Hepatic GSH Content Research Articles

No documentable role for xanthine oxidase in the pathogenesis of hepatic in vivo ischaemia/reperfusion injury

An investigation was made into the possible involvement of the enzyme xanthine oxidase (XO) (EC 1.1.3.22), both reversible (XOrev) and irreversible (XOirr), in damage observed after short-term in vivo hepatic ischaemia/reperfusion (60 or 120 min I and 15 min R) in fasted rats with: (i) a physiological content of XO (25%); and (ii) higher XO percentage (45%). In the latter the hepatic XO physiological percentage was increased by diethylmaleate treatment (300 mg kg −1) that depleted the cytosolic glutathione (GSH) to 14% of the controls. It was shown that, in animals with physiological content of XO, 60 and 120 min of hepatic ischaemia followed by 15 min reperfusion results in decreased GSH levels, and significantly increased alanine aminotransferase (ALT) and aspartate aminotransferase (AST) serum levels, without any modification of either the percentages of XO (XOirr and XOrev) or the hepatic thiobarbituric acid reactive substances (TBARS). Sixty minutes of ischaemia/reperfusion in rats with the higher XO level and lower hepatic GSH content led to further conversion of XDH to XOrev, with no increase in XOirr. In addition, the ALT and AST serum levels in these animals rose to the same extent as in normal rats after 120 min ischaemia and 15 min reperfusion, this extent being observed to be associated with a moderate increase in thiobarbituric acid reactive substances (TBARS). However, the administration of allopurinol, at a dose of 50 mg kg 1, which almost completely inhibits XO activity, did not lead to any decrease in liver damage or TBARS. These findings exclude any role of XO in liver damage in the short term following ischaemia/reperfusion events, also when marked GSH depletion could increase the enzymatic physiological XO level.

Protective effect of antioxidants against free radical-mediated lipid peroxidation induced by DON or T-2 toxin.

An experiment was conducted on rats to investigate the capacity of antioxidants to protect against acute toxicity caused by DON or T-2 toxin. Male rats were fed two different feeds. One group received a feed deficient in vitamins C and E and selenium, whereas the other group was fed with a feed enriched in antioxidants. After two weeks, selected groups of rats were administered orally a single dose of DON or T-2 toxin. After the treatment with mycotoxins, all rats were decapitated. The livers were analyzed for TBARS values, hepatic GSH content and for the activities of CyP-450, CAT, SOD and GSH-TR. Increases in lipid peroxides of 21% and 268% were observed in those rats which did not receive the supplement of antioxidants and which were administered DON or T-2 toxin, respectively. There was no significant increases in the TBARS values in the groups receiving DON with selenium and vitamins, but increases of 57% and 79% were recorded in the groups administered T-2 toxin and antioxidants. Furthermore, in the groups fed the deficient feed and administered DON or T-2 toxin, the lipid peroxidation increased by 33% and 307%, respectively. No mortality, and a lower number of intoxicated animals were observed in rats fed a diet supplemented with antioxidants. Significant decreases of GSH, CAT, SOD, CyP-450 and GSH-TR were recorded in treated rats receiving the deficient feed. The results of this study demonstrate that trichothecenes stimulate lipid peroxidation with consequent decrease of GSH content, but that the dietary use of selenium, alpha-tocopherol and ascorbic acid provides protection against acute toxicosis caused by DON or T-2 toxin.

Age-related Changes in Glutathione Concentration, Glutathione Peroxidase, Glutathione-S-Transferase, and Superoxide Dismutase Activities in Senescence Accelerated Mice

Age-associated changes in several antioxidative factors were studied in blood and liver from mice prone to accelerated senescence aged 0.7 to 1.4 years and mice resistant to accelerated senescence aged 1.0 to 2.1 years. In both strains, activities of hepatic glutathione peroxidase (GSH-Px) and glutathione-S-transferase (GST) decreased significantly with age, and the activity of superoxide dismutase (SOD), and the concentration of glutathione (GSH) in liver and blood cells tended to decrease with age. The GSH concentration in the plasma and blood cells, and the hepatic GST and SOD activities were significantly lower in the mice prone to senescence than other strain, but the hepatic GSH concentration and GSH-Px activity were similar in the two strains. These results suggest that GSH and hepatic antioxidative enzymes are associated to some extent with senescence in mice with accelerated senescence.

