Acetaminophen (APAP), an over-the-counter analgesic and antipyretic, can cause hepatotoxicity when ingested in large overdoses. APAP has multiple formulations including immediate-release (IR) and extended-release (ER) preparations. A recently published consensus statement on the management of APAP poisoning indicated that management of APAP-ER overdose is the same as that for APAP-IR overdose. Consistent with this consensus, it was previously reported that quantitative systems toxicology (QST) modeling using DILIsym predicted similar pharmacokinetic (PK) and hepatic biomarker profiles for the APAP-ER and APAP-IR formulations after overdose in healthy adults. Hepatic injury from APAP is caused by the reactive metabolite, N-acetyl-ρ-benzoquinone imine (NAPQI), which is formed predominantly by CYP2E1-mediated metabolism and eliminated by hepatic glutathione. As such, conditions that can increase NAPQI production (e.g., CYP2E1 induction by alcohol) or decrease hepatic glutathione stores (e.g., underling liver disease) may impact PK and susceptibility to hepatotoxicity after overdose of APAP-IR and APAP-ER. In the current study, APAP-IR and APAP-ER models in chronic alcohol users and individuals with low hepatic glutathione were developed and verified within DILIsym. Simulations using verified models predicted similar PK and hepatic biomarker profiles for the APAP-ER and APAP-IR formulations in moderate and excessive chronic alcohol users and adults with low hepatic glutathione levels after single acute overdoses up to ~100 g and repeat supratherapeutic ingestions (up to 7.8 g/day for 10 days). These results further support that approaches to manage APAP-IR overdoses can be applied to manage APAP-ER overdoses in adults with chronic alcohol consumption or lower hepatic glutathione levels.

Alcoholic liver disease (ALD) is one of the most common health problems worldwide, especially in developing countries caused by chronic consumption of alcohol on a daily basis. The ALD spectrum is initiated with the early stages of alcoholic fatty liver (steatosis), progressing to alcoholic steatohepatitis, followed by the later stages of fibrosis and in some cases, cirrhosis and hepatocellular carcinoma (HCC). The Wnt/β-catenin signaling required for healthy liver development, function, and regeneration is found to be aberrated in ALD, attributed to its progression. This review is to elucidate the association of Wnt/β-catenin signaling with various stages of ALD progression. Alcohol causes downregulation of Wnt/β-catenin signaling components and thereby suppressing the pathway. Reports have been published that aberrated Wnt/β-catenin signaling, especially the absence of β-catenin, results in decreased alcohol metabolism, causing steatosis followed by steatohepatitis via lipid accumulation, lipid peroxidation, liver injury, increased oxidative stress and apoptosis of hepatocytes, contributing to the advancement of ALD. Contrastingly, the progression of later stages of ALD like fibrosis and HCC depends on the increased activation of Wnt/β-catenin signaling and its components. Existing studies reveal the varied expression of Wnt/β-catenin signaling in ALD. However, the dual role of the Wnt/β-catenin pathway in earlier and later stages of ALD is not clear. Therefore, studies on the Wnt/β-catenin pathway and its components in various manifestations of ALD might provide insight in targeting the Wnt/β-catenin pathway in ALD treatment.

Exposure to hepatotoxic chemicals is involved in liver disease-related morbidity and mortality worldwide. The liver responds to damage by triggering compensatory hepatic regeneration. Physical agent or chemical-induced liver damage disrupts hepatocyte proteostasis, including endoplasmic reticulum (ER) homeostasis. Post-liver injury ER experiences a homeostatic imbalance, followed by active ER stress response signaling. Activated ER stress response causes selective upregulation of stress response genes and downregulation of many hepatocyte genes. Acetaminophen overdose, carbon tetrachloride, acute and chronic alcohol exposure, and physical injury activate the ER stress response, but details about the cellular consequences of the ER stress response on liver regeneration remain unclear. The current data indicate that inhibiting the ER stress response after partial hepatectomy-induced liver damage promotes liver regeneration, whereas inhibiting the ER stress response after chemical-induced hepatotoxicity impairs liver regeneration. This review summarizes key findings and emphasizes the knowledge gaps in the role of ER stress in injury and regeneration.

