Brain mitochondrial cytochrome oxidase and respiratory activities were compared after in vivo and in vitro exposure to cyanide. For the in vivo studies, mice were exposed to a non-lethal (4 mg kg-1) or lethal (20 mg kg-1) dose of KCN. From these mice, purified brain mitochondria were prepared and cytochrome oxidase and respiratory activities measured. Results of these experiments revealed greater inhibition of cytochrome oxidase activity following a lethal (20 mg kg-1) than a non-lethal (4 mg kg-1) KCN dose (57 and 45% inhibition, respectively). Respiration states 3 and 4 of brain mitochondria prepared from mice that received 4 mg kg-1 KCN were inhibited by 15 and 20%, respectively. In mice that received a lethal 20 mg kg-1 KCN dose, respiration states 3 and 4 were each inhibited by ca. 30% (P < 0.05). In vitro, mitochondrial cytochrome oxidase activity was inhibited in a concentration-dependent fashion at cyanide concentrations of 10(-6)-10(-2) M. A biphasic inhibition of ADP-stimulated (state 3) respiration was observed. Cyanide concentrations of 10(-6)-10(-4) M produced only a 25% inhibition of respiration state 3, whereas 10(-3) M produced 80% inhibition. Because this dramatic inhibition only occurred at cyanide concentrations that caused > 50% inhibition of mitochondrial cytochrome oxidase activity, these findings suggest that a large proportion of cytochrome oxidase activity may be functional reserve and that cyanide poisoning likely involves other mechanisms in addition to inhibition of cytochrome oxidase.

Adenosine Diphosphate↗

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Dyrene toxicity and lack of carboxylesterase inhibition in mice and rats.

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Aniline Compounds↗

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A comparative study of mouse liver proteins arylated by reactive metabolites of acetaminophen and its nonhepatotoxic regioisomer, 3'-hydroxyacetanilide.

Acetaminophen (4'-hydroxyacetanilide), a widely used analgesic/antipyretic drug, is hepatotoxic in large doses, whereas the m-hydroxy isomer of acetaminophen, 3'-hydroxyacetanilide, is not hepatotoxic. Both are oxidized by mouse liver cytochromes P-450 to reactive metabolites that bind covalently to hepatic proteins. Because previous studies have shown that peak levels of liver protein adducts formed after the administration of each of these compounds to mice are nearly equivalent, and because liver protein adduct formation correlates with hepatotoxicity caused by acetaminophen in mice, we investigated the abundance and patterns of protein adducts formed by acetaminophen and its regioisomer for significant differences. Hepatotoxic doses of acetaminophen to mice significantly altered the abundances of several liver proteins 2 h after dosing as revealed by densitometric analysis of two-dimensional electrophoretic patterns of these proteins. The same analysis after the administration to mice of 3'-hydroxyacetanilide indicated that this nonhepatotoxic regioisomer of acetaminophen caused several similar changes in protein patterns, but also revealed some significant differences. Binding of radiolabeled acetaminophen and 3'-hydroxyacetanilide to hepatic proteins corroborated and extended these results. Two hours after the administration of 14C-labeled analogs of these two compounds to mice, at a time when their extent of total covalent binding to hepatic proteins is approximately equivalent, there are many similarities but also some differences in selectivity of proteins that are adducted, as revealed by both one-dimensional and two-dimensional gel electrophoresis followed by phosphorimage analysis of radiolabel bound to protein bands. Moreover, protein adducts formed from 3'-hydroxyacetanilide were found to be less stable than those formed from acetaminophen under the conditions of electrophoretic analysis. Furthermore, a comparison of radiodetection and immunodetection of protein adducts formed from acetaminophen with an antibody specific for acetaminophen protein adducts indicates that the antibody detects most of the same proteins that are radiolabeled and that the relative quantitative contribution of various adducts to the overall covalent binding of acetaminophen to proteins is approximately the same by both methods. Thus, 3'-hydroxyacetanilide should prove to be a useful tool to aid in the discrimination of hepatic acetaminophen protein adducts that may be critical or noncritical to survival of hepatocytes.

Acetaminophen↗

PubMedDate not supplied

Ribose cysteine protects against acetaminophen-induced hepatic and renal toxicity.

