Free radicals and antioxidant status in rat liver after dietary exposure of environmental mercury

Potential health effect of dietary exposure to environmental mercury was examined in this study. Dietary exposure significantly increased content of reduced glutathione (GSH) and activity of glutathione peroxidase (GSH-Px) in rat liver at 7 or 20 days (P < 0.05; P < 0.01), but parameters droped to normal levels after 90 days of exposure. The early increases of the two antioxidants were partly associated with the co-accumulated selenium. However, activity of superoxide dismutase (SOD) was observed significantly decreased after 30 and 90 days of exposure (P < 0.05, P < 0.05). Changes of antioxidants were paralleled by the induction and aggravation of free radicals in rat liver at 30 and 90 days (P < 0.01, P < 0.01), increased nitric oxide (NO) content at 90 days (P < 0.01). The excess availability of free radicals and the decreased levels of antioxidants resulted in a significant increase of malonyldialdehyde (MDA) after 90 days of exposure, indicating the aggravation of hepatic oxidative status. A number of biomarkers were required to monitor and minimize the health risk for the local population.     

Intracellular distribution of inorganic and organic mercury in rat liver after exposure to methylmercury salts

The intracellular distribution of inorganic and organic mercury in rat liver after exposure to methylmercury salts has been reported. The results have been discussed comparing intracellular distribution of mercury after exposure to various mercury compounds. After exposure to methylmercury salts the lysosomes/peroxisomes contain the highest concentration of mercury followed by microsomes and mitochondria. Differences in distribution between various mercury compounds seem to be related to the stability of the carbon-mercury bond, lysosomes accumulating inorganic mercury by preference either injected as such, or released in vivo from the intact organomercurial. Toxicological and pharmacokinetic implications of these conclusions have been discussed.     

The effects of vigabatrin on rat liver antioxidant status

The anti-epileptic drug vigabatrin was developed as an inhibitor of gamma-aminobutyric acid transaminase, and its ability to increase inhibition in the central nervous system led to its testing in an animal model. In animal models chronic use of vigabatrin is associated with irreversible myelin vacuolation. Antioxidant drugs change the antioxidant capacity of the body. Oxidative stress of the body increased when valproic acid and carbamazepine were used chronically. To assess whether vigabatrin may affect protein oxidation and lipid peroxidation, glutathione, glutathione peroxidase (GPx), and glutathione-S-transferase (GST) levels were studied in the livers of 57 rat fetuses after administration of vigabatrin to the mothers (19 in the first week of pregnancy, 20 in the second week, and 18 in the third week) and in 19 control rat fetuses without vigabatrin. We compared the results of administration of vigabatrin in each group with the controls. Rat fetus protein oxidation in group I (0.686 nmol/mg protein) and group II (0.723 nmol/mg protein) was higher than in the control group (0.388 nmol/mg protein). Lipid peroxidation (0.209, 0.224, 0.253 nmol/mg protein, respectively) and GPx levels (345.4, 329.0, 283.2 nmol/mg protein, respectively) of groups I, II, and III were higher than in the control group (0.104, 167.2 nmol/mg protein, respectively). GST in group II (79.2 nmol/mg protein) and group III (77.8 nmol/mg protein) were not different from that in the control group (78 nmol/mg protein). It was found that vigabatrin affected all the parameters that were studied, especially in group I, which was given the drug in the first week of pregnancy.     

Mercury distribution in the rat brain after mercury vapor exposure.

Brown Norwegian rats were exposed to mercury vapor at a concentration of approximately 1 mg/m3 for 5 weeks 24 hr/day 7 days a week and 6 hr/day 3 days a week, respectively. The total mercury absorption was calculated to 264 and 35 micrograms per week and 100 g body weight. The mean blood mercury concentration was 0.25 +/- 0.03 and 0.09 +/- 0.01 microgram/g, and the total concentration in the brain was 5.03 +/- 0.73 and 0.71 +/- 0.10 microgram/g tissue, respectively. The mercury distribution in the brains was examined using a method based on chemographic principles. Mercury was found primarily in the neocortex, in the basal nuclei, and in the cerebellar Purkinje cells. This distribution pattern corresponded to the pattern of inorganic mercury described after exposure to methyl mercury. Distribution of mercury after administration of different mercury compounds is discussed.     

