Immunohistochemical localization and quantification of the
3-(cystein-S-yl)-acetaminophen protein adduct in acetaminophen
Roberts DW; Bucci TJ; Benson RW; Warbritton AR; McRae TA; Pumford
Division of Biochemical Toxicology' National Center for Toxicological
Research' Jefferson' AR 72079-9502.
Am J Pathol, 138(2):359-71 1991 Feb
acetaminophen overdose causes severe hepatotoxicity in humans
laboratory animals' presumably by metabolism to N-acetyl-p-benzoquinone
imine: and binding to cysteine groups as
3-(cystein-S-yl)acetaminophen-protein adduct. Antiserum specific
the adduct was used immunohistochemically to demonstrate the
distribution' and concentration of this specific adduct in livers
treated mice and was correlated with cell inJury as a function
and time. Within the liver lobule' immunohistochemically demonstrable
adduct occurred in a temporally progressive' central-to-peripheral
pattern. There was concordance between immunohistochemical staining
quantification of the adduct in hepatic 10'000g supernate' using
quantitative particle concentration fluorescence immunoassay.
include: 1) immunochemically detectable adduct before the appearance
centrilobular necrosis' 2) distinctive lobular zones of adduct
localization with subsequent depletion during the progression
toxicity' 3) drug-protein binding in hepatocytes at subhepatotoxic
doses and before depletion of total hepatic glutathione' 4)
immunohistochemical evidence of drug binding in the nucleus'
adduct in metabolically active and dividing hepatocytes and in
macrophagelike cells in the regenerating liver.
Peroxidase-catalyzed oxidation of 3'5-dimethyl acetaminophen
cell death by selective protein thiol modification in isolated
Weis M; Rundgren M; Nelson S; Mold]eus P
Institute of Environmental Medicine' Division of Toxicology'
Institute' Stockholm' Sweden.
Chem Biol Interact, 100(3):255-65 1996 May 6
In this study we used a peroxidase model system (glucose/glucose
oxidase and horseradish peroxidase) to investigate the effect
extracellularly generated reactive metabolites of 3'5-Me2-acetaminophen
on cell viability and on cellular thiol levels. Incubation of
hepatocytes with 3'5-Me2-acetaminophen in the presence of
glucose/glucose oxidase and horseradish peroxidase caused a
concentration-dependent loss of cell viability. Loss of viability
associated with decreased protein thiol levels. Addition of the
reducing agent DTT' but not catalase' during the incubation restored
cellular protein thiol levels and arrested the cell killing.
thiol depletion occurred selectively to the mitochondrial and
microsomal fractions and was specific for a very limited number
protein bands. The data suggest that the oxidative modification
individual protein cysteine residues within the latter two organelle
fractions is critically involved in the mechanism of toxicity.
Protection from oxidative damage in mouse liver cells.
Harman AW; Adamson GM; Shaw SG
Department of Pharmacology' University of Western Australia'
Toxicol Lett, 64-65 Spec No():581-7 1992 Dec
Paracetamol toxicity in mouse hepatocytes involved oxidative
initiated by the formation of NAPQI. This oxidative component
paracetamol inJury is associated with the latter stages of the
poisoning process. Ebselen' a drug with GSH-peroxidase activity'
effective in ameliorating these oxidative events.
[Protective effect of the fulvotomentosides on paracetamol-induced
hepatotoxicity in mice
Shi JZ; Liu GT
Guizhou Institute of Traditional Chinese Medicine' Guiyang.
Yao Hsueh Hsueh Pao, 30(4):311-4 1995
Fulvotomentoside alpha-hederin (H) and sapindoside B (S) are
components of the fulvotomentosides reported to decrease the
of a number of hepatotoxins in mice. We studied the effects of
mixture of alpha-hederin and sapindoside B (HS) on paracetamol
(Par)-induced liver inJury. HS (20 mg.kg-1 sc twice) was shown
decrease the mortality produced by Par. The liver inJury produced
Par was remarkably decreased as indicated by decrease of sGPT
histopathological examination. Additionally' HS mitigated the
GSH depletion caused by Par. Par excretion into urine was increased
[Protective action of seed oil of Hippophae rhamnoides L. (HR)
experimental liver inJury in mice
Department of Pharmacology' Lanzhou Medical College.
