Disulfiram prevents acetaminophen hepatotoxicity in rats

Hepatic necrosis due to an oral acetaminophen overdose (4.25 g/kg b.wt.) was prevented by pretreatment with disulfiram 100 mg/kg, given for 3 weeks or as a single dose. Twenty-four hours after acetaminophen the impairment of hepatic function, measured as prothrombin index, and the depletion of hepatic glutathione were prevented. Hepatic cytochrome P-450 levels were unchanged but cytochrome P-450 mediated p-nitroanisole demethylation was reduced by disulfiram pretreatment. Disulfiram pretreatment reduced 24 hour urinary excretion of acetaminophen-mercapturate and- cysteine while excretion of -sulfate and -glucuronide was unchanged. After 72 hours acetaminophen induced hepatic necrosis were prevented. Identical observations were made in animals pretreated with disulfiram for 3 weeks. Five hours after acetaminophen overdose its irreversible binding to hepatic proteins was not changed. After 24 hours, however, it was increased in animals pretreated with a single disulfiram dose and unchanged in animals pretreated for 3 weeks. The protective mechanism of disulfiram after acetaminophen overdose is not mediated via a change in overall irreversible binding of acetaminophen to hepatic protein.     

双环醇对大鼠黄曲霉毒素B1代谢和肝毒性的影响

目的:研究抗肝炎新药双环醇对大鼠黄曲霉毒素B_1(AFB_1)代谢和肝毒性的影响.方法:大鼠灌胃双环醇300 mg·kg~(-1)·d~(-1),连服三日后腹腔注射黄曲霉毒素B_1 1.5 mg·kg~(-1).给黄曲霉毒素B_1 16小时后观察双环醇对黄曲霉毒素B_1引起肝损伤的防护作用以及对体外代谢的影响.结果:双环醇(300 mg·kg~(-1)·d~(-1),连服三日)可明显降低黄曲霉毒素B_1引起的大鼠血清转氨酶和肝脏MDA的升高,增加低毒代谢产物AFQ_1的生成.双环醇还可增加大鼠肝脏细胞色素P450总量和胞浆谷胱甘肽含量,诱导P450 CYP2B1介导的7-戊氧基香豆素脱烃酶和谷胱甘肽疏基转移酶的活性.此外,双环醇对P450 CYP3A介导的红霉素脱甲基酶和 P450 CYP1A介导的7-乙氧基香豆素脱烃酶也有诱导作用.结论:双环醇可通过增加大鼠肝脏对AFB_1代谢的解毒功能起到肝保护作用.     

Effects and mechanisms of rifampin on hepatotoxicity of acetaminophen in mice

This study examined the effects and possible mechanisms of rifampin against acetaminophen-induced hepatotoxicity in mice. Rifampin significantly enhanced the biotransformation of acetaminophen, evidenced by the increase in p-aminophenol formation in rifampin-treated microsomes and the increase in plasma clearance rate of acetaminophen. Pretreatment with rifampin significantly decreased serum alanine transaminase (ALT) activities, aspartate transaminase (AST) activities and prevented severe liver necrosis following acetaminophen overdose. The contents and activities of microsomal drug-metabolizing enzyme were less affected in rifampin-pretreated mice in comparison to the animals treated with acetaminophen alone. Rifampin was capable of increasing glutathione (GSH) level and GSH reductase activity and reducing GSH depletion and the decrease in GSH reductase activity by acetaminophen in mice. In addition, it was found that the microsomal Ca²⁺-ATPase activity was not directly related to acetaminophen toxic species generated in the P450 enzyme system in vitro. These findings suggest that rifampin has species-specific effects on the liver against acetaminophen-induced hepatotoxicity in mice, which increase the level of GSH by promoting GSH regeneration.     