The prominent role of gut in regulating the physiology of different organs in a human body is increasingly acknowledged, to which the bidirectional communication between gut and liver is no exception. Liver health is modulated via different key components of gut-liver axis. The gut-derived products mainly generated from dietary components, microbial metabolites, toxins, or other antigens are sensed and transported to the liver through portal vein to which liver responds by secreting bile acids and antibodies. Therefore, maintaining a healthy gut microbiome can promote homeostasis of this gut-liver axis by regulating the intestinal barrier function and reducing the antigenic molecules. Conversely, liver secretions also regulate the gut microbiome composition. Disturbed homeostasis allows luminal antigens to reach liver leading to impaired liver functioning and instigating liver disorders. The perturbations in gut microbiome, permeability, and bile acid pool have been associated with several liver disorders, although precise mechanisms remain largely unresolved. Herein, we discuss functional fingerprints of a healthy gut-liver axis while contemplating mechanistic understanding of pathophysiology of liver diseases and plausible role of gut dysbiosis in different diseased states of liver. Further, novel therapeutic approaches to prevent the severity of liver disorders are discussed in this review.

Alcohol-associated liver disease (ALD) is a global health problem caused, among other factors, by oxidative stress from the formation of reactive oxygen species (ROS). One important source of ROS is microsomal ethanol metabolism catalyzed by cytochrome P450 2E1 (CYP2E1), which is induced by chronic ethanol consumption. Inhibition of CYP2E1 by clomethiazole (CMZ) decreases oxidative stress in cell cultures and improves ALD in animal studies. Our study aimed to assess the benefits of a CYP2E1 inhibitor (clomethiazole) in detoxification of patients with ALD.

Open label, randomized controlled clinical trial to study whether CYP2E1 inhibition improves ALD in the patients with alcohol use disorders admitted for alcohol detoxification therapy (ADT). Patients had to have a serum aspartate aminotransferase (AST) activity exceeding twice the upper normal limit at time of admission and be non-cirrhotic defined by fibroscan value <12 kPa. Sixty patients were randomly assigned to ADT with either CMZ or clorazepate (CZP) for 7-10 days in a 1:1 ratio. The chlorzoxazone test of CYP2E1 activity was performed at enrolment and at 2 points during the study.

ADT improved hepatic steatosis (controlled attenuation parameter) in both groups significantly. A trend towards a greater improvement in hepatic fat content during ADT (-21.5%) was observed in the CMZ group (252 ± 48 dB/m vs. 321 ± 38 dB/m; P < 0.0001) compared with the CZP group (-13.9%; 273 ± 38 dB/m vs. 317 ± 39 dB/m; P < 0.0001). As already reported, serum AST (P < 0.004) and alanine aminotransferase (ALT) activities (P < 0.0006) significantly decreased in CMZ patients as compared with patients on CZP by the end of hospitalization. A significant correlation was found between AST (P = 0.023), ALT (P = 0.009), GGT (P = 0.039) and CAP.

This study demonstrates that CMZ improves clinical biomarkers for ALD in humans most likely due to its inhibitory effect on CYP2E1. Because of its addictive potential, CMZ can only be given for a short period of time and therefore other CYP2E1 inhibitors to treat ALD are needed.

Alcoholic liver disease (ALD) due to chronic alcohol consumption is a significant global disease burden and a leading cause of mortality. Alcohol abuse induces a myriad of aberrant changes in hepatocytes at both the cellular and molecular level. Although the disease spectrum of ALD is widely recognized, the precise triggers for disease progression are still to be fully elucidated. Oxidative stress, mitochondrial dysfunction, gut dysbiosis and altered immune system response plays an important role in disease pathogenesis, triggering the activation of inflammatory pathways and apoptosis. Despite many recent clinical studies treatment options for ALD are limited, especially at the alcoholic hepatitis stage. We have therefore reviewed some of the key pathways involved in the pathogenesis of ALD and highlighted current trials for treating patients.

Complement is known to play a role in alcoholic fatty liver disease (AFLD), but the underlying mechanisms are poorly understood, thereby constraining the development of a rational approach for therapeutic intervention in the complement system. C3 deficiency has been shown to impart protective effects against ethanol-induced hepatic steatosis and inflammation. Here we demonstrate a protection effect in wild-type mice by treatment with CR2-Crry, a specific inhibitor of C3 activation. The expression of glycine transfer (t) RNA-derived fragments (Gly-tRFs) is upregulated in ethanol-fed mice and inhibition of Gly-tRFs in vivo decreases chronic ethanol feeding-induced hepatosteatosis without affecting inflammation. The expression of Gly-tRF was downregulated in C3-deficient or CR2-Crry-treated mice, but not in C5-deficient mice; Gly-tRF expression was restored by the C3 activation products C3a or Asp (C3a-des-Arg) via the regulation of CYP2E1. Transcriptome profiling of hepatic tissues showed that Gly-tRF inhibitors upregulate the expression of sirtuin1 (Sirt1) and subsequently affect downstream lipogenesis and β-oxidation pathways. Mechanistically, Gly-tRF interacts with AGO3 to downregulate Sirt1 expression via sequence complementarity in the 3' UTR. Notably, the expression levels of C3d, CYP2E1 and Gly-tRF are upregulated, whereas Sirt1 is decreased in AFLD patients compared to healthy controls. Collectively, our findings suggest that C3 activation products contribute to hepatosteatosis by regulating the expression of Gly-tRF. Complement inhibition at the C3 activation step and treatment with Gly-tRF inhibitors may be potential and precise therapeutic approaches for AFLD.

