CYP2E1 Research Articles

Pathogenesis of Alcoholic Fatty Liver a Narrative Review.

Alcohol effect hepatic lipid metabolism through various mechanisms, leading synergistically to an accumulation of fatty acids (FA) and triglycerides. Obesity, as well as dietary fat (saturated fatty acids (FA) versus poly-unsaturated fatty acids (PUFA)) may modulate the hepatic fat. Alcohol inhibits adenosine monophosphate activated kinase (AMPK). AMPK activates peroxisome proliferator activated receptor a (PPARα) and leads to a decreased activation of sterol regulatory element binding protein 1c (SRABP1c). The inhibition of AMPK, and thus of PPARα, results in an inhibition of FA oxidation. This ß-oxidation is further reduced due to mitochondrial damage induced through cytochrome P4502E1 (CYP2E1)-driven oxidative stress. Furthermore, the synthesis of FAs is stimulated through an activation of SHREP1. In addition, alcohol consumption leads to a reduced production of adiponectin in adipocytes due to oxidative stress and to an increased mobilization of FAs from adipose tissue and from the gut as chylomicrons. On the other side, the secretion of FAs via very-low-density lipoproteins (VLDL) from the liver is inhibited by alcohol. Alcohol also affects signal pathways such as early growth response 1 (Egr-1) associated with the expression of tumour necrosis factor α (TNF α), and the mammalian target of rapamycin (mTOR) a key regulator of autophagy. Both have influence the pathogenesis of alcoholic fatty liver. Alcohol-induced gut dysbiosis contributes to the severity of ALD by increasing the metabolism of ethanol in the gut and promoting intestinal dysfunction. Moreover, pathogen-associated molecular patterns (PAMPS) via specific Toll-like receptor (TLR) bacterial overgrowth leads to the translocation of bacteria. Endotoxins and toxic ethanol metabolites enter the enterohepatic circulation, reaching the liver and inducing the activation of the nuclear factor kappa-B (NFκB) pathway. Pro-inflammatory cytokines released in the process contribute to inflammation and fibrosis. In addition, cellular apoptosis is inhibited in favour of necrosis.

CYP2E1 triggered GRP78/ATF6/CHOP signaling axis inhibit apoptosis and promotes progression of hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is the leading cause of cancer-related death worldwide. Cytochrome P450 2E1 (CYP2E1) is an enzyme, primarily involved in the metabolism of xenobiotics and procarcinogens. The present study was designed to investigate the potential role of CYP2E1 triggered endoplasmic reticulum stress in the progression of HCC through inhibition of apoptosis. In vitro CYP2E1 promotes HepG2 cell migration, reduced chromatin condensation, enhanced intracellular ROS accumulation and induce cell cycle progression. Conversely this effect was averted by CYP2E1 siRNA, selective inhibitor Diallyl sulphide (DAS) and antioxidants (vitamin C and E). In vivo Diethylnitrosamine (DEN) induced HCC rats showed decreased body weight and increased relative liver weight. Moreover, macro trabecular-massive HCC (MTM-HCC) histological subtyping showed pathological features like well-differentiated tumors, micro-trabecular and pseudo glandular patterns, megakaryocytes and cholestasis. Masson's trichrome staining revealed an intensive accumulation of collagen fibers in the extracellular matrix (ECM). Increased CYP2E1, VEGF and PCNA enhance the carcinogenicity as revealed in immunohistochemistry results. Immunoblot analysis showed reduced expression of copper-zinc superoxide dismutase (CuZnSOD) and manganese superoxide dismutase (MnSOD) in cytosolic as well as mitochondrial fraction of rat liver tissue respectively. Also, increased level of CYP2E1 stimulated the upregulation of unfolded proteins response (UPR) and ER stress-related proteins such as Glucose regulatory protein 78 (GRP78), activating transcription factor 6 (ATF6) and CCAAT enhancer-binding protein (C/EBP) homologous protein (CHOP). Meanwhile, CYP2E1 stimulated ER-stress reduces BCL2 and downregulates the cleaved caspase 3 thus suppresses apoptosis. in. Furthermore, immunofluorescence revealed increased expression level of α-SMA in the HCC rat liver tissue. The level of CYP2E1 mRNA was significantly increased. Altogether, these findings indicate that CYP2E1 has a dynamic role in the pathogenesis of HCC and might be a budding agent in liver carcinogenesis therapy.

