Feed intake and milk production in dairy cows fed whole fat soybeans]

Although the effect of hyperoxia on antioxidant enzymes is well known, the effect of subtoxic levels of hyperoxia on gamma-glutamyltransferase (gamma-GT), involved in the degradation and uptake of extracellular GSH for intracellular GSH synthesis, is unknown. The aim of the study was to investigate (1) the effects of in vitro hyperoxia on gamma-GT activity of type II cells and (2) the effects of the lazaroid U-74389G and N-acetylcysteine (NAC) on the hyperoxia-induced changes in gamma-GT and antioxidant enzyme activities. At 48 h after isolation, rat type II cells were exposed for 2 days to air, 60% O2 or 85% O2 with or without 30 microM U-74389G or 100 microM NAC. After the exposure, the cells were harvested and assayed for superoxide dismutase (SOD), glutathione peroxidase (GPx), gamma-GT activity, and GSH levels. In another series of experiments 85% O2-exposed cells, with or without U-74389G, were used for Northern blotting of gamma-GT mRNA. Exposure to 60% O2 decreased gamma-GT and GSH by -47 and -34%, respectively, while SOD and GPx activities remained unchanged. After 85% O2-exposure gamma-GT decreased by -55%, SOD and GPx increased by +55 and +87%, respectively, while GSH decreased by -35%. NAC treatment decreased gamma-GT activity by -42% in the air-exposed cells. After 60% O2, U-74389G led to significantly higher gamma-GT (+117%) and GSH (+26%) while NAC only led to higher GSH (+28%) compared to the oxygen-exposed cells not treated with NAC or U-74389G. After 85% O2 U-74389G increased gamma-GT, SOD, and GSH by +72, +58, and +68%, respectively, while NAC only increased SOD (+49%) and GSH (+26%) compared to the oxygen-exposed cells not treated with NAC or U-74389G. The 85% O2 exposure, with or without U-74389G, had no effect on gamma-GT mRNA levels. The results show that hyperoxia decreases rat type II cell gamma-GT activity in vitro. This effect was not related to an altered regulation at mRNA level and it was not associated with the hyperoxia-induced decrease in intracellular GSH, since restoration of the GSH levels by NAC did not restore gamma-GT activity. The lazaroid U-74389G with vitamin E-like properties effectively prevented the decrease in gamma-GT and GSH, so that direct inactivation of the membrane-bound gamma-GT by hyperoxia is the most likely mechanism.     

Multi-center European evaluation of HIV testing on serum and saliva samples

Sulphur dioxide (SO2) is an air pollutant implicated in the initiation of asthmatic symptoms. Glutathione (GSH) has been proposed to play a role in detoxification of SO2 through the sulfitolysis of glutathione disulphide (GSSG) to S-sulphoglutathione (GSSO3-). Rats were exposed to concentrations of SO2 between 5 and 100 ppm for 5 hr a day between 7 and 28 days. Lung injury as assessed by bronchoalveolar lavage and tissue GSH status were evaluated. SO2 5 ppm failed to elicit any lung injury or inflammatory response but did deplete GSH pools in lung, liver, heart and kidney. Activities of gamma-glutamylcysteine synthetase (GCS), glutathione peroxidase (GPx), glutathione S-transferase (GST) and glutathione reductase (GRed) in lung were lowered relative to those in control animals. In liver, GRed activity was decreased. SO2 50 ppm exposure also failed to elicit injury or inflammation but did lower inflammatory cell numbers in the circulation. Rats exposed to 50 ppm SO2 maintained tissue GSH status, but activities of GCS, GPx, GRed and gamma-glutamyltranspeptidase in lung and hepatic GRed and GPx were significantly lower than in control rats. Unaltered GST activity in lung and liver was suggestive of an impairment of the sulfitolysis reaction in these animals, perhaps through lower substrate flux through the GPx reaction, as GSSO3- is a known inhibitor of GST in the rat. Rats exposed to 100 ppm SO2 exhibited evidence of inflammation (120-fold increase in neutrophil numbers recovered in lavage fluid) and like the 5 ppm exposed rats had lower tissue GSH concentrations and GSH-related enzyme activities in lung. We conclude that sulfitolysis of GSSG does occur in vivo during SO2 exposure and that SO2, even in the absence of pulmonary injury, is a potent glutathione depleting agent.     