Massive acetaminophen (N-acetyl-p-aminophenol; APAP) ingestion is characterized by a rapid onset of mitochondrial dysfunction, including metabolic acidosis, lactemia, and altered mental status without hepatotoxicity which may not respond to the standard doses of N-acetylcysteine (NAC). A 64-year-old woman without medical history presented comatose after an ingestion of 208 tablets of Tylenol PM™ (APAP 500 mg and diphenhydramine 25 mg). The initial APAP concentration measured 1,017 µg/mL (therapeutic range 10-30 µg/mL), and elevated anion gap metabolic acidosis, lactemia, and 5-oxoprolinemia were detected. High-dose intravenous (IV) NAC, 4-methylpyrazole (4-MP), and hemodialysis (HD) were initiated. She was transferred to a liver transplant center and continued both NAC and HD therapies until complete resolution of metabolic acidosis and coma without developing hepatitis. She was discharged without sequelae. This is the fourth highest APAP concentration recorded in a surviving patient. Moreover, this is the first report of a novel "triple therapy" using NAC, 4-MP, and HD in the setting of massive APAP ingestion that presents with coma, elevated anion gap metabolic acidosis, and lactemia. Emergency physicians should recognize these critically ill patients and consider high-dose NAC, 4-MP, and HD to be initiated in the emergency department (ED).

Cytochrome p4502E1 (CYP2E1) autoantibodies are biomarkers for drug-induced hepatitis and chronic hepatitis C. However, major histocompatibility-restricted CYP2E1 epitopes associated with these diseases have not been identified. We hypothesized that CYP2E1 epitopes associated with different types of hepatitis may be shared and may impact immune responses and metabolism. SYFPEITHI epitope prediction identified CYP2E1 candidate epitopes that would be recognized by MHC II haplotypes. Candidate epitopes were tested for induction of hepatitis and CYP2E1 autoantibodies in mice and recognition by sera from patients with anesthetic drug-induced and viral hepatitis. Human liver cells treated with epitope hybridoma serum were analyzed for mitochondrial stress. CYP2E1 activity was measured in human microsomes similarly treated. Epitope antibodies in viral hepatitis sera were analyzed using linear regression to uncover associations with liver pathology. A P value of <0.05 was considered significant. One epitope (Gly113-Leu135) induced hepatitis and CYP2E1 autoantibodies in mice after modification of Lys123 (P < 0.05). Gly113-Leu135 antiserum recognized mitochondria and endoplasmic reticula (P < 0.05), upregulated HSP27 (P < 0.01) and mitochondrial oxidative stress via complex 1 inhibition (P < 0.001), and inhibited CYP2E1 activity. Gly113-Leu135 IgG4 detected in viral hepatitis sera was associated with severe hepatic fibrosis (P = 0.0142). We found a novel CYP2E1 epitope that was detected in anesthetic and viral hepatitis and that triggered hepatitis in mice. Our findings may improve understanding of hepatic immune responses triggered by metabolism or viruses.IMPORTANCE Drug-induced hepatitis is the leading reason that an approved drug is removed from the commercial market. Halogenated anesthetics can induce hepatitis in susceptible persons, and cytochrome p4502E1 (CYP2E1) enzymes responsible for their metabolism induce antibodies in addition to hepatitis. CYP2E1 antibodies detected in anesthetic hepatitis patients have been detected in patients with viral hepatitis, suggesting that these different forms of hepatitis could develop immune reactions to a common segment or epitope of CYP2E1. We have found a common MHC-restricted CYP2E1 epitope in anesthetic and viral hepatitis that is a dominant epitope in anesthetic hepatitis and is significantly associated with fibrosis in patients with viral hepatitis. Along with conformational epitopes, our identification of MHC-restricted CYP2E1 epitopes can be used to develop specific diagnostic tests for drug-induced or viral hepatitis or associated fibrosis or to predict individuals at risk for developing these diseases or their sequelae.