Co-occurrence of Mycotoxin-Induced Hepatotoxicity in Mice Inhibited by Lycopene: Mitochondrial Impairment and Early Hepatic Fibrosis.

Mycotoxins co-contamination of agricultural products poses a serious threat to human and animal health, especially hepatic dysfunction. Zearalenone (ZEN), deoxynivalenol (DON), and aflatoxin B1 (AFB1) are three commonly co-occurring mycotoxins. This study is to determine whether lycopene (LYC) can alleviate hepatic toxicity induced by the co-occurrence of ZEN, DON, and AFB1 in mice. Eighty 6-week-old male ICR mice are divided into four groups: CON group (solvent control), LYC group (10 mgkg-1 LYC), Co-M group (10 mgkg-1 ZEN + 1 mgkg-1 DON + 0.5 mgkg-1 AFB1), and LYC+Co-M group (10 mgkg-1 LYC + 10 mgkg-1 ZEN + 1 mgkg-1 DON + 0.5 mgkg-1 AFB1). The results show that LYC can suppress the co-occurrence of mycotoxin-induced mitochondrial swelling and vacuolization accompanied by dysregulation of indices of mitochondrial dynamics (Mitofusin 1 (Mfn1), Mfn2, Optic atrophy 1 (Opa1), Dynamin-related protein 1 (Drp1), Fission 1 (Fis1) at the mRNA level; DRP1 and FIS1 at the protein level). LYC effectively inhibits co-occurrence of mycotoxin-induced activation of Cytochrome P450 2E1, and early fibrosis, as determined by staining with Masson's trichrome and α-SMA protein. LYC successfully attenuates early hepatic fibrosis mainly through antioxidant activities and prevented mitochondrial injury.

Synergistic Effects of Vitis vinifera L. and Centella asiatica against CCl4-Induced Liver Injury in Mice.

Liver injury can be acute or chronic, resulting from a variety of factors, including viral hepatitis, drug overdose, idiosyncratic drug reaction, or toxins, while the progression of pathogenesis in the liver rises due to the involvement of numerous cytokines and growth factor mediators. Thus, the identification of more effective biomarker-based active phytochemicals isolated from medicinal plants is a promising strategy to protect against CCl4-induced liver injury. Vitis vinifera L. (VE) and Centella asiatica (CE) are well-known medicinal plants that possess anti-inflammatory and antioxidant properties. However, synergism between the two has not previously been studied. Here, we investigated the synergistic effects of a V. vinifera L. (VE) leaf, C. asiatica (CE) extract combination (VCEC) against CCl4-induced liver injury. Acute liver injury was induced by a single intraperitoneal administration of CCl4 (1 mL/kg). VCEC was administered orally for three consecutive days at various concentrations (100 and 200 mg/kg) prior to CCl4 injection. The extent of liver injury and the protective effects of VCEC were evaluated by biochemical analysis and histopathological studies. Oxidative stress was evaluated by measuring malondialdehyde (MDA) and glutathione (GSH) levels and Western blotting. VCEC treatment significantly reduced serum transaminase levels (AST and ALT), tumor necrosis factor-α (TNF-α), and reactive oxygen species (ROS). CCl4- induced apoptosis was inhibited by VCEC treatment by reducing cleaved caspase-3 and Bcl2-associated X protein (Bax). VCEC-treated mice significantly restored cytochrome P450 2E1, nuclear factor erythroid 2-related factor 2 (Nrf2), and heme oxygenase-1 (HO-1) expression in CCl4-treated mice. In addition, VCEC downregulated overexpression of proinflammatory cytokines and hepatic nuclear factor kappa B (NF-κB) and inhibited CCl4-mediated apoptosis. Collectively, VCEC exhibited synergistic protective effects against liver injury through its antioxidant, anti-inflammatory, and antiapoptotic ability against oxidative stress, inflammation, and apoptosis. Therefore, VCEC appears promising as a potential therapeutic agent for CCl4-induced acute liver injury in mice.

