Effects of dithiocarb and (+)-cyanidanol-3 on the hepatotoxicity and metabolism of vinylidene chloride in rats.

The acute hepatotoxic effects of vinylidene chloride (VDC) were evidenced by measurement of the increase in the serum levels of the aminotransferase (GPT) and sorbitol dehydrogenase (SDH), hepatic glutathione (GSH) depletion and histological examinations in rats. The hepatoprotective agents dithiocarb and (+)-cyanidanol-3 proved well able to antagonize these toxic effects of VDC. While dithiocarb inhibited the in vivo metabolism of VDC in a closed exposure system, (+)-cyanidanol-3 had no influence at all. These findings substantiate the role of the microsomal monooxygenase system in the metabolism and hepatotoxicity of VDC. The mechanisms by which dithiocarb and (+)-cyanidanol-3 act as antihepatotoxic agents are different: the inhibition of the metabolic activation by dithiocarb and free radical-scavenging by (+)-cyanidanol-3.     

Cytotoxic and cytoprotective activities of curcumin. Effects on paracetamol-induced cytotoxicity, lipid peroxidation and glutathione depletion in rat hepatocytes

The cytoprotective effect of curcumin, a natural constituent of Curcuma longa, on the cytotoxicity of paracetamol in rat hepatocytes was studied. Paracetamol was selected as a model-toxin, since it is known to be bioactivated by 3-methylcholanthrene inducible cytochromes P450 presumably to N-acetyl-p-benzoquinone imine (NAPQI), a reactive metabolite which upon overdosage causes protein- and non-protein thiol-depletion, lipid peroxidation and cytotoxicity measured as LDH-leakage. At low concentrations curcumin was found to protect significantly against paracetamol-induced lipid peroxidation, without protection against paracetamol-induced LDH-leakage and without protection against paracetamol-induced GSH-depletion. At a 100 times higher concentration of curcumin the observed protective effect on lipid peroxidation was accompanied with a tendency to increase cellular GSH-depletion and LDH-leakage. No time-dependency was found as to the curcumin-induced effects: treatment of the hepatocytes 1 hr before, concomitantly or 1 hr after the addition of paracetamol to the cells had similar effects. In contrast to what was expected on the basis of previous in vivo experiments, at higher concentrations curcumin itself was found to be slightly cytotoxic. Curcumin-induced LDH-leakage was accompanied by a significant depletion of GSH. It has been concluded that the observed cytoprotective and cytotoxic activities of curcumin may be explained by a strong anti-oxidant capacity of curcumin and the capability of curcumin to conjugate with GSH. Furthermore, it has been concluded that lipid peroxidation is not playing a causal role in cell-death induced by paracetamol or by curcumin.     

Role of lipid peroxidation on renal dysfunction associated with glutathione depletion. Effects of vitamin E.

This study was designed to investigate the role of lipid peroxidation in the pathogenesis of renal dysfunction in glutathione (GSH)-depleted rats. Renal function parameters and acid-base status were analyzed in diethylmaleate (DEM)-treated rats previously injected with vitamin E (Vit.E). Vit.E was effective in inhibiting the elevation in renal lipid peroxidation found in GSH-depleted rats. Vit.E also ameliorated the renal response to the metabolic acidosis without modification in lactate production induced by DEM administration. The increase in sodium and water urine excretion and the diminution of the urine to plasma osmolalities ratio were not reversed in these animals. These results lead us to conclude that lipid peroxidation is associated with distal acidification impairment observed with GSH-depletion, but it is not related to the sodium reabsorption alteration in the ascending loop of Henle.     

Time-course of changes in hepatic lipid peroxidation and glutathione metabolism in rats with carbon tetrachloride-induced cirrhosis

