damage caused by Al-induced oxidative stress, together with its low toxicity, make melatonin worthy of investigation as a potential supplement to be included in the treatment of neurological disorders in which the oxidative effects must be minimized.

Interactions in developmental toxicology: effects of concurrent exposure to lead, organic mercury, and arsenic in pregnant mice.

The development toxicity of lead nitrate (25 mg/kg, SC), methylmercury chloride (12.5 mg/kg, PO), and sodium arsenite (6 mg/kg, SC) was assessed in CD1 mice following administration on gestation day 10 of these chemicals separately or in their binary and ternary combinations. Cesarean sections were performed on day 18 of gestation, and fetuses were examined for malformations and variations. Three fetuses from each dam were used for whole-body analyses of Pb, Hg, and As. Maternal toxic effects were more remarkable in the group concurrently exposed to Pb, Hg, and As than in those given binary combinations of the elements. In turn, maternal toxicity was more notable in these groups than in those given separately the test compounds. With regard to developmental toxicity, the most relevant effects (decreased fetal weight, cleft palate) corresponded to the Hg-treated groups. It is in agreement with the finding that in all experimental groups the levels of Pb and As in whole fetuses were under their respective detection limits. In general terms, the present data suggests that at the current doses, the interactive effects of Pb and As on Hg-induced developmental toxicity were not greater than additive. In contrast, exposure of pregnant mice to Pb and As at doses that were practically nontoxic to dams, concurrently with organic Hg at a toxic dose, caused supra-additive interactions in maternal toxicity.

Interactions in developmental toxicology: combined action of restraint stress, caffeine, and aspirin in pregnant mice.

BACKGROUND: Stress can result in an increased use of substances such as caffeine and aspirin. The effect of maternal stress on concurrent exposure to caffeine and aspirin on prenatal development was assessed in mice. METHODS: On gestational day 9, mice were assigned to three treatment groups orally exposed to caffeine (30 mg/kg), aspirin (250 mg/kg), or a combination of caffeine (30 mg/kg) and aspirin (250 mg/kg). Three additional groups of pregnant animals received similar caffeine and aspirin doses and were immediately subjected to restraint for 14 hr. Control groups included unrestrained and restrained pregnant mice not exposed to caffeine or aspirin. All dams were euthanized on gestational day 18. Live fetuses were evaluated for sex, body weight, and external, internal, and skeletal malformations and variations. RESULTS: A single oral dose of caffeine or aspirin did not cause significant maternal toxicity. However, coadministration of these drugs with restraint produced some adverse maternal effects (i.e., reduction in maternal weight gain and food consumption on gestational days 9-11). In relation to embryo/fetal toxicity, the incidence of some skeletal defects was significantly increased after exposure to caffeine, aspirin, or maternal restraint, and their binary and ternary combinations. CONCLUSIONS: Although caffeine and aspirin were given in a single dose in this study, the results suggest that prenatal stress could slightly exacerbate the maternal and developmental toxicity of the combination of these drugs in mice.

Haptoglobin gene expression in human glioblastoma cell lines.

Increases in cerebrospinal fluid (CSF) levels of the acute phase protein haptoglobin (Hp) occur in central nervous system (CNS) disorders such as Alzheimer's disease. To establish if Hp CSF level increases can be associated with Hp expression in brain, reverse transcription-polymerase chain reaction (RT-PCR) experiments were conducted to determine if the Hp mRNA transcript is expressed in human glioblastoma cells. Furthermore, Western blots and immunoprecipitations were performed to elucidate if Hp protein is synthesized and secreted by human glioblastoma cells. The Hp mRNA (alpha2beta) transcript (1155 bp) was detected both in U-87MG and U-138MG cells, and was positively verified by nested PCR in which a part of the beta sequence (482 bp) was targeted for amplification. Despite the presence of Hp mRNA, Hp protein was not secreted by U-87MG cells as compared to the hepatoma cell line, HepG2, where Hp protein (approximately 46 kDa) was detected in the media. The results suggest the expression of Hp protein by glioblastoma cells is possible since the Hp mRNA transcript exist, but whether or not Hp mRNA is contained in a storage pool requiring a specific signal for translation or is transiently expressed remains to be uncovered in future studies.

Effects of oral aluminum on essential trace elements metabolism during pregnancy.