Lipid peroxides, glutathione, gamma-glutamylcysteine synthetase and gamma-glutamyltranspeptidase activities in several tissues of rats following water-immersion stress

The effects of water-immersion restraint (WIR) stress on lipid peroxide, glutathione (GSH), glutathione peroxidase (GSH-Px), gamma-glutamylcysteine synthetase (gamma-GCS) and gamma-glutamyltranspeptidase (gamma-GT) activities in several tissues of rats were investigated. Hepatic and intestinal lipid peroxide levels were increased significantly in the WIR stress group. In both tissues, GSH levels were significantly decreased and gamma-GCS activity was significantly increased. In addition, gamma-GT activities remained unchanged in both tissues following WIR stress. However, lipid peroxide and GSH levels did not change in the stomach and brain in the WIR stress group compared to the control group. These results suggest that lipid peroxidation, but not the depression of GSH synthesis and/or the increase of GSH breakdown may be a factor in hepatic and intestinal GSH reduction following WIR stress.     

Effect of co-planar polychlorinated biphenyl on the hepatic glutathione peroxidase redox system in rats and guinea pigs

The effect of 3,3',4,4',5-pentachlorobiphenyl (PCB 126) on hepatic glutathione peroxidase (GPx) redox system was studied in vivo in rats and guinea pigs. PCB 126 treatment caused significant reduction of Se-dependent and -non-dependent GPx activity in rats. In agreement with this, the content of glutathione (GSH) and the activities of GSH reductase (GR) and gamma-glutamyl transpeptidase (gamma-GTP) were also decreased in this species. On the contrary, guinea pig liver Se-non-dependent GPx activity was significantly enhanced by PCB 126 treatment, while no effect on Se-dependent activity was observed. Neither the content of GSH nor the enzyme activities responsible for GSH supply in guinea pig liver was affected by PCB 126. These result suggested that the damage on GPx redox system is, at least, one of mechanisms by which co-planar PCB induces the toxicity in rats. However, in guinea pigs, this is not the case, and different mechanism from the damage on active oxygen quenching system is likely to be involved.     

Exon-intron structure of the fet5+ gene of Schizosaccharomyces pombe and physical mapping of genome encompassing regions]

Groups of young male adult guinea pigs were fed a diet devoid in supplemental ascorbic acid (AA) or the same diet supplemented with 0.1 or 2.5% AA for four weeks. The animals were then euthanized and Phase I and Phase II drug metabolizing components in the liver were determined. Phase I components are those related to the metabolism of xenobiotics and include microsomal cytochrome P-450 and mixed function oxygenase activities. Phase II components are those related to conjugation and detoxification reactions of xenobiotics and their metabolites and include glutathione-S-transferases (GST), glutathione (GSH), UDP-glucuronyl transferase (UDP-GT) and DT-diaphorase (quinone reductase, QR). Tissue levels of AA increased progressively with increase in AA intake. The Phase I components increased in response to increased intake of AA from 0 to 0.1%, but were unaffected by further increase in AA intake to 2.5%. However, the Phase II components increased with increased intake of AA except for GST. In vitro metabolism of aflatoxin B1 (AFB1) using liver microsomes showed tendency towards increased production of aflatoxin M1 (AFM1) with increase in AA intake. The production of aflatoxin P1 (AFP1) was not affected by AA intake. AFB1-DNA production was increased when AA intake was increased to 0.1%. It was however lowered with further increase in AA intake to 2.5%.     

Postnatal effect of smokeless tobacco on phytic acid or the butylated hydroxyanisole-modulated hepatic detoxication system and antioxidant defense mechanism in suckling neonates and lactating mice

The present study evaluates the potential of smokeless tobacco to translactationally modify the chemopreventive efficacy of phytic acid and butylated hydroxyanisole (BHA) via modulation of the hepatic xenobiotic detoxication system and antioxidant defense mechanism in the murine system. Phytic acid (1000 mg/kg b.w./day) by gavage while BHA (1% w/w) in diet induced a significant increase in the levels of glutathione-S-transferase (GST), acid soluble sulfhydryl (-SH), cytochrome b5 (Cyt. b5) and cytochrome P-450 (Cyt. P-450) in lactating dams and suckling pups. The hepatic levels of GST and -SH were significantly depressed whereas microsomal Cyt. b5, Cyt. P-450 and MDA levels were elevated in groups treated with smokeless tobacco (50 or 100 mg/kg b.w./day). The data reveals the inhibitory potential of smokeless tobacco on phytic acid-induced GST/GSH system efficiency besides the significant augmentation by smokeless tobacco on phytic acid or BHA-induced microsomal phase I enzymes. The direct or translactational modulation in the levels of xenobiotic detoxication system enzymes suggests the potential of smokeless tobacco to modify the chemopreventive efficacy of phytic acid or BHA.     