1. The aims of the present study were to assess: (i) the temporal relationships between hepatic lipid peroxidation, changes in the glutathione detoxification system and the onset/development of cirrhosis in CCl4-treated rats; and (ii) the effects of oral zinc administration on these parameters. 2. Cirrhosis was induced in 120 rats by intraperitoneal injections of CCl4 twice a week over 9 weeks. One hundred and twenty additional animals were used as controls. Both groups were further subdivided to receive either a standard diet or one supplemented with zinc. Subsets of 10 animals each were killed at weeks 1, 2, 3, 5, 7 and 9 from the start of the study. 3. Induction of cirrhosis produced a decrease in the components of the hepatic glutathione anti-oxidant system: glutathione transferase activity decreased from week 1, the concentration of reduced glutathione (GSH) decreased from week 5 and glutathione peroxidase (GPx) activity decreased from week 7. This impairment was chronologically related to an increase in free radical generation. Hepatic lipid peroxidation was significantly correlated with GPx activity (r = -0.47; P < 0.001) in CCl4-treated rats. Zinc administration did not produce any significant improvement of the hepatic glutathione system. 4. In conclusion, cirrhosis induction in rats by CCl4 administration produced a decrease in the hepatic glutathione antioxidant system that was related to an increase in free radical production. Furthermore, zinc supplementation produced a reduction in the degree of hepatic injury and a normalization of lipid peroxidation, but not an improvement of the hepatic GSH anti-oxidant system.     

Inhibition of the renin-angiotensin system attenuates the development of liver fibrosis and oxidative stress in rats

1. The present study was designed to investigate the potential antifibrotic and anti-oxidant effects of lisinopril, fosinopril and losartan in an experimental rat model of liver injury using carbon tetrachloride (CCl(4)). 2. First, the potential hepatoprotective dose of each drug was screened against CCl(4)-induced acute hepatotoxicity. Then, we chose the minimum hepatoprotective dose of each drug to further investigate the mechanisms involved in the hepatoprotection using a chronic model of hepatotoxicity induced by CCl(4). 3. Liver function was assessed in addition to histopathological examination. Furthermore, oxidative stress markers (reduced glutathione (GSH) and lipid peroxides levels) and markers of fibrosis (hydroxyproline content and liver fibrosis area) were assessed. 4. It was found that treatment of animals with different drugs concomitantly with CCl(4) significantly counteracted the changes in liver function induced by CCl(4) (except fosinopril). In addition, the drugs ameliorated the histopathological changes induced by CCl(4). All drugs significantly counteracted lipid peroxidation and GSH depletion (except fosinopril) compared with the CCl(4)-intoxicated group. Moreover, the drugs studied significantly reduced liver hydroxyproline levels and the area of fibrosis compared with the CCl(4)-intoxicated group. 5. In conclusion, the present study provides evidence for the hepatoprotective effect of lisinopril, fosinopril and losartan. Both lisinopril and losartan was found to have better hepatoprotective potential than fosinopril against CCl(4)-induced hepatotoxicity. These hepatoprotective effects can be explained on the basis of anti-oxidant and antifibrotic mechanisms, mainly enhancement of GSH and reduction of lipid peroxidation and fibrosis.     

Glutathione depletion by methyl chloride and association with lipid peroxidation in mice and rats

Inhalation of methyl chloride (CH3Cl) by male B6C3F1 mice resulted in a concentration-dependent depletion of glutathione (GSH) in liver, kidney, and brain. Exposure for 6 hr to 100 ppm CH3Cl decreased the concentration of GSH in mouse liver by 45%, while exposure to 2500 ppm for 6 hr lowered liver GSH to approximately 2% of control levels. For those exposures which decreased liver GSH to less than 20% of control levels, the extent of liver GSH depletion was closely correlated with the capacity of a 9000g supernatant fraction from the liver to undergo lipid peroxidation in vitro. GSH was depleted to a lesser extent in mouse brain and kidney, compared to liver, and no relationship to peroxidation was observed for single exposures to CH3Cl. A dose-dependent decrease in liver GSH was also produced by diethyl maleate, although a nearly lethal amount (2 ml/kg) was required to lower liver GSH to less than 10% of control levels. Under these conditions the amount of lipid peroxidation was 3.5-fold less than in mice exposed to 2000 ppm CH3Cl. Exposure of rats to 2000 ppm CH3Cl reduced liver GSH to 20% of control levels, compared to 4.5% in mice similarly exposed, and under these exposure conditions the amount of lipid peroxidation measured in vitro was 40-fold greater in mouse liver than in rat liver. During exposure of mice to 2500 ppm CH3Cl, ethane expiration increased to an extent comparable to that produced by administration of 2 ml/kg of CCl4. These findings suggest that GSH depletion in liver may be an important component of CH3Cl-induced hepatotoxicity.     