Chung Hua Yu Fang I Hsueh Tsa Chih, 26(4):227-9 1992 Jul
Protection by seed oil of HR against hepatic inJury induced by
ethyl alcohol and acetaminophen (AAP) on mice was studied in
paper. It was found that seed oil of HR markedly inhibited MDA
formation of liver induced by CCl4' AAP and ethyl alcohol' seed
4.75 g/kg could lower SGPT levels induced by CCl4 and AAP. It
also depletion of GSH damaged liver induced by AAP. The mechanism
heptoprotective actions of HR seed oil might be related to anti-lipid
p-aminophenol nephrotoxicity: biosynthesis of toxic glutathione
Klos C; Koob M; Kramer C; Dekant W
Institut f ur Toxikologie' Universit at W urzburg' Germany.
Toxicol Appl Pharmacol, 115(1):98-106 1992 Jul
p-Aminophenol causes necrosis of the pars recta of the proximal
in rats' and its nephrotoxicity may be due to glutathione-dependent
bioactivation reactions. We have investigated the hepatic metabolism
p-aminophenol in Wistar rats and the cytotoxicity of formed glutathione
S-conJugates in rat renal epithelial cells. After ip application
p-aminophenol (100 mg/kg)' the following metabolites were identified
rat bile: 4-amino-2-(glutathion-S-yl)phenol'
6)-tris(glutathion-S-yl)phenol' an aminophenol conJugate (likely
sulfate or glucuronide)' acetaminophen glucuronide' and
4-amino-2'5-bis(glutathion-S-yl)phenol' and 4-amino-2'3'5(or
6)-tris(glutathion-S-yl)phenol induced a dose- and time-dependent
of cell viability in rat kidney cortical cells. Cell killing
significantly reduced by inhibition of gamma-glutamyl transpeptidase
with Acivicin. p-Aminophenol was also toxic to renal epithelial
Coincubation of p-aminophenol with tetraethylammonium bromide'
competitive inhibitor of the organic cation transporter' and
SKF-525A' an inhibitor of cytochrome P450' protected cells from
p-aminophenol-induced toxicity. p-Aminophenol would thus be accumulated
in the kidney mainly by organic cation transport systems' which
concentrated in the S-1 segment of the proximal tubule. However'
p-aminophenol toxicity in vivo is directed toward the S-2 and
segments' which are rich in gamma-glutamyl transpeptidase. These
results and the observation that biliary cannulation and glutathione
depletion reduce p-aminophenol nephrotoxicity suggest that the
biosynthesis of toxic glutathione conJugates is responsible for
p-aminophenol nephrotoxicity in vivo. The aminophenol glutathione
S-conJugates formed induce p-aminophenol nephrotoxicity by a
dependent on gamma-glutamyl transpeptidase.
Managing patients with acute myocardial ischemia and reperfusion
Stewart S; Ryan C; Poropat S
Department of Cardiology' Queen Elizabeth Hospital' Woodville'
Dimens Crit Care Nurs, 16(3):122-31 1997 May-Jun
Previously administered in cases of acetaminophen toxicity'
N-Acetylcysteine (NAC) is now also being used in the management
acute myocardial ischemia and reperfusion inJury. NAC potentiates
beneficial effects of nitrates such as nitroglycerin and reduces
oxidative stress on the heart. The critical care nurse plays
important role in optimizing the therapeutic benefits of NAC
minimizing its potential harmful effects.