The present study was conducted to assess in rats the effects of oral aluminum (Al) exposure on calcium (Ca), magnesium (Mg), manganese (Mn), copper (Cu), zinc (Zn), and iron (Fe) accumulation and urinary excretion. Three groups of plug-positive Sprague-Dawley (SD) rats were given by gavage 0, 200, and 400 mg/kg/d of Al(OH)3 on gestational days 1-20. Three groups of nonpregnant female SD rats of the same age received Al(OH)3 by gavage at the same doses for 20 consecutive days. At the end of the treatment period, 24-h urine samples were collected for analysis of Al and essential elements. Subsequently, all animals were sacrificed and samples of liver, bone, spleen, kidneys, and brain were removed for metal analyses. With some exceptions, the urinary amounts of Al, Mn, and Cu excreted by pregnant animals as well as the urinary levels of Al excreted by nonpregnant rats were higher in the Al-treated groups than in the respective control groups. Although higher Al levels were found in the liver of pregnant rats, the concentrations of Al in the brain of these animals were lower than those found in the same tissues of nonpregnant rats. With regard to the essential elements, tissue accumulation was most affected in pregnant than in nonpregnant animals. In pregnant rats, the hepatic and renal concentrations of Ca, Mg, Mn, Cu, Zn, and Fe, as well as the levels of Ca in bone, and the concentrations of Cu in brain were significantly higher in the Al-exposed groups than in the control group. According to the current results, oral Al exposure during pregnancy can produce significant changes in the tissue distribution of a number of essential elements.

Nephrotoxicity of simultaneous exposure to mercury and uranium in comparison to individual effects of these metals in rats.

Both inorganic mercury and uranium are known nephrotoxicants in mammals. In this study, the renal toxicity of a concurrent exposure to inorganic mercury and uranium was compared with the nephrotoxic effects of the individual metals in a rat model. Eight groups of rats, 10 animals per group, were subcutaneously given a single administration of mercuric chloride (HgCl2, 0.34 mg/kg and 0.68 mg/kg), uranyl acetate dihydrate (UAD, 2.5 mg/kg and 5 mg/kg), or combinations of both compounds at the same doses. A ninth group of rats received sc injections of 0.9% saline and was designated as the control group. Necrosis of proximal tubules, which was observed in all experimental groups, was the most relevant morphologic abnormality. Marked changes, which were remarkably greater than those induced by the individual elements, were noted in some urinary parameters in the groups concurrently exposed to HgCl2 and UAD. It could be an indicator of a synergistic interaction between mercury and uranium. In contrast, compared with the urinary levels found after individual administration of the highest doses of mercury and uranium, significant reductions in the urinary concentrations of these elements were noted following simultaneous exposure to both metals. At these doses, the reduction in the urinary metal excretion was also accompanied by significant decreases in the renal content of mercury and uranium. Whereas the results of some parameters pointed out a possible synergistic interaction between mercury and uranium, other measures hinted that an antagonistic interaction between these elements is also present.

A novel class of herpesvirus-encoded membrane-bound E3 ubiquitin ligases regulates endocytosis of proteins involved in immune recognition.

Kaposi's sarcoma-associated herpesvirus encodes two transmembrane proteins (modulator of immune recognition [MIR]1 and MIR2) that downregulate cell surface molecules (MHC-I, B7.2, and ICAM-1) involved in the immune recognition of infected cells. This downregulation results from enhanced endocytosis and subsequent endolysosomal degradation of the target proteins. Here, we show that expression of MIR1 and MIR2 leads to ubiquitination of the cytosolic tail of their target proteins and that ubiquitination is essential for their removal from the cell surface. MIR1 and MIR2 both contain cytosolic zinc fingers of the PHD subfamily, and these structures are required for this activity. In vitro, addition of a MIR2-glutathione S-transferase (GST) fusion protein to purified E1 and E2 enzymes leads to transfer of ubiquitin (Ub) to GST-containing targets in an ATP- and E2-dependent fashion; this reaction is abolished by mutation of the Zn-coordinating residues of the PHD domain. Thus, MIR2 defines a novel class of membrane-bound E3 Ub ligases that modulates the trafficking of host cell membrane proteins.

Evaluation of the protective activity of deferiprone, an aluminum chelator, on aluminum-induced developmental toxicity in mice.

