Catalase-dependent ethanol metabolism in vivo in deermice lacking alcohol dehydrogenase

2-Nitroimidazoles, such as pimonidazole, are reduced in cells with low oxygen tension and are, therefore, used as hypoxia markers. However, the effect of the pyridine nucleotide redox state on pimonidazole reduction is not known. Therefore, livers from fed or fasted rats were perfused with oxygen-saturated buffer containing pimonidazole (400 µM) in the presence and absence of an inhibitor of the mitochondrial respiratory chain, potassium cyanide ; these treatments were used to modulate the mitochondrial and cytosolic pyridine nucleotide redox states. Pimonidazole-induced increases in oxygen uptake over basal values were as follows: fed, 15.1Ϯ 2.4 ; fasted, 4.2 Ϯ 0.8; fedϩ KCN, 32.1Ϯ0.9 ; fastedϩKCN, 0.2 Ϯ 0.2 µmol · g Ϫ1 · h Ϫ1 . However, if NADPH was added in excess, microsomal oxygen uptake due to oxidative metabolism of pimonidazole was independent of treatment. These results indicate that pimonidazole-stimulated O 2 uptake, due predominantly to N-oxidation and glucuronidation, is dependent on the NADPH redox state. In contrast, reduced pimonidazole adducts, detected immunochemically, accumulated in pericentral regions in liver. Increasing the NADH redox state by inhibiting the mitochondrial respiratory chain with KCN decreased protein-bound pimonidazole adducts. Concomitantly, the average O 2 tension of the liver was increased at least 30%. However, KCN had no effect on total pimonidazole adducts detected by ELISA, although both cytosolic (lactate/pyruvate) and mitochondrial (3-hydroxybutyrate/acetoacetate) NADH redox states were elevated by at least a factor of eight. These results indicate that, unlike oxidative metabolism, the pyridine nucleotide redox state does not determine the rate of reductive metabolism of pimonidazole. Instead, the cellular oxygen tension regulates this process. Therefore, even in cases where the supply of reducing equivalents is increased (e.g., ethanol metabolism), accumulation of the reduced bound product of pimonidazole is oxygen dependent in liver.

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“…8 B). to sacrifice leads to suicide inhibition of catalase [24]. Treatment with aminotriazole also increased O 2 uptake significantly by a…”

Section: Resultsmentioning

confidence: 95%

Reductive metabolism of the hypoxia marker pimonidazole is regulated by oxygen tension independent of the pyridine nucleotide redox state

Arteel

Thurman

Raleigh

1998

European Journal of Biochemistry

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157

125

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“…Handler et al [40] reported that 3-AT (1.5 g/kg, i.p.) rapidly inhibits not only catalase activity but also CYP2E1-dependent aniline hydroxylation.…”

Section: Resultsmentioning

confidence: 99%

Protection against acetaminophen hepatotoxicity by clofibrate pretreatment: Role of catalase induction

Chen

Hennig

Whiteley

et al. 2002

J Biochem & Molecular Tox

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Mice pretreated with the peroxisome proliferator clofibrate (CFB) are highly resistant to acetaminophen (APAP)-induced hepatotoxicity. The objective of the present study was to investigate whether the increase in hepatic catalase activity following CFB pretreatment plays a role in this hepatoprotection. An irreversible inhibitor, 3-amino-1,2,4-triazole (3-AT), was used to modulate catalase activity. Hepatic catalase activity in mice pretreated with CFB (500 mg/kg, i.p., for 10 days) was significantly inhibited by 3-AT (100 or 500 mg/kg, i.p.). In addition, the lower dose of 3-AT (100 mg/kg) had minimal effect on biliary and urinary excretion of APAP metabolites generated from a nontoxic dose, suggesting that APAP metabolism was not modulated by this dose of 3-AT. The mortality rate of corn-oil-pretreated mice challenged with APAP (800 mg/kg, p.o.) was significantly increased by 3-AT (100 mg/kg, i.p.) given 1 h before APAP. As expected, CFB pretreatment conferred full protection against APAP-induced hepatotoxicity. The same 3-AT treatment, however, did not abolish hepatoprotection in CFB-pretreated mice, despite the marked inhibition of hepatic catalase activity. In conclusion, these results indicate that elevated catalase activity in mice exposed to CFB does not appear to mediate the hepatoprotection, suggesting that other cellular defense mechanisms are involved.