Effects of butylated hydroxyanisole on acetaminophen hepatotoxicity and glucuronidation in vivo

The present study examined the effects of butylated hydroxyanisole (BHA) on acetaminophen-induced hepatotoxicity and metabolism in vivo with emphasis on possible changes in the glucuronidation pathway. Female Swiss-Webster mice received BHA in the diet (1% w/w) for 12 days (600 to 800 mg/kg/day). BHA prevented acetaminophen hepatotoxicity (600 mg/kg, ip), based on serum alanine and aspartate aminotransferase activities and histopathological examination. The rate of elimination of acetaminophen from blood was 10-fold higher in BHA-fed mice (clearance, 49 ml/min/kg) than in controls (4.4 ml/min/kg). In general, the urinary metabolite excretion patterns in control and BHA-treated mice were the same. However, the rates of acetaminophen conjugation via the sulfation, glucuronidation, and mercapturic acid pathways were enhanced with the rate of glucuronide formation, the major biotransformation pathway of acetaminophen, increased sevenfold in BHA-treated mice (0.041 min-1) compared to controls (0.006 min-1). BHA increased hepatic UDP-glucuronosyltransferase activity twofold, as well as hepatic UDP-glucuronic acid concentrations. In addition, after acetaminophen administration, UDP-glucuronic acid in BHA-treated mice was depleted to a lesser extent and returned to control values more rapidly than in untreated animals. BHA had a similar but less pronounced effect on hepatic glutathione levels. The findings indicate that the rate of acetaminophen glucuronidation is increased in vivo during BHA feeding to mice. This effect appears to play a role in the enhanced excretion of acetaminophen as well as protection against acetaminophen-induced hepatotoxicity.     

In vivo and in vitro hepatotoxicity and metabolism of acetaminophen in Syrian hamsters

The purpose of this investigation was to correlate the in vitro and in vivo toxicity of the hepatotoxicant, acetaminophen. Hamsters were pretreated with either phenobarbital (70 mg/kg) or 3-methylcholanthrene (20 mg/kg) or an appropriate vehicle for 3 days. In non-pretreated hamsters, single doses of acetaminophen (200-400 mg/kg i.p.) caused elevations in serum alanine aminotransferase and sorbitol dehydrogenase activities in a dose-related manner. 3-Methylcholanthrene significantly potentiated, while phenobarbital significantly reduced acetaminophen-induced elevations in serum liver enzyme activities. Both phenobarbital and 3-methylcholanthrene significantly reduced acetaminophen plasma T1/2 while only 3-methylcholanthrene increased APAP clearance. Phenobarbital pretreatment increased the urinary excretion of APAP-glucuronide. Exposure of isolated hepatocytes to acetaminophen (0.01-2.0 mM) resulted in concentration-related decreases in hepatocyte viability. Cells from 3-methylcholanthrene-pretreated hamsters were more markedly susceptible to acetaminophen toxicity than cells isolated from non-induced animals. Hepatocytes isolated from phenobarbitol pretreated animals were slightly but significantly more susceptible to acetaminophen toxicity than cells from control animals. Hepatocytes isolated from 3-methylcholanthrene pretreated animals had increased formation of an acetaminophen-glutathione conjugate compared to control. Pre-treatment with either phenobarbital or 3-methylcholanthrene enhanced glucuronidation of acetaminophen in vitro. These data demonstrate a lack of correlation between in vivo hepatotoxicity and in vitro cytotoxicity in that phenobarbital pre-treatment protected hamsters from acetaminophen-induced liver toxicity, but failed to protect hepatocytes exposed to acetaminophen in vitro.     

Strain differences in susceptibility of normal and diabetic rats to acetaminophen hepatotoxicity

The effects of streptozotocin (STZ)-induced diabetes on acetaminophen metabolism and hepatotoxicity in male Sprague-Dawley (SD) and Long Evans Hooded (LEH) rats were compared. In agreement with earlier studies, normal SD rats were more resistant to acetaminophen-induced hepatic necrosis than normal LEH rats. In contrast to LEH rats, the diabetic state did not protect SD rats from liver injury. Pharmacokinetic studies revealed that normal SD rats eliminated acetaminophen faster than normal LEH rats, and that the diabetic state further enhanced elimination in both strains of rats; however, the effect was much greater in LEH rats. Normal SD rats had a greater capacity to metabolize acetaminophen to nontoxic glucuronide and sulfate conjugates than normal LEH rats. In LEH rats, the diabetic state enhanced acetaminophen glucuronidation and sulfation, whereas in SD rats the diabetic state increased only sulfation; glucuronidation was unaffected. Additional studies revealed that the difference in the glucuronidation capacities between normal LEH and normal SD rats was not due to differences in either the amount of the enzyme, glucuronyl transferase, or basal hepatic levels of the cofactor, UDPGA. Similarly, the diabetes-induced enhancement of glucuronidation in LEH rats was not due to differences in predrug levels of either glucuronyl transferase or UDPGA. Thus, the major difference in susceptibility of the two strains of normal rats to acetaminophen hepatotoxicity appears to be due to the capacity to clear the drug through nontoxic pathways. The greater glucuronidation capacity seen in diabetic LEH rats and in normal and diabetic SD rats as compared to normal LEH rats, appears to be due to a greater ability to produce UDPGA in response to the metabolic demand.     