Ribose cysteine (RibCys) is a cysteine prodrug that increases both hepatic and renal glutathione with documented antagonism of acetaminophen (APAP)-induced hepatotoxicity. To determine if RibCys could also protect against APAP-induced kidney damage, mice were injected with APAP (600 mg/kg) or APAP and RibCys (1.0 g/kg) (APAP/RIB) followed by additional RibCys injections 1 and 2 hours later. Mice were euthanatized 10-12 hours after APAP administration, and liver and kidney toxicity were assessed by plasma sorbitol dehydrogenase (SDH) activity and blood urea nitrogen (BUN), respectively, and by histopathology. APAP treatment resulted in elevation of SDH activity and BUN to 2,490 U/ml and 47 mg/dl, respectively. By contrast, SDH and BUN values for APAP/RIB-treated mice were not different from controls, 0 U/ml and 31 mg/dl, respectively. Histopathologic examination revealed moderate to severe hepatic centrilobular necrosis in 9/11 and renal proximal tubular necrosis in 10/11 APAP-treated mice. However, no evidence of hepatic or renal toxicity was noted in any of the 12 APAP/RIB-treated mice. Utilizing the same treatment regimen, APAP covalent binding to hepatic and renal cytosolic proteins was assessed 4 hours after APAP challenge. RibCys cotreatment decreased covalent binding to the 58-kDa acetaminophen-binding protein in both liver and kidney. RibCys decreased both toxicity and covalent binding after APAP administration, and in addition to protecting the liver, this cysteine prodrug can also effectively protect the kidney from APAP-induced injury.

Acetaminophen↗

PubMedDate not supplied

Selective protein arylation and acetaminophen-induced hepatotoxicity.

More than 20 years have passed since the early reports of acute hepatotoxicity with APAP overdose. During that period investigative research to discover the "mechanism" underlying the toxicity has been conducted in many species and strains of intact animals as well as in a variety of in vitro and culture systems. Such work has clarified the primary role of biotransformation and the protective role of GSH. Understanding the former provides explanations for the toxic interactions which may occur with alcohol or other xenobiotics, while understanding of the latter led to the development of antidotes for the treatment of acute poisoning. Acetaminophen (APAP)-induced hepatotoxicity: roles for protein arylation. Initiating events in toxicity require biotransformation of APAP to NAPQI followed by arylation of several important proteins with subsequent alteration of protein structure and function. The immediate consequence of the alterations is detectable in several organelles and these may represent multiple initiating events which are depicted as acting in concert to cause cell injury (large arrowheads). Arylation of cytosolic 58-ABP with subsequent translocation to the nucleus is depicted as a possible signaling mechanism for determining outcome at the cell or organ level (within dotted boundary). For simplicity NAPQI's potentials for oxidizing protein sulfhydryls and direct binding to DNA have been omitted. Significant light has also been shed on the biochemical and cellular events which accompany APAP-induced hepatotoxicity. However, such studies have not identified a unique mechanism of toxicity that is universally accepted. The recent identification of several protein targets which become arylated during toxicity--along with the findings that arylation of some of those target proteins results in loss of protein function--demonstrates that covalent binding does, indeed, have biological consequences and is not merely an indicator of the fleeting presence of reactive electrophiles. These observations further suggest that multiple independent insults to the cell may be involved in toxicity. it is now apparent that the concept of a multistage process that involves both initiation and progression events is appropriate for APAP toxicity, and it is unlikely that a unique initiating event will ever be identified. In light of recent findings it is more likely that a number of such cellular events occur very early after toxic overdosage, and that they collectively set in motion and perpetuate the biochemical, cellular, and molecular processes which will determine outcome. The importance of 58-ABP arylation with early, apparently selective, translocation to the nucleus remains to be elucidated. To date there is nothing to suggest that this represents an initiating event in toxicity. rather it is plausible that the translocation may play a role in signaling electrophile presence and in calling for cellular defense against electrophile insult. This is reflected in the hypothetical model presented in Fig. 3. Critical experimental testing of this model will advance our understanding of the cellular and molecular responses to toxic electrophile insult.

Acetaminophen↗