Excessive alcohol consumption is a global healthcare problem. The liver sustains the greatest degree of tissue injury by heavy drinking because it is the primary site of ethanol metabolism. Chronic and excessive alcohol consumption produces a wide spectrum of hepatic lesions, the most characteristic of which are steatosis, hepatitis, and fibrosis/cirrhosis. Steatosis is the earliest response to heavy drinking and is characterized by the deposition of fat in hepatocytes. Steatosis can progress to steatohepatitis, which is a more severe, inflammatory type of liver injury. This stage of liver disease can lead to the development of fibrosis, during which there is excessive deposition of extracellular matrix proteins. The fibrotic response begins with active pericellular fibrosis, which may progress to cirrhosis, characterized by excessive liver scarring, vascular alterations, and eventual liver failure. Among problem drinkers, about 35 percent develop advanced liver disease because a number of disease modifiers exacerbate, slow, or prevent alcoholic liver disease progression. There are still no FDA-approved pharmacological or nutritional therapies for treating patients with alcoholic liver disease. Cessation of drinking (i.e., abstinence) is an integral part of therapy. Liver transplantation remains the life-saving strategy for patients with end-stage alcoholic liver disease.

Enhanced drug elimination in alcoholics remains largely indefinable. In contrast, the reduced elimination of drugs in patients with advanced alcoholic liver disease (ALD) is normally owing to hepatic end-stage disease such as cirrhosis. We here study the mRNA expression of various hepatic drug metabolizing enzymes and transporters in association with liver stiffness (LS) being a novel noninvasive parameter for the assessment of cirrhosis to unravel the dynamic relationship between ALD and determinants of pharmacokinetics such as drug metabolizing enzymes and transporters.

We quantified mRNA expression levels of various cytochrome P-450 isoenzymes (CYPs) and drug transporters in 26 liver specimens of chronic alcoholics and 5 controls by quantitative polymerase chain reaction. In addition, liver histology, clinical data, and LS evaluated by transient elastography (Fibroscan) were obtained.

Eighteen patients had a normal or moderate LS < 8 kPa (69.2%), while in the remaining 8 patients (30.7%) advanced F3 or F4 fibrosis could be established with an LS > 8 kPa. Overall, CYP3A4, CYP2E1, and solute carrier organic anion transporter 1B1 (SLCO1B1) were negatively correlated with increasing LS. CYPs and drug transporters tended to be up-regulated in alcoholics without advanced fibrosis (LS < 8.0 kPa) compared to healthy controls supporting data of boosted drug elimination in alcoholics without advanced ALD. However, in alcoholics with severely increased LS (>8 kPa), expression levels of CYP2E1, SLC22A2, and SLCO1B1 were significantly lower.

In conclusion, CYPs and drug transporters seem to be induced in chronic alcoholics without irreversible liver damage but decline in case of manifest cirrhosis. Our study also suggests that noninvasive measurements of LS could be useful for pharmacokinetic predictions and individualized pharmacotherapy.

To validate the induction of blood lymphocyte cytochrome P450 2E1 (CYP2E1) expression in alcoholic liver cirrhosis and mRNA and protein expression of CYP2E1 in freshly prepared blood lymphocytes of alcoholic liver cirrhotic (ACP), nonalcoholic cirrhotic patients (NACP), alcoholic controls (ACs), and nonalcoholic controls (NACs) were investigated. Registered ACP and NACP patients at Sanjay Gandhi Postgraduate Institute of Medical Science, Lucknow, India along with NACs and ACs were included in the study. Real time polymerase chain reaction, enzyme-linked immunosorbent assay, and CYP2E1-dependent enzyme activity were determined in blood lymphocytes isolated from cases and controls. Significant increases in CYP2E1 mRNA and protein expression were observed in freshly prepared blood lymphocytes isolated from ACs and ACP patients as compared with respective NACs or NACP patients. A concomitant increase in N-nitrosodimethyamine demethylase activity was evident in the blood lymphocytes of ACs and ACP patients. Interestingly, the comparative increase observed in CYP2E1 expression was of greater magnitude in the blood lymphocytes isolated from ACP patients, although they abstained from alcohol drinking. Findings suggest that significant increase in the CYP2E1 mRNA and protein expression in the blood lymphocytes, isolated from early stage ACP patients, can be used to predict alcohol-induced toxicity.

Nowadays, there are numerous commercial applications of microalgae, and they have been used to enhance the nutritional value of food and animal feed owing to their chemical composition. They are cultivated as a source of highly nutritional and valuable source. Recently, microalgae have been reported to use as a potent source for food additive, nutraceutical, or pharmaceuticals. According to the criteria of nutritional quality and cost, variety of marine organisms has been investigated for their suitability to be applied in the production of protein hydrolysates in functional foods. Recently, a great deal of interest has been expressed regarding marine-derived bioactive peptides because of their numerous health benefits. In addition, many studies have been reported that marine bioactive peptides can be used as functional foods, nutraceuticals, or pharmaceuticals due to their therapeutic potential in the treatment or prevention of various diseases. Hence, in this chapter, we discussed the importance of marine microalgae in relation to their medicinal value.