Effects of dietary protein source on plasma lipids, HMG CoA reductase activity, and hepatic glutathione levels in gerbils

It has recently been postulated that soy protein-induced hypocholesterolemia is induced by increased production of reduced glutathione (GSH). The objective of the current study was to determine the influences of dietary proteins and amino acids on plasma lipid concentrations, hepatic GSH, and HMG CoA reductase activity. Twenty-eight adult male, inbred gerbils were fed diets similar in all respects except that dietary protein source was from either soy protein isolate (SOY), casein (CAS), L-amino acid patterned after SOY (SAA), or L-amino acid patterned after CAS (CAA). Results indicated that gerbils fed SOY had lower plasma total cholesterol concentrations than gerbils fed CAS or CAA. Low density lipoprotein-cholesterol was also lowest in SOY-fed animals compared with all other groups. High density lipoprotein (HDL)-cholesterol concentrations were highest in CAA-fed animals. Plasma triglycerides were unaffected by dietary treatment. Hepatic GSH concentrations were lowest in SOY-fed gerbils, but were not different between SAA- and CAA-fed groups. HMG CoA reductase activity was highest in gerbils fed SOY compared with CAS; however, when amino acids were fed the opposite occurred-reductase activity was higher in gerbils fed CAA compared with animals fed SAA. Correlation analysis showed no association between GSH and HMG CoA reductase activity. Positive correlations were present between GSH and total and HDL-cholesterol concentrations. These results indicate that lipid metabolism is influenced by dietary protein source, but these changes are not likely to be mediated by a GHS-mediated modification of cholesterol biosynthesis.

Influence of thyroid hormone administration on hepatic glutathione content and basolateral γ-glutamyltransferase ectoactivity in the isolated perfused rat liver

The effect of thyroid hormone administration on liver glutathione (GSH) content and γ-glutamyltransferase activity in the isolated perfused liver was studied for a period of 1–7 days in fed rats following a single dose of 0.1 mg 3,5,3'- l-triiodothyronine (T 3)/kg. T 3 elicited an early and transient calorigenic response, together with GSH depletion at 1 day after treatment. Recovery of hepatic GSH content and enhancement in total basolateral γ-glutamyltransferase activity occurred in parallel 2–3 days after T 3 treatment, parameters that were normalized in the 4- to 7-day time interval studied. The increase in total basolateral γ-glutamyltransferase activity by T 3 at early times after treatment was due mainly to increments in its transpeptidation mechanism, and was characterized by increments in the apparent maximum velocities without changes in the apparent Michaelis constant ( K m ) for the substrate γ-glutamyl- p-nitroanilide. Data presented suggest that the elevation in sinusoidal γ-glutamyltransferase activity could be related to the recovery of hepatic GSH content after depletion by T 3 treatment, by supplying the precursors for intracellular GSH synthesis, an effect that seems to be mediated by enhanced synthesis of the enzyme.

Effects of aurothioglucose and dietary Se on glutathione S-transferase activities and glutathione concentrations in chick tissues.

Experiments were conducted to determine whether the increased glutathione S-transferase (GSH-T) activity associated with selenium (Se) deficiency is necessarily related to losses in the activity of Se-dependent glutathione peroxidase (SeGSHpx) in chicks. Nutritional Se status was altered in two ways: by treatment with an antagonist of Se utilization, aurothioglucose (AuTG), and by feeding diets containing excess Se. Chicks given AuTG (10-30 mg AU/kg, sc) had growth rates and hepatic GSH concentrations that were comparable to those of saline-treated controls; however, their plasma GSH levels exceeded those of either Se-deficient (6-fold) or -adequate (3-fold) saline-treated chicks. Hepatic SeGSHpx activities of AuTG-treated chicks were half those of controls under conditions of Se-adequacy; however, this effect was not detected when Se was deficient. Hepatic GSH-TCDNB (assayed with 1-chloro-2,4-dinitrobenzene) activities of AuTG-treated chicks were significantly greater than those of controls when Se was deficient (i.e., when SeGSHpx activity was 12% of the Se-adequate level); however, deprivation of Se did not affect GSH-TCDNB activity in the absence of AuTG. Chicks fed excess Se (6-20 ppm as Na2SeO3) in diets containing either low (2 IU/kg) or adequate (100 IU/kg) VE, showed hepatic GSH-TCDNB activities and GSH concentrations greater than those of Se-adequate (0.2 ppm Se) chicks by 100% and 40%, respectively. That increased hepatic GSH-TCDNB activity can occur because of either AuTG or excess Se status under conditions wherein SeGSHpx activity is not affected indicates that the transferase response is not directly related to changes in the peroxidase.