Impaired aquaporins expression in the gastrointestinal tract of rat after mercury exposure

The main route of exposure to mercury in humans is through the diet. Consequently, the gastrointestinal mucosa is exposed to the mercurial forms, where they cause intestinal fluid accumulation, mucosal injuries and diarrhea. The relationship between inorganic mercury (HgCl₂) and methylmercury (CH₃HgCl) exposure and water movement in the gastrointestinal tract is still unexplored. The leading role of aquaporins (AQPs) in the rapid bidirectional movement of fluid in the gastrointestinal tract of mammals is well established. The present study evaluates the effect of HgCl₂ and CH₃HgCl exposure on AQP expression in different portions of the gastrointestinal tract of rats treated by gavage (5 mg kg^–1 of mercury species, single dose, 4 days). The results show that mercury species reduce mRNA and protein levels of AQPs in different parts of the gastrointestinal tract. In the stomach, treated rats show a significant reduction of expression of AQP3 (80–90% for mRNA and 50% for protein) and AQP4 (95–99% for mRNA and 20–40% for protein). In the small and large intestine, treated rats experience a significant reduction of AQP3 and AQP7 expression. Protein contents of both AQPs are reduced in similar proportions in jejunum (AQP3: 40–50%; AQP7: 45–50%) and colon (AQP3: 35–40%; AQP7: 45–60%), regardless of the treatment. Our results indicate that some AQPs are downregulated in the rat gastrointestinal tract by mercury exposure, suggesting a possible role of AQPs in the development of mercury gastrointestinal symptoms. Copyright © 2015 John Wiley & Sons, Ltd.     

Dimethoate-induced effects on antioxidant status of liver and brain of rats following subchronic exposure

Dimethoate, an organophosphate pesticide, is used in controlling the pests of a variety of crops. The study was carried out to understand the role of dimethoate in inducing oxidative stress leading to generation of free radicals and alterations in antioxidant enzymes and scavengers of oxygen free radicals. The effects of subchronic exposure of dimethoate in the production of oxidative stress were evaluated in male Wistar rats in the present study. Dimethoate was administered orally at doses 0.6, 6, and 30 mg/kg for 30 days in these rats. The results indicated an increase in levels of hepatic Cytochrome P450, lipid peroxidation, catalase, superoxide dismutase, glutathione peroxidase and glutathione reductase in liver and brain at doses 6 and 30 mg/kg. There were no significant changes in the level of glucose-6-phosphate dehydrogenase activity except in liver at 30 mg/kg. A decrease in glutathione was observed at 30 and 6 mg/kg in both liver and brain. Glutathione-S-transferase increased at 30 and 6 mg/kg in liver and 30 mg/kg in brain. Erythrocyte acetylcholinesterase was inhibited at 30 and 6 mg/kg doses. Dose-dependent histopathological changes were seen in both liver and brain. This study concludes that oxidative stress due to dimethoate may be ascribed to induction of Cytochrome P450, inhibition of AChE and disturbance in activities of GSH and GST enzymes causing lipid peroxidation and histological and electron microscopic changes in liver and brain.     

Lipidomic changes in rat liver after long-term exposure to ethanol

Alcoholic liver disease (ALD) is a serious health problem with significant morbidity and mortality. In this study we examined the progression of ALD along with lipidomic changes in rats fed ethanol for 2 and 3 months to understand the mechanism, and identify possible biomarkers. Male Fischer 344 rats were fed 5% ethanol or caloric equivalent of maltose-dextrin in a Lieber-DeCarli diet. Animals were killed at the end of 2 and 3 months and plasma and livers were collected. Portions of the liver were fixed for histological and immunohistological studies. Plasma and the liver lipids were extracted and analyzed by nuclear magnetic resonance (NMR) spectroscopy. A time dependent fatty infiltration was observed in the livers of ethanol-fed rats. Mild inflammation and oxidative stress were observed in some ethanol-fed rats at 3 months. The multivariate and principal component analysis of proton and phosphorus NMR spectroscopy data of extracted lipids from the plasma and livers showed segregation of ethanol-fed groups from the pair-fed controls. Significant hepatic lipids that were increased by ethanol exposure included fatty acids and triglycerides, whereas phosphatidylcholine (PC) decreased. However, both free fatty acids and PC decreased in the plasma. In liver lipids unsaturation of fatty acyl chains increased, contrary to plasma, where it decreased. Our studies confirm that over-accumulation of lipids in ethanol-induced liver steatosis accompanied by mild inflammation on long duration of ethanol exposure. Identified metabolic profile using NMR lipidomics could be further explored to establish biomarker signatures representing the etiopathogenesis, progression and/or severity of ALD.     