The present paper is an update of the data on the effects of diseases and environmental factors on the expression and/or activity of human cytochrome P450 (CYP) enzymes and transporters. The data are presented in tabular form (Tables 1 and 2) and are a continuation of previously published summaries on the effects of drugs and other chemicals on CYP enzymes (Rendic, S.; Di Carlo, F. Drug Metab. Rev., 1997, 29(1-2), 413-580., Rendic, S. Drug Metab. Rev., 2002, 34(1-2), 83-448.). The collected information presented here is as stated by the cited author(s), and in cases when several references are cited the latest published information is included. Inconsistent results and conclusions obtained by different authors are highlighted, followed by discussion of the major findings. The searchable database is available as an Excel file, for information about file availability contact the corresponding author.

To investigate associations between the Rsa I polymorphism of CYP2E1 and risk of colorectal cancer.

A case-control study was conducted with 315 colorectal cancer cases (105 colon, 210 rectal) and 439 population-based controls in Jiangsu Province of China. Genomic DNA samples were assayed for restriction fragment length polymorphisms in CYP2E1 by PCR amplification followed by digestion with Rsa I. Information on smoking and alcohol drinking was collected using a questionnaire. Odds ratios (ORs) were estimated with an unconditional logistic model.

The proportional distribution of the CYP2E1 Rsa I c1/c1, c1/c2 and c2/c2 genotypes were 61.4%, 35.6% and 3.0% in controls, 60.6%, 33.7% and 5.8% in colon cancer cases, and 58.4%, 34.0% and 7.7% in rectal cancer cases, respectively. A significant difference was noted between controls and rectal cancer cases (P = 0.029), the c2/c2 genotype being associated with elevated OR (adjusted age, sex and status of the smoking and alcohol drinking) for rectal cancer (1.64, 95% CI, 1.12-2.41, vs c1 allele carriers), but not for colon cancer. In interaction analysis between the CYP2E1 Rsa I genotype and smoking and drinking habits, we found a significant cooperative action between the c2/c2 genotype and alcohol drinking in the sex-, age-adjusted ORs for both colon (4.74, 95% CI, 1.10-20.40) and rectal (5.75, 95% CI, 1.65-20.05) cancers. Among non-smokers, the CYP2E1 Rsa I c2/c2 genotype was also associated with elevated ORs in the two sites (1.95, 95% CI, 0.99-3.86 and 2.30, 95% CI, 1.32-3.99).

The results of the present study suggest that the CYP2E1 c2/c2 genotype increases susceptibility to rectal cancer and the gene-environmental interactions between the CYP2E1 polymorphism and smoking or alcohol drinking exist for colorectal neoplasia in general.

Advanced glycation end products (AGEs), e.g., carboxymethyllysine (CML) or imidazolone are involved in several age-related disorders. Concerning their accumulation, the importance of hepatic and renal function is controversially discussed. To test whether impairment of hepatic or renal function will affect their accumulation, both AGEs have been measured in various populations, such as 52 patients with liver disease [viral hepatitis C without (n = 19) and with (n = 10) fatty liver; nonalcoholic fatty liver (n = 13), nonalcoholic steatohepatitis (n = 10)]. Serum concentrations of both AGEs have been compared to those in 20 healthy controls and 24 patients with moderate renal impairment (creatinine clearance 23-55 ml/min). Concerning CML (95% C.I. 803-1200 ng/ml), no differences between the various groups could be observed. Likewise, serum levels of imidazolone (95% C.I. 1.3-5.6 units) were similar in all populations. In conclusion, moderate impairment in hepatic or in renal function did not affect serum levels of CML and imidazolone. Apparently, any increase observed in severe cirrhosis or renal failure seems to be rather a consequence than a cause of both disorders.

A physiologically based pharmacokinetic model for investigating inter-individual and inter-racial variability in ethanol pharmacokinetics is presented. The model is a substantial modification of an existing model which described some genetic polymorphisms in the hepatic alcohol dehydrogenase enzymes. The model was modified to incorporate a description of ethanol absorption from the stomach and gastro-intestinal tract and the retardation of gastric emptying due to a concentration-dependent inhibition of gastric peristalsis. In addition, intra-venous and intra-arterial routes of administration were added to investigate whether the biological structure of the model provided a core which may be easily adapted for any route of exposure. The model is proposed as suitable for the investigation of the effects of both acute and chronic ethanol exposure.