Integration of proteomics and network toxicology reveals the mechanism of mercury chloride induced hepatotoxicity, in mice and HepG2 cells.

Mercury is one heavy metal toxin that could cause severe health impairments. Mercury exposure has become a global environmental issue. Mercury chloride (HgCl2) is one of mercury's main chemical forms, but it lacks detailed hepatotoxicity data. The present study aimed to investigate the mechanism of hepatotoxicity induced by HgCl2 through proteomics and network toxicology at the animal and cellular levels. HgCl2 showed apparent hepatotoxicity after being administrated with C57BL/6 mice (16mg/kg.bw, oral once a day, 28 days) and HepG2 cells (100μmol/L, 12h). Otherwise, oxidative stress, mitochondrial dysfunction and inflammatory infiltration play an important role in HgCl2-induced hepatotoxicity. The differentially expressed proteins (DEPs) after HgCl2 treatment and enriched pathways were obtained through proteomics and network toxicology. Western blot and qRT-PCR results showed acyl-CoA thioesterase 1 (ACOT1), acyl-CoA synthetase short chain family member 3 (ACSS3), epidermal growth factor receptor (EGFR), apolipoprotein B (APOB), signal transducer and activator of transcription 3 (STAT3), alanine--glyoxylate aminotransferase (AGXT), cytochrome P450 3A5 (CYP3A5), CYP2E1 and CYP1A2 may be the major biomarkers for HgCl2-induced hepatotoxicity, which involved chemical carcinogenesis, fatty acid metabolism, CYPs-mediated metabolism, GSH metabolism and others. Therefore, this study can provide scientific evidence for the biomarkers and mechanism of HgCl2-induced hepatotoxicity.

The CYP2E1 inhibitor Q11 ameliorates LPS-induced sepsis in mice by suppressing oxidative stress and NLRP3 activation

Sepsis is an infection-induced, multi-organ system failure with a pathophysiology related to inflammation and oxidative stress. Increasing evidence indicates that cytochrome P450 2E1 (CYP2E1) is involved in the incidence and development of inflammatory diseases. However, a role for CYP2E1 in lipopolysaccharide (LPS)-induced sepsis has not been completely explored. Here we use Cyp2e1 knockout (cyp2e1-/-) mice to determine if CYP2E1 could be a therapeutic target for sepsis. We also evaluated the ability of Q11, a new specific CYP2E1 inhibitor, to prevent and ameliorate LPS-induced sepsis in mice and in LPS-treated J774A.1 and RAW264.7 cells. Cyp2e1 deletion significantly reduced hypothermia, multi-organ dysfunction and histological abnormalities in LPS-treated mice; consistent with this finding, the CYP2E1 inhibitor Q11 significantly prolonged the survival time of septic mice and ameliorated multi-organ injury induced by LPS. CYP2E1 activity in liver correlated with indicators of multi-organ injury, such as the level of lactate dehydrogenase (LDH) and blood urea nitrogen (BUN) (P < 0.05). Q11 significantly suppressed the expression of NLRP3 in tissues after LPS injection; in vitro studies revealed that activation of NLRP3 signaling and increase of ROS was attenuated by Q11 in LPS-stimulated macrophages, which was reflected by reduced expression of caspase-1 and formation of ASC specks. Overall, our results indicate that Q11 improves the survival of mice with LPS-induced sepsis and attenuates sepsis-induced multiple-organ injury, suggesting that CYP2E1 could be a therapeutic target for sepsis.

Identification of Cytochrome P450 2E1 as a Novel Target in Glioma and Development of Its Inhibitor as an Anti-Tumor Agent.

