Behaviour of cyclic nucleotides and Ca2+ levels in liver tissue of rats poisoned by white phosphorus and trichlorobromomethane

The content of hepatic cyclic AMP was increased soon after intoxication by white phosphorus. Its level reached a maximum 4 h after poisoning, but in subsequent phases tended to return to normal. In contrast, the cyclic GMP concentration was altered only 24 and 36 h after treatment with the same hepatotoxin. Similar modifications of cAMP and cGMP content were also detected after poisoning by trichlorobromomethane (CBrCl3). As a consequence, an altered cGMP/cAMP ratio was found in both experimental conditions. Further, the modification of cAMP content after white phosphorus was detected prior to liver damage (steatosis and necrosis), while the highest concentration of the cyclic nucleotide in CBrCl3-poisoned rats was found when fatty liver was already evident. In addition, in phosphorus-poisoned rats, the hepatic content of Ca2+ was found to be unmodified during all phases of the intoxication, while after CBrCl3 a phasic increase of the Ca2+ level was observed at 4, 24 and 36 h.     

Modifications of liver content of cyclic nucleotides in the rat poisoned with white phosphorus]

In rat liver following white phorphorus poisoning a biphasic increase in cyclic AMP concentration was observed. After a lag period of 1 hour the cyclic AMP content rose to a first peak at 4 hours and to a second peak at 12 hours of intoxication. The cyclic AMP level fell to normal after 24 hours, by which time the cyclic nucleotide concentration was approaching control values. On the contrary, cyclic GMP content was found to the normal level during the different stages of intoxication. Only at 36 hours the cyclic GMP amount appeared significantly increased above the control values. Serum activity of alanine- and aspartate-amino transferases was found changed from 8 hours to 24 hours after poisoning. The serum level of the two enzymes was overlapping the control values after 36 hours. These results are discussed in relation to hepatocyte necrosis following white phosphorus intoxication.     

Age-dependent changes in rat liver lipid peroxidation and glutathione content induced by acute ethanol ingestion

The study of the influence of the age of animals (13 to 53 weeks) on total liver thiobarbituric acid reactive substances (TBAR) content showed an increase which is maximal in rats of 39 weeks of age compared to young animals (13 weeks), followed by a dimunition in the 53 weeks old group. In this situation, the content of hepatic GSH and total GSH equivalents as well as the GSH/GSSG ratio were decreased with ageing, while GSSG levels were enhanced in the oldest group studied. Acute ethanol intoxication resulted in a marked increase in liver TBAR content in young animals, together with a decline in GSH, total GSH equivalents and GSH/GSSG ratio, and an enhancement in GSSG. These changes elicited by ethanol intake were reduced with ageing. It is concluded that ethanol-induced oxidative stress in the liver is diminished during ageing, despite the progressive decrease in the glutathione content of the tissue observed in control animals.     

Changes in Na+, K+-adenosinetriphosphatase and 5'-nucleotidase activities in cell membrane isolated from rat liver after acute white phosphorus poisoning

Na+, K+-ATPase and 5'-Nucleotidase activities in rat liver plasmamembranes after "in vivo" intoxication with a single dose of white phosphorus (10 mg/kg b.w. "per os") are investigated. Na+, K+-ATPase activity is significantly increased 1 hour and inhibited 12 hours after intoxication. 5'-Nucleotidase is strongly increased at 1, 2 and 4 hours after poisoning and is significantly decreased at 12 hours. The enhancement of both the enzymatic activities is evident prior to triglyceride accumulation in rat liver. Our results suggest that lipid fluidity of cell membrane is early and mildly affected during white phosphorus poisoning.     

Lindane-induced liver oxidative stress.