Cephaloridine-induced biochemical changes and cytotoxicity in suspensions of rabbit isolated proximal tubules

Cephalosporin antibiotics, such as cephaloridine (Cld), are known to be nephrotoxic in vivo and in vitro. In vivo, Cld causes proximal tubule necrosis in rabbits which is preceded by glutathione (GSH) depletion and, under certain conditions, inhibition of mitochondrial function. In vitro, Cld causes GSH depletion, lipid peroxidation, and inhibition of rat kidney slice organic ion uptake. The present investigations were designed to evaluate the temporal relationships of the biochemical "lesions" caused by Cld to the onset of lethal cell injury in suspensions of isolated rabbit proximal tubules. Cld was cytotoxic to suspensions of rabbit proximal tubules (EC50 = 1.10 +/- 0.33 mM) in the absence of amino acids (to support GSH synthesis). In this model, Cld also caused GSH and ATP depletion, lipid peroxidation (malondialdehyde formation), and inhibition of tubule respiration. Probenecid prevented Cld accumulation, tubule injury, ATP depletion, and lipid peroxidation and markedly attenuated the GSH depletion. Addition of glycine, cystine, and glutamate to the incubation buffer to support GSH synthesis decreased the tubule accumulation of Cld (due solely to the presence of glutamate) and blocked Cld-induced tubule lethality, lipid peroxidation, ATP depletion, and GSH depletion. Glycine or glutamate alone had no effect on Cld-induced cytotoxicity, whereas cystine was cytoprotective. Buthionine sulfoximine partially reversed the amino acid protection against Cld-induced tubule injury. Thus amino acid-induced protection of tubules from Cld cytotoxicity was due to the combination of a high intracellular GSH content and cytoprotection by cystine. The antioxidant N-N'-diphenyl-p-phenylenediamine (DPPD) blocked tubule injury, ATP depletion, and lipid peroxidation but had no effect on Cld-induced GSH depletion when tubules were incubated for 3 hr. However, when incubations were allowed to run for up to 8 hr, DPPD had no effect on Cld cytotoxicity, despite continued inhibition of lipid peroxidation. These data demonstrate that Cld-induced tubule injury in short-term (3 hr) incubations in vitro occurs by a mechanism probably involving lipid peroxidation and occurs only in the absence of amino acids to support GSH synthesis. Inhibition of tubule respiration and ATP depletion could not clearly be causally linked to the onset of cell death in this model. The mechanism of the peroxidation-independent Cld toxicity in tubules incubated for 8 hr or longer is not known at this time.     

绞股蓝在大鼠肝的抗脂质过氧化作用

目的：观察绞股蓝（GP）对四氯化碳（CCl4)所致肝脏脂质过氧化的干预作用。方法：将Wistar大鼠随机分为4组：A（对照组）;B（CCL4）;C（GP);D(CCL4＋CP）。结果：CCL4组的脂质过氧化物（LPO）含量明显高于对照组,谷胱甘肽过氧化物酶（GSH－Px）活性低于对照组;而同时给予GP的D组可减弱CCL4诱发的上述作用。结论：GP能减弱CCL4对肝脏的损害。     

Stimulation of lipid peroxidation and impairment of glutathione-dependent defense system in Wistar rats treated with cryptopine, a rare non-narcotic opium alkaloid.

The present study was designed to evaluate the effect of repeated oral administration of cryptopine at differential dosing regimens (50, 100, 150, 200 mg/kg bwt) in vivo on lipid peroxide measures, glutathione levels (GSH) and activity of glutathione S-transferase (GST) and glutathione reductase (GR) in the liver, spleen, kidney and lung of Male Wistar rats after a 5 day treatment period. In all the tissues examined, we observed an increase in lipid peroxidation and a decline in glutathione content and activity of glutathione S-transferase and glutathione reductase in a dose-dependent manner. The decrease in GSH content did not definitively correlate with a concomitant increase of lipid peroxidation in all the tissues. Our results ensemble that the enhancement of lipid peroxidation in the tissues investigated is a consequence of depletion of glutathione to certain critical levels and impairment of the glutathione-dependent enzyme systems viz. GST and GR. Our study potentiates that decreased levels of GSH may lead to lipid peroxidation, one of the key events in cellular damage. The inhibition of GST also suggests that the detoxification of the alkaloid could be suppressed following acute exposures. Conclusively, it appears that cryptopine in vivo disturbs the cellular defense system, so that it tips in the direction of autoxidative lipid peroxidation, producing cytotoxicity.     