Glioblastoma (GBM) is a devastating inflammation-related cancer for which novel therapeutic targets are urgently required. Previous studies of the authors indicate Cytochrome P450 2E1 (CYP2E1) as a novel inflammatory target and develop a specific inhibitor Q11. Here it is demonstrated that CYP2E1 overexpression is closely related to higher malignancy in GBM patients. CYP2E1 activity is positively correlated with tumor weight in GBM rats. Significantly higher CYP2E1 expression accompanied by increased inflammation is detected in a mouse GBM model. Q11, 1-(4-methyl-5-thialzolyl) ethenone, a newly developed specific inhibitor of CYP2E1 here remarkably attenuates tumor growth and prolongs survival in vivo. Q11 does not directly affect tumor cells but blocks the tumor-promoting effect of microglia/macrophage (M/Mφ) in the tumor microenvironment through PPARγ-mediated activation of the STAT-1 and NF-κB pathways and inhibition of the STAT-3 and STAT-6 pathways. The effectiveness and safety of targeting CYP2E1 in GBM are further supported by studies with Cyp2e1 knockout rodents. In conclusion, a pro-GBM mechanism in which CYP2E1-PPARγ-STAT-1/NF-κB/STAT-3/STAT-6 axis fueled tumorigenesis by reprogramming M/Mφ and Q11 as a promising anti-inflammatory agent for GBM treatment is uncovered.

Stabilization of glutathione redox dynamics and CYP2E1 by green synthesized Moringa oleifera-mediated zinc oxide nanoparticles against acrylamide induced hepatotoxicity in rat model: Morphometric and molecular perspectives.

The terrible reality is that acrylamide (AA) is a common food contaminant found in a wide variety of commonly consumed foods. This research involves the advancement of a more dependable technique for the bio-fabrication of zinc oxide nanoparticles (ZNPs) through the green method using Moringa Oleifera extract (MO-ZNPs) as an efficient chelating agent for acrylamide (AA). The effects of AA on glutathione redox dynamics, liver function, lipid profile, and zinc residues in Sprague Dawley rats are investigated. Finally, the microarchitecture and immunohistochemical staining of Caspase-3 and CYP2E1 were determined in the liver tissue of rats. Four separate groups, including control, MO-ZNPs (10mg/kg b. wt), AA (20mg/kg b. wt), and AA+MO-ZNPs for 60 days. The results revealed a suppressed activity of glutathione redox enzymes (GSH, GPX,and GSR) on both molecular and biochemical levels. Also, AA caused elevated liver enzymes, hepatosomatic index, and immunohistochemical staining of caspase-3 and CYP2E1 expression. MO-ZNPs co-treatment, on the other hand, stabilized glutathione-related enzyme gene expression, normalized hepatocellular enzyme levels, and restored hepatic tissue microarchitectures. It could be assumed that MO-ZNPs is a promising hepatoprotective molecule for alleviating AA-induced hepatotoxicity. We witnessed changes in glutathione redox dynamics to be restorative. Glutathione and cytochrome P450 2E1 play crucial roles in AA detoxification, so maintaining a healthy glutathione redox cycle is necessary for disposing of AA toxicity.

Lipid nanoparticle delivery of siRNA targeting Cyp2e1 gene attenuates subacute alcoholic liver injury in mice.