Identification of the mechanical impedance at the human finger tip

The present study evaluates the potential of smokeless tobacco to modify the chemopreventive efficacy of minor dietary constituents, including garlic, mace or black mustard, via modulating the competing pathways of hepatic detoxication system and antioxidant defense mechanism in murine system. Garlic (100 mg/kg b.w. per day) by gavage and mace (1% w/w) or black mustard (1% w/w) in diet induced a significant increase in the levels of glutathione-S-transferase (GST), acid-soluble sulfhydryl (-SH), cytochrome b5 (Cyt.b5) and cytochrome P-450 (Cyt.P-450) in murine liver. The hepatic levels of GST and -SH were significantly depressed whereas microsomal Cyt.b5, Cyt.P-450 and MDA levels were elevated in groups treated with smokeless tobacco (50 or 100 mg/kg b.w. per day). The data revealed the inhibitory potential of smokeless tobacco on garlic-induced hepatic GST/GSH system besides the significant augmentation by smokeless tobacco on garlic or mace or black mustard-induced microsomal cytochromes. The possible implications of modulation in competing bioactivation and detoxication pathways in the process of chemical carcinogenesis are discussed.     

The mouse estrogen receptor-related orphan receptor alpha 1: molecular cloning and estrogen responsiveness

We observed that glutathione (GSH) status regulates the Ah receptor inducible cytochrome P4501A (CYP1A) gene expression and catalytic activity in 3,3',4,4'-tetrachlorobiphenyl (TCB) exposed rainbow trout. Tissue GSH status of TCB (1 mg/kg body weight, in corn oil) injected fish was manipulated by a) injecting (i.p.) GSH (0.25 g/kg), b) arresting GSH synthesis by L-buthionine-[S,R]-sulfoximine (BSO; 6 mmol/kg) injection for 3 and 6 days. Our attempt to manipulate GSH levels by lipoate supplementation (16 mg/kg) was not productive. Both BSO- and lipoate-supplemented fish maintained a low tissue redox (GSSG/GSH) ratio. Activities of glutathione peroxidase and glutathione reductase were elevated following 3 days of GSH supplementation in GSH rich tissues. Low activities of these enzymes were observed in BSO treated GSH deficient tissues. TCB injection markedly induced hepatic and renal CYP1A catalytic (ethoxyresorufin O-deethylase [EROD]) activities. This effect was further potentiated (3-fold) in GSH-supplemented fish tissues. In contrast, EROD induction by TCB was markedly suppressed in GSH deficient (BSO-treated) and lipoate-supplemented fish. The suppression of CYP1A catalytic activities in GSH deficient and lipoate-supplemented fish was consistently associated with a suppression of TCB induced CYP1A mRNA and protein expressions in these groups. In glutathione-supplemented fish, TCB induced CYP1A protein expression was markedly higher following 3 days of GSH supplementation. Results of our study suggest that tissue thiol status modulates cytochrome P450 CYP1A gene expression and catalytic activity.     

The effect of 1,2,3,4-tetrachlorodibenzo-p-dioxin on drug-metabolizing enzymes in the rat liver

