Induction Of CYP2E1 Research Articles

Alleles of alcohol and acetaldehyde metabolism genes modulate susceptibility to oesophageal cancer from alcohol consumption

Alleles of alcohol and acetaldehyde metabolism genes modulate susceptibility to oesophageal cancer from alcohol consumption

Ethanol-induced cytochrome P450 2E1 causes carcinogenic etheno-DNA lesions in alcoholic liver disease

Aims: Oxidative stress is thought to play a major role in the pathogenesis of hepatocellular cancer (HCC), the most frequent complication of alcoholic liver disease (ALD). However, its underlying mechanisms are poorly understood. In hepatocytes of patients with ALD, we could recently detect significant levels of carcinogenic exocyclic etheno-DNA adducts that are formed by the reaction of the lipid peroxidation product 4-hydroxynonenal (4-HNE) with nucleobases. Methods and Results: We here show in patients with ALD that 4-HNE and exocyclic etheno-DNA adducts closely correlate with the expression of CYP2E1 expression induced by chronic consumption of alcohol (r=0.93, p<0.01). Increased amounts of exocyclic DNA-adducts were also detectable in the livers of alcohol-fed lean Zucker rats (fa/fa-) and the number of positive nuclei correlated with CYP2E1-expression (r=0.63, p=0.03). To further assess the role of CYP2E1 in the formation of etheno-DNA adducts, HepG2 cells stably transfected with CYP2E1 or vector mock transfected control cells were treated with different concentrations of ethanol with or without the CYP2E1 inhibitor chlormethiazole (CMZ). Ethanol treatment increased etheno-DNA adducts in the nuclei of CYP2E1-transfected HepG2 cells in a dose and time dependent manner, but not in control cells. In addition, CMZ almost completely blocked the generation of etheno-DNA adducts (p<0.01). Conclusion: CYP2E1 plays an important role in ethanol-induced DNA damage. Therefore ethanol-mediated induction of hepatic CYP2E1 may be a central mechanism in HCC development in patients with ALD.

Chronic Alcohol Feeding Stimulates the Formation of Mutagenic Etheno-Adducts in a Rodent model of NASH Via Induction of Cytochrome P450 2E1

Background and Aims: Chronic alcohol consumption may cause liver fibrosis and hepatocellular carcinoma (HCC) in non-alcoholic fatty liver disease (NAFLD). Synergistic effects of insulin resistance and alcohol triggering fibrogenesis and HCC are implicated. Formation of mutagenic etheno adducts binding to adenosin (EdA) generated via induced CYP2E1 has been suggested as a key mechanisms of hepatocarcinogenesis. Methods: Obese Zucker rats and lean littermates (4 groups) received Lieber-De Carli liquid diets containing either ethanol or carbohydrates for 3 months. Fibrosis and steatosis were quantified by Sirius Red and Oil Red O, respectively. EdA immunohistology was performed using a monoclonal antibody, and fibrosis-related gene and protein expression and hydroxyproline quantified. CYP2E1 and 4A2 were assayed at transcription and protein level. Results: Obese rats had more steatosis than lean animals but showed no aggravating impact of chronic alcohol intake. Histology and liver hydroxyproline showed similar extent of fibrosis in all groups. Procollagen alpha1(I), alpha smooth muscle actin, and TGFbeta-1 gene expression were not different. CYP2E1 protein was induced by 35% in both groups of obese rats, while corresponding mRNA levels were upregulated 3.5-fold in alcoholic lean rats. CYP4A2 was lower in obese rats, but upregulated 2-fold in those fed alcohol. In alcoholic obese and lean rats hepatocyte nuclei stained positive for EdA in 36% and 23%, respectively, but only 14% in obese rats without alcohol. EdA immunostaining was absent in lean non-alcoholic rats. EdA labelling correlated positively with CYP2E1 and 4-HNE immunostaining. Conclusions: Chronic alcohol feeding in obese, insulin-resistant rats exerts no effect on liver fibrosis formation, but potentiates the formation of mutagenic EdA possibly due to CYP2E1 induction. This observation may explain the increased risk of developing HCC in patients with NAFLD who chronically drink alcohol.