The cytochrome P450 (P450) CYP2E1 enzyme metabolizes and activates a wide array of toxicological substrates, including alcohols, the widely used analgesic acetaminophen, acetone, benzene, halothane, and carcinogens such as azoxymethane and dimethylhydrazine. Most studies on the biochemical and pharmacological actions of CYP2E1 are derived from studies with rodents, rabbits, and cultured hepatocytes; therefore, extrapolation of the results to humans can be difficult. Creating "humanized" mice by introducing the human CYP2E1 gene into Cyp2e1-null mice can circumvent this disadvantage. A transgenic mouse line expressing the human CYP2E1 gene was established. Western blot and high-performance liquid chromatography/mass spectrometry analyses revealed human CYP2E1 protein expression and enzymatic activity in the liver of CYP2E1-humanized mice. Treatment of mice with the CYP2E1 inducer acetone demonstrated that human CYP2E1 was inducible in this transgenic model. The response to the CYP2E1 substrate acetaminophen was explored in the CYP2E1-humanized mice. Hepatotoxicity, resulting from the CYP2E1-mediated activation of acetaminophen, was demonstrated in the livers of CYP2E1-humanized mice by elevated serum alanine aminotransferase levels, increased hepatocyte necrosis, and decreased P450 levels. These data establish that in this humanized mouse model, human CYP2E1 is functional and can metabolize and activate different CYP2E1 substrates such as chlorzoxazone, p-nitrophenol, acetaminophen, and acetone. CYP2E1-humanized mice will be of great value for delineating the role of human CYP2E1 in ethanol-induced oxidative stress and alcoholic liver damage. They will also function as an important in vivo tool for predicting drug metabolism and disposition and drug-drug interactions of chemicals that are substrates for human CYP2E1.

Acetaminophen has been used safely and effectively for many years to manage pain and/or fever in patients of all ages. It is commonly recommended as first-line therapy for a variety of patients and conditions, including the elderly, children with viral illnesses, and patients with osteoarthritis, gastrointestinal conditions, bleeding disorders, cardiovascular disease, or renal disease. However, its use is often avoided in patients with chronic liver disease. The perception that acetaminophen should be avoided in such patients arose from awareness of the association between massive acetaminophen overdose and hepatotoxicity, combined with a lack of understanding of the metabolism of acetaminophen in patients with liver disease. There are various theoretical mechanisms of acetaminophen hepatotoxicity in chronic liver disease including: altered metabolism and depleted glutathione stores that would be expected to increase accumulation of the hepatotoxic intermediate, N-acetyl-p-benzoquinone imine (NAPQI). Available studies in patients with chronic liver disease, however, have shown that although the half-life of acetaminophen may be prolonged, cytochrome P-450 activity is not increased and glutathione stores are not depleted to critical levels in those taking recommended doses. Furthermore, acetaminophen has been studied in a variety of liver diseases without evidence of increased risk of hepatotoxicity at currently recommended doses. Therefore, acetaminophen can be used safely in patients with liver disease and is a preferred analgesic/antipyretic because of the absence of the platelet impairment, gastrointestinal toxicity, and nephrotoxicity associated with nonsteroidal antiinflammatory drugs.

Chlorzoxazone is metabolized by cytochrome P450 2E1 (CYP2E1) to a single oxidized metabolite, 6-hydroxychlorzoxazone. The aim of the study was to test the robustness of chlorzoxazone as an in vivo probe of CYP2E1 activity in humans, with emphasis on investigating short-term and long-term intra-individual variabilities and effects of different doses of the drug. In addition, the influences of body build, drug metabolizing enzyme genotype, blood sampling time, and moderate recent ethanol intake were investigated.

The 6-hydroxychlorzoxazone:chlorzoxazone (metabolic) ratio in plasma was measured at 2 h in 28 male and nine female volunteers following a single oral dose of 500 mg chlorzoxazone. Similarly, the metabolic ratios at 4 h and 6 h were measured in 20 of the males. The metabolic ratio at 2 h was also determined 1.5 and 2.5 years later in 13 and seven males, respectively, and weekly for 3 weeks in seven males, after a dose of 500 mg, once at higher (750 mg) and lower (250 mg) doses, and once (500 mg) following moderate ethanol intake (0.5 g kg(-1) body weight) the preceding evening. Genotypes were determined for CYP2E1 as well as for N-acetyltransferase 2 and glutathione transferase M1.

Excluding an outlier (ratio = 1.6) the metabolic ratio at 2 h ranged from 0.12 to 0.61 (n = 36). A positive correlation with body weight (r = 0.61, P < 0.001) suggested dose-dependent metabolism of chlorzoxazone. The metabolic ratio decreased with increasing chlorzoxazone dose (P = 0.01), again suggesting dose-dependent metabolism. Long-term (yearly intervals) and short-term (weekly intervals) intra- and interindividual variabilities in metabolic ratio were similar (30% and 63%vs 28% and 54%, respectively). Both inter- and intra-individual variabilities tended to decrease with increasing dose of chlorzoxazone. There was no significant influence of moderate ethanol intake the preceding evening, or of CYP2E1 genotype on the metabolic ratio.