The effects of 1,2,3,4-tetrachlorodibenzo-p-dioxin (1,2,3,4-TCDD) on drug-metabolizing enzymes were studied in male and female rats. 1,2,3,4-TCDD (25, 50, 100 and 200 mumol/kg) was administered by i.p. injection once. Among the cytochrome P-450 (P450)-mediated monooxygenase activities tested, 7-ethoxyresorufin O-deethylase (EROD) activities in both male and female rats, which are associated with CYP1A1, were remarkably induced by all doses of 1,2,3,4-TCDD. The relative induction to each control activity were from 3.0- to 24.5-fold and from 2.2- to 16.5-fold, respectively. Also, 1,2,3,4-TCDD increased other CYP1A-mediated monooxygenase activities such as 7-ethoxycoumarin O-deethylase (ECOD) and 7-methoxyresorufin O-demethylase (MROD) in male and female rats dose-dependently (1.4- to 4.3-fold). Western immunoblotting showed that the levels of CYP1A1 and CYP1A2 proteins in liver microsomes were increased by 1,2,3,4-TCDD. Although the activities of other P450-mediated monooxygenases, namely 7-pentoxyresorufin O-depentylase (PROD), 7-benzyloxyresorufin O-debenzylase (BROD), aminopyrine N-demethylase (APND) and nitrosodimethylamine N-demethylase (NDAND) in both male and female rats were induced at high doses (> or = 50 mumol/kg) of 1,2,3,4-TCDD, the relative level was low compared with those of the CYP1A-mediated monooxygenase such as EROD, ECOD or MROD. In addition to P450-mediated monooxygenase, there was significant induction in the activities of the Phase II drug-metabolizing enzymes, UDP-glucuronyltransferase (UGT) activities towards 4-nitrophenol (4-NP) and 7-hydroxycoumarin (7-HC) and glutathione S-transferase (GST) towards 1-chloro-2,4-dinitrobenzene (CDNB), 1,2-dichloro-4-nitrobenzene (DCNB) and DT-diaphorase. These results indicate that 1,2,3,4-TCDD induces both Phase I (CYP1A-mediated monooxygenase) and Phase II drug-metabolizing enzymes (UGT, GST, DT-diaphorase) in the male and female rat liver, and that the alterations of drug-metabolizing enzyme are characteristic of PCDD toxicity.     

Effects of singly administered betaine on hepatotoxicity of chloroform in mice

Effects of a single dose of betaine on the chloroform-induced hepatotoxicity were examined in adult male ICR mice. Administration of betaine (1000 mg/kg, ip) 1 to 7 hr prior to a chloroform challenge (0.25 ml/kg, ip) resulted in remarkable enhancement of hepatotoxicity as indicated by increases in serum sorbitol dehydrogenase (SDH), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities. The potentiation of hepatotoxicity was most significant when mice were treated with betaine 4 hr earlier than chloroform. However, a 24 hr prior administration of betaine protected the animals from induction of the chloroform hepatotoxicity. Thus, its effect appeared to be highly dependent on the time lapse from the betaine pretreatment to the challenge of mice with chloroform. Betaine treated either 4 or 24 hr prior to sacrifice did not alter the hepatic contents of cytochrome P-450, cytochrome b5, or NADPH cytochrome P-450 reductase activity. Accordingly the hepatic microsomal p-nitroanisole O-demethylase, aminopyrine N-demethylase, or p-nitrophenol hydroxylase activities were not influenced by the betaine pretreatment. Betaine was shown not to affect any of the enzyme activities associated with glutathione (GSH) conjugation reaction, such as glutathione S-transferases (GSTs), glutathione disulfide (GSSG) reductase and GSH peroxidase irrespective of the time of its administration. When betaine was administered to mice 2-6 hr prior to sacrifice, hepatic GSH level, but not plasma GSH, was decreased significantly. Enhancement of the chloroform hepatotoxicity by betaine correlated well with the reduction in hepatic GSH levels. Both hepatic and plasma GSH levels were elevated in mice 24 hr following the betaine treatment. The results suggest that betaine affects induction of the chloroform hepatotoxicity by modulating the availability of hepatic GSH, which appears to be associated with its role in the transsulfuration pathway in the liver.     

Effects of peroxisome proliferators and/or hypothyroidism on xenobiotic-metabolizing enzymes in rat testis

The objectives of the present work were to study the effects of certain peroxisome proliferators on xenobiotic-metabolizing enzyme activities in the testes of normal and hypothyroid rats, i.e. phenol sulfotransferases (pST), phenol UDP-glucuronosyl transferases (pUDPGT), glutathione transferases (GST), catalase, epoxide hydrolase (EH), glutathione peroxidase (GPX) and NAD(P)H quinone oxidoreductase (QR). Adult male rats (normal and hypothyroid) were treated for 10 days with clofibrate (0.5%), perfluorooctanoic acid (0.5%, PFOA), acetylsalisylic acid (1%, ASA) and di(2-ethylhexyl)phthalate (2%, DEHP) in their diet. The results show that treatment of normal rats with peroxisome proliferators dramatically affects the activities of xenobiotic-metabolizing enzymes (40-60% reduction). The highest effects are seen in catalase activity (50-60% with PFOA and ASA), pUDPGT (55% with PFOA), pST (55% with PFOA) and QR (50% with DEHP). These effects are not seen or are weaker after induction of hypothyroidism. Taken together, it is concluded that different classes of peroxisome proliferators have different effects on rat testicular xenobiotic-metabolizing enzymes.     