Saponins isolated from the root of Platycodon grandiflorum protect against acute ethanol-induced hepatotoxicity in mice

The protective effects of saponins isolated from the root of Platycodon grandiflorum (Changkil saponins: CKS) against alcoholic steatosis in liver injury induced by acute ethanol administration were investigated. Pretreatment with CKS prior to ethanol administration significantly prevented the increases in serum alanine aminotransferase activity, hepatic TNF-α level, hepatic lipid peroxidation and hepatic triglyceride level. CKS prevented ethanol-induced steatosis and necrosis, as indicated by liver histopathological studies. Additionally, CKS protected against ethanol-induced depletion of hepatic glutathione levels. CYP2E1 has been suggested as a major contributor to ethanol-induced oxidative stress and liver injury. The concurrent administration of CKS efficaciously abrogated the CYP2E1 induction and CYP2E1-dependents hydroxylation of aniline as compared to the individual treatment at higher doses. These findings suggest that CKS may prevent ethanol-induced acute liver injury, possibly through its ability to block CYP2El-mediated ethanol bioactivation and its free radical scavenging effects.

Role of nitric oxide and nuclear factor-κB in the CYP2E1 potentiation of tumor necrosis factor α hepatotoxicity in mice

Induction of CYP2E1 by pyrazole (PY) potentiated the hepatotoxicity induced by TNFα in mice. We evaluated the role of nitrosative and oxidative stress and the NF-κB activation pathway in this liver injury. The iNOS inhibitor N-(3-aminomethyl)benzylacetamindine (1400W) or the antioxidant N-acetyl-l-cysteine (NAC) prevented this liver injury. TNFα plus PY treatment triggered radical stress in the liver with increased lipid peroxidation and decreased glutathione and caused mitochondrial damage as reflected by elevated membrane swelling and cytochrome c release. The radical stress and mitochondrial damage were prevented by 1400W and NAC. TNFα plus PY treatment elevated 3-nitrotyrosine adduct formation and induced NOS2 in the liver; 1400W and NAC blocked these changes. A lower extent of liver injury and oxidative stress was found in NOS2 −/− mice treated with TNFα plus PY compared with wild-type controls. Neither 1400W nor NAC modified CYP2E1 activity or protein. Activation of JNK and p38MAPK was weaker in TNFα plus PY-treated NOS2 −/− mice and 1400W and NAC blocked the activation of JNK and p38MAPK in wild-type mice. IKKα/β protein levels were decreased by TNFα plus PY treatment, whereas IκBα and IκBβ protein levels were elevated compared with saline, PY, or TNFα alone. NF-κB DNA binding activity was increased by TNFα alone but lowered by TNFα plus PY. All these changes were blocked by 1400W and NAC. NF-κB activation products such as Bcl-2, Bcl-X L, cFLIP S, cFLIP L, and Mn-SOD were reduced by TNFα plus PY and restored by 1400W or NAC. We conclude that TNFα plus CYP2E1 induces oxidative/nitrosative stress, which plays a role in the activation of JNK or p38MAPK and mitochondrial damage. These effects combine with the blunting of the NF-κB activation pathways and the synthesis of protective factors to cause liver injury.