To investigate the effect and mechanism of lipid nanoparticle (LNP) delivery of small interfering RNA (siRNA) targeting Cyp2e1 gene on subacute alcoholic liver injury in mice. siRNA targeting Cyp2e1 gene was encapsulated in LNP (si-Cyp2e1 LNP) by microfluidic technique and the resulting LNPs were characterized. The optimal dose of si-Cyp2e1 LNP administration was screened. Forty female C57BL/6N mice were randomly divided into blank control group, model control group, si-Cyp2e1 LNP group, LNP control group and metadoxine group. The subacute alcoholic liver injury mouse model was induced by ethanol feeding for 10 d plus ethanol gavage for the last 3 d. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, and the superoxide dismutase (SOD) activity as well as malondialdehyde, reactive oxygen species, glutathione, triacylglycerol, total cholesterol contents in liver tissue were measured in each group, and liver index was calculated. The expression of genes related to oxidative stress, lipid synthesis and inflammation in each group of mice were measured by realtime RT-PCR. Compared with the model control group, the levels of liver index, serum ALT, AST activities, malondialdehyde, reactive oxygen species, triacylglycerol, total cholesterol contents in liver tissue decreased, but the SOD activity as well as glutathione increased in the si-Cyp2e1 LNP group (all P<0.01). Hematoxylin-eosin staining result showed disorganized hepatocytes with sparse cytoplasm and a large number of fat vacuoles and necrosis in the model control group, while the si-Cyp2e1 LNP group had uniformly sized and arranged hepatocytes with normal liver tissue morphology and structure. Oil red O staining result showed si-Cyp2e1 LNP group had lower fat content of the liver compared to the model control group (P<0.01), and no fat droplets accumulated. Anti-F4/80 monoclonal antibody fluorescence immunohistochemistry showed that the si-Cyp2e1 LNP group had lower cumulative optical density values compared to the model control group (P<0.01) and no significant inflammatory reaction. Compared with the model control group, the expression of catalytic genes P47phox, P67phox and Gp91phox were reduced (all P<0.01), while the expression of the antioxidant enzyme genes Sod1, Gsh-rd and Gsh-px were increased (all P<0.01). The mRNA expression of the lipid metabolism genes Pgc-1α and Cpt1 were increased (all P<0.01) and the lipid synthesis-related genes Srebp1c, Acc and Fasn were decreased (all P<0.01); the expression of liver inflammation-related genes Tgf-β, Tnf-α and Il-6 were decreased (all P<0.01). The si-Cyp2e1 LNP may attenuate subacute alcoholic liver injury in mice mainly by reducing reactive oxygen levels, increasing antioxidant activity, blocking oxidative stress pathways and reducing ethanol-induced steatosis and inflammation.

Sex- and lifestyle-related factors are associated with altered hepatic CYP protein levels in people diagnosed with mental disorders.

In this study, we used human postmortem tissue to investigate hepatic protein expression levels of CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4 by LC‒MS/MS in a population of people suffering from mental disorders (n=171). We report hepatic protein levels of these six CYP isoforms in 171 individuals in total, and define a focused population dataset of 116 individuals after excluding 55 samples due to low Microsomal Protein Per Gram of Liver (MPPGL) yield. Postmortem decay was most likely the reason for the low MPPGL yield in the 55 samples. In the focused population, we found women to have significantly higher protein levels of CYP3A4 than men in addition to decreased CYP3A4 protein levels among obese individuals. Furthermore, MPPGL was negatively correlated with body mass index (BMI). An increase in CYP1A2 protein levels was observed among smokers and increased CYP2E1 protein levels were observed among individuals with a history of alcohol abuse. Finally, individuals who received phenobarbital (CYP3A4 inducer) had significantly higher CYP3A4 levels. In conclusion, lifestyle-related factors prevalent among people suffering from mental disorders are associated with altered CYP protein levels which may alter drug metabolism and affect the efficacy of commonly prescribed drugs. Furthermore, this investigation demonstrates that postmortem hepatic tissue can be used to study how lifestyle and effectors affect hepatic CYP-levels in a large cohort of patients. Significance Statement Using a large number of postmortem hepatic tissue specimens (n=116) originating from the autopsy of individuals diagnosed with mental disorders, we were able to show that hepatic CYP-levels were affected by alcohol, smoking, BMI, and sex and that MPPGL was affected by BMI. Theses lifestyle-related changes may alter drug metabolism and affect the efficacy of commonly prescribed drugs. It is a novel approach to use a large postmortem cohort to investigate how lifestyle and effectors affect hepatic CYP-levels.

Walnut (Juglans regia L.) Oligopeptides Alleviate Alcohol-Induced Acute Liver Injury through the Inhibition of Inflammation and Oxidative Stress in Rats.