Effect of methanol or modifications of the hepatic glutathione concentration on the metabolism of dichloromethane to carbon monoxide in rats.

1. The metabolism of dichloromethane (DCM) to carbon monoxide as measured by the carboxyhaemoglobin (COHb) level in the blood is stimulated by pretreatment with methanol (MET). After simultaneous administration of both DCM, 6.2 mmol kg-1 p.o., and MET, > 148 mmol kg-1 p.o., the COHb formation is inhibited. 2. MET ingestion results in a transient decrease of the glutathione (GSH) content of the liver. In rats treated with GSH-depleting chemicals such as diethylmaleate (DEM), phorone (PHO), or buthionine sulphoximine (BSO) there were no enhancements of the carboxyhaemoglobinaemia caused by DCM. The COHb formation was not influenced by an increase of the hepatic GSH concentration due to repeated administration of butylated hydroxyanisole (BHA). 3. It is concluded that cytochrome P450 IIE1 (CYP 2E1) is responsible for the metabolic interaction of both DCM and MET, and MET may be an inducer of CYP 2E1. The two pathways of DCM, the oxidative via CYP 2E1 and the metabolism via GSH/GSH-S-transferase seem to be independent.

Enhancement of renal and hepatic glutathione metabolism by dimethylthiourea

Dimethylthiourea (DMTU), a free radical scavenger, was administered to rats to study its effect on renal and hepatic glutathione metabolism, since it is a potential sulfhydryl donor. Six hours following DMTU, renal GSH content was significantly ( P < 0.05) increased (10%), and was increased further after 24 h (28%) ( P < 0.001). Hepatic GSH content was also significantly ( P < 0.001) elevated at 6 and 24 h (5 and 33%, respectively). Seven days of daily DMTU therapy significantly ( P < 0.001) increased renal and hepatic GSH content by 36 and 54%, respectively, which was associated with a significant ( P < 0.001) increase in the renal activities of glutathione peroxidase (GP) by 38%, glutathione transferase (GT) by 92%, and glutathione reductase (GR) by 19% ( P < 0.05). Significantly increased activities of hepatic GP by 84% ( P < 0.01) and GT by 101% ( P < 0.001) also occurred in DMTU-treated rats after 7 days of continuous therapy. From these data, we conclude that DMTU stimulates renal and hepatic GSH metabolism, which may be important in mediating DMTU's protective effect against free radical-induced tissue injury.

Effects of feeding and fasting on hepatolobular distribution of glutathione and cadmium-induced hepatotoxicity

Relationship between hepatolobular distribution profile of glutathione (GSH) and cadmium (Cd)-induced hepatotoxicity was examined in both fed and fasted rats by computerized densitometry of histochemically stained GSH in the liver sections using an image analyzer system. In fed rats, density gradient distribution of hepatolobular GSH, which was higher in the periportal region than in the perivenous one, was always observed even at a diurnally minimal concentration of GSH. This heterogeneous distribution of GSH, however, disappeared in fasted rats, even though the hepatic GSH concentration recovered to 81% of the control level in rats fasted for 48 h. In histopathological examination on livers 24 h after oral treatment of fed and fasted rats with 60 mg Cd/kg, zonal necrotic changes were observed from the perivenous to midlobular region but not in the periportal one in fed rats even at a diurnally minimal concentration of hepatic GSH. On the other hand, necrotic changes in the liver extended to the panlobular region including the periportal one in fasted rats. These necrotic changes were greater with a longer duration of fasting. These results suggest that the density gradient distribution of hepatic GSH but not the actual concentration of the compound plays an important role in protecting rats against acute hepatotoxicity of Cd.