Subcellular distribution of mercury in the rat kidney cortex after exposure to mercuric chloride

The pharmacologic effects and plasma concentrations produced by the ip administration of high doses of thymidine were determined in CDF₁ mice. Within 15 min after the injection of thymidine at a dose of 4.075 g/kg (between LD0 and LD50), mean arterial blood pressure and heart rate fell precipitously and remained depressed for over 6 hr. During the experimental period, sedation and anuresis were also consistently observed in thymidine-treated mice. After a dose of 4.7 g/kg, millimolar plasma thymidine concentrations were maintained for 16 to 20 hr; by contrast, at a lower but nonlethal dose (3.8 g/kg), millimolar plasma thymidine concentrations were maintained for only 9 hr. It is suggested that sustained elevation of circulating thymidine in the range 1–10 mm for periods greater than $\text{1}\text{2}$ day may be associated with toxicity in mice.     

Effect of lupeol, a pentacyclic triterpene, on the lipid peroxidation and antioxidant status in rat kidney after chronic cadmium exposure

This study pertains to the role of lupeol, a pentacyclic triterpene, against the toxic manifestations of chronic cadmium exposure. Cadmium is a potent nephrotoxin on prolonged exposure. Cadmium (as cadmium chloride) at a dose of 1 mg kg(-1) body weight was administered subcutaneously for 15 days to rats. This led to an increase in the level of lipid peroxides and a decrease in the level of antioxidants in the kidney. Lupeol, when supplemented at a dosage of 40 mg kg(-1) body weight concurrent with cadmium administration, showed an improvement in the antioxidant status. The level of lipid peroxides also showed a significant decrease, which shows the nephroprotective action of the drug.     

Distribution and retention of mercury in metallothionen-null mice after exposure to mercury vapor

We studied the role of metallothionein (MT) in the distribution and retention of mercury in the brain, lung, liver and kidney of MT-null and wild-type mice after exposure to mercury (Hg0) vapor. Mice were exposed to Hg0 vapor at 5.5-6.7 mg/m3 for 3 h and killed at 1, 24, 72 or 168 h after exposure. One hour after exposure to Hg0 vapor, there were no differences in mercury concentrations in these organs from MT-null and wild-type mice. However, the elimination rate of mercury from the organs, except the brain, were remarkably faster in MT-null mice than in wild-type mice. MT-I and -II levels in the lung and kidney were increased significantly in wild-type mice but not in MT-null mice at 24 h after exposure to Hg0 vapor. At this time point, over 65% of the mercury was retained in the MT fraction of the cytosol of organs from wild-type mice. In contrast, mercury appeared mainly in the high-molecular-weight protein fractions in the cytosol of organs from MT-null mice. In the brain, a large amount of mercury was bound to MT in both strains of mice immediately after exposure. No difference was observed in the elimination rate of mercury from the brain between both strains of mice. Brain MT levels were elevated slightly in wild-type mice at 168 h after exposure but could not be detected in MT-null mice. These data suggest that no detectable MT-I and -II levels were found in the brain of MT-null mice and that mercury was apparently bound to MT-III. Using MT-null mice, we showed also that MT-III may play an important role in the retention of mercury in the brain.     

Modes of cell death in rat liver after monocrotaline exposure.