Evaluation of acetaminophen-induced developmental toxicity using
Fort DJ; Rayburn JR; Bantle JA
Stover Biometric Laboratories' Inc.' Stillwater' Oklahoma 74076.
Drug Chem Toxicol, 15(4):329-50 1992
Potential mechanisms of acetaminophen-induced developmental toxicity
were evaluated using FETAX (Frog Embryo Teratogenesis Assay-Xenopus).
Early Xenopus laevis embryos were exposed to acetaminophen for
two definitive concentrations-response assays with and without
exogenous metabolic activation system (MAS). Two static renewal
of acetaminophen and the MAS treated with carbon monoxide' cimetidine'
ellipticine' diethyl maleate' and supplemented with glutathione
also performed. Addition of the MAS decreased the 96-h LC50 and
(malformation) values of unactivated acetaminophen 3.9-fold and
7.1-fold' respectively. Addition of the carbon monoxide- and
ellipticine-inhibited MAS' as well as the glutathione-supplemented
decreased the developmental toxicity of activated acetaminophen
levels near that of the unactivated parent compound.
Cimetidine-inhibited MAS also reduced the developmental toxicity
acetaminophen' but not to the extent observed with the carbon
and ellipticine-inhibited' or glutathione-supplemented MAS. Addition
the diethyl maleate-treated MAS substantially increased the
developmental toxicity of acetaminophen. Results indicate that
reactive intermediate formed as the result of MFO-mediated metabolism
(possibly P-448) significantly increased the developmental toxicity
acetaminophen. glutathione was also found to play a maJor role
intermediate detoxification in vitro.
Sex-related differences in mouse renal metabolism and toxicity
Hu JJ; Lee MJ; Vapiwala M; Reuhl K; Thomas PE; Yang CS
Department of Chemical Biology and Pharmacognosy' College of
Rutgers University' Piscataway' New Jersey 08855-0789.
Toxicol Appl Pharmacol, 122(1):16-26 1993 Sep
The obJective of this study is to elucidate the role of cytochrome
2E1 in the metabolic activation of acetaminophen (APAP) in mouse
kidneys. With the kidney microsomes from C3H/HeJ mice' a significant
sex-related difference was observed in the NADPH-dependent formation
a reactive APAP metabolite which was trapped as a glutathione
conJugate. The enzyme activity in male mouse kidney was about
50-fold higher than that in the females at substrate concentrations
and 500 microM' respectively' and treatment of female mice with
testosterone significantly induced the enzyme activity in the
kidney. No sex-related difference in this enzyme activity' however'
observed in the livers. The oxidative metabolism of APAP in control
male mouse kidney microsomes displayed an apparent low Km of
microM and an apparent high Km of 603-702 microM. Studies using
monoclonal antibodies against P450 2E1 showed that P450 2E1 accounted
for about half of the activity in the metabolic activation of
both mouse liver and kidney. Furthermore' there was as strong
correlation between the renal P450 2E1 content (measured by Western
blot analysis) and the rate of APAP oxidation by renal microsomes.
test the hypothesis that the susceptibility of renal tissue to
toxicity is determined by the distribution and level of P450
toxicity studies were conducted. A time-dependent depletion of
and renal cortical glutathione was observed in both male and
mice following a dose of 1200 mg/kg APAP. At 4 hr after APAP
the level of renal cortical glutathione depletion in male mice
significantly greater than that in the females. The level of
creatinine elevation in male mice was higher than that in the
hr after APAP dosing. Histopathology studies by light and electron
microscopic assessments demonstrated that renal damage by APAP
restricted mostly to the epithelial cells of the proximal convoluted
tubules where P450 2E1 was localized. The renal proximal tubular
necrosis induced by APAP was more severe in males than that in
females. Results from this study suggest that P450 2E1 plays
important role in the metabolic activation of APAP and is a key
in determining the sex-related difference of APAP-mediated toxicity
the mouse kidney.