Varied Protein Intake Alters Glutathione Metabolism in Rats

We tested the possibility that protein consumption greater than needed for optimum growth of young adult rats might increase either their hepatic glutathione (GSH) content or plasma GSH turnover. Additional aims were to characterize the relationship between hepatic GSH content and plasma turnover under physiologic conditions and to evaluate the ability of values obtained by noninvasive blood sampling to accurately predict hepatic GSH content. Male Sprague-Dawley rats (300 g) were adapted to purified diets containing 0, 5, 10, 20 or 40% casein. Plasma GSH and amino acid concentrations, hepatic GSH content and [35S]GSH-determined plasma GSH turnover were measured in the anesthetized, intact animals. As dietary protein increased from 0 to 20% casein, liver weight and liver GSH concentration (μmol/g wet wt) both increased. In rats fed the 40% casein diet, liver weight increased even further while liver GSH concentration decreased, with the net result that total liver GSH content of the 40% casein-fed group was not significantly different from that of the 20% casein-fed group. Plasma urea, cysteine, methionine and GSH concentrations increased with increasing protein intake, but with the exception of plasma urea, which increased by 60%, there was no further increase at the 40% casein level. Plasma GSH turnover also increased as dietary casein increased from 0 to 20% but was not significantly increased further by the 40% casein diet. A sigmoid function best described the relationship between plasma GSH turnover and hepatic GSH content (r = 0.80, P < 0.0001). The best indirect predictor of liver GSH content was not plasma GSH concentration (r = 0.56, P < 0.0001), but rather plasma cysteine (r = 0.84, P < 0.0001).

Glutathione homeostasis in rats chronically treated with ethanol. Evidence for an increased hepatic GSH export in vivo.

The influence of chronic ethanol feeding to rats on the hepatic glutathione (GSH and GSSG) system (synthesis, catabolism, export) and on the GSH and GSSG concentrations in extrahepatic tissues was investigated. Histological examination of livers from ethanol pretreated rats revealed a minor dilatation of the hepatic sinusoids. After ethanol administration the distribution pattern of gamma-glutamyltranspeptidase (enzymehistochemistry) was nearly unchanged, but the hepatic activity of this enzyme was increased. The ethanol pretreatment led to a decrease in hepatic GSH content. The hepatic activity of the GSSG-reductase were increased after ethanol treatment whereas the activities of the GSH synthesizing enzymes (gamma-glutamyl-cysteinyl-synthetase and GSH-synthetase) were not affected. A strong increase in sinusoidal GSH export was found in the ethanol-pretreated rats. The GSH- and GSSG concentrations of brain, lung, kidney and skeletal muscle were unchanged. It can be concluded that the ethanol-induced alteration of the hepatic GSH metabolism is caused mainly by changes of the sinusoidal membrane of the hepatocytes (direct effect of ethanol on the sinusoidal GSH carrier) leading to an increased GSH export into plasma. This effect should not due to an increased extrahepatic requirement for GSH.

The protective role of glutathione in chlorothalonil-induced stoxicity to channel catfish