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“…These conclusions contrast with conclusions from reports where the catalase inhibitor, aminotriazole, did not decrease rates of ethanol elimination in vivo [52 -541; however, these previous studies did not control for the extent of inhibition of the peroxide-forming activity of catalase by monitoring catalase-H 2 0 2 . When catalase was effectively inhibited by aminotriazole, as demonstrated by a lack of effect of ethanol on catalase-H202 measured continuously in the perfused liver, rates of ethanol elimination were decreased significantly [32]. Because the contribution of catalase-H202 to hepatic ethanol metabolism has previously been underestimated, due to submaximal rates of H 2 0 2 generation, more work will be needed in the future to quantify the role of catalase-H202 in ethanol oxidation under physiological conditions in the presence of fatty acids.…”

Section: E Lmentioning

confidence: 99%

“…The results indicate that H 2 0 2 is indeed generated from fatty acid metabolism at rates sufficient to support high rates of ethanol oxidation via catalase-H202. Preliminary accounts of this work have been presented elsewhere [18,191.…”

mentioning

confidence: 99%

Catalase‐dependent ethanol oxidation in perfused rat liver

Handler

Thurman

1988

European Journal of Biochemistry

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The purpose of this study was to measure rates of catalase-dependent ethanol uptake and rates of HzOz generation in perfused rat livers in the presence of fatty acids of varying chain length. Rates of ethanol uptake in livers from fasted rats, perfused in a recirculating system, of about 80 pmol g-' h-' were decreased to about 10 pmol g-' h-' by the addition of an inhibitor of alcohol dehydrogenase (ADH), 4-methylpyrazole. The medium-chain-length fatty acid, laurate (12 : 0; 1 mM), increased rates of 4-methylpyrazole-insensitive ethanol uptake maximally to [80][81][82][83][84][85] Oleate did not increase rates of hydroxylation of p-nitrophenol, a substrate for the ethanol-inducible form of cytochrome P-450, indicating that the stimulation of ethanol uptake by fatty acids was not due to increased mixed-function oxidation. The increase of ethanol uptake was also not due to displacement of 4-methylpyrazole from ADH by fatty acids, since oleate stimulated ethanol uptake by about 50% in perfused livers from deermice genetically deficient in ADH. The increase in 4-methylpyrazole-insensitive ethanol uptake by fatty acids was blocked by the catalase inhibitor, aminotriazole, indicating the involvement of catalase. Rates of H z 0 2 generation by livers perfused in a non-recirculating system with 1.7% albumin were increased from 6 f 1 to 23 f. 5 pmol g-' h-' by oleate (1 mM). Because of the discrepancy between rates of ethanol metabolism and HzOz production, methods were developed to measure H z 0 2 production in a recirculating perfusion system. H 2 0 2 generation was determined from the time necessary for steady-state level of catalase-H20z, measured spectrophotometrically (660-640 nm) through a lobe of the liver, to return to basal values after the addition of a known quantity of methanol, which is not metabolized by ADH in the rat. Basal rates of H 2 0 2 generation measued under these conditions were 12 f 5 pmol g-' h-', values similar to basal rates of 4-methylpyrazole-insensitive ethanol uptake.Rates of HzOz generation were stimulated maximally by laurate to about 80 -85 pmol g-' h-' , and decreased as the fatty acid chain length was decreased or increased, analogous to the stimulation of 4-methylpyrazoleinsensitive ethanol uptake by fatty acids. There was an excellent correlation (r = 0.99) between rates of HzOz generation and rates of 4-methylpyrazole-insensitive ethanol uptake with all fatty acids studied. Thus, increases in H 2 0 2 generation and ethanol oxidation have similar chain-length specificities for fatty acids. Rates of H 2 0 2 generation from oleate of about 60 pmol g-' h-' were decreased 50% by an inhibitor of peroxisomal p-oxidation, thioridazine (0.02 mM). Rates were also decreased by 60% by decreasing the albumin concentration in the recirculating system from 4.0% to 1.7%. Rates of ketone body production from oleate were half-maximal at about 0.13 mM oleate; maximal rates were about 90 pmol g-' h-'. In contrast, rates of H 2 0 2 production required higher concentrations of fatty acids (e.g., half-...

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Catalase-dependent ethanol metabolism in vivo in deermice lacking alcohol dehydrogenase

Cited by 39 publications

References 22 publications

Reductive metabolism of the hypoxia marker pimonidazole is regulated by oxygen tension independent of the pyridine nucleotide redox state

Reductive metabolism of the hypoxia marker pimonidazole is regulated by oxygen tension independent of the pyridine nucleotide redox state

Protection against acetaminophen hepatotoxicity by clofibrate pretreatment: Role of catalase induction

Catalase‐dependent ethanol oxidation in perfused rat liver

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