Monocrotaline (MCT) is a pyrrolizidine alkaloid (PA) plant toxin that produces sinusoidal endothelial cell (SEC) injury, hemorrhage, fibrin deposition, and coagulative hepatic parenchymal cell (HPC) oncosis in centrilobular regions of rat livers. Cells with apoptotic morphology have been observed in the livers of animals exposed to other PAs. Whether apoptosis occurs in the livers of MCT-treated animals and whether it is required for full manifestation of pathological changes is not known. To determine this, rats were treated with 300 mg MCT/kg, and apoptosis was detected by transmission electron microscopy and the TUNEL (TdT-mediated dUTP nick end labeling) assay. MCT produced significant apoptosis in the liver by 4 h after treatment. To determine if MCT kills cultured HPCs by apoptosis, HPCs were isolated from the livers of rats and exposed to MCT. MCT caused a concentration-dependent release of alanine aminotransferase (ALT), a marker of HPC injury. Furthermore, caspase 3 was activated and TUNEL staining increased in MCT-treated HPCs. MCT-induced TUNEL staining and release of ALT into the medium were completely prevented by the pancaspase inhibitors z-VAD.fmk and IDN-7314, suggesting that MCT kills cultured HPCs by apoptosis. To determine if caspase inhibition prevents MCT-induced apoptosis in the liver, rats were cotreated with MCT and IDN-7314. IDN-7314 reduced MCT-induced TUNEL staining in the liver and release of ALT into the plasma. Morphometric analysis confirmed that IDN-7314 reduced HPC oncosis in the liver by approximately 50%. Inasmuch as HPC hypoxia occurred in the livers of MCT-treated animals, upregulation of the hypoxia-regulated cell-death factor, BNIP3 (Bcl2/adenovirus EIB 19kD-interacting protein 3), was examined. BNIP3 was increased in the livers of mice treated 24 h earlier with MCT. Results from these studies show that MCT kills cultured HPCs by apoptosis but causes both oncosis and apoptosis in the liver in vivo. Furthermore, caspase inhibition reduces both apoptosis and HPC oncosis in the liver after MCT exposure.     

Garlic-induced alteration in rat liver and kidney morphology and associated changes in endogenous antioxidant status.

The effects of chronic garlic intake on various endogenous antioxidant enzymes and lipid peroxidation on two major organs, the liver (L) and kidneys (K), were investigated. Wistar albino rats were fed with fresh garlic homogenate daily by gavage in three different doses (250, 500 and 1000 mg/kg/day) for 30 days. After this period, rats were sacrificed and liver and kidneys were harvested for biochemical estimation. In comparison to saline-treated rats, the 250 mg/kg/day dose significantly (P<0.02) reduced thiobarbituric acid reactive substances (TBARS) (L: 187.48+/-9.23 vs 150.66+/-11.45; K: 177.38 15.88 vs 120.66+/-9.39 nmol/g wet. weight) and glutathione peroxidase (GPx) (L: 0.2438+/-0.05 vs 0.0046+/-0.0005; K: 0.1459+/-0.034 vs 0.0055+/-0.0003 U/mg protein). There was no change in catalase and reduced glutathione (GSH) but superoxide dismutase (SOD) increased significantly (P<0.01) (L: 5.49+/-0.76 vs 18.38+/-2.26; K: 11.47+/-1.48 vs 21.22+/-3.19 U/mg protein). Both 500 and 1000 mg/kg/day doses significantly (P<0.05) reduced endogenous antioxidants (catalase and SOD) without altering TBARS. A 1000 mg/kg/day dose of garlic caused marked histopathological and ultrastructural changes in both liver and kidneys. The results suggest that garlic in low doses has the potential to enhance the endogenous antioxidant status, although at higher doses a reversal of these effects is observed. The present study also highlights the potential ability of a high dose of garlic to induce morphological changes in the liver and kidneys, indicating the need to identify a safe dose range for garlic.     

Effect of 4-methylpyrazole on antioxidant enzyme status and lipid peroxidation in the liver of rats after exposure to ethylene glycol and ethyl alcohol