The study was aimed at investigating the effects of walnut oligopeptides (WOPs) on alcohol-induced acute liver injury and its underlying mechanisms. Male Sprague Dawley (SD) rats were randomly assigned to six groups: normal control, alcohol control, whey protein (440 mg/kg.bw), and three WOPs (220 mg/kg.bw, 440 mg/kg.bw, 880 mg/kg.bw) groups. After 30 days of gavage, ethanol with a volume fraction of 50%, administered at a dose of 7 g/kg.bw., caused acute liver injury. A righting reflex experiment and a blood ethanol concentration evaluation were then performed. Serum biochemical parameters, inflammatory cytokines, liver alcohol metabolism enzymes, oxidative stress biomarkers, liver nuclear factor-κB (NF-κB p65), and cytochrome P4502E1 expression were determined. The results revealed that the intervention of 440 mg/kg and 880 mg/kg WOPs could alleviate the degree of intoxication, decrease blood ethanol concentration, alleviate alcohol-induced hepatic steatosis, enhance the activity of hepatic ethanol metabolizing enzymes and antioxidant capacity, reduce lipid oxidation products and pro-inflammatory factor contents, and inhibit the expression of NF-κBp65 in the livers of rats. The outcomes of the study suggest that WOPs have beneficial effects on liver damage caused by acute ethanol binge drinking, with the high-dose WOPs (880 mg/kg.bw) exerting the most pronounced hepatoprotective effect.

Indole-3-carbinol ameliorated the thioacetamide-induced hepatic encephalopathy in rats

Indole-3-carbinol (I3C) is reported to have hepatic and neuroprotective properties. However, the I3C role in the protection of the liver and brain in the pathological condition of hepatic encephalopathy has not been investigated. Therefore, in the present study, we have assessed the hepatic and neuroprotective roles of I3C against thioacetamide (TAA)- induced hepatic encephalopathy in Wistar rats. TAA (300 mg/kg) was intraperitoneally administered to Wistar rats to induce hepatic encephalopathy. The elevated levels of ammonia in the blood, liver, and brain were substantially lowered by I3C treatment (25, 50, and 100 mg/kg, oral, 7 days). I3C significantly ameliorated the TAA-induced liver dysfunction by decreasing the alanine transaminase, aspartate transaminase, and alkaline phosphatase enzymes and reduced the elevated cytochrome P4502E1 (CYP2E1) activity in the liver and brain. Further, I3C alleviated mitochondrial dysfunction and oxidative stress in the brain. I3C treatment improved the anti-inflammatory cytokine interleukin (IL)− 10 while reducing inflammatory cytokines such as tumor necrosis factor-1 and IL-6 in hepatic encephalopathy rats. I3C reduced the levels of apoptotic indicators mediated by the mitochondria, including cytochrome c, caspase 9, and caspase 3. Concurrently, I3C mitigated the liver and brain histological abnormalities in hepatic encephalopathy rats. Therefore, the present study concluded that the I3C protected the liver and brain from TAA-induced hepatic encephalopathy injury by inhibiting CYP2E1 enzyme activity and decreasing ammonia, oxidative stress, inflammation, and apoptosis. The present study provides preclinical validation of I3C use as hepatic and neuroprotective for hepatic encephalopathy management.

Hepatitis Delta Virus Antigens Trigger Oxidative Stress, Activate Antioxidant Nrf2/ARE Pathway, and Induce Unfolded Protein Response.

Hepatitis delta virus (HDV) is a viroid-like satellite that may co-infect individuals together with hepatitis B virus (HBV), as well as cause superinfection by infecting patients with chronic hepatitis B (CHB). Being a defective virus, HDV requires HBV structural proteins for virion production. Although the virus encodes just two forms of its single antigen, it enhances the progression of liver disease to cirrhosis in CHB patients and increases the incidence of hepatocellular carcinoma. HDV pathogenesis so far has been attributed to virus-induced humoral and cellular immune responses, while other factors have been neglected. Here, we evaluated the impact of the virus on the redox status of hepatocytes, as oxidative stress is believed to contribute to the pathogenesis of various viruses, including HBV and hepatitis C virus (HCV). We show that the overexpression of large HDV antigen (L-HDAg) or autonomous replication of the viral genome in cells leads to increased production of reactive oxygen species (ROS). It also leads to the upregulated expression of NADPH oxidases 1 and 4, cytochrome P450 2E1, and ER oxidoreductin 1α, which have previously been shown to mediate oxidative stress induced by HCV. Both HDV antigens also activated the Nrf2/ARE pathway, which controls the expression of a spectrum of antioxidant enzymes. Finally, HDV and its large antigen also induced endoplasmic reticulum (ER) stress and the concomitant unfolded protein response (UPR). In conclusion, HDV may enhance oxidative and ER stress induced by HBV, thus aggravating HBV-associated pathologies, including inflammation, liver fibrosis, and the development of cirrhosis and hepatocellular carcinoma.