BACKGROUND: Since deferiprone can be an effective chelating agent for the treatment of aluminum (Al) overload, in the present study we investigated whether this chelator could protect against Al-induced maternal and developmental toxicity in mice. METHODS: A single oral dose of Al nitrate nonahydrate (1,327 mg/kg) was given on gestation day 12, the most sensitive time for Al-induced maternal and developmental toxic effects in mice. At 2, 24, 48, and 72 hr thereafter, deferiprone was given by gavage at 0 and 24 mg/kg. Cesarean sections were performed on day 18 of gestation and fetuses were examined for malformations and variations. RESULTS: Aluminum-induced maternal toxicity was evidenced by significant reductions in body weight gain, corrected body weight change, and food consumption. Developmental toxicity was evidenced by a significant decrease in fetal weight per litter and an increase in the total number of fetuses and litters showing bone retardation. No beneficial effects of deferiprone on these adverse effects could be observed. By contrast, a more pronounced decrease in maternal weight gain and corrected body weight change, as well as a higher number of litters with fetuses showing skeletal variations was noted in the group exposed to Al nitrate and treated with deferiprone at 24 mg/kg. CONCLUSIONS: According to the current results, deferiprone would not be effective to prevent Al-induced maternal and embryo/fetal toxicity in mice.

Behavioral effects of aluminum in mice: influence of restraint stress.

The influence of restraint stress on potential aluminum (Al)-induced behavioral changes was assessed in CD-1 mice. Three groups of adult mice were given 0, 300 and 600 mg Al/kg body weight per day in drinking water for 2 weeks. One-half of the animals in each group were concurrently subjected to restraint stress during 1 h per day throughout the study. After cessation of treatment, open-field activity, active avoidance learning, and motor resistance and coordination of the animals were evaluated. At the end of the behavioral testing period, mice were killed and Al concentrations were determined in a number of tissues. There were no remarkable effects of Al, restraint stress or their combined administration on either open-field activity or on the number of avoidances in an automatic reflex conditioner. However, a lower motor resistance and coordination in a rotarod were observed following exposure to Al at 600 mg/kg/day, restraint alone or concurrent administration of Al (300 and 600 mg/kg/day) plus restraint stress. The levels of Al in whole brain and cerebellum were significantly enhanced in mice exposed to Al plus restraint. Although the present results scarcely show Al-induced neurobehavioral effects, the influence of restraint stress on Al levels in whole brain and cerebellum can be the basis for further studies on the potential role of this element in certain neurological disorders.

Prevention by sodium 4,5-dihydroxybenzene-1,3-disulfonate (Tiron) of vanadium-induced behavioral toxicity in rats.

Recent studies have shown that oral vanadate (V5+) administration results in behavioral toxicity in rats. The chelating agent Tiron (sodium 4,5-dihydroxybenzene-1,3-disulfonate) is an effective antidote in the removal of vanadium from vanadium-loaded rats. In this study, the protective activity of Tiron on vanadate-induced behavioral toxicity was evaluated in adult rats. Intraperitoneal treatment with Tiron at 235 or 470 mg/kg was initiated after 6 wk of oral sodium metavanadate administration (16 mg/kg/d) and continued for 2 wk. Although vanadate exposure did not result in a significant reduction in the general activity of the animals in an open field, a lower active avoidance acquisition could be observed. However, the vanadate-induced behavioral deficit was reverted by Tiron administration at 470 mg/kg. The present results suggest that Tiron may protect, at least in part, against metavanadate-induced behavioral toxicity.

Lack of protective effects of dietary silicon on aluminium-induced maternal and developmental toxicity in mice.

In recent years, it has been demonstrated that oral aluminium (Al) exposure can produce growth retardation, delayed ossification and an increased incidence of foetal abnormalities in rats and mice. On the other hand, it has been also suggested that silicon may have a protective effect in limiting oral Al absorption. The aim of the present study was to assess whether dietary silicon could prevent against Al-induced maternal and developmental toxicity in mice. On gestation days 6-15, Al nitrate nonahydrate (398 mg/kg/day) was given by gavage to three groups of pregnant animals, which also received silicon in drinking water at concentrations of 0, 118 and 236 mg/l on days 7-18 of gestation. Three additional groups of pregnant mice received respectively: 270.6 mg/kg of sodium nitrate (gavage), and silicon in drinking water at 118 and 236 mg/l. Although silicon administration at 236 mg/l significantly reduced the percentage of Al-induced deaths, abortions and early deliveries, neither 118 nor 236 mg/l of silicon produced significant ameliorations on Al-induced foetotoxicity. Under the current experimental conditions dietary silicon was not effective in protecting against Al-induced developmental toxicity.

