Acetaminophen and protein thiol modification.
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Acetaminophen↗
Biomedical subjects
M A Tirmenstein
Publications and source records attributed to M A Tirmenstein.
Acetaminophen-induced oxidation of protein thiols. Contribution of impaired thiol-metabolizing enzymes and the breakdown of adenine nucleotides.
The administration of a hepatotoxic dose of acetaminophen (250 mg/kg) to mice induced the loss of protein thiols in mouse liver. Our data suggest that a significant portion of this loss was due to protein thiol oxidation. The administration of the nonhepatotoxic regioisomer, 3'-hydroxyacetanilide (600 mg/kg) did not produce a similar decrease in liver protein thiols despite producing similar levels of covalent binding. Mice treated with acetaminophen exhibited decreased glutathione peroxidase activity, decreased thioltransferase activity, and decreased adenine nucleotide concentrations in the liver. The increase in urinary allantoin after the administration of acetaminophen suggests that the decrease in adenine nucleotides was due to their degradation in the liver. Acetaminophen also promoted the conversion of the enzyme xanthine dehydrogenase to the oxidase form, and pretreatment of mice with allopurinol, an inhibitor of xanthine oxidase, significantly decreased acetaminophen-mediated hepatotoxicity. The conversion of xanthine dehydrogenase to the oxidase form may lead to a transient increase in the production of activated oxygen species. The increase in activated oxygen species coupled with decreases in glutathione peroxidase and thioltransferase activity may be responsible in part for the increased levels of oxidized protein thiols observed following acetaminophen administration.
Acetaminophen↗
Subcellular binding and effects on calcium homeostasis produced by acetaminophen and a nonhepatotoxic regioisomer, 3'-hydroxyacetanilide, in mouse liver.
Acetaminophen (250 mg/kg) administered intraperitoneally to fasted, phenobarbital-induced mice produced hepatotoxicity. No hepatotoxicity was observed after the administration of the regioisomer 3'-hydroxyacetanilide (600 mg/kg). Similar levels of covalent binding to liver homogenates occurred in mice receiving either acetaminophen or 3'-hydroxyacetanilide at these doses. However, subcellular fractionation techniques revealed that the acetaminophen treatment produced greater levels of covalent binding to mitochondrial proteins than 3'-hydroxyacetanilide. In addition, acetaminophen depleted mitochondrial glutathione levels more extensively than 3'-hydroxyacetanilide. Plasma membrane calcium-ATPase activity was reduced to 79.8% and 55.7% of control values at 1 h and 6 h, respectively, following the administration of acetaminophen. No inhibition of this enzyme was detected in mice receiving 3'-hydroxyacetanilide. Acetaminophen also induced alterations in mitochondrial calcium levels and decreased the ability of isolated mitochondria to sequester calcium. These effects were not produced by 3'-hydroxyacetanilide. Our results indicate that acetaminophen induces alterations in calcium homeostasis while 3'-hydroxyacetanilide does not.
Acetaminophen↗
Role of a partially purified glutathione S-transferase from rat liver nuclei in the inhibition of nuclear lipid peroxidation.
Glutathione protects isolated rat liver nuclei against lipid peroxidation by inducing a lag period prior to the onset of peroxidation. This GSH-dependent protection was abolished by exposing isolated nuclei to the glutathione S-transferase inhibitor S-octylglutathione. In incubations containing 0.2 mM S-octylglutathione, the GSH-induced lag period was reduced from 30 to 5 min. S-Octylglutathione (0.2 mM) also completely inhibited nuclear glutathione S-transferase activity and reduced glutathione peroxidase activity by 85%. About 70% of the glutathione S-transferase activity associated with isolated nuclei was solubilized with 0.3% Triton X-100. This solubilized glutathione S-transferase activity was partially purified by utilizing a S-hexylglutathione affinity column. The partially purified nuclear glutathione S-transferase exhibited glutathione peroxidase activity towards lipid hydroperoxides in solution. The data from the present study indicate that a glutathione S-transferase associated with the nucleus may contribute to glutathione-dependent protection of isolated nuclei against lipid peroxidation. Evidence was obtained which indicates that this enzyme is distinct from the microsomal glutathione S-transferase.
Animals↗
The glutathione status of rat kidney nuclei following administration of buthionine sulfoximine.
Rat kidney nuclei were isolated by techniques designed to limit the loss of small water-soluble metabolites. Lyophilized rat kidney powder was disrupted with a Tekmar Tissumizer, and nuclei were purified by discontinuous gradient centrifugation through a glycerol-metrizamide solution at 4 degrees C. Purified nuclei exhibited similar DNA to protein ratios as reported for other rat nuclei isolated by nonaqueous isolation techniques. Results suggest that in control animals the concentration of glutathione in the kidney nucleus is similar to that in the cytoplasm. However, following treatment with buthionine sulfoximine, rat kidney nuclear glutathione levels were less than the corresponding cytoplasmic glutathione levels.
Animals↗
Characterization of glutathione-dependent inhibition of lipid peroxidation of isolated rat liver nuclei.
Glutathione (GSH) is known to play an important role in protecting cells against oxidative stress. The present study was undertaken to assess the ability of GSH to protect isolated rat liver nuclei against NADPH-induced peroxidation. Nuclei were isolated from rat liver homogenates by discontinuous sucrose gradient centrifugation, and lipid peroxidation was induced by 1.7 mM ADP, 0.11 mM EDTA, 0.1 mM FeCl3, and either 1 mM NADPH or 0.5 mM ascorbate. The amount of lipid peroxidation was determined by measuring the formation of thiobarbituric acid-reactive products and the disappearance of lipid unsaturated fatty acid moieties. The addition of GSH (0.1 to 1.0 mM) produced a concentration-dependent lag period prior to the onset of lipid peroxidation. This GSH-induced lag period was abolished by pretreatment of nuclei with trypsin, thiol modifying reagents, disulfides, or heating nuclei at 60 degrees C for 15 min. Nuclei which were incubated with GSH also catalyzed the conversion of cumene hydroperoxide to cumyl alcohol. Similarly, this activity was also inhibited by thiol modifying reagents, disulfides, and heating nuclei at 60 degrees C for 15 min. The data suggest that a GSH-dependent peroxidase activity is associated with rat liver nuclear membranes which are capable of inhibiting lipid peroxidation.
Animals↗
Hepatotoxicity after 3'-hydroxyacetanilide administration to buthionine sulfoximine pretreated mice.
The administration of 3'-hydroxyacetanilide, a regioisomer of acetaminophen, to mice failed to produce hepatotoxicity even after the administration of diethyl maleate. In contrast, hepatotoxicity did occur when 3'-hydroxyacetanilide was administered to buthionine sulfoximine pretreated mice. Although the administration of 3'-hydroxyacetanilide in conjunction with either diethyl maleate or buthionine sulfoximine depleted total hepatic glutathione, only the combined buthionine sulfoximine-3'-hydroxyacetanilide treatment decreased hepatic mitochondrial glutathione concentrations to below 20% of control values. In addition, pretreatment with buthionine sulfoximine increased the amount of 3'-hydroxyacetanilide bound to mitochondrial proteins. These results, in conjunction without previous results on the involvement of mitochondrial damage in the pathogenesis of hepatotoxicity caused by acetaminophen, suggest a probable relationship between mitochondrial damage caused by the buthionine sulfoximine-3'-hydroxyacetanilide treatment and hepatotoxicity caused by this treatment.
Acetaminophen↗