Mechanism for the protective effects of silymarin against carbon tetrachloride-induced lipid peroxidation and hepatotoxicity in mice. Evidence that silymarin acts both as an inhibitor of metabolic activation and as a chain-breaking antioxidant

Administration of silymarin (800 mg/kg i.p.) 30 min before carbon tetrachloride (18 microL/kg i.p.) did not modify total hepatic levels of CCl4 and metabolites in mice, but decreased by 40% the in vivo covalent binding of CCl4 metabolites to hepatic lipids at 2 hr. This pretreatment decreased by 60% the exhalation of ethane during the first hour after CCl4, and decreased by 50% the incidence of liver cell necrosis. In vitro, silymarin (800 micrograms/mL) decreased by 50 to 70% various monooxygenase activities, and decreased by 20% the covalent binding of CCl4 metabolites to microsomal proteins. Silymarin (800 micrograms/mL) decreased by 70% in vitro lipid peroxidation mediated by CCl4 metabolites, and decreased by 90% peroxidation mediated by NADPH alone. Silibinin, one of the three isomers composing silymarin, also decreased carbon tetrachloride-induced lipid peroxidation; this effect, however, was less than that of silymarin in vitro, and was more transient in vivo. Pretreatment with silibinin (800 mg/kg i.p.) 30 min before CCl4 (18 microL/kg i.p.) did not improve SGPT activity or liver histology at 24 hr. We conclude that silymarin prevents carbon tetrachloride-induced lipid peroxidation and hepatotoxicity in mice, firstly, by decreasing the metabolic activation of CCl4, and, secondly, by acting as a chain-breaking antioxidant.     

Suppressive effect of tritoqualine on lipid peroxidation and enzyme leakage induced by carbon tetrachloride in rat hepatocytes

Since tritoqualine (TRQ) is effective in suppressing the increase of serum transaminases in acute hepatic injured rats induced by some hepatotoxins, protection of the hepatocyte membrane is suggested to be one of the pharmacological effects of TRQ. In the present study, we investigated the effects of TRQ on lipid peroxidation and enzyme leakage caused by carbon tetrachloride (CCl4) exposure in isolated hepatocytes and the liver in vivo, compared with vitamin E. The results were as follows: Hepatocytes isolated from TRQ-administered rats showed less enzyme leakage than those from control rats after CCl4 addition. TRQ displayed strong inhibition of lipid peroxidation in isolated hepatocytes. In comparison with vitamin E, TRQ showed almost the same inhibitory action on lipid peroxidation, but a stronger suppression of enzyme leakage. Vitamin E showed a weaker protection from increase of glutamic oxaloacetic transaminase than TRQ, in spite of its stronger inhibition of lipid peroxidation in vivo. From these results, it is suggested that the membrane protecting action of TRQ is partially derived from its suppression of lipid peroxidation, but "another action" may also play an important role in protecting the fragile membrane.     

In vitro tests of 1,3-dithia-2-thioxo-cyclopent-4-ene to evaluate the mechanisms of its hepatoprotective action

Some in vitro tests were performed using 1,3-dithia-2-thioxo-cyclopent-4-ene (DT827A) to evaluate its mode of action on hepatoprotection. Experiments with leucine, uridine and thymidine uptakes using L-132 cells showed no increase in leucine uptake, but did show a tendency toward increases in both uridine and thymidine uptakes, which suggested a stimulating effect on RNA and DNA synthesis in vitro. DT827A did not influence GSH levels in the HepG2 cultured cell system, and did not show inhibition of cytochrome P450 2El in microsomes obtained from mouse liver. There may be no possibility for DT827A to protect a liver injury through its influence on liver GSH levels and inhibiting metabolic activation of CCl4 in this in vitro system. However, the inhibitory effect of DT827A on lipid peroxidation was observed in vitro, the same as the observation in vivo. Furthermore, when DT827A was incubated with Alamar Blue in the presence of a mixture of mouse liver S9 and mitochondrial fractions for 20 hr, DT827A showed more reducibility than it did in a case of non-incubation, and it was suggested that DT827A would be metabolized in the S9-mitochondrial fraction in vitro to show an increase of Alamar Blue reducibility. The protective effect of DT827A on a liver injury is due neither to its influence on liver GSH levels nor inhibition of the metabolic activation of CCl4, but a possible mechanism of action for the DT827 series of compounds to indicate an antioxidative effect would be brought about by the role of the compounds as a radical scavenger as well as its reductive effect.     