Induction of brain CYP2E1 changes the effects of ethanol on dopamine release in nucleus accumbens shell

CYP2E1 is an important enzyme involved in the brain metabolism of ethanol that can be induced by chronic consumption of alcohol. Recent works have highlighted the importance of this system in the context of the behavioural effects of ethanol. Unfortunately, the underlying neurochemical events for these behavioural changes, has not been yet explored. In this work, we have started this exploration by analyzing the possible changes in the neurochemical response of the mesolimbic system to ethanol after pharmacological induction of brain CYP2E1. We have used the dopamine extracellular levels in nucleus accumbens (NAc) core and shell, measured by means of microdialysis in vivo, as an index of the effects of ethanol. Acetone 1% in the tap water was used to induce brain CYP2E1. Efficacy of the induction protocol was assessed by immunoblotting. Intravenous administration of 1.5 g/kg of ethanol in control rats provoked a significant increase of the dopamine levels in both the core (up to 127% of baseline) and the shell (up to 122% of baseline) of the NAc. However, the same dose of ethanol in acetone-treated rats only increased the dopamine extracellular levels in the core (up to 142% of baseline) whereas dopamine levels in the shell subregion remain unaltered relative to baseline. The results of this study indicate that induction of CYP2E1 changes the response of the mesolimbic system to ethanol in a region-dependent manner. Two hypotheses are postulated to explain the observed effects.

Effects of Oral Rutaecarpine on the Pharmacokinetics of Intravenous Chlorzoxazone in Rats.

It has been reported that hepatic microsomal cytochrome P450 (CYP) 2E1 is responsible for the metabolism of chlorzoxazone (CZX) to 6-hydroxychlorzoxazone. The present study was undertaken to assess the possible interaction of rutaecarpine, an alkaloid originally isolated from the unripe fruit of Evodia rutaecarpa, with CZX. Male Spraque-Dawley rats were administered with 80 mg/kg/day of oral rutaecarpine for three consecutive days. Twenty four hr after the pre-treatment with rutaecarpine, the rats were treated with 20 mg/kg of intravenous CZX Rat hepatic microsomes isolated from rutaecarpine-treated rats showed greater (50% increase) activity of p-nitrophenol hydroxylase (a marker of CYP2E1) when compared with the control rats. Compared with control rats, the AUC of CZX was significantly smaller (84% decrease) possibly due to significantly faster CL (646% increase) in rats pretreated with rutaecarpine. This could be, at least partially, due to induction of CYP2E1 by rutaecarpine.

Inhibition of rat hepatic CYP2E1 by quinacrine: molecular modeling investigation and effects on 4-(methyl nitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced mutagenicity

Increased activity of CYP2E1 has been associated with increased risk of chemically-mediated cancers, through enhanced activation of a variety of procarcinogens. In this context, inhibition of CYP2E1 is potentially of significance in xenobiotic toxicity. The aim of the present study was to test the hypothesis that quinacrine inhibits hepatic CYP2E1. For this purpose, disulfiram (75 mg/kg i.p) as an inhibitor and isoniazid (100 mg/kg i.p) as an inducer of CYP2E1, as well as quinacrine (50 mg/kg i.p) were administered to Wistar rats and the hepatic activity of CYP2E1 was measured. The expression of CYP2E1 was further assessed by Western blot analysis. As expected, disulfiram inhibited, while isoniazid induced the activity and expression of the enzyme. Interestingly, treatment with quinacrine resulted in a significant decrease of CYP2E1 activity and expression. To investigate any similarities in the inhibition of CYP2E1 by quinacrine and disulfiram, molecular modeling techniques were adopted and revealed that quinacrine molecule anchors inside the same binding pocket of the protein where disulfiram is also attached. Finally, as assessed by the sister chromatid exchanges (SCE) assay, quinacrine was demonstrated to reduce the mutagenic effects of the tobacco-specific N-nitrosamine 4-(methyl nitrosamino)-1-(3-pyridyl)-1-butanone (NNK), which is known to be converted to active mutagen in the liver principally through CYP2E1. We suggest that these antimutagenic effects of quinacrine could be possibly attributed, at least in part, to its ability to block the bioactivation of NNK, mainly by the inhibition of CYP2E1. Our results, even preliminary, indicate that quinacrine as an inhibitor of CYP2E1 might be protective against chemically-induced toxicities such as NNK-induced mutagenicity.