Chlorzoxazone reduced the paracetamol-induced toxicity via competitive inhibition of CYP2E1-mediated metabolism

BackgroundDrug metabolism is crucial to attaining the therapeutic index of any drug. The metabolism and elimination of the drugs are governed mainly by P-glycoprotein (P-gp) and Cytochrome P450 (CYP). Paracetamol is mostly used as analgesic and antipyretic agent. The metabolism of paracetamol is primarily via Glucuronidation and sulphation at therapeutic doses. About 5–10% of paracetamol is metabolized via CYP mediated pathway. Cytochrome P450 2E1 (CYP2E1) is primarily responsible for forming a toxic metabolite of paracetamol called N-acetyl-p-benzoquinoneimine (NAPQI). Even at therapeutic doses, long-term usage of paracetamol leads to the hepatic and nephrotoxicity because of NAPQI. Several in-vitro and in-vivo studies conducted by different research groups and reported that chlorzoxazone is a substrate and inhibitor of CYP2E1. However, the effect of chlorzoxazone on the paracetamol (CYP2E1 substrate) metabolism via the CYP2E1 has not yet been reported. This study investigated the effect of chlorzoxazone on the CYP2E1-mediated metabolism of Paracetamol and NAPQI formation in Wistar rats.ResultsFor 15 days, animals were orally administered with Paracetamol (300 mg/kg) with and without Silymarin (100 mg/kg) (standard CYP2E1 inhibitor) and Chlorzoxazone (50 and 100 mg/kg). Analysis was performed using RP-HPLC on the 15th day to determine paracetamol and NAPQI concentration in the plasma. Paracetamol combination with chlorzoxazone (50 and 100 mg/kg) showed a dose-dependent increase in the AUC0–∞ and the peak plasma concentration (Cmax) of Paracetamol and a dose-dependent decrease of AUC0–∞ and Cmax of NAPQI compared to paracetamol control (p < 0.001). Chlorzoxazone significantly decreased the elevated liver and renal markers compared to paracetamol control. Simultaneously, Hepatic and nephrotic tissue studies showed that compared to the paracetamol control group, the combination of chlorzoxazone significantly ameliorated paracetamol-induced hepatotoxicity and nephrotoxicity.ConclusionFinally, this study revealed that paracetamol in combination with chlorzoxazone led to a significant decrease in the plasma levels of NAPQI and enhanced absorption of paracetamol in rats via the inhibition of CYP2E1- mediated metabolism. In addition, chlorzoxazone significantly ameliorated paracetamol-induced hepatotoxicity and nephrotoxicity.

The Wnt/β-catenin signaling pathway plays a role in drug-induced liver injury by regulating cytochrome P450 2E1 expression.

The online version contains supplementary material available at 10.1007/s43188-023-00180-6.

Ethanol Exposure to Ethanol-Oxidizing HEPG2 Cells Induces Intracellular Protein Aggregation.

Aggresomes are collections of intracellular protein aggregates. In liver cells of patients with alcoholic hepatitis, aggresomes appear histologically as cellular inclusions known as Mallory-Denk (M-D) bodies. The proteasome is a multicatalytic intracellular protease that catalyzes the degradation of both normal (native) and abnormal (misfolded and/or damaged) proteins. The enzyme minimizes intracellular protein aggregate formation by rapidly degrading abnormal proteins before they form aggregates. When proteasome activity is blocked, either by specific inhibitors or by intracellular oxidants (e.g., peroxynitrite, acetaldehyde), aggresome formation is enhanced. Here, we sought to verify whether inhibition of proteasome activity by ethanol exposure enhances protein aggregate formation in VL-