The relatively low intra-individual variability in the metabolism of chlorzoxazone suggests that a single-sample procedure may suffice to assess CYP2E1 activity in vivo. However, chlorzoxazone metabolism is dose-dependent at commonly used doses and it is therefore advisable to adjust the dose for body weight. Moderate intake of ethanol the preceding evening did not significantly affect the chlorzoxazone metabolic ratio.

Examination of the pharmacokinetics of acetaminophen can decrease misconceptions involved in clinical evaluation. Enzyme patterns and acetaminophen levels must be related to time and known metabolic phenomena. A careful look at ethanol and nutrition, especially fasting demonstrates that therapeutic doses of acetaminophen do not place patients at a greater risk in either of these instances. An overdose of acetaminophen in a chronic alcohol abuser may result in more severe hepatotoxicity than in the nonalcoholic. CYP2E1 and glutathione must be evaluated simultaneously rather than in isolation. Glucuronidation capacity in humans is not a factor except in massively overdosed patients.

Cytochrome P450-2E1 (CYP2E1) is one of the major hepatic enzymes involved in the metabolism of procarcinogen. Our study aimed to investigate the differential expression level of CYP2E1 and its clinicopathological significance in hepatocellular carcinoma (HCC). CYP2E1 revealed low level of expression in 70% of the tumor tissues, when compared to the adjacent nontumor tissues, at both mRNA and protein levels. The low expression of CYP2E1 was significantly correlated with the aggressive tumor phenotype, including poor differentiation status (by the Edmondson grading system) (p=0.038), absence of tumor capsule (p=0.030) and younger age of the patients (p=0.002). Multivariate analysis indicated that CYP2E1 expression level and pTNM stage were independent prognostic factors for disease-free survival. CYP2E1 was also shown to have a differential expression level in different liver tissues. The level of CYP2E1 was significantly higher in nontumor tissues from HCC patients compared to the intermediate level in cirrhosis livers from noncancer patients and normal livers from healthy persons. Tumor tissues were shown to have the lowest expression level. In conclusion, our results have shown that CYP2E1 is upregulated in the nontumor tissue and downregulated in tumor tissue, which is associated with aggressive tumor type and poor prognosis of the patients. It suggested that the differential expression of CYP2E1 may play an important role in HCC tumorigenesis.

Alcohol and xenobiotics share the same oxidative microsomal pathway, which is mainly located in the endoplasmic reticulum of hepatocytes. This pathway involves enzymes that belong to the super family of cytochrome P450 and allows to explain a lot of pharmacokinetic or toxic interactions between alcohol and xenobiotics. Cytochrome P450 2E1 (CYP2E1) is the key enzyme of the microsomal pathway of ethanol oxidation. It is inducible by chronic ethanol consumption and its activity is increased by three to five fold in liver from alcoholics subjects. This induction involves to a lesser extent cytochromes P450 3A4 and 1A2 and contributes to the metabolic tolerance of alcohol and drugs observed in alcoholics. The metabolic tolerance persits several days after ethanol withdrawal. Furthermore, CYP2E1 has a high capacity to activate numerous xenobiotics into toxic or carcinogenic compounds. Drugs currently used such as paracetamol, anesthetics (enflurane, halothane), industrial solvents (benzene or its derivatives), halogenated solvents (CCl4, trichlorethylene) and nitrosamines which are present in food or tobacco smoke are included. Therefore, heavy consumption of alcohol, which results in CYP2E1 induction, increases individual susceptibility to the toxic or carcinogenic effects of these xenobiotics.

Humans are exposed to over 70,000 man-made chemicals including drugs, food additives, herbicides, pesticides, and industrial agents. It is well established that environmental chemicals are the cause of numerous human diseases including cancer. In most cases, chemical carcinogens require metabolic activation, which is mainly achieved by P450s enzymes. CYP2E1 is of clinical relevance because it is inducible by ethanol, and it metabolizes many common organic solvents such as benzene, alcohols and halogenated solvents. Therefore, alteration in the level of CYP2E1 might influence the health effects of the environmental pollutants. This hypothesis needs to be validated by epidemiological studies and the objective of the "Biomed-2" project was to develop new tests to assess the individual metabolic capacity of workers exposed to volatile organic compounds in order to predict their occupational risk. In vivo chlorzoxazone 6-hydroxylation was validated as a non-invasive and selective test for the determination of liver CYP2E1 activity. Preliminary data in workers exposed to organic solvents indicated that chlorzoxazone metabolism may be a biomarker of occupational exposure to organic solvents. Other approaches, such as use of salicylate as catalytic probe or measurement of catalytic activity in lymphocytes, were not conclusive. Attempts to use CYP2E1 genotyping for estimating human risks from chemical exposure did not bring convincing data as genetic polymorphism of CYP2E1 could not be clearly related to its catalytic activity.