Induction of several cytochrome P450 genes by doxorubicin in H9c2 cells

Doxorubicin is a potent anti-neoplastic antibiotic used to treat a wide variety of malignancies; however, its use is limited by dose dependent cardiotoxicity. There is indirect evidence suggesting that doxorubicin cardiotoxicity is CYP-mediated. In the current study, we investigated the effect of doxorubicin on hypertrophic markers, and different CYP gene expression in cardiac derived H9c2 cells. H9c2 cells were incubated with increasing concentrations of doxorubicin and the expressions of different genes were determined by real-time PCR. Our results demonstrate that multiple CYP genes are expressed in H9c2 cells and the level of expression from the highest to the lowest were; CYP1B1, CYP2B1, CYP2J3, CYP1A1, CYP2C11, CYP2C23, CYP2E1, CYP1A2, and CYP2B2. Doxorubicin treatment caused an induction of the hypertrophic markers, ANP and BNP. In addition, doxorubicin caused a significant induction of CYP1A1, CYP1A2, CYP1B1, CYP2B2, CYP2E1, and CYP2J3 gene expression in a concentration-dependent manner. However, only the highest concentration tested, 10 μM, caused an induction of CYP2C11; whereas, CYP2B1 and CYP2C23 were not altered. Our findings demonstrate that doxorubicin induces the hypertrophic markers, ANP and BNP as well as several CYP genes in H9c2 cells. Doxorubicin-mediated CYP induction may represent a novel mechanism by which this drug induces cardiotoxicity.

Growth retardation of fetal rats exposed to nicotine in utero: Possible involvement of CYP1A1, CYP2E1, and P‐glycoprotein

To elucidate the possible metabolic mechanism of intrauterine growth retardation induced by nicotine, this study determines the effects of prenatal nicotine exposure on fetal development and cytochrome P4501A1 (CYP1A1), CYP2E1, and P-glycoprotein (Pgp) expression in maternal liver and placenta. Pregnant rats were given 1.0 mg/kg nicotine subcutaneously twice a day from gestational day (GD) 8 to GD 15, 18, or 21. In nicotine-treated groups, fetal developmental parameters including body weight were significantly lower. The activities of CYP1A1 and CYP2E1 in maternal liver microsomes in nicotine-treated groups increased significantly with progressing gestation when compared with the corresponding control, but returned to the level similar to the control in late pregnancy. Nicotine-treated groups induced pathological changes and increased malondialdehyde (MDA) content in the placenta when compared with the control. The gene expressions of CYP1A1 and CYP2E1 in the placenta increased significantly in nicotine-treated groups on GD 15 and GD 18, but returned to the level similar to the corresponding control on GD 21. In nicotine group, there was a decrease of mdr1a expression on GD 15, GD 18, and GD 21, with the most significant decrease on GD 15. In contrast, no significant difference was found in mdr1b mRNA expression between the nicotine-treated animals and the corresponding control. In comparison with the corresponding control, the placental Pgp protein significantly decreased on GD 15 and GD 18. Our results showed that prenatal nicotine exposure resulted in inhibition of fetal growth significantly. The induction of CYP2E1 and CYP1A1 gene expression by nicotine in the maternal liver and placenta may be involved with the observed increase in oxidative stress and lipid peroxidation. The inhibition of the placental Pgp expression by nicotine may also contribute to an increased susceptibility of the fetus to environmental toxins.