The purpose of this investigation was to examine the nature of the protective role of glutathione (GSH) in liver and gills of channel catfish ( Ictalurus punctatus) exposed to chlorothalonil (2,4,5,6 tetrachloroiso-phthalonitrile). In an initial experiment, the acute toxicity of chlorothalonil was enhanced three-fold after channel catfish were depleted of liver and gill GSH by l-buthionine S.R-sulfoximine (BSO. 1000 mg/kg i.p.) and diethyl maleate (DEM, 0.6 ml/kg i.p.), indicating that GSH plays a critical role in mitigating acute chlorothalonil toxicity in this species. In a subsequent experiment, sublethal exposure to chlorothalonil elicited significant increases ( p < 0.05) in hepatic GSH concentrations at 72 h and 144 h of exposure. Elevations in hepatic GSH were accompanied by increases in hepatic cysteine concentrations at 72 h and also apparent induction of hepatic gammaglutamylcysteine synthetase (GCS) activity at 144 h. Gill GSH in chlorothalonil-exposed catfish was also significantly increased at 72 h and 144 h. as were increased gill cysteine concentrations. Gill GCS activity was somewhat elevated in chlorothalonil-exposed catfish at 144 h; however, this increase was not statistically significant ( p > 0.05). These results indicate that the mechanism of GSH induction in liver and gills of catfish exposed to chlorothalonil involves increased cysteine uptake and also increased GCS activity. The results of this study support the hypothesis that simultaneous exposure to multiple environmental chemicals that utilize or deplete tissue GSH may predis pose fish to acute toxicity.

A comparison of glutathione-dependent enzymes in liver, gills and posterior kidney of channel catfish ( ictalurus punctatus)

1. Six enzymes which collectively catalyze a number of glutathione-dependent synthetic, catabolic and detoxification reactions were examined along with glutathione status in liver, gills, and posterior kidney of channel catfish ( Ictalurus punctatus). 2. Hepatic GSH concentrations were higher than those in kidney or gills. Oxidized glutathione (GSSG) concentrations were similar among the three tissues. 3. Specific (per unit protein) gamma-glutamylcysteine synthetase (GCS) activity was greater in the gills than in liver or posterior kidney. However, total organ GCS activity was greatest in the liver. 4. Specific and total hepatic glutathione peroxidase (GSH peroxidase) activities were substantially greater than those of gills or kidney. 5. Similar specific glutathione reductase (GSSG reductase) activities were observed among all three tissues. 6. All three tissues exhibited glutathione S-transferase (GST) activity towards 1-chloro-2,4-dinitrobenzene (CDNB). Specific and total organ GST activities were highest in the liver, followed by the posterior kidney and gills. 7. Gamma-glutamyltranspeptidase (GGT) activity was present in the posterior kidney, but was undetectable in the gills or liver.

Effects of buthionine sulfoximine and diethyl maleate on glutathione turnover in the channel catfish

Despite the growing use of fish in toxicological studies, little is known regarding glutathione (GSH) metabolism and turnover in these aquatic species. Therefore, we examined GSH metabolism in the liver and gills of channel catfish ( Ictalurus punctatus), a commonly employed aquatic toxicological model. Treatment of channel catfish with l-buthionine- S, R-sulfoximine (BSO, 400 or 1000 mg/kg, i.p.), an inhibitor of GSH biosynthesis, did not deplete hepatic GSH in channel catfish. In addition, hepatic GSH concentrations did not fluctuate in catfish starved for 3 days, indicating relatively slow turnover of hepatic GSH. However, hepatic GSH concentrations were reduced significantly (P < 0.05) after 7 days of starvation. Administration of the thiol alkylating agent diethyl maleate (DEM, 0.6 mL/kg, i.p.) resulted in depletion of 85% of hepatic GSH at 6 hr post-DEM, with complete GSH recovery observed at 24 hr post-DEM. Co-administration of BSO and DEM (1000 mg/kg, 0.6 mL/kg, respectively) substantially depleted gill GSH and eliminated detectable liver GSH. Following BSO/DEM, GSH recovery in hepatic mitochondria occurred more rapidly than did liver cytosolic GSH. γ-Glutamylcysteine synthetase (GCS) activities were comparable in the 10,000 g supematants of catfish liver and gills (204 ± 21 and 268 ± 20 nmol/min/mg protein, respectively) whereas γ-glutamyltranspeptidase (GGT) activity was not detected in the 600g post-nuclear fraction of either liver or gills. In conclusion, i.p. administration of DEM was an effective means for achieving short-term hepatic GSH depletion in channel catfish, whereas co-administration of BSO and DEM elicited prolonged and extensive hepatic GSH depletion in this species. Like rodents, channel catfish maintained physiologically distinct hepatic mitochondrial and cytosolic GSH pools, and also regulated hepatic GSH levels by in situ hepatic GSH biosynthesis. However, unlike rodents, there was no evidence for a labile hepatic cytosolic GSH pool in channel catfish. These similarities and differences need to be considered when designing toxicological studies involving the GSH pathway in channel catfish and possibly other fish species.