The effects of acute administration of dietary levels of ethanol and the garlic oil extract, diallyl sulphide (DAS), on cytochrome P450 2E1 (CYP2E1) activity in volunteers were studied using the selective probe substrate, chlorzoxazone (CZX). The ratio of the CZX metabolite 6- hydroxychlorzoxazone (6-OHCZX) to CZX was taken to indicate CYP2E1 activity. The mean differences between the baseline and DAS-treated (0.2 mg/kg) CYP2E1 activities were significantly different (two-tailed p value = 0.0242, n = 8). Likewise, the mean differences between the baseline and ethanol-treated (0.8 g/kg) CYP2E1 activities were also significantly different (two-tailed p value = 0.0005, n = 7). The reduction in in vivo CYP2E1 activity by DAS is consistent with reported inhibition observed in vitro. The marked reduction in CYP2E1 activity following acute ingestion of ethanol is consistent with a competitive inhibition mechanism of CZX metabolism. The inhibitory effect of DAS maybe additive with daily consumption of Allium vegetables in particular. This may explain the lower 6-OHCZX/CZX metabolic ratios measured in various European and Mexican cohorts and is consistent with the lower incidence of stomach, liver and colon cancers observed in southern Europeans.

Cytochrome (CYP) P450 2E1 is clinically and toxicologically important and it is constitutively expressed in the liver and many other tissues. In contrast to many other CYP isoenzymes, indisputable evidence for a functionally important polymorphism of CYP2E1 in the human population is lacking. CYP2E1 metabolizes a wide variety of chemicals with different structures, in particular small and hydrophobic compounds, including potential cytotoxic and carcinogenic agents. In addition, chlorzoxazone and trimethadione metabolism are good CYP2E1 probes for liver disease in vivo and in vitro. In the future, methods for fully analysing the function of CYP2E1 using knockout mice will be established. This article reviews recent advances in our understanding of the role of human CYP2E1 in drug metabolism.

It is claimed that chronic alcoholics are at increased risk of paracetamol (acetaminophen) hepatotoxicity not only following overdosage but also with its therapeutic use. Increased susceptibility is supposed to be due to induction of liver microsomal enzymes by ethanol with increased formation of the toxic metabolite of paracetamol. However, the clinical evidence in support of these claims is anecdotal and the same liver damage after overdosage occurs in patients who are not chronic alcoholics. Many alcoholic patients reported to have liver damage after taking paracetamol with 'therapeutic intent' had clearly taken substantial overdoses. No proper clinical studies have been carried out to investigate the alleged paracetamol-alcohol interaction and acute liver damage has never been produced by therapeutic doses of paracetamol given as a challenge to a chronic alcoholic. The paracetamol-alcohol interaction is complex; acute and chronic ethanol have opposite effects. In animals, chronic ethanol causes induction of hepatic microsomal enzymes and increases paracetamol hepatotoxicity as expected (ethanol primarily induces CYP2E1 and this isoform is important in the oxidative metabolism of paracetamol). However, in man, chronic alcohol ingestion causes only modest (about twofold) and short-lived induction of CYP2E1, and there is no corresponding increase (as claimed) in the toxic metabolic activation of paracetamol. The paracetamol-ethanol interaction is not specific for any one isoform of cytochrome P450, and it seems that isoenzymes other than CYP2E1 are primarily responsible for the oxidative metabolism of paracetamol in man. Acute ethanol inhibits the microsomal oxidation of paracetamol both in animals and man. This protects against liver damage in animals and there is evidence that it also does so in man. The protective effect disappears when ethanol is eliminated and the relative timing of ethanol and paracetamol intake is critical. In many of the reports where it is alleged that paracetamol hepatotoxicity was enhanced in chronic alcoholics, the reverse should have been the case because alcohol was actually taken at the same time as the paracetamol. Chronic alcoholics are likely to be most vulnerable to the toxic effects of paracetamol during the first few days of withdrawal but maximum therapeutic doses given at this time have no adverse effect on liver function tests. Although the possibility remains that chronic consumption of alcohol does increase the risk of paracetamol hepatotoxicity in man (perhaps by impairing glutathione synthesis), there is insufficient evidence to support the alleged major toxic interaction. It is astonishing that clinicians and others have unquestion-ingly accepted this supposed interaction in man for so long with such scant regard for scientific objectivity.