Effect of dietary oils, cholesterol and antioxidant vitamin supplementation on liver microsomal fluidity and xenobiotic-metabolizing enzymes in rats

This study was undertaken to compare the effects of four oils: corn (C), olive (O), hazelnut (H) or fish (F), and the intake of two supplements: cholesterol, 1% (Ch) or dl-alpha-tocopherol acetate, 500 mg/kg, and beta-carotene, 30 mg/kg (V), on liver microsomal fluidity, cyt P450 content and aniline hydroxylase (AH), aminopyrine-N-dimethylase (AND) and UDP-glucuronyltransferase (UDP-GT) activities. Male Sprague-Dawley rats (n = 6/group) were fed semipurified diets containing 15% oil, without or with Ch or V, for 20 days. Dietary intake and feed efficiency were lower in rats fed F. Relative liver weight was higher in animals fed F, similar in O and H, and lower in the group fed C. The intake of V increased feed intake in C+V group and decreased the relative liver weight of F+V group, which also decreased with the intake of F+Ch. Ch intake increased the relative liver weight in all groups consuming vegetable oils. Cyt P450 content was higher in rats fed F. Decreased cyt P450 content was observed in C+Ch and F+Ch groups, while it augmented in H+Ch group. Mixture V increased cyt P450 in rats fed C+V, F+V and O+V. The highest membrane fluidity was observed in rats fed F. Fluidity was also higher in group H versus O or C. The intake of Ch decreased microsomal fluidity in all groups, while V induced an increase in microsomal fluidity in group O+V. Rats fed F exhibited higher enzyme activities. AND activity increased with V only in rats fed H+V, while AH activity increased with V intake in groups F+V and O+V. In the C+V group, fluidity was not affected by V, while the cyt P450 content and UDP-GT activity increased. The O+V group exhibited lower UDP-GT activity and higher fluidity and cyt P450 content. The activity of AH decreased in groups F+Ch and C+Ch. UDP-GT activity was higher in rats fed F. It diminished after the intake of Ch in H+Ch and F+Ch. These results indicate that although AH and AND act in the same microsomal metabolic pathway, their localization into the membrane may be determinant of their activity and the response to dietary lipids. It is shown that F intake exerts the most significant effects upon liver microsomal properties, e.g. higher fluidity, cyt P450 content and enzymatic activities, an effect that prevails over the intake of the supplements tested.     

Mechanism of inhibition of benzo[a]pyrene-induced forestomach cancer in mice by dietary curcumin

Curcumin (diferuloylmethane), the major yellow pigment in turmeric, has been shown to inhibit benzo[a]pyrene (BaP)-induced forestomach cancer in mice through mechanism(s) not fully understood. It is well known that while cytochrome P4501A1 (CYP1A1) and epoxide hydrolase (EH) are important in the conversion of BaP to its activated form, (+)-anti-7,8-dihydroxy-9,10-oxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(+)-anti-BaPDE], the detoxification of (+)-anti-BaPDE is accomplished by glutathione (GSH) S-transferases (GST). Therefore, it seems reasonable to postulate that curcumin may exert anti-carcinogenic activity either by inhibiting activation of BaP or (and) by enhancing the detoxification of (+)-anti-BaPDE. Administration p.o. of 2% curcumin in the diet to female A/J mice for 14 days, which has been shown to cause a significant inhibition in BaP-induced forestomach tumorigenesis, resulted in a modest but statistically significant reduction in hepatic ethoxyresorufin O-deethylase (EROD) activity, a reaction preferentially catalyzed by CYP1A1. While EROD activity could not be detected in the forestomach of either control or treated mice, curcumin feeding caused a statistically significant increase (approximately 2.3-fold) in hepatic EH and GST activities. Hepatic and forestomach GSH levels, and forestomach EH and GST activities were not affected by curcumin treatment. Even though the levels of various hepatic GST isoenzymes were significantly increased upon curcumin feeding, maximum induction was noticed for the pi class isoenzyme (mGSTP1-1), which among murine hepatic GSTs is highly efficient in the detoxification of (+)-anti-BaPDE. In conclusion, the results of the present study suggest that curcumin may inhibit BaP-induced forestomach cancer in mice by affecting both activation as well as inactivation pathways of BaP metabolism in the liver.     