The development of an oxidative stress condition in the liver by lindane intoxication is discussed as a possible hepatotoxic mechanism of the insecticide. Lindane is metabolized by liver microsomal enzymes to a variety of metabolites, which are susceptible of conjugation for proper elimination. In addition, the interaction of lindane with the liver tissue results in the induction of the microsomal cytochrome P-450 system, together with enhanced rates of superoxide radical generation and a significant increase in indicators of lipid peroxidation. Concomitantly, lindane intoxication induces a derangement of some antioxidant mechanisms of the liver cell, including decreased superoxide dismutase and catalase activities and alterations in reduced glutathione content leading to depressed GSH/GSSG ratios. The time course study of the changes in hepatic lipid peroxidation and antioxidant parameters are closely interrelated and coincide with the onset and progression of morphological lesions.     

Administration of glutathione and lipid peroxidation induced during fasting

It is well known that lipid peroxidation may be initiated or exaggerated by conditions leading to hepatic GSH depletion or altered GSH/GSSG ratio. In our study we evaluated the effects of GSH administration on hepatic, bile and plasma GSH, GSSG and MDA in rats depleted of the tripeptide by a prolonged. fasting. An exteriorized biliary-duodenal fistula was established and GSH or saline solution was administered i.p. for a period of 6h. Rats treated with GSH exhibited an increased GSH and decreased GSSG biliary excretion. Whereas in control rats an opposite pattern was observed, namely enhanced GSSG and decreased GSH biliary excretion. While hepatic GSH and GSSG concentrations were comparable in the two groups, a significant increase in liver and plasma MDA production was found in controls compared to GSH treated rats. Our data suggest a protective role of GSH against the production of lipoperoxidation as evidenced by the decrease of hepatic, biliary and plasma MDA levels and by a decreased percentage of biliary GSSG. In addition, the significant increase of biliary GSH excretion, observed in rats treated with GSH compared to controls, may be due to an increased supply of the tripeptide which is known to be preferentially excreted into bile in the reduced form.     

Increased loss and decreased synthesis of hepatic glutathione after acute ethanol administration. Turnover studies.

The effect of acute ethanol administration on rates of synthesis and utilization of hepatic glutathione (GSH) was studied in rats after a pulse of [35S]cysteine. A 35% decrease in hepatic GSH content 5h after administration of 4 g of ethanol/kg body wt. was accompanied by a 33% increase in the rate of GSH utilization. The decrease occurred without increases in hepatic oxidized glutathione (GSSG) or in the GSH/GSSG ratio. The rate of non-enzymic condensation of GSH with acetaldehyde could account for only 6% of the rate of hepatic GSH disappearance. The increased loss of [35S]GSH induced by ethanol was not accompanied by an increased turnover; rather, a 30% inhibition of GSH synthesis balanced the increased rate of loss, leaving the turnover rate unchanged. The rate of acetaldehyde condensation with cysteine in vitro occurred at about one-third of the rate of GSH loss in ethanol-treated animals. However, ethanol induced only a minor decrease in liver cysteine content, which did not precede, but followed, the decrease in GSH. The characteristics of 2-methylthiazolidine-4-carboxylic acid, the condensation product between acetaldehyde and cysteine, were studied and methodologies were developed to determine its presence in tissues. It was not found in the liver of ethanol-treated animals. Ethanol administration led to a marked increase (47%) in plasma GSH in the post-hepatic inferior vena cava, but not in its pre-hepatic segment. Data suggest that an increased loss of GSH from the liver constitutes an important mechanism for the decrease in GSH induced by ethanol. In addition, an inhibition of GSH synthesis is observed.     

Effects of fluoride on hepatic antioxidant system and transcription of Cu/Zn SOD gene in young pigs

Thirty-two barrows (Duroc x Landrace x Yorkshire) were randomly divided into four groups, each of which included eight pigs. The groups received the same basal diet supplemented with 0, 100, 250 and 400mg/kg fluoride, respectively. The malondialdehyde (MDA) and glutathione (GSH) levels, antioxidant enzymes activities and zinc/copper superoxide dismutase (Cu/Zn SOD) mRNA content in the liver were determined to evaluate the fluoride hepatic intoxication. Results showed the increased lipid peroxides (LPO) level and the reduced GSH content, along with a concomitant decrease in the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px). Moreover, the level of hepatic Cu/Zn SOD mRNA was also significantly reduced. We suggest the mechanism of fluoride injuring the liver as follows: fluoride causes a decrease in Cu/Zn SOD mRNA and the reduced activities of antioxidant enzymes, leads to the declined ability of scavenging free radicals with excessive production of LPO, which seriously damages the hepatic structure and function.     