The influence of antagonists of poly(adp-ribose) metabolism on acetaminophen hepatotoxicity

An array of therapeutically used analgetic and antirheumatic drugs causes severe liver damage. The present study investigates the hepatoprotective effects of inhibitors of NAD-dependent adenoribosylation reactions in analgesics-induced hepatic injury. Male NMRI mice were treated perorally with 500 mg/kg of acetaminophen, and the activities of both glutamate-oxaloacetate transaminase (GOT) and glutamate-pyruvate transaminase (GPT) were determined in serum. In addition, the activity of poly(ADP-ribose)polymerase (PARP) was quantified in liver cell nuclei. While the PARP-activity remained essentially unchanged, the acetaminophen-induced release of both GOT and GPT from injured liver cells could be inhibited by 90–99%, when mice were injected additionally with the selective PARP-inhibitors nicotinic acid amide, benzamide, caffeine, theophyline, and thymidine, respectively. We see the main application of inhibitors of adenoribosylation reactions as for the combinational use in pharmaceutical preparations of analgesics and antirheumatic drugs in order to avoid hepatic damage.     

Effects of treatment with enalapril on hepatotoxicity induced by acetaminophen in mice

There is a current need for new therapeutic options for acetaminophen (APAP)-induced hepatotoxicity. Herein, we assessed the effects of prophylactic and therapeutic treatment with the angiotensin-converting enzyme (ACE) inhibitor, enalapril, on APAP-caused hepatotoxicity. Male and female C57BL/6?J mice were used, and hepatotoxicity was induced by a single application of APAP (400?mg/kg, i.p.). Macroscopic and histological liver alterations, serum alanine transaminase (ALT) and aspartate transaminase (AST) activity, liver catalase activity (CAT), reduced glutathione concentrations (GSH), hepatic measurement of neutrophil migration (myeloperoxidase, MPO activity), and caspase-3 liver expression were evaluated. The prophylactic and the therapeutic treatments with enalapril were able to markedly reduce the macroscopic and histological liver alterations as well as the caspase-3 immunopositivity. Both schedules of treatment were also effective in reducing GSH concentrations as well as neutrophil migration. Conversely, only the pre-treatment (but not the post-administration) with enalapril significantly reversed APAP-induced CAT decrease. Furthermore, the pre- or the post-treatment with enalapril largely reduced ALT and AST serum activity in APAP-intoxicated mice. The hepatoprotective effects of enalapril were comparable to those obtained with the clinically used compound N-acetylcysteine (NAC) when given in a therapeutic regimen. Data obtained with the prophylactic protocol of treatment might indicate that individuals under treatment with ACE inhibitors are less susceptible to the toxic effects of APAP. Additionally, the therapeutic approach allows us to suggest that enalapril might represent an innovative tool for treating APAP intoxication.     

Metabolic phenotyping applied to pre-clinical and clinical studies of acetaminophen metabolism and hepatotoxicity

Abstract

Acetaminophen (APAP, paracetamol, N-acetyl-p-aminophenol) is a widely used analgesic that is safe at therapeutic doses but is a major cause of acute liver failure (ALF) following overdose. APAP-induced hepatotoxicity is related to the formation of an electrophilic reactive metabolite, N-acetyl-p-benzoquinone imine (NAPQI), which is detoxified through conjugation with reduced glutathione (GSH). One method that has been applied to study APAP metabolism and hepatotoxicity is that of metabolic phenotyping, which involves the study of the small molecule complement of complex biological samples. This approach involves the use of high-resolution analytical platforms such as NMR spectroscopy and mass spectrometry to generate information-rich metabolic profiles that reflect both genetic and environmental influences and capture both endogenous and xenobiotic metabolites. Data modeling and mining and the subsequent identification of panels of candidate biomarkers are typically approached with multivariate statistical tools. We review the application of multi-platform metabolic profiling for the study of APAP metabolism in both in vivo models and humans. We also review the application of metabolic profiling for the study of endogenous metabolic pathway perturbations in response to APAP hepatotoxicity, with a particular focus on metabolites involved in the biosynthesis of GSH and those that reflect mitochondrial function such as long-chain acylcarnitines. Taken together, this body of work sheds much light on the mechanism of APAP-induced hepatotoxicity and provides candidate biomarkers that may prove of translational relevance for improved stratification of APAP-induced ALF.     