Estrés oxidativo hepatocitario y hepatopatía alcohólica

Acute and chronic alcohol consumption increases the production of reactive oxygen species (ROS), and enhances lipid peroxidation of lipids, proteins, and DNA. The mechanism by which alcohol causes cell injury is still not clear but a major role for ROS and lipid peroxidation-end products is considered. Many pathways have been suggested to play a role on how ethanol induces a state of "oxidative stress", including redox-state changes, acetaldehyde production, damage to the mitochondria, membrane injury, apoptosis, ethanol-induced hypoxia, effects on the immune system and altered cytokine production, increased endotoxin levels and activation of Kupffer cells, mobilization of iron, changes in the antioxidant defense, particularly mitochondrial glutathione (GSH), one electron oxidation of ethanol to 1-hydroxy-ethyl radical, and induction of CYP2E1. These pathways are not exclusive of one another and it is likely that several, indeed many systems contribute to the ability of ethanol to induce a state of oxidative stress.

Efficacious treatment of alcoholic steatosis by a pharmaceutical composition comprising metadoxine and garlic oil

Alcoholic steatosis is the earliest and common response to heavy alcohol intake, and may precede severe forms of liver injury. As an effort to develop an efficacious pharmaceutical composition, this study evaluated the effects of metadoxine, garlic oil or their combination on alcoholic steatosis. Feeding rats an alcohol‐containing diet for 4 weeks increased hepatic triglyceride content with CYP2E1 induction. Concurrent administration of metadoxine and garlic oil (MG, 1:1 w/w ratio) to rats synergistically abrogated both fat accumulation and CYP2E1 induction compared to the individual treatment. Histopathology confirmed the ability of MG combination to inhibit lipid accumulation. Blood biochemistry verified improvement of liver function in MG‐treated rats. Alcohol administration resulted in decreases in the AMPKα and ΑCC phosphorylations, which was restored by MG treatments. Hepatic fatty acid synthase (FAS) expression was decreased after alcohol consumption, which correlated with a decrease in AMPK activity and a commensurate increase in lipid content. Combined MG treatments allowed FAS expression restored. These results demonstrate that a pharmaceutical composition comprising MG effectively treats alcoholic steatosis, which may be associated with CYP2E1 repression and recovery of AMPK activity, promising pharmaceutical therapy for alcoholic steatosis.Supported by a contract fund from the Seoul R&amp;BD Program (2005–2010).

Alleviation of acute ethanol-induced liver injury and impaired metabolomics of S-containing substances by betaine supplementation

Oxidative stress is suggested to play a key role in the development of alcoholic liver injury. We investigated the induction of oxidative damage in association with changes in hepatic concentrations of sulfur-containing substances in mice challenged with binge-like ethanol administration. Also the protective effect of dietary betaine against ethanol-induced liver injury was determined. Serum alanine aminotransferase activity, TNFα level, and hepatic malondialdehyde level were increased significantly by ethanol administration. Hepatic Cyp2e1 was induced to 250% of control. Ethanol administration decreased hepatic S-adenosylmethionine, cysteine, and glutathione, but elevated hypotaurine and taurine levels. Betaine supplied in drinking water for 2 weeks attenuated the induction of alcoholic liver injury and Cyp2e1 significantly. Reduction of hepatic S-adenosylmethionine and glutathione was alleviated, and elevation of hypotaurine and taurine was depressed. The results suggest that betaine may protect the liver against ethanol-induced oxidative injury most probably via its effects on the sulfur–amino acid metabolism.

Ethanol enhanced the genotoxicity of acrylamide in human, metabolically competent HepG2 cells by CYP2E1 induction and glutathione depletion