BackgroundThe aim of the conducted studies was to evaluate the effect of 4-methylpyrazole, increasingly used in detoxifying treatments after ethylene glycol poisoning, on the activity of some antioxidant enzymes and lipid peroxidation formation in the liver of rats after experimental co-exposure to ethylene glycol and ethyl alcohol.MethodsThe trials were conducted on adult male Wistar rats. Ethylene glycol (EG) at the dose of 3.83 g/kg bw and ethyl alcohol (EA) at the dose of 1 g/kg bw were administered po, and 4-methylpyrazole (4-MP) at the dose of 0.01 g/kg bw was administered ip. Parameters of antioxidant balance were evaluated in hepatic cytosol, including the activity of the following enzymes: glutathione S-transferase (GST), glutathione reductase (GR), glutathione peroxidase (GPx) and lipid peroxidation level (TBARS).ResultsThe results suggest that evaluation of the effects of administrated 4-MP after co-exposure to EG and EA in the liver revealed statistically significant changes on antioxidant enzyme system and malondialdehyde formation.ConclusionThe changes in biomarkers activity indicate a greater production of free radicals which exceeds the capability of antioxidant system, appearing with oxidative stress in the group of animals treated by 4-MP combined with EG and EA.     

The effect of various dietary fibres on tissue concentration and chemical form of mercury after methylmercury exposure in mice

The whole-body retention of mercury after exposure of BALB/c mice to methylmercury was measured in animals fed fibre-free, 5% pectin, 5% cellulose or 5, 15 or 30% wheat bran diets. The rate of elimination of mercury was dependent on the diet fed, with dietary bran increasing the rate of elimination. The incorporation of 15 or 30% bran in the diet of the mice decreased the total mercury concentration in the brain, blood and small intestine, although the effects were significant only in those animals on 30% bran diet. The fibres had little effect on mercury levels in other tissues. The proportion of mercury found in the mercuric form was significantly greater in liver, kidneys and gut of mice fed bran. The results suggest that dietary bran may reduce the levels of mercury in the brain after methylmercury exposure and may therefore reduce the neurotoxic effects of the organomercurial. We suggest that wheat bran exerts its effects on mercury retention and brain level via a modification of the metabolic activity of the gut microflora.     

低剂量无机汞宫内及哺乳期暴露对仔大鼠肝、肾和脑组织中谷胱甘肽抗氧化物酶系统的影响

目的　研究低剂量无机汞子宫内及哺乳期暴露对仔大鼠肝脏、肾脏及脑组织中谷胱甘肽抗氧化物酶系统的影响 ,探讨低剂量无机汞对子代发育潜在的毒性。方法　对怀孕d 0的母大鼠进行 0 .2mg·L- 1Hg2 +饮水染毒直至仔鼠出生后d 2 0断乳期。研究仔鼠肝脏、肾脏及脑区 (大脑、小脑、脑干、海马及其余脑组织 )中汞的分布及汞暴露对上述组织中总巯基含量、还原型谷胱甘肽 (GSH)水平及谷胱甘肽过氧化物酶 (GSH Px)活性的影响。结果汞暴露后仔鼠肾脏中汞的含量最高 ,其次是肝脏和脑组织。在所研究的仔鼠各脑区中 ,海马汞含量最高 ,其次是小脑和大脑。汞暴露组仔鼠肝脏和肾脏中巯基含量比对照组升高了 2倍 ,而脑区中汞暴露组巯基含量与对照组相比则降低至 1/ 2～ 1/ 3。汞暴露组仔鼠肝脏中GSH水平与对照组相比升高了6 5 % ,而在肾脏和脑组织中GSH的水平与对照组相比则显著降低 ,其中 ,汞暴露组仔鼠各脑区中GSH水平降低幅度与对照组相比均超过了 5 8%。除肝脏外 ,汞暴露组仔鼠肾脏和脑区中GSH Px活性明显下降。在脑组织中 ,除海马外 ,GSH Px活性在汞暴露组其他 4个脑区中的下降幅度均超过 80 %。结论　低剂量无机汞子宫内及哺乳期暴露对仔鼠肾脏 ,特别是对脑组织的正常发育造成了一定程度的危害 ;但上述指标对肝脏的     

Assessment of drug exposure in rat dietary studies

1. The objective of this study was to justify the evaluation of exposure of animals to chemical substances on the basis of only three blood samples taken during a 24-h period, but still with acceptable accuracy.

2. Fischer rats were fed a diet mixed with either paracetamol, 100mg.kg^-1 (short half-life compound), antipyrine, 100mg.kg^-1 (medium half-life compound), or phenylbutazone, 50mg kg^-1 (long half-life compound) for 3 weeks. It had been shown in a preliminary study that these compounds when administered at these dose levels did not influence feeding behaviour. At the end of 3 weeks, five rats were sampled every 3?h beginning and ending at 19.00?h (45 rats in total) and plasma concentrations were measured using?h.p.l.c.