First-pass metabolism of acetaminophen in thyroxine treated rats

The study on the first-pass metabolism of acetaminophen was carried out in normal and thyroxine-treated rats, administered 30 mg/kg by three routes of intravenous, intraperitoneal, and oral one. Unconjugated acetaminophen and two major metabolites, glucuronide and sulfate in the plasma and urine were then measured 5 and 24 h after the administration, respectively. It was found that there was no difference in total percentage of excreted amount, independent of the routes for administration, between normal and thyroxine-treated rats. This fact shows that acetaminophen is absorbed completely from the gastrointestinal tract. However, it was also found that the extraction ratio of gastrointestinal tract in thyroxine-treated rats became smaller, and that the volume of distribution and total body clearance became larger than those in normal rats. The first-pass metabolism of acetaminophen was found to be influenced by the continuous administration of thyroxine.     

Potentiated hepatotoxicity from concurrent administration of acetaminophen and allyl alcohol to rats

Female Wistar rats were treated with acetaminophen 3.0 g/kg BW and allyl alcohol 75 microliter/kg BW by gastric tube. Hepatic function, measured as galactose elimination capacity and prothrombin index, was reduced to about 0.40 times control value. Plasma alanine transferase activity was elevated more and earlier after treatment with acetaminophen and allyl alcohol compared to administration of acetaminophen alone. Also prothrombin index was reduced more and earlier from the combination. Hepatic glutathione was depleted to a lower level 3 hr after administration of the combination of toxins, compared to administration of acetaminophen alone, after 6 hr there was no difference. Excretion of acetaminophen metabolites, especially the acetaminophen mercapturate, into urine was not changed from the combination. After administration of the toxic combination histological changes in the liver were minor. The results indicate that the two toxins potentiate each other's action. The potentiation is proposed to be due to prevention of compensatory hyperfunction of non-necrotic liver cells rather than to direct metabolic interaction of the toxins.     

Effect of a concomitant single dose of ethanol on the hepatotoxicity and metabolism of acetaminophen in mice

A concomitant single dose of ethanol (1 g/kg) protected mice from hepatic injury induced by acetaminophen (250 mg/kg) as evidenced by the lowering of plasma transaminases. Pharmacokinetic studies with [¹⁴C]acetaminophen indicated that ethanol enhanced the initial blood concentrations of radiolabel and its rate of elimination. A tissue distribution study suggested that these effects were probably due to an ethanol-induced inhibition of the biliary clearance of acetaminophen from the blood. Examination of the urinary and biliary metabolites indicated that ethanol inhibited the excretion of the degradation products derived from the glutathione-deactivated hepatotoxic acetaminophen intermediate. The decrease in acetaminophen induced hepatotoxicity was therefore attributed to an inhibitory effect of ethanol on the biotransformation of acetaminophen to the toxic intermediate.     

Age-dependent changes in first-pass metabolism of acetaminophen in rats

The contribution of gastrointestinal tract (GIT), liver, and lung towards the first-pass metabolism of acetaminophen was examined using 3-week-old, 10-week-old and 1-year-old rats after administration of 30 mg kg-1 doses by intra-arterial, intravenous, intraperitoneal, and oral routes. Plasma concentrations of acetaminophen and its two major metabolites, acetaminophen glucuronide and acetaminophen sulfate, were measured for about 5h after drug administration. Total oral extraction of acetaminophen was extensive in 10-week-old and 1-year-old rats (Eo = 0.46) and the major contribution to the overall first-pass metabolism was due to the GIT (Eg = 0.50-0.53). Oral extraction in 3-week-old rats was minimal (Eo = 0.10) and there did not appear to be an extraction by the GIT (Eg = 0.00). These results suggest that the ability of GIT to metabolize acetaminophen to glucuronide and sulfate is undeveloped in the infant rats. No changes in the contribution of different organs to the first-pass metabolism of acetaminophen was observed 10 weeks after birth. Pharmacokinetic parameters for acetaminophen in infant rats (3-week-old) and 10-week-old rats were similar after drug administration by the intra-arterial and intravenous routes.     