In the present study, the genotoxicity of acrylamide (AA) was investigated in HepG2 cells using SCGE. Additionally, the influence of ethanol on the modulation of AA-induced DNA-migration caused by CYP2E1-upregulation and/or GSH-depletion was examined in the same cell line. For the ethanol/AA combination assays, the cells were treated with ethanol for 24 h prior to exposure to 5 mM AA for another 24 h. 1.25 to 10 mM AA-induced DNA migration (OTM) in HepG2 cells in a concentration-dependent manner, e.g., exposure to 10 mM AA, resulted in an 8-fold increase of DNA migration compared to the negative control. Treatment with 120 mM ethanol prior to exposure to 5 mM AA increased the level of DNA migration more than 2-fold as compared to cells treated with 5 mM AA alone. Immunoblotting showed a clear ethanol-induced increase of CYP2E1, which plays a pivotal role in AA toxification. Additionally, intracellular GSH levels were significantly reduced after ethanol or AA treatment. In the ethanol/AA combination experiments, GSH depletion was comparable to the additive effect of the single compounds. No induction of apoptosis (ssDNA assay), but necrosis was identified as responsible for the reduction of viability with increasing compound concentration. The data clearly show a higher genotoxic potential of ethanol/AA combination treatment compared to AA treatment alone. In conclusion, both the ethanol-mediated induction of CYP2E1 and the depletion of GSH provide a mechanistic explanation for the over-additive effects of ethanol and AA. Even though the concentrations used in this study were rather high, consequences for the dietary intake of AA-containing food and alcoholic beverages should be discussed.

Induction of Hepatic CYP2E1 by a Subtoxic Dose of Acetaminophen in Rats: Increase in Dichloromethane Metabolism and Carboxyhemoglobin Elevation

Dichloromethane (DCM) is metabolically converted to carbon monoxide mostly by CYP2E1 in liver, resulting in elevation of blood carboxyhemoglobin (COHb) levels. We investigated the effects of a subtoxic dose of acetaminophen (APAP) on the metabolic elimination of DCM and COHb elevation in adult female rats. APAP, at 500 mg/kg i.p., was not hepatotoxic as measured by a lack of change in serum aspartate aminotransferase, alanine aminotransferase, and sorbitol dehydrogenase activities. In rats pretreated with APAP at this dose, the COHb elevation resulting from administration of DCM (3 mmol/kg i.p.) was enhanced significantly. Also blood DCM levels were reduced, and its disappearance from blood appeared to be increased. Hepatic CYP2E1-mediated activities measured with chlorzoxazone, p-nitrophenol, and p-nitroanisole as substrates were all induced markedly in microsomes of rats treated with APAP. Aminopyrine N-demethylase activity was also increased slightly, but significantly. Western blot analysis showed that APAP treatment induced the expression of CYP2E1 and CYP3A proteins. Neither hepatic glutathione contents nor glutathione S-transferase activity was changed by the dose of APAP used. The results indicate that, contrary to the well known hepatotoxic effects of this drug at large doses, a subtoxic dose of APAP may induce CYP2E1, and to a lesser degree, CYP3A expression. This is the first report that APAP can increase cytochrome P450 (P450)-mediated hepatic metabolism and the resulting toxicity of a xenobiotic in the whole animal. The pharmacological/toxicological significance of induction of P450s by a subtoxic dose of APAP is discussed.

Combined metadoxine and garlic oil treatment efficaciously abrogates alcoholic steatosis and CYP2E1 induction in rat liver with restoration of AMPK activity

Alcoholic steatosis is the earliest and most common response to heavy alcohol intake, and may precede more severe forms of liver injury. Accumulation of fat, largely triglyceride, in hepatocytes results from the inhibition of fatty acid oxidation and excessive oxidative stress involving CYP2E1. This study evaluated the therapeutic effects of metadoxine, garlic oil or their combination on alcoholic steatosis. Feeding rats an alcohol-containing diet for 4 weeks elicited an increase in hepatic triglyceride content and induced CYP2E1. The concurrent administration of metadoxine and garlic oil (MG) to rats during the last week of the diet feeding efficaciously abrogated both fat accumulation and CYP2E1 induction as compared to the individual treatment at higher doses. Histopathology confirmed the ability of MG combination to inhibit lipid accumulation. Blood biochemistry verified improvement of liver function in rats treated with MG. Alcohol administration resulted in a decrease in AMP-activated protein kinase-α (AMPKα) phosphorylation, which was restored by MG treatments. Recovery of AMPK activity by MG was supported by an increase in acetyl-CoA carboxylase phosphorylation. Hepatic fatty acid synthase (FAS) expression was markedly decreased after alcohol consumption, which correlated with a decrease in AMPK activity and a commensurate increase in lipid content. Combined MG treatments caused restoration of the FAS level. These results demonstrate that the combination of MG effectively treats alcoholic steatosis with CYP2E1 inhibition, which may be associated with the recovery of AMPK activity, promising that the combination therapy may constitute an advance in the development of clinical candidates for alcoholic steatosis.