Biochemical changes and essential metals concentration in lead-intoxicated rats pre-exposed to ethanol

The effects of chronic lead exposure on some hematopoietic and hepatic biochemical indices and urine, feces, and tissue essential metal concentration were investigated in rats pre-exposed to different doses of ethanol. Exposure to ethanol (0.5, 1.0, and 2.0 g/kg intraperitoneally, once daily) for 4 weeks produced an inhibition of blood δ-aminolevulinic acid dehydratase (ALAD) activity and a decrease in hepatic glutathione (GSH) concentration. Ethanol ingestion also produced a dose-dependent elevation of hepatic lipid peroxidation. Blood and hepatic calcium and hepatic magnesium contents decreased and urinary Ca and fecal Ca and Mg contents increased significantly following 4-week exposure to ethanol (2 g/kg). Lead administration (10 mg/kg, orally) for 4 weeks in ethanol pre-exposed (2 g/kg) animals produced a more pronounced inhibition of blood ALAD and elevation of urinary δ-aminolevulinic acid (ALA) excretion and hepatic GSH contents. Hepatic GSH contents decreased and hepatic lipid peroxidation increased significantly in rats given lead and pre-exposed to ethanol (2 g/kg). A more pronounced depletion of blood Ca and Mg and hepatic Mg was observed along with significant elevation of urinary Mg and fecal Ca excretion in animals administered lead and pre-exposed to ethanol (2 g/kg). The results suggest that nutritional deficiencies, particularly depletion of body Ca and Mg levels, play an important role in increasing susceptibility to lead intoxication in the rat.

Diurnal variation in hepatic glutathione content as a function of age

The purpose of this study was to investigate the influence of aging on the diurnal varitaion in glutathione (GSH) content of mouse liver. Young (7 months) and old (27 months) C57BL mice were housed under standarized conditions with lighting from 0800 to 2000 and free access to food and water. GSH was determined in acid extracts of liver homogenates by HPLC separation and fluorometric detection of the o-phthaldialdehyde derivative. A diurnal rhythm in hepatic GSH concentration was evident in young and old mice of both sexes. Peak levels of GSH occurred at 1000 and were unaffected by age or gender; however, age and sex-dependent differences were observed in the lowest concentration of GSH and in the amplitude of the GSH cycle. Hepatic GSH levels at 2200 and 0600 were 30–35% lower in old males than in young males and 25–40% lower in young females than in young males. The results may provide an explanation for the inconsistencies in previous findings about the influence of aging on hepatic GSH concentrations. © 1992 Wiley-Liss, Inc.

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 4hr, once) or low dose (500 ppm for 6hr, 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 (< 40-fold) and hsp90 (< 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.

The effects of bucillamine on glutathione and glutathione-related enzymes in the mouse

The effect of bucillamine (BA) on glutathione (GSH) and GSH-related enzymes was investigated in C57 mouse. Administration of high doses of BA (150–400 mg/kg) produced a dose-dependent depletion (20–44%) of hepatic GSH, which was similar in magnitude to that produced by equimolar doses of other sulphydryl drugs studied previously. GSH depletion after acute BA administration correlated well with the elevation of serum glutamic-pyruvic transaminase (SGPT) (6–9-fold increase above control). The increase in SGPT after chronic administration (7 days), although significantly higher than the controls, was however much less than after acute administration. The hepatic GSH concentrations of mice given 7 days of BA were similar to the controls, again correlating well with SGPT activity. Administration of BA (150–400 mg/kg) caused also a significant dose-dependent increase in the oxidized glutathione (GSSG) in blood by 2–7-fold, as well as a dose-dependent increase in blood glutathione S-transferase (GST) activity (2–13-fold). In an in vitro experiment, hepatic GST activity was activated by various concentrations of BA (1 μM–1 mM). There was little or no effect on GSSG reductase and on glutathione peroxidase (GSH-Px) after acute administration of BA. Chronic administration of BA had no effect on hepatic GSSG reductase and GSH-Px, but GSSG reductase activity in blood was increased significantly by 4-fold. It is possible that BA may affect the redox status through auto-oxidation and oxidation with endogenous thiols such as glutathione, affecting GSH concentrations and the GSH/GSSG ratio in tissues and, thus, having both metabolic and toxicological consequences. Whether or not the induction of GST activity in vivo in blood and in vitro in liver enzyme preparations shared the same underlying mechanism(s) requires further investigation.