Effect of hypoxia on the metabolism and hepatotoxicity of carbon tetrachloride and vinylidene chloride in rats

The metabolism of vinylidene chloride (VDC) and carbon tetrachloride (CCl4) was investigated by measuring the removal of the compounds from the atmosphere of a closed exposure system occupied with male rats. Hepatotoxicity was evidenced in the same rats by determining serum enzyme activities of the aminotransferases (GOT, GPT) and sorbitol dehydrogenase (SDH) before, at the end of the exposure time and 24 hrs later. Control rats exposed to VDC concentrations up to 2000 p.p.m. showed only slight increases of serum aminotransferase- and SDH-activities, which were not at all observed under hypoxic conditions. Hypoxia evoked a small, but significant reduction of the VDC metabolism at 500 p.p.m., but not at 2000 p.p.m. exposure concentration. In contrast to VDC CCl4-metabolism (150 p.p.m.) was increased under hypoxia and consequently hepatotoxicity was aggravated.     

Role of glutathione depletion in the cytotoxicity of acetaminophen in a primary culture system of rat hepatocytes

A primary culture system of postnatal rat hepatocytes was utilized to study the cytotoxicity of acetaminophen and the toxicological significance of glutathione (GSH) depletion. The relative time of onset and magnitude of GSH depletion, lipid peroxidation and cytotoxicity were contrasted in order to gain insight into their interrelationships. Exposure of the hepatocytes to acetaminophen resulted in time- and dose-dependent depletion of cellular GSH. The acetaminophen-induced GSH depletion and ensuing lactate dehydrogenase (LDH) leakage were quite modest and delayed in onset, in contrast to that caused by iodoacetamide (IAA) and by diethylmaleate (DEM), 2 well-known depletors of GSH. There was comparable LDH leakage, irrespective of drug treatment, when GSH levels decreased to about 20% of normal. Reduction of GSH levels below the 20% threshold by IAA treatment resulted in marked LDH leakage and loss of viability. Maximal LDH leakage in response to IAA and acetaminophen preceded maximal malondialdehyde (MDA) formation, suggesting that lipid peroxidation may be a consequence of cell damage as well as GSH depletion. IAA and DEM produced a comparable, modest accumulation of MDA, yet IAA was much more cytotoxic. These findings indicate that lipid peroxidation does not play a central role in hepatocellular injury by compounds which deplete GSH, although it may contribute to degeneration of the cell. As events in the cultured postnatal hepatocytes paralleled those reported in vivo, the system can be a useful and valid model with which to study mechanisms of chemical toxicity.     

Effect of iron overload on spontaneous and xenobiotic-induced lipid peroxidation in vivo

To study the effect of iron-overload on hepatic lipid peroxidation, two rat models of haemochromatosis were employed: in the first model resembling secondary haemochromatosis, repeated i.p. injections with Fe-dextran led to an accumulation of Fe in Kupffer cells, while in the second model resembling hereditary haemochromatosis, iron was located mainly in periportal hepatocytes after feeding on a diet containing 3.5% Fe-fumarate for 3 weeks. In both models, total hepatic iron content was elevated four- to fivefold over controls. In vivo lipid peroxidation (ethane exhalation) was enhanced only in the second model, indicating that the hepatocytes are the main targets of Fe-induced lipid peroxidation. Low hepatotoxicity was observed in the second model. Additional treatment of the rats with hepatotoxic agents led to different results: with ethanol and bromobenzene, lipid peroxidation was only evident in both models of iron-overload, while paracetamol-induced lipid peroxidation was seen only in Fe-fumarate-fed rats. CCl4-induced lipid peroxidation was strongly enhanced in both models of haemochromatosis. Hepatotoxicity was enhanced by iron overload only in the case of CCl4-treated, Fe-fumarate-fed rats. The activities of phase I and phase II enzymes of xenobiotic metabolism were not markedly altered in livers of iron-overloaded rats. This implies that neither the bioactivation nor the detoxification of the agents studied were affected in experimental haemochromatosis.     