3. The area under the curve over 24h (AUC₂₄), calculated using all nine concentrations was considered to be the true AUC₂₄. Subsequently, estimates of this parameter were made using different combinations of concentrations at three or even two selected time points.

4. For each compound, the highest concentration occurred at 07.00?h. It was shown that using the concentrations at 07.00, 10.00 and 16.00?h the estimate of the AUC₂₄ was within 15% of the true value.

5. In comparison with a gavage study in the same rat (strain and age), bioavailability was lower in the diet study with relative bioavailabilities of 27, 22 and 61% for paracetamol, antipyrine and phenylbutazone, respectively.

6. In conclusion, drug exposure as expressed by AUC₂₄ and C_max can be accurately determined in rat studies using compound administration in the diet by measuring concentrations at three selected time points for compounds with elimination half-lives ranging from about 1 to 5?h.     

Induction of glutathione-S-transferase and heat-shock proteins in rat liver after ethylene oxide exposure

Defense mechanisms in rat liver against depletion of glutathione (GSH) and cellular injuries induced by ethylene oxide (EO) were studied. Rats were exposed to EO under either high dose (1300 ppm for 4 hr, once) or low dose (500 ppm for 6 hr, three times a week for 6 weeks) conditions. The hepatic content of GSH decreased dramatically after EO treatment, probably due to detoxication of EO. After the high dose treatment the hepatic GSH content fell by 90% of the control values but recovered within 10 to 15 hr. EO reacts directly with a variety of cellular macromolecules but all rats survived the exposure. Since the metabolites of EO are ethylene glycol and GSH-conjugates, the enzymatic activities of epoxide hydrolase and glutathione-S-transferase (GST) were determined. Only GST activity was found to occur after low dose chronic exposure. The defense mechanism at mRNA level was investigated using probes for GST and several heat-shock proteins (hsps). Enhanced accumulation of GST mRNA was detectable during the recovery period of rats after both high and low dose exposure to EO. Interestingly, both hsp32 (less than 40-fold) and hsp90 (less than 3-fold) mRNA increased after high dose exposure but the mRNA level of one of the major heat-shock proteins, hsp70, did not change under these conditions. Diethylmaleate, which is known to be a GSH depleter in liver, induced hsp32 mRNA only in rat liver, while hsp70 and hsp90 mRNA levels did not change when GSH was depleted. These results suggest that individual heat-shock proteins are induced in different ways under unphysiological conditions such as EO exposure.     

Changes in lymphocyte oxidant/antioxidant parameters after carbonyl and antioxidant exposure

During normal B- and T-cell life, processes including activation, proliferation, signaling pathways and apoptosis are markedly dependent on ROS generation. However, these cells can also suffer the effect of oxidant overproduction. Thus, the purpose of the present study was to examine the possible pro-oxidant effects of MGO/high glucose and antioxidant effects of astaxanthin associated with vitamin C on some oxidative and antioxidant parameters of human lymphocytes in vitro. Lymphocytes from healthy subjects were treated with 20 mM of glucose and 30 μM MGO followed or not by the addition of the antioxidants astaxanthin (2 μM) and vitamin C (100 μM) for up to 24 h. We examined superoxide dismutase (SOD), catalase, glutathione peroxidase, glutathione reductase, and glucose-6-phosphate dehydrogenase (G6PDH) activities, GSH/GSSG ratio and total thiol and carbonyl content. Oxidative parameters included superoxide anion, hydrogen peroxide and nitric oxide production. The association of astaxanthin and vitamin C proved to be a powerful antioxidant in human lymphocytes as showed by the marked reduction in superoxide anion, and hydrogen peroxide production as well as increased GSH content, GSH/GSSG ratio, GPx and GR activities. The antioxidant association showed to be more potent than their individual application. High glucose and methylglyoxal did not promote oxidative stress in human lymphocytes, since neither the oxidative parameters nor the antioxidant defense system was altered. According to these results, new therapies with the association of astaxanthin and vitamin C may be helpful to improve the immune function of patients with exacerbated production of ROS.