Enzymic evaluation of thiamin, riboflavin and pyridoxine status of parturient mothers and their newborn infants in a Mediterranean area of Spain.

OBJECTIVE: To determine the biochemical status of thiamin, riboflavin and pyridoxine in parturient mothers and their newborn infants in a Mediterranean region. DESIGN: Transveral study. SETTING: St Joan University Hospital and Faculty of Medicine & Health Sciences, Universitat Rovira i Virgili, Reus, Spain. SUBJECTS: 131 healthy parturient mothers, with normal pregnancies and deliveries in St Joan University Hospital, and their newborn infants. INTERVENTIONS: Erythrocyte haemolysates were prepared from maternal blood at delivery and infants' umbilical cord blood and used to measure micronutrient status using the transketolase, glutathione reductase and aspartate aminotransferase coenzyme stimulation tests. RESULTS: Maternal and infant coenzyme activities were significantly correlated, but infant coenzyme status was better than maternal, with significantly higher basal and stimulated activity (P < 0.001) and significantly lower activation coefficients (P < 0.001). Inadequate thiamin, riboflavin or pyridoxine status occured in 38.2 62.6% (50-82) of the mothers and 3.1-37.4% (4 49) of the infants; 85.2% (46/54), 12.9% (4/31) and 24.1% (12/54) of infants born to mothers with biochemical deficiency of either thiamin, riboflavin or pyridoxine, respectively also had inadequate status. Maternal deficiencies in more than one vitamin further increased the risk of infant thiamin and pyridoxine deficiency. Maternal and infant riboflavin status were significantly correlated with fetal development (e.g. length at birth, P < 0.001). The incidence of thiamin deficiency in paturient mothers in Spain was the highest out of a 12-country comparison. CONCLUSIONS: Inadequate status for each vitamin was evident in mothers and infants. Maternal status of each individual vitamin, but especially riboflavin, was affected by maternal status of the other vitamins. Infant thiamin status was the most adversely affected by maternal deficiencies in more than one vitamin. Infant riboflavin status, however, was apparently protected from adverse maternal status.

Prenatal effects of caffeine and restraint stress in mice.

The maternal and developmental toxicity of combined exposure to restraint stress and caffeine was assessed in mice. On Day 9 of gestation, six groups of pregnant mice were treated (p.o.) with a single dose of 30, 60, or 120 mg/kg of caffeine. Immediately after caffeine administration, three of these groups were subjected to restraint for 14 hr. Control groups included unrestrained and restrained pregnant mice not exposed to caffeine. An additional group of animals (unrestrained and not exposed to caffeine) was deprived of food for 14 hr. A two-way (caffeine dose x restraint) analysis of variance revealed an overall effect (reduction) of restraint and caffeine exposure on maternal body weight gain and food consumption on gestation Days 9-11. Significant reductions were also observed in body weight at termination and corrected body weight change of dams concurrently exposed to 120 mg/kg of caffeine and restraint. By contrast, no significant effects of caffeine, restraint, or caffeine plus restraint on embryo/fetal development were noted. The doses of caffeine administered here are much higher than those usually consumed by the general population. Under the current experimental conditions, caffeine alone or combined with restraint stress was not embryotoxic or teratogenic in mice.

Effects of simultaneous administration of desferrioxamine and tacrine in rats.

Tacrine (1,2,3,4-tetrahydro-9-aminoacridine), a reversible cholinesterase inhibitor, was effective in the treatment of Alzheimer's disease (AD). In turn, desferrioxamine (DFO), a chelating agent with ability to chelate iron and aluminum (Al), produced a 50% decrease in the rate of cognitive decline in patients with AD. Since combined therapy with tacrine and DFO might be more effective than individual administration of these drugs for the treatment of AD patients, this study evaluated the toxic effects of concomitant administration of tacrine and DFO to rats. Three groups of 8 rats each received the following treatments for 8 w: 80 mg DFO/kg/d i.m., 7.5 mg tacrine/kg/d po, or 80 mg DFO/kg/d i.m. +7.5 mg tacrine/kg/d po. A control group received distilled water by gavage daily and a 0.9% saline injection i.m. The administration of DFO + tacrine for 8 w did not increase most of the side effects caused by the individual DFO or tacrine administrations. These results open the possibility of considering the effectiveness of simultaneous administration of DFO and tacrine as a palliative treatment for AD patients.

Silicon reduces aluminum accumulation in rats: relevance to the aluminum hypothesis of Alzheimer disease.