Effects of novel antioxidants on carbon tetrachloride-induced lipid peroxidation and toxicity in precision-cut rat liver slices

Previous studies in rat liver microsomes have demonstrated the effectiveness of the 21-aminosteroid, U-74,006F, the troloxamine, U-78,517G, and N,N'-diphenyl-p-phenylenediamine (DPPD) in preventing carbon tetrachloride (CCl4)-induced lipid peroxidation. Studies reported here utilized liver slices to assess whether these antioxidants could prevent lipid peroxidation and ensuing toxicity in a more complete/complex system. Liver slices prepared from Aroclor 1254-induced SD rats were incubated in Dulbecco's modified eagle media, 37 degrees C, for up to 9 hr. Slices were preincubated with test compounds for 30 min prior to addition of CCl4. Lipid peroxidation, as measured by the formation of thiobarbituric acid-reactive substances and ethane evolution, was decreased by U-74,006F (100 microM), U-78,517G (100 microM), and DPPD (1 microM). CCl4 (2.5 microliters) decreased intracellular K+ content, intracellular lactate dehydrogenase (LDH), and intracellular isocitrate dehydrogenase (ICD) activities over a 9-hr incubation period. Despite the marked effects on lipid peroxidation, U-74,006F showed no protection against K+ or LDH loss and only moderate protection against ICD loss. U-78,517G showed no protection against K+ loss but substantial protection against enzyme loss. DPPD demonstrated slight protection against K+ and marked protection against enzyme loss. All three compounds inhibited CCl4-induced lipid peroxidation; U-78,517G being most effective, followed by DPPD and U-74,006F. Inhibition of lipid peroxidation provided protection to the membrane structure as indicated by inhibition of LDH and ICD loss. The antioxidants failed to protect against CCl4-induced toxicity (K+ loss). These results suggest that CCl4-induced lipid peroxidation and toxicity may be dissociable.     

Lethal effects of CCl4 and its metabolism by Mongolian gerbils pretreated with chlordecone, phenobarbital, or mirex.

Gerbils are much more sensitive to the hepatotoxic and lethal effects of CCl4 than rats as indicated by 48-hr LD50 values (0.08 vs 2.8 ml/kg). On the other hand, gerbils are refractory to chlordecone (CD) potentiation of CCl4 toxicity. To investigate the possible mechanism underlying the high sensitivity of gerbils to CCl4 lethality, the metabolism of CCl4 was studied in gerbils pretreated with dietary CD, phenobarbital (PB), or mirex (M) at 10, 225, and 10 ppm, respectively. The hepatic content of 14CCl4, the expiration of 14CCl4 and 14CCl4-derived 14CO2, and lipid peroxidation were measured and the results were compared with the previous data for rats. After the 15-day dietary pretreatment, male gerbils (60-80 g) received 14CCl4 (0.08 ml/kg; sp act 0.04 mCi/mmol) ip in corn oil and the radioactivity present in the expired air was collected for 6 hr. More than 80% of the parent compound as represented by the 14C-label in the toluene trap was expired in 6 hr regardless of the pretreatments. Expiration of 14CO2 measured during the 6 hr after 14CCl4 administration in control gerbils was 3.5-fold more than that in rats and was significantly increased in pretreated groups (M greater than PB greater than CD). PB and M pretreatments resulted in a significant increase of 14C-label bound to the nonlipid fraction of the liver as compared with CD-treated or control gerbils. The radiolabel present in the livers of control gerbils was 5-fold higher than that of rats. In vivo lipid peroxidation measured as diene conjugation in lipid extracts from the livers was lower in gerbils than in rats, and none of the pretreatments significantly affected lipid peroxidation. The serum alanine aminotransferase and aspartate aminotransferase were significantly elevated at 6 hr after CCl4 injection in all groups of gerbils. These data indicate that the more extensive metabolism of CCl4, as represented by 14CO2 formation and 14C-label bound to hepatic tissue, in gerbils as compared with rats, may partially explain the high sensitivity of gerbils to CCl4 toxicity. However, the enhanced metabolism of CCl4 found in CD-, PB-, or M-pretreated gerbils did not lead to amplified hepatotoxic and lethal effects of CCl4. The reason gerbils may be refractory to CD amplification of CCl4 injury might be associated with other factors yet to be investigated.     