The effect of acetaminophen (four grams a day for three consecutive days) on hepatic tests in alcoholic patients – a multicenter randomized study

BackgroundHepatic failure has been associated with reported therapeutic use of acetaminophen by alcoholic patients. The highest risk period for alcoholic patients is immediately after discontinuation of alcohol intake. This period exhibits the largest increase in CYP2E1 induction and lowest glutathione levels. Our hypothesis was that common liver tests would be unaffected by administration of the maximum recommended daily dosage of acetaminophen for 3 consecutive days to newly-abstinent alcoholic subjects.MethodsAdult alcoholic subjects entering two alcohol detoxification centers were enrolled in a prospective double-blind, randomized, placebo-controlled trial. Subjects were randomized to acetaminophen, 4 g/day, or placebo for 3 consecutive days. The study had 95% probability of detecting a 15 IU/L difference in serum ALT.ResultsA total of 443 subjects were enrolled: 308 (258 completed) received acetaminophen and 135 subjects (114 completed) received placebo. Study groups did not differ in demographics, alcohol consumption, nutritional status or baseline laboratory assessments. The peak mean ALT activity was 57 ± 45 IU/L and 55 ± 48 IU/L in the acetaminophen and placebo groups, respectively. Subgroup analyses for subjects presenting with an elevated ALT, subjects fulfilling a diagnosis of alcoholic hepatitis and subjects attaining a peak ALT greater than 200 IU/L showed no statistical difference between the acetaminophen and control groups. The one participant developing an increased international normalized ratio was in the placebo group.ConclusionAlcoholic patients treated with the maximum recommended daily dose of acetaminophen for 3 consecutive days did not develop increases in serum transaminase or other measures of liver injury. Treatment of pain or fever for 3 days with acetaminophen appears safe in newly-abstinent alcoholic patients, such as those presenting for acute medical care.

Undernutrition enhances alcohol-induced hepatocyte proliferation in the liver of rats fed via total enteral nutrition

To assess the relative contributions of undernutrition and ethanol (EtOH) exposure to alcohol-induced hepatotoxicity, female Sprague-Dawley rats were intragastrically infused liquid diets containing 187 or 154 kcal.kg(-3/4).day(-1) with or without 11 g.kg(-1).day(-1) EtOH. EtOH clearance was impaired in the 154 kcal.kg(-3/4).day(-1) EtOH group (P < or = 0.05). A combination of undernutrition and EtOH also increased the induction of hepatic cytochrome P-450 (CYP)2E1 and CYP4A1 mRNA, apoprotein, and activities (P < or = 0.05). This was accompanied by increased oxidative stress (P < or = 0.05). The severity of liver steatosis, macrophage infiltration, and focal necrosis was comparable in both EtOH groups. Alanine aminotransferase levels were elevated (P < or = 0.05) but did not significantly differ between the two EtOH groups. TUNEL analysis also demonstrated a comparable increase in apoptosis in the two EtOH groups (P < or = 0.05). The development of alcohol-induced liver pathology was accompanied by little change in fatty acid (FA) synthesis or degradation at 187 kcal.kg(-3/4).day(-1) but at 154 kcal.kg(-3/4).day(-1) was accompanied by decreased expression of FA synthesis genes and increased expression of peroxisome proliferator-activated receptor-alpha (PPAR-alpha)-regulated FA degradation pathways (P < or = 0.05). In addition, 154 kcal.kg(-3/4).day(-1) EtOH group livers exhibited greater hepatocyte proliferation (P < or = 0.05). We conclude that undernutrition does not exacerbate alcoholic steatohepatitis despite additional oxidative stress produced by an increased induction of CYP2E1 and CYP4A1. However, enhanced ethanol-induced cellular proliferation, perhaps as a result of enhanced PPAR-alpha signaling, may contribute to an increased risk of hepatocellular carcinoma in undernourished alcoholics.