Decreased Antioxidant Defense Mechanisms in Rat Liver after Methanol Intoxication

The primary metabolic fate of methanol is oxidation to formaldehyde and then to formate by enzymes of the liver. Cytochrome P-450 and a role for the hydroxyl radical have been implicated in this process. The aim of the paper was to study the liver antioxidant defense system in methanol intoxication, in doses of 1.5, 3.0 and 6.0 g/kg b.w., after methanol administration to rats. In liver homogenates, the activities of Cu, Zn-superoxide dismutase and catalase were significantly increased after 6 h following methanol ingestion in doses of 3.0 and 6.0 g/kg b.w. and persisted up to 2-5 days, accompanied by significant decrease of glutathione reductase and glutathione peroxidase activities. The content of GSH was significantly decreased during 6 hours to 5 days. The liver ascorbate level was significantly diminished, too, while MDA levels were considerably increased after 1.5, 3.0 and 6.0 g/kg b.w. methanol intoxication. Changes due to methanol ingestion may indicate impaired antioxidant defense mechanisms in the liver tissue.     

Experimental hyperprolinemia induces mild oxidative stress, metabolic changes, and tissue adaptation in rat liver

The present study investigated the effects of chronic hyperprolinemia on oxidative and metabolic status in liver and serum of rats. Wistar rats received daily subcutaneous injections of proline from their 6th to 28th day of life. Twelve hours after the last injection the rats were sacrificed and liver and serum were collected. Results showed that hyperprolinemia induced a significant reduction in total antioxidant potential and thiobarbituric acid-reactive substances. The activities of the antioxidant enzymes catalase and superoxide dismutase were significantly increased after chronic proline administration, while glutathione (GSH) peroxidase activity, dichlorofluorescin oxidation, GSH, sulfhydryl, and carbonyl content remained unaltered. Histological analyses of the liver revealed that proline treatment induced changes of the hepatic microarchitecture and increased the number of inflammatory cells and the glycogen content. Biochemical determination also demonstrated an increase in glycogen concentration, as well as a higher synthesis of glycogen in liver of hyperprolinemic rats. Regarding to hepatic metabolism, it was observed an increase on glucose oxidation and a decrease on lipid synthesis from glucose. However, hepatic lipid content and serum glucose levels were not changed. Proline administration did not alter the aminotransferases activities and serum markers of hepatic injury. Our findings suggest that hyperprolinemia alters the liver homeostasis possibly by induction of a mild degree of oxidative stress and metabolic changes. The hepatic alterations caused by proline probably do not implicate in substantial hepatic tissue damage, but rather demonstrate a process of adaptation of this tissue to oxidative stress. However, the biological significance of these findings requires additional investigation.     

Protective effect of naringenin on hepatic and renal dysfunction and oxidative stress in arsenic intoxicated rats

Arsenic has a long history as a potent human poison, chronic exposure over a period of time may result in the manifestation of toxicity in practically all systems of the body. In the present investigation the efficacy of naringenin (NRG), a naturally occurring citrus flavanone against arsenic-induced hepatotoxic and nephrotoxic manifestations have been studied in rats. Arsenic trioxide was administered orally at the dose of 2 mg/kg/day with or without combination of NRG (20 or 50 mg/kg/day) for 28 days. At the end of the experimental period the hepatic and renal dysfunction was evaluated by histological examination, serum biomarkers and markers of oxidative stress; lipid peroxidation (LPO), reduced glutathione (GSH) and antioxidant enzymes. Arsenic intoxication increased serum bilirubin, urea, uric acid and creatinine levels, additionally enhanced the activities of hepatic marker enzymes aspartate transaminase, alanine transaminase and alkaline phosphatase. Also, the hepatic and renal tissues showed a marked elevation in LPO levels with a decrease in GSH content and the activities of antioxidant enzymes such as superoxide dismutase, catalase, glutathione peroxidase and glutathione-S-transferase on arsenic treatment. Simultaneous treatment with NRG restored the activities of serum biomarkers and antioxidant enzymes in the tissues in a dose-dependent manner. Furthermore, the histopathological studies confirmed the protective effect of NRG co-treatment by reducing the pathological changes due to arsenic intoxication in both liver and kidney. Thus, our present study demonstrates that NRG has a potential to protect arsenic-induced oxidative hepatic and renal dysfunction.     