Preventive effect of neutropenia on carbon tetrachloride-induced hepatotoxicity in rats

The preventive effect of neutropenia on carbon tetrachloride (CCl4)-induced hepatotoxicity was examined in rats. In rats treated once with CCl4 (1 ml kg(-1), i.p.), the serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), indices of liver cell damage, and the hepatic activity of myeloperoxidase (MPO), an index of tissue neutrophil infiltration, increased at 6 h after the intoxication and further increased at 24 h. The liver of CCl4 -treated rats showed an increase in the concentration of thiobarbituric acid reactive substances (TBARS), an index of lipid peroxidation, and decreases in superoxide dismutase (SOD) activity and reduced glutathione (GSH) concentration at 6 h after the intoxication followed by a further increase in TBARS concentration and further decreases in SOD activity and GSH concentration at 24 h with increased xanthine oxidase (XO) activity at 24 h. Neutropenic treatment with anti-rat neutrophil antiserum (2 ml kg(-1), i.p.) at 0.5 h after CCl4 intoxication attenuated the increases in serum ALT and AST activities and hepatic MPO activity and TBARS concentration and the decreases in hepatic SOD activity and GSH concentration found at 6 and 24 h after CCl4 intoxication and the increase in hepatic XO activity found at 24 h after the intoxication. This neutropenia reduced the necrotic and degenerative changes with inflammatory cell infiltration in the liver cell of CCl4 -treated rats. These results indicate that neutropenia prevents CCl4 -induced hepatotoxicity in rats by attenuating the disruption of hepatic reactive oxygen species metabolism mediated by neutrophils accumulating in the liver tissue.     

The role of iron in the NADPH-dependent lipid peroxidation due to glutathione depletion by phorone

Enhanced NADPH-dependent LPO in rat liver postmitochondrial supernatants in vitro due to depletion of GSH by treatment with phorone (diisopropylidene acetone) in vivo was inhibited by Fe2+-chelating agents (desferrioxamine, DETAPAC), but not by scavengers of .O2-, H2O2, singlet oxygen or .OH-radicals, indicating that a perferryl ion (Fe2+ . O2) is needed as an initiating factor. In vivo, rats treated with phorone (250 mg/kg i.p.) exhaled 1.4 times as much ethane within 4 h as compared to controls. Pretreatment with FeSO4 resulted in a 6.9-fold enhancement of LPO as compared to Fe2+-pretreated controls. Our results indicate that GSH-depletion results in a strong enhancement of NADPH-dependent LPO also in vivo, provided that an initiating factor is present.     

Activation of the microsomal glutathione-S-transferase and reduction of the glutathione dependent protection against lipid peroxidation by acrolein

Allyl alcohol is hepatotoxic. It is generally believed that acrolein, generated out of allyl alcohol by cytosolic alcohol dehydrogenase, is responsible for this toxicity. The effect of acrolein in vitro and in vivo on the glutathione (GSH) dependent protection of liver microsomes against lipid peroxidation, and on the microsomal GSH-S-transferase (GSH-tr) in the rat was determined. In vitro incubation of liver microsomes with 5 mM acrolein for 30 sec resulted in a 2-fold activation of the GSH-tr. This activation probably proceeds via alkylation of the thiol group of the GSH-tr. In vivo administration of 1.1 mmol allyl alcohol/kg to rats did also result in a 2-fold stimulation of the GSH-tr activity. Administration of 375 mg pyrazole/kg, an inhibitor of the alcohol dehydrogenase, thus reducing the acrolein formation, prevented the in vivo stimulation of GSH-tr by allyl alcohol. This indicates that the activation of GSH-tr in vivo by allyl alcohol probably also proceeds via alkylation of the thiol group of the GSH-tr by acrolein. GSH protects liver microsomes against lipid peroxidation, probably via a free radical reductase that reduces vitamin E radicals at the expense of GSH. Incubating liver microsomes for 30 min with 0.1 mM acrolein reduced the GSH dependent protection against lipid peroxidation, probably because an essential thiol group(s) on the free radical reductase is alkylated. In vivo administration of allyl alcohol did not reduce the GSH dependent protection of the microsomes. Probably the thiol group(s) located on the free radical reductase is less accessible or less reactive than the thiol group on the GSH-tr. After administration of allyl alcohol we found no evidence for in vivo lipid peroxidation. Therefore we could not evaluate the importance of the GSH dependent protection against lipid peroxidation in vivo.