Induction of CYP1A1 and CYP2E1 in rat liver by histamine: binding and kinetic studies

Histamine (HA) may bind to cytochrome P450 (CYP450) in rat liver microsomes. The CYP450-HA complex seems to regulate some cellular processes such as proliferation. In the present work, it is shown that HA increases the activity and protein level of CYP1A1 and CYP2E1, in vivo. CYP1A1 is associated with polycyclic aromatic hydrocarbon-mediated carcinogenesis and CYP2E1 with liver damage by oxidative stress. Studies of enzyme kinetics and binding with rat liver microsomes and supersomes were carried out to determine whether HA is a substrate of CYP1A1 and/or CYP2E1. The lack of NADPH oxidation in the presence of HA showed that it is not a substrate for CYP1A1. Activity measurements using the O-dealkylation of ethoxyresorufin indicated that HA is a mixed-type inhibitor of CYP1A1 in both microsomes and supersomes. On the other hand, HA induced a significant NADPH oxidation catalyzed by CYP2E1 supersomes, strongly suggesting that HA is a substrate for this isoform. Furthermore, HA is consumed in the presence of CYP2E1-induced microsomes and supersomes, as determined by o-phtalaldehyde complexes with HA by HPLC. The present findings may contribute to understand better the physiological function of CYP450 in relation with inflammation and other physiological processes in which HA may have a relevant role.

Propolis protects CYP 2E1 enzymatic activity and oxidative stress induced by carbon tetrachloride

Induction of CYP 2E1 by carbon tetrachloride (CCl(4)) is one of the central pathways by which CCl(4) generates oxidative stress in hepatocytes. Experimental liver injury was induced in rats by CCl(4) to determine toxicological actions on CYP 2E1 by microsomal drug metabolizing enzymes. In this report, ethanolic extract of propolis at a dose of 200 mg/kg (po) was used after 24 h of toxicant administration to validate its protective potential. Intraperitoneal injection of CCl(4) (1.5 ml/kg) induced hepatotoxicity after 24 h of its administration that was associated with elevated malonyldialdehyde (index of lipid peroxidation), lactate dehydrogenase and gamma-glutamyl transpeptidase release (index of a cytotoxic effect). Hepatic microsomal drug metabolizing enzymes of CYP 2E1 showed sharp depletion as assessed by estimating aniline hydroxylase and amidopyrine N-demethylase activity after CCl(4) exposure. Toxic effect of CCl(4) was evident on CYP 2E1 activity by increased hexobarbitone induced sleep time and bromosulphalein retention. Propolis extract showed significant improvement in the activity of both enzymes and suppressed toxicant induced increase in sleep time and bromosulphalein retention. Choleretic activity of liver did not show any sign of toxicity after propolis treatment at a dose of 200 mg/kg (id). Histopathological evaluation of the liver revealed that propolis reduced the incidence of liver lesions including hepatocyte swelling and lymphocytic infiltrations induced by CCl(4). Electron microscopic observations also showed improvement in ultrastructure of liver and substantiated recovery in biochemical parameters. Protective activity of propolis at 200 mg/kg dose was statistically compared with positive control silymarin (50 mg/kg, po), a known hepatoprotective drug seems to be better in preventing hepatic CYP 2E1 activity deviated by CCl(4). These results lead us to speculate that propolis may play hepatoprotective role via improved CYP 2E1 activity and reduced oxidative stress in living system.