Acetaminophen-induced liver oxidative stress and hepatotoxicity: influence of Kupffer cell activity assessed in the isolated perfused rat liver

Summary

The influence of acetaminophen (APAP) treatment (400 mg/kg) on Kupffer cell function was studied in the isolated perfused liver by colloidal carbon infusion, concomitantly with parameters related to oxidative stress (thiobarbituric acid reactants (TBARS) formation and glutathione (GSH) content) and tissue injury (sinusoidal efflux of lactate dehydrogenase (LDH)). APAP led to increased rates of hepatic TBARS formation, GSH depletion, and higher sinusoidal LDH efflux compared to control values, without changes in the basal rate of O₂ consumption. In addition, APAP significantly enhanced the rate of carbon uptake by perfused livers and the associated carbon-induced O₂ consumption, with carbon-induced LDH effluxes being increased by 411% over control values or by 124% compared to basal LDH release in APAP-treated rats. APAP-induced changes in liver TBARS formation and GSH levels were attenuated by gadolinium chloride (GdCl₃) pretreatment, whereas those in carbon uptake, carbon-induced respiration, and LDH efflux were abolished. GdCl₃ pretreatment decreased liver O₂ consumption irrespectively of APAP treatment, an effect that seems to be due to depression of mitochondrial respiration. It is concluded that APAP intoxication enhances Kupffer cell function as assessed in the intact liver, which may represent an important source of reactive O₂ species and chemical mediators conditioning the increased oxidative stress status and the tissue injury which developed.     

Glutathione depletion and lipid peroxidation in the mouse brain after bromobenzene poisoning

Intoxication of NMRI Albino mice with bromobenzene is often followed by the appearance of neurological symptoms. The possibility was investigated that the intoxication results in glutathione (GSH) depletion in central nervous systems as seen in other tissues, and that such a depletion is followed by the development of lipid peroxidation. 18-20 hours after bromobenzene administration (15 mmoles/Kg, p.o.) GSH content of prosencephalic and metencephalic regions was depleted by 39 and 55%, respectively. Lipid peroxidation (measured by the tissue content of malonildialdehyde) was observed only when GSH content reached a threshold value, which was different for prosencephalon as compared to metencephalon (2-1.5 mumoles GSH/g and 1.2-0.7 mumoles GSH/g, respectively). Possible mechanisms underlying the phenomenon are discussed.     

Liver and biliary levels of glutathione and thiobarbituric acid reactants after acute lindane intoxication

The i.p. administration of 60 mg kg-1 body weight of lindane, the gamma-isomer of hexachlorocyclohexane, to fed rats led to an enhancement of hepatic lipid peroxidation after 24 h of treatment. This was evidenced by significant increases in the hepatic production and biliary release of thiobarbituric acid reactive substances, and in the biliary release of glutathione disulphide. Under these conditions, the content of cytochrome P450 was enhanced concomitantly with increases in the total microsomal oxygen uptake, superoxide radical generation and (+)-catechin (cyanid-3-ol) sensitive respiration. The glutathione status of hepatocytes was altered by lindane as the content and biliary release of glutathione disulphide was drastically augmented, leading to a decrease in the cellular and biliary GSH/GSSG ratios. It is suggested that lindane treatment leads to an induced oxidative capacity, which, in turn, alters the glutathione status of the liver tissue.     

Regulation of delta-aminolevulinate synthase activity during the development of oxidative stress.