Changes in GSH-antioxidant system induced by daunorubicin in human normal and diabetic fibroblasts

We investigated the effect of daunorubicin on glutathione content and activity of GSH-related enzymes in cultured normal and diabetic human fibroblasts. Cells were incubated with 4 microM daunorubicin (DNR) for 2 h followed by culture in drug-free medium for up to 72 h. Treatment of diabetic cells with the drug caused a time-dependent depletion of intracellular GSH and a decrease of the GSH to total glutathione ratio. GSH depletion was accompanied by apoptotic changes in morphology of the nucleus. Analysis of GSH-related enzymes showed a significant increase of the activities of Se-dependent and Se-independent peroxidases and glutathione S-transferase. In contrast, glutathione reductase activity was reduced by 50%. Significant differences between normal and diabetic cells exposed to DNR were observed in the level of GST and Se-dependent glutathione peroxidase activities. These findings indicated that daunorubicin efficiently affects the GSH antioxidant defense system both in normal and diabetic fibroblasts leading to disturbances in glutathione content as well as in the activity of GSH-related enzymes.     

Risk factors of severe hypoglycaemia in adult patients with Type I diabetes--a prospective population based study

Novel thiazolidine prodrugs were prepared by the condensation of L-cysteine with aldose disaccharides. Using a disaccharide in prodrug construction allows for a terminal cyclic sugar moiety to be present on the prodrug, which may allow the delivery of the agent to specific receptors, such as the asialoglycoprotein receptor (ASGPR) of hepatocytes, that require specific structural motifs for recognition. Three L-cysteine prodrugs were synthesized with a pendant cyclic galactose moiety; two related glucose-bearing prodrugs were synthesized for comparison. The prodrugs were designed to release L-cysteine, which is then available to support glutathione (GSH) biosynthesis and provide cytoprotection against a variety of toxic insults. Protection studies in Swiss-Webster mice used acetaminophen (575 mg/kg), a well-documented hepatotoxin which depletes GSH at overdose. Three prodrugs performed exceptionally well against acetaminophen-induced hepatotoxicity, as measured by increased survival and improved histological profiles of liver tissue after 48 h. In further experimentation, two of the disaccharide-based prodrugs, prepared from alpha- and beta-lactose, were compared with the monosaccharide-based compound prepared from ribose. Co-administration of the selected prodrugs with a 400 mg/kg dose of acetaminophen to Swiss-Webster mice prevented the short-term depletion in hepatic GSH and also reduced hepatotoxicity as determined by histological damage and serum levels of alanine aminotransferase. A single dose of the prodrugs alone had no effect on hepatic drug metabolizing enzymes [glutathione S-transferase (GST), NAD(P)H:quinone oxidoreductase (QOR), UDP-glucuronosyltransferase (UGT), and cytochrome P450], but, concordant with the reduction of hepatotoxicity, the latentiated forms prevented the significant elevation in QOR activity and mRNA and GST mRNA elicited by acetaminophen itself. GST activity, UGT activity and mRNA, and cytochrome P450 concentration were all unaffected by acetaminophen or the prodrugs. These studies identified novel L-cysteine prodrugs with potentially useful hepatoprotective activity. However, no structure-activity relationships were obvious. In addition, the occurrence of targeted delivery to hepatocytes remains ambiguous.     

Induced abortion as an independent risk factor for breast cancer: a comprehensive review and meta-analysis

Effects of continuous feeding flavonoids (flavone, flavanone, and tangeretin) on drug-metabolizing enzymes in rat liver were investigated to ascertain how long feeding is required to reach maximal induction and to determine whether maximal induction is maintained for a long period of feeding. In the first experiment rats received a diet containing 10 mmol flavonoid/kg dry matter for 4, 8, 16, or 32 d. The second experiment was designed to examine the time course for induction during the first 4 d. The kinetics of induction depended on the chemical structure of the flavonoid and was different from one enzyme to another. Flavone increased P450 1A and P450 2B apoproteins and stimulated many enzyme activities. A significant increase of P450 1A1/2 proteins, ethoxyresorufin O-deethylase (EROD), and methoxyresorufin O-demethylase (MROD) activities occurred as early as 6 h after the first administration, and a gradual increase was observed up to 4 d of feeding. P450 2B1/2 proteins and pentoxyresorufin O-depentylase (PROD) activity were also increased but after a lag period when compared with P450 1A1/2 proteins. EROD and MROD activities declined after 4 d, whereas PROD activity remained steady during 32 d of flavone feeding. Glutathione transferase (GST) and p-nitrophenol UDP-glucuronosyl transferase (UGT) activities were also increased. The maximal induction was reached by 4 d of feeding for UGT and after a longer duration of feeding (16 d) for GST. Flavanone treatment induced mostly P450 2B1/2 proteins and PROD, GST, and UGT activites. After 4 d of feeding, P450 2B1/2 proteins and PROD activity declined whereas GST and UGT activities remained steady. Tangeretin treatment produced changes similar to flavone but of lesser magnitude and after a longer delay.     