Outpatient N-acetylcysteine treatment for acetaminophen poisoning:
ethical dilemma or a new financial mandate?
Dean BS; Bricker JD; Krenzelok EP
Pittsburgh Poison Center' Children`s Hospital of Pittsburgh'
Vet Hum Toxicol, 38(3):222-4 1996 Jun
The mainstay of treatment for acetaminophen-induced hepatotoxicity'
produced by the accumulation of the toxic metabolite
N-acetylbenzoquinoneimine' is an enteral 18-dose course of
N-acetylcysteine (NAC). However' absence of characteristic
symptomatology is a frequent reason for premature cessation of
early discharge of the toxic acetaminophen poisoned patient.
a series of confirmed acetaminophen poisonings who were discharged
early with NAC and instructions to self-administer. All cases
acetaminophen poisoning without concomitant drugs' reported to
certified Regional Poison Information Center for a 3-mo period
were reviewed. Inclusion criteria included patients who were
with orders to complete the course of NAC outside of a hospital'
despite toxic serum acetaminophen concentrations. Data parameters
evaluated included age' amount taken' symptoms' laboratory results'
treatment' and medical outcome. 131 cases of confirmed toxic
acetaminophen poisoning yielded 6 patients who received 4 to
6 doses of
NAC during hospitalization' but were discharged to home with
remaining 11-13 doses. Patients` ages ranged from 16-28 y (mean
y). Serum acetaminophen concentrations measured at 4 h post-ingestion
ranged from 171-198 mcg/ml (mean 182 mcg/ml). Follow-up by the
certified Regional Poison Information Center at 1-3 w post-discharge
determined dosing compliance to be 83%. All 6 patients remained
asymptomatic with normal liver function testing. Since health
reform encourages practitioners to reconsider established approaches
the delivery of health care' perhaps home delivery of NAC would
only be clinically preferred to premature cessation of the antidote'
but also offer cost savings. Self-administration of NAC in the
setting may be representative of a new era in America`s health
acetaminophen structure-toxicity studies: in vivo covalent binding
nonhepatotoxic analog' 3-hydroxyacetanilide.
Roberts SA; Price VF; Jollow DJ
Department of Pharmacology' Medical University of South Carolina'
Toxicol Appl Pharmacol, 105(2):195-208 1990 Sep 1
High doses of 3-hydroxyacetanilide (3HAA)' a structural isomer
acetaminophen' do not produce hepatocellular necrosis in normal
hamsters or in those sensitized to acetaminophen-induced liver
by pretreatment with a combination of 3-methylcholanthrene' borneol'
and diethyl maleate. Although 3HAA was not hepatotoxic' the
administration of acetyl-labeled [3H or 14C 3HAA (400 mg/kg'
produced levels of covalently bound radiolabel that were similar
those observed after an equimolar' hepatotoxic dose of [G-3H
acetaminophen. The covalent nature of 3HAA binding was demonstrated
retention of the binding after repetitive organic solvent extraction
following protease digestion. Hepatic and renal covalent binding
3HAA was approximately linear with both dose and time. In addition'
3HAA produced only a modest depletion of hepatic glutathione'
suggesting the lack of a glutathione threshold. 3-Methylcholanthrene
pretreatment increased and pretreatment with cobalt chloride
piperonyl butoxide decreased the hepatic covalent binding of
indicating the involvement of cytochrome P450 in the formation
3HAA reactive metabolite. The administration of multiple doses
single dose of [ring-3H 3HAA to hamsters pretreated with a combination
of 3-methylcholanthrene' borneol' and diethyl maleate produced
levels of 3HAA covalent binding that were in excess of those
after a single' hepatotoxic acetaminophen dose. These data suggest
the nature and/or the intracellular processing of the reactive
metabolites of acetaminophen and 3HAA are different. These data
demonstrate that absolute levels of covalently bound xenobiotic
metabolites cannot be utilized as absolute predictors of cytotoxic