Activities of rat liver delta-aminolevulinate synthetase (delta-ALAS), glutathione reductase (GR), and glucose-6-phosphate dehydrogenase (G6PDH), GSH content in the liver, and the absorption spectrum of blood serum were investigated after CoCl2, HgCl2, or beta-adrenoblocker (propranolol) injection and after CoCl2 and propranolol co-administration. Inhibition of the activity of the key heme biosynthesis enzyme delta-ALAS was most pronounced and prolonged during the first hours after CoCl2 and CoCl2 plus propranolol injections; this was associated with accumulation of Co2+--protoporphyrin-containing products of hemolysis. Inhibition of delta-ALAS after propranolol injection is not mediated by hemolysis. A decrease in GSH content precedes the induction of heme biosynthesis only in the case of HgCl2 administration, and this was associated with inhibition of GR and G6PDH. The decreased GSH content during the first hours after injection of propranolol and co-administration of CoCl2 and propranolol was not followed by increase in delta-ALAS activity 24 h after the injection. The mechanisms of the increase in the free heme content in the liver during the early stages of oxidative stress and the regulation of the key heme biosynthesis enzyme are discussed.     

Modification of hepatic vitamin E stores in vivo. I. Alterations in plasma and liver vitamin E content by methyl ethyl ketone peroxide

Since experiments with freshly isolated rat hepatocytes have shown that cellular vitamin E is consumed in response to insult by compounds that induce an oxidative stress only after cellular glutathione (GSH) concentrations have been substantially depleted, experiments were performed to determine whether this sequence of events occurred in response to oxidative insult in vivo. The role that plasma vitamin E plays in the response to chemically induced oxidative injury in vivo was also assessed. Treatments with 40 mg/kg of methyl ethyl ketone peroxide (MEKP) quickly induced lipid peroxidation in vivo and from one to 4 h after treatment caused a depression in the plasma content of vitamin E and the liver content of GSH, as well as signs of toxicity (elevations in serum activities of alanine and aspartate aminotransferases). At these time points however, the liver content of vitamin E was either indistinguishable from or slightly elevated from controls. By 12 to 24 h after treatment the liver content of vitamin E was reduced by 20-25% whereas values for all other indicators had returned toward control levels. Pretreatment of rats with L-buthionine-S,R-sulfoximine, an inhibitor of GSH by 4 or 24 h after treatment, did not alter the time course or extent of hepatic vitamin E depletion that was observed after treatment with MEKP. Other compounds that induce oxidative stress and lipid peroxidation to the liver, carbon tetrachloride and menadione, did not provoke an alteration in hepatic vitamin E levels as compared to controls 1 day after treatment. These findings indicate that depletion of hepatic vitamin E may not occur as an immediate consequence of oxidative insult to the liver and that the depletion of hepatic vitamin E levels may not be related to the extent of prior GSH depletion. Moreover, these findings suggest that alterations in the plasma concentration of vitamin E may not reflect concurrent alterations in hepatic vitamin E levels. A mechanism whereby liver vitamin E stores are mobilized for the maintenance of plasma vitamin E levels is proposed.     

Depletion of liver glutathione induced by iodobenzene poisoning and its relation to lipid peroxidation and necrosis

The mechanisms underlying iodobenzene hepatotoxicity were investigated in Albino mice in which the hepatic glutathione (GSH) content had been decreased by nearly 50% by starvation for 16 h before poisoning. After iodobenzene administration (9 mmol/Kg, p.o.) the hepatic GSH content decreased progressively and liver necrosis, as measured by the plasma transaminase (GPT, GOT) levels, occurred in many animals at 12 and 16 h. A clear cut necrosis was evident only when the hepatic GSH depletion reached a threshold value (3.5-2.5 nmol/mg protein). The same threshold value was evident for the occurrence of lipid peroxidation (measured as both carbonyl functions and conjugated dienes in liver phospholipids). The highly significant correlation found between lipid peroxidation and liver necrosis supports the possibility of a cause-effect relationship between the two phenomena.     

