Contribution of acetaminophen-cysteine to acetaminophen nephrotoxicity II. Possible involvement of the γ-glutamyl cycle
Introduction
Acetaminophen (APAP) is a commonly used over-the-counter analgesic and antipyretic. Though acetaminophen is recognized as a safe and efficacious drug, overdose can result in both nephrotoxicity and hepatotoxicity (Curry et al., 1982, Proudfoot and Wright, 1970). In a recent review of pediatric overdose cases, 9% of the patients had renal injury (Boutis and Shannon, 2001). Though acetaminophen-induced nephrotoxicity is less common than hepatotoxicity, the fact that nephrotoxicity can occur in the absence of hepatotoxicity requires that kidney function always be monitored following overdose (Davenport and Finn, 1988, Kher and Maker, 1987). The apparent independence of acetaminophen-induced nephrotoxicity and hepatotoxicity suggests that different mechanisms may be involved. Our earlier investigations demonstrated the possible role of glutathione-conjugate-derived APAP metabolites in APAP nephrotoxicity. Thus, pretreatment with acivicin, an inhibitor of γ-glutamyl transpeptidase (γ-glutamyl transpeptidase; γ-GT), or probenecid, an organic anion transport inhibitor, protected against APAP nephrotoxicity but not hepatotoxicity in male CD-1 mice (Emeigh et al., 1996). Our accompanying report (Stern et al., 2004) demonstrated the ability of acetaminophen-cysteine (APAP-CYS) to potentiate APAP-induced renal injury. The present report provides a possible mechanism for the observed potentiation.
In the accompanying manuscript, APAP-CYS administration to male CD-1 mice resulted in a dose-dependent decrease in renal GSH levels, while sparing hepatic thiols (Stern et al., 2004). Established mechanisms for other nephrotoxic glutathione conjugates have involved a kidney-selective uptake or bioactivation (Dekant, 2001) and cannot adequately explain the APAP-CYS phenomenon. Recently, Mutlib et al. (2001) proposed a novel mechanism for renal GSH depletion by benzylamine. In this mechanism, benzylamine functions as a γ-glutamyl acceptor substrate resulting in enhancement of GSH catabolism by γ-GT and perturbation of the γ-glutamyl cycle (Scheme 1). The γ-glutamyl cycle is responsible for maintenance of GSH homeostasis and may play a role in amino acid transport (Meister et al., 1979, Smith et al., 1991). Administration of γ-glutamyl acceptor substrates (e.g., glycylglycine) to mice and rats increases transpeptidation by brush border γ-GT and results in catabolism of intracellular GSH, presumably following luminal translocation (Griffith et al., 1978, Palekar et al., 1975). Organs such as the liver, with much lower γ-GT levels, were considerably less susceptible to such GSH depletion after these substrates. Because APAP-CYS is similar to benzylamine and amino acid γ-glutamyl acceptors, it may also selectively deplete renal GSH by acting as a γ-glutamyl acceptor (Scheme 1). If APAP-CYS were to function as a γ-glutamyl acceptor and decrease renal GSH stores, this could explain how APAP-GSH-derived metabolites contribute to APAP nephrotoxicity. The present study explores the hypothesized role of APAP-CYS as a γ-glutamyl acceptor substrate with consequent implications for the mode of action in APAP-induced kidney toxicity.
Section snippets
Materials
Acivicin, 2-amino-methyl-1,3-propanediol, acetaminophen (APAP), bovine renal γ-glutamyl transpeptidase (γ-GT), bovine serum albumin (BSA), l-cysteine, 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB), l-glutamic acid γ (p-nitroanilide), glutathione (reduced), glutathione reductase, glycylglycine, l-lysine, reduced nicotinamide adenine dinucleotide phosphate (NADPH), disodium ethylinediaminetetraacetic acid (Na2EDTA), potassium carbonate, propylene glycol, o-phthaldehyde, sodium phosphate (monobasic
Role of γ-GT in APAP-CYS-induced reduction in renal GSH
APAP-CYS administration to male CD-1 mice resulted in a dose-dependent decrease in renal GSH levels while sparing hepatic thiols (Stern et al., 2004). To verify that the effect of APAP-CYS on renal GSH is mediated by γ-GT, groups of mice were pretreated with the γ-GT inhibitor acivicin (50 mg kg−1, ip), 30 min before administration of APAP-CYS (Reed et al., 1980). If the mechanism behind APAP-CYS-induced renal GSH depletion involved enhancement of γ-GT-dependent GSH catabolism, then acivicin
Discussion
We have documented that APAP-CYS administration to male CD-1 mice resulted in a dose-dependent decrease in renal GSH levels, while sparing hepatic thiols (Stern et al., 2004). The present study demonstrates that this APAP-CYS-induced renal GSH depletion is antagonized by the γ-GT inhibitor acivicin (Fig. 1). There are many established mechanisms by which GSH-conjugate-derived APAP metabolites may alter renal homeostasis. In searching for potential mechanisms that could explain APAP-CYS-induced
Acknowledgment
Supported in part by NIH ES07163 and the University of Utah College of Pharmacy. Underlying support for facilities from NCI grant 5 P30 CA42014 and NIH grant 1 S10 RR06262 is greatly appreciated.
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- 1
Present address: Department of Drug Delivery and Disposition, School of Pharmacy, University of North Carolina, Chapel Hill, NC 27599, USA.
- 2
Present address: School of Pharmacy, University of Wisconsin, Madison, WI 53705-2222, USA.