Effects of erdosteine on cyclosporine-A-induced hepatotoxicity in rats

Cyclosporine A (CsA) is a potent immunosuppressive agent used for organ transplantations and various autoimmune disorders. However, hepatotoxicity due to CsA remains one of the major side effects. The use of antioxidants reduces the adverse effects of CsA. The aim of this study was to determine the protective effects of erdosteine on CsA-induced liver injury through tissue oxidant/antioxidant parameters and to evaluate light microscopic alterations in rat-liver tissues. Rats were randomly divided into four experimental groups: The control group received sunflower oil (2?mL/kg/day, per orally; p.o.), while the other groups were treated with CsA (25?mg/kg/day, p.o.) or erdosteine (10?mg/kg/day, p.o.) or CsA+erdosteine, respectively. Serum aspartate aminotransferase and alanine aminotransferase levels, tissue malondialdehyde and nitric oxide levels, and superoxide dismutase, glutathione peroxidase and catalase enzyme activities were measured. Histological examination was performed. CsA caused a significant deterioration in the hepatic function tests, morphology, and gave rise to severe oxidative stress in the liver. Erdostein significantly improved the functional and histological parameters and attenuated the oxidative stresss induced by CsA. Erdostein protects liver tissue against oxygen free radicals and prevents hepatic dysfunction and morphological abnormalities associated with chronic CsA administration.     

Comparative hepatotoxicity and metabolism of N-methylformamide in rats and mice

N-methylformamide (NMF) produced dose-dependent zone 3 haemorrhagic necrosis in mice; the threshold dose was 100–200 mg/kg. In rats a dose of 1000 mg/kg caused hepatic damage in some animals and slight elevations of plasma transaminases. A species difference in susceptibility to NMF-induced hepatotoxicity is clearly indicated. NMF depleted liver non-protein sulphydryl (NPSH) in a dose-dependent manner in mice, but not in rats. Depletion of liver glutathione by buthionine sulphoximine or diethylmaleate potentiated the hepatotoxicity of NMF in mice. [¹⁴C]-methyl NMF was metabolised by mice and rats and a number of urinary metabolites including an N-acetylcysteine conjugate, methylamine and N-hydroxymethylformamide were detected. There were no qualitative differences in the metabolites between rats and mice but mice metabolised NMF much faster and more extensively than rats.     

Effects of dimethylsulfoxide on metabolism and toxicity of acetaminophen in mice

Effects of dimethylsulfoxide (DMSO) on metabolism and toxicity of acetaminophen (APAP) were examined using male mice. A dose of DMSO (1 ml/kg, i.p.) inhibited the induction of APAP hepatotoxicity almost completely as indicated by changes in serum hepatotoxic parameters. Quantification of major APAP metabolites in plasma showed that APAP-glutathione (GSH), a conjugate generated via metabolic activation of APAP, was reduced significantly while APAP-sulfate and APAP-glucuronide, detoxified metabolites both produced directly from the parent drug, were increased in mice pretreated with DMSO. However, microsomal CYP2E1 activity measured with p-nitrophenol and p-nitroanisole as substrates was increased by DMSO treatment. Generation of APAP-GSH in microsomes from control mice was inhibited by DMSO in a dose-dependent manner. Lineweaver-Burk plot analysis indicated that the inhibition pattern produced by DMSO was competitive in nature. A 10000 g supernatant was reconstituted with the cytosolic fraction and microsomes from DMSO- or saline-treated animals. APAP-GSH production was increased significantly when the cytosolic fraction from saline-treated mice and/or microsomes from DMSO-treated mice were used. The results indicate that DMSO induces the enzyme activity responsible for oxidative metabolism of APAP, but its direct inhibitory effect on the enzymatic interaction with this drug decreases the overall production of a reactive metabolite, resulting in reduction of the hepatotoxicity. It is suggested that DMSO effects on metabolism of a xenobiotic would vary depending on its potential to inhibit the interaction of enzyme(s) and the xenobiotic.