Cytotoxicity and cell-proliferation induced by the nephrocarcinogen hydroquinone and its nephrotoxic metabolite 2,3,5-(tris-glutathion-S-yl)hydroquinone

Hydroquinone, an intermediate used in the chemical industry and a metabolite of benzene, is a nephrocarcinogen in the 2-year National Toxicology Program bioassay in male Fischer 344 rats. Current evidence suggests that certain chemicals may induce carcinogenesis by a mechanism involving cytotoxicity, followed by sustained regenerative hyperplasia and ultimately tumor formation. Glutathione (GSH) conjugates of a variety of hydroquinones are potent nephrotoxicants, and we now report on the effect of hydroquinone and 2,3,5-(tris-glutathion-S-yl)hydroquinone, on site-selective cytotoxicity and cell proliferation in rat kidney. Male Fischer 344 rats (160-200 g) were treated with hydroquinone (1.8 mmol/kg or 4.5 mmol/kg, p.o.) or 2,3,5-(tris-glutathion-S-yl)hydroquinone (7.5 micromol/kg; 1.2-1.5 micromol/rat, i.v.), and blood urea nitrogen (BUN), urinary gamma-glutamyl transpeptidase (gamma-GT), alkaline phosphatase (ALP), glutathione-S-transferase (GST) and glucose were measured as indices of nephrotoxicity. Hydroquinone (1.8 mmol/kg, p.o.) is nephrotoxic in some rats, but not others, but cell proliferation (BrDU incorporation) in proximal tubular cells of the S3M region correlates with the degree of toxicity in individual rats. At 4.5 mmol/kg, hydroquinone causes significant increases in the urinary excretion of gamma-GT, ALP and GST. Pretreatment of rats with acivicin prevents hydroquinone-mediated nephrotoxicity, indicating that toxicity is dependent on the formation of metabolites that require processing by gamma-GT. Consistent with this view, 2,3,5-(tris-glutathion-S-yl)hydroquinone, a metabolite of hydroquinone, causes increases in BUN, urinary gamma-GT and ALP, all of which are maximal 12 h after administration of 2,3,5-(tris-glutathion-S-yl)hydroquinone. In contrast, the maximal excretion of GST and glucose occurs after 24 h. By 72 h, BUN and glucose concentrations return to control levels, while gamma-GT, ALP and GST remain slightly elevated. Examination of kidney slices by light microscopy revealed the presence of tubular necrosis in the S3M segment of the proximal tubule, extending into the medullary rays. Cell proliferation rates in this region were 2.4, 6.9, 15.3 and 14.3% after 12, 24, 48 and 72 h, respectively, compared to 0.8-2.4% in vehicle controls. Together with the metabolic data, the results indicate a role for hydroquinone-thioether metabolites in hydroquinone toxicity and carcinogenicity.     

Effects of auditory stimuli on intracranial pressure and cerebral perfusion pressure in traumatic brain injury