In recent years, a possible relation between the aluminum and silicon levels in drinking water and the risk of Alzheimer disease (AD) has been established. It has been suggested that silicon may have a protective effect in limiting oral aluminum absorption. The present study was undertaken to examine the influence of supplementing silicon in the diet to prevent tissue aluminum retention in rats exposed to oral aluminum. Three groups of adult male rats were given by gavage 450 mg/kg/day of aluminum nitrate nonahydrate 5 days a week for 5 weeks. Concurrently, animals received silicon in the drinking water at 0 (positive control), 59, and 118 mg Si/L. A fourth group (-Al, - Si) was designated as a negative control group. At the end of the period of aluminum and silicon administration, urines were collected for 4 consecutive days, and the urinary aluminum levels were determined. The aluminum concentrations in the brain (various regions), liver, bone, spleen, and kidney were also measured. For all tissues, aluminum levels were significantly lower in the groups exposed to 59 and 118 mg Si/L than in the positive control group; significant reductions in the urinary aluminum levels of the same groups were also found. The current results corroborate that silicon effectively prevents gastrointestinal aluminum absorption, which may be of concern in protecting against the neurotoxic effects of aluminum.

Effects of vanadium on activity and learning in rats.

The effects of vanadate administration on activity and learning were assessed in rats. Four groups of adult male rats were given by gavage 0, 4.1, 8.2, and 16.4 mg/kg/day of sodium metavanadate for eight consecutive weeks. Three weeks after the cessation of the treatment, general motor activity of all animals was measured in an open-field. Rats were also tested for two-way shock avoidance learning in an automatic reflex conditioner. At the end of the testing period, rats were killed and vanadium concentration was determined in a number of tissues. Vanadium exposure caused an observable but not significant effect on body weight gain, while a persistent presence of vanadium was observed in all tissues measured. The results of the behavioral testing show that oral vanadate administration resulted in significant reductions in both general activity and learning.

Concentrations of some essential elements in the brain of aluminum-exposed rats in relation to the age of exposure.

This study evaluated the influence of age on the aluminum (Al) interaction with calcium (Ca), magnesium (Mg), iron (Fe), manganese (Mn), copper (Cu) and zinc (Zn) in the brain of rats. Since both Al and aging have been associated with neurobehavioral deficits in mammals, the brain was chosen to assess that influence. Male young (21 day), adult (8 months), and old (16 months) rats were given 0, 50, and 100 mg/kg per day of aluminum administered as aluminum nitrate in drinking water for 6.5 months. During that period, citric acid (0, 355 and 710 mg/kg per day) was also added to the drinking water. After 6.5 months of Al exposure, Al, Ca, Mg, Fe, Mn, Cu and Zn concentrations were determined in brain tissue as well as in a number of cerebral regions: cortex, hippocampus, striatum, cerebellum, thalamus, olfactory bulb, and rachidical bulb. While no significant age-related differences were found for Ca concentrations in the seven cerebral regions analyzed, most Mg, Fe, Mn and Zn levels were significantly higher in young than in adult and old rats. In turn, Al concentrations were mostly higher in the cerebral regions of young rats than in the same regions of adult and old rats. In contrast, Cu levels were lower in most brain regions of old animals than in those of young rats. According to the results of the present study, the age-related changes in brain Al, Ca, Mg, Fe, Mn, Cu and Zn concentrations induced by Al and aging would not suggest any influence on Al-induced neurobehavioral deficits.

The effect of age on aluminum retention in rats.

The present study was designed to assess potential changes in aluminum (Al) retention during advanced age. Young (21 day old), adult (8 months), and old (16 months) rats were exposed to 0, 50, and 100 mg Al/kg/day administered as aluminum nitrate in drinking water for a period of 6.5 months. Urinary Al levels were measured after 3 and 6.5 months of Al exposure. Organ weights and tissue Al concentrations were examined at 6.5 months of Al administration. Differences in the tissue accumulation of Al with age included higher liver, kidneys, spleen, bone and testes levels in old rats than in tissues of both young or adult animals. In contrast, brain concentrations were higher in young rats. Urinary Al levels of young, adult or old Al-exposed rats showed different trends at 6.5 months of Al exposure: compared with young values adult values declined, while those of old rats tended to increase further. The current results show that tissue Al retention patterns may be significantly altered depending on the age at Al exposure. This finding may be of concern for future investigations on the potential role of Al in certain neurological disorders.