S-adenosylmethionine (SAMe) protects against acute alcohol induced hepatotoxicity in mice small star, filled

Although S-Adenosylmethionine (SAMe) has beneficial effects in many hepatic disorders, the effects of SAMe on acute alcohol-induced liver injury are unknown. In the present study, we investigated effects of SAMe on liver injury in mice induced by acute alcohol administration. Male C57BL/6 mice received ethanol (5 g/kg BW) by gavage every 12 hrs for a total of 3 doses. SAMe (5 mg/kg BW) was administrated i.p. once a day for three days before ethanol administration. Subsequent serum ALT level, hepatic lipid peroxidation, enzymatic activity of CYP2E1 and hepatic mitochondrial glutathione levels were measured colorimetrically. Intracellular SAMe concentration was measured by high-performance liquid chromatography (HPLC). Histopathological changes were assessed by H&E staining. Our results showed that acute ethanol administration caused prominent microvesicular steatosis with mild necrosis and an elevation of serum ALT activity. SAMe treatment significantly attenuated the liver injury. In association with the hepatocyte injury, acute alcohol administration induced significant decreases in both hepatic SAMe and mitochondrial GSH levels along with enhanced lipid peroxidation. SAMe treatment attenuated hepatic SAMe and mitochondrial GSH depletion and lipid peroxidation following acute alcohol exposure. These results demonstrate that SAMe protects against the liver injury and attenuates the mitochondrial GSH depletion caused by acute alcohol administration. SAMe may prove to be an effective therapeutic agent in many toxin-induced liver injuries including those induced by alcohol.     

Relation between lipid peroxidation and iron concentration in mouse liver after acute and subacute cadmium intoxication

In this study the effect of acute and subacute cadmium (Cd) intoxication on iron (Fe) concentration and lipid peroxidation (LPO) was investigated in the livers of Swiss mice. Animals were divided into two groups: the Cd group – mice intoxicated with Cd and controls. In acute time-response studies, Fe and malondialdehyde (MDA) levels were determined at 4, 6, 12, 24 and 48 h after a single oral dose of Cd (20 mg Cd/kg b.w.). In the subacute experiment, mice were given 10 mg Cd/kg b.w. orally every day for 14 days; Fe and MDA contents were determined in liver after 1 and 2 weeks. Acute Cd intoxication induced a significantly increased hepatic Fe content after 4 and 6 h, and a statistically significant increase in MDA 6, 12 and 24 h after Cd administration, although a significantly decreased MDA level was observed after 48 h. The results suggest development of early oxidative stress in livers of mice after acute intoxication with Cd. The decreased MDA observed after 48 h occurred presumably due to the adaptive response of the organism. Subacute Cd intoxication induced a significant decrease of hepatic Fe and MDA levels at both investigated time intervals compared with control. These results indicate a positive correlation between hepatic Fe and MDA content and suggest that prolonged Cd intoxication decreases hepatic LPO indirectly, by reducing the Fe content of mouse liver.     

Mechanistic aspects of enhanced lipid peroxidation following glutathione depletion in vivo

Several agents known to conjugate with glutathione (GSH) were administered to phenobarbital-induced rats resulting in a more or less pronounced depletion of hepatic GSH. In vitro incubations showed that a large enhancement of spontaneous lipid peroxidation was observed when the GSH content was below 1 μmol/g liver. This effect was inhibited by addition of exogenous GSH in a concentration-dependent manner, the GSH-concentration yielding 50% inhibition (I₅₀) being 1 μM. Using phorone (diisopropylidene acetone), which proved to be the most potent GSH-depletor, the time- and dose-dependence of the GSH-depletion and the consequent lipid peroxidation was studied. Again it was assured that the GSH concentration must reach a critical value of about 20% of the initial hepatic GSH content, before an enhanced lipid peroxidation is seen. Employing scavengers of excited oxygen species no evidence was found for the involvement of free oxygen radicals. Hepatoprotective agents and inhibitors of mixed-function oxidases exerted a more or less pronounced inhibitory action. Our findings are further support of our previous postulate that GSH depletion per se might lead to an increased lipid peroxidation, possibly due to its lack as a part of the cellular defence system against endogenous toxic intermediates.