In the kidney, ischaemia-reperfusion results in both hypoxic and oxidant cellular injury which is most marked in the tubules of cortex and outer medulla. These contrasting conditions may have opposite effects on the expression of enzymes that reduce or repair oxidant damage. To investigate this, the activities of CuZn and Mn superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione S-transferase (GST) were measured after 4 h and 3, 6, and 10 days of reperfusion following sham surgery or 45- or 90-min left renal artery occlusion. The right kidney served as internal control. Sham surgery had no effect on Mn SOD or GPx, but caused small (p < 0.05) reductions in CuZn SOD and GST activities. Forty-five minutes of ischaemia had no net effect on Mn SOD, increased GPx activity (maximum at 6 days, p < 0.01), and reduced CuZn SOD (nadir 3 days, p < 0.02) and GST (nadir 6 days, p < 0.02) activities. Ninety minutes of ischaemia again had no net effect on Mn SOD, prevented the induction of GPx, and further suppressed the activities of CuZn SOD and GST. The activity of the non-anti-oxidant enzyme lactate dehydrogenase was equal in left and right kidneys after 45 min of ischaemia, but different (p < 0.01) 10 days following 90-min injury, due to a combination of reduced activity in the ischaemic kidney and an increase of activity in the internal control. The immediate effect of ischaemia-reperfusion injury on the kidney is to reduce the activity of intracellular anti-oxidant enzymes in proportion to the severity of the ischaemic insult. Recovery or net induction of enzyme activity paralleled tubular regeneration. Protection resulting in acquired resistance to a second ischaemic event is unlikely to be due to induction of anti-oxidant enzymes if it occurs within 6 days.     

Effect of P-450 inducers on biliary excretion of glutathione and its hydrolysis products. Correlation between hepatic gamma-glutamyltranspeptidase activity and the proportion of glutathione hydrolysis products in bile

This study was designed to determine if a relationship exists between hepatic gamma-glutamyltranspeptidase (gamma-GT) activity and the biliary excretion of glutathione (GSH) and its hydrolysis products. Rats were pretreated with the following microsomal enzyme inducers: pregnenolone-16 alpha-carbonitrile (PCN), dexamethasone (DEX), 3-methylcholanthrene, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), phenobarbital (PB), ethanol (ETOH), trans-stilbene oxide (TSO), butylated hydroxyanisole (BHA), isosafrole (ISF), clofibrate, and benzo(a)pyrene. Hepatic gamma-GT activity was quantitated spectrophotometrically; bile and liver samples were analyzed by HPLC for reduced and oxidized GSH and their hydrolysis products (cysteine, cysteinylglycine, and cysteinylglycine disulfide). Administration of the inducers had only minor effects on hepatic GSH concentration, as BHA was the only agent to increase GSH concentration. However, these inducers had a pronounced effect on the biliary excretion of total thiol-derived sulfur as PCN, PB, and ISF produced an increase, whereas TCDD, ETOH, and TSO caused a decrease. The relative amount of the GSH hydrolysis products in bile was highly dependent on gamma-GT activity. For example, hepatic gamma-GT activity was increased by PCN, DEX, BHA, TSO, and ISF. They also increased the GSH hydrolysis products to total thiol-derived sulfur ratio in bile. In conclusion, the ratio of GSH hydrolysis products to total thiol-derived sulfur excreted in rat bile reflects the hepatic gamma-GT activity.     

Effects of the agrochemicals butachlor, pretilachlor and isoprothiolane on rat liver xenobiotic-metabolizing enzymes

1. The herbicides butachlor (2-chloro-2',6',diethyl-N-[buthoxymethyl] acetanilide) and pretilachlor (2-chloro-2',6'-diethyl-N-[2-propoxyethyl] acetanilide) are widely used in Asia, South America, Europe and Africa. Isoprothiolane (diisopropyl-1,3-dithiolan-2-ylidenemalonate) is used as a fungicide and an insecticide in rice paddies. We administered these agrochemicals to the male rat and examined their effects on cytochrome P450 (P450), glutathione S-transferase (GST), UDP-glucuronosyltransferase (UDPGT), and NAD(P)H-quinone oxidoreductase 1 (NQO1)-related metabolism in the liver. 2. Administration of isoprothiolane, butachlor or pretilachlor to rat induced hepatic P4502B subfamily-dependent enzyme activities (pentoxyresorufin O-depentylation and testosterone 16 beta-hydroxylation) up to 271-413% of control, which coincided with the increase in expression levels of the P4502B apoprotein. 3. Activities of GST toward 1-chloro-2,4-nitrobenzene and 3,4-dichloronitrobenzene were slightly induced (127-133% of control) in the liver of the rat treated with these pesticides. On the other hand, marked elevations of UDPGT activities toward p-nitrophenol (164-281% of control) were observed. NQO1-related metabolism (menadione reductase activity) was also induced (123-176% of control) in the liver of rat treated with these agrochemicals. 4. These results indicate that some of the agrochemicals currently in use are capable of inducing phase I and II xenobiotic-metabolizing enzyme activities in an isozyme selective manner. The induction of these activities may disrupt normal physiologic functions related to these enzymes in exposed animals.