Effect of ascorbic acid esters on hepatic glutathione levels in mice treated with a hepatotoxic dose of acetaminophen

Acetaminophen (APAP) with or without ascorbyl stearate (AS) or ascorbyl palmitate (AP) was administered by gavage to male Swiss-Webster mice at a dose of 600 mg/kg for each chemical. The biochemical markers of hepatotoxicity, serum transaminases (serum glutamate pyruvate transaminase [SGPT], serum glutamate oxaloacetic transaminase [SGOT]) and serum isocitrate dehydrogenase (SICD) activities were monitored after APAP and APAP + AP or AS dosing. There were significant reductions in serum transaminase and SICD activities in the APAP- + ascorbate ester-treated animals as compared to APAP-positive controls. Oral coadministration of APAP with AP or AS did not prevent the initial hepatic GSH depletion (15 min-4 hr postdosing). However, hepatic GSH content began to rise in the APAP + AS or AP-treated animals at 4 hr and reached control values within 12 hr postdosing. Urinary mercapturate conjugates were also significantly higher in the APAP + AP or AS-treated animals as compared to APAP alone when measured over a 60-min postdosing period. Plasma sulfobromophthalein (BSP) retention was approximately eight times higher in APAP-treated animals as compared to the APAP + ascorbate ester treatments indicating maintenance of hepatic excretory functions in presence of AP or AS. Prior depletion of hepatic GSH by diethyl maleate (DEM) did not alter hepatoprotective effects of AP or AS in the presence of APAP. Hepatic ascorbate levels also peaked at 4 hours after APAP + AP or AS treatments. The possible role of L-ascorbic acid esters in GSH regeneration following co-administration of a hepatotoxic dose and APAP is discussed.

Oltipraz-induced amelioration of acetaminophen hepatotoxicity in hamsters: II. Competitive shunt in metabolism via glucuronidation

Previously, we demonstrated that oltipraz [OTP: 5-(2-pyrazinyl)-4-methyl-1,2-dithiol-3-thione] prevented the hepatotoxicity of acetaminophen (AAP) in hamsters and that the observed protection was not related to increases in hepatic reduced glutathione (GSH) levels. These experiments were designed to elucidate the mechanism of OTP-induced protection with respect to an apparent non-GSH-dependent system. Marked differences in the relative amounts of hepatic GSH content depleted by AAP in control vs OTP-treated hamsters occurred. Urinary recoveries of AAP and metabolites indicated that more AAP-glucuronide was formed at the expense of other major metabolites (AAP-GSH, - N-acetylcysteine, and -sulfate) in OTP-treated hamsters, while plasma toxicokinetic modeling suggested a greater rate of AAP systemic clearance. An increased apparent formation rate constant for AAP glucuronidation (135%), in concert with significantly lower apparent formation rate constants for those metabolites which reflect the production of the reactive intermediate from AAP (glutathione and N-acetylcysteine), provide the rationale for this shift of metabolism. The biochemical basis for metabolic shunting is significantly elevated hepatic UDP-glucuronic acid content, an increased calculated UDP-glucuronic acid synthetic rate, and an increased liver microsomal UDP-glucuronyl transferase activity in OTP-treated animals. These changes in AAP conjugation were concomitant with decreased fractional clearance of AAP via bioactivation and less in vivo AAP covalent binding. These data support the hypothesis that OTP provides a protecting effect from AAP hepatotoxicity due to an augmented and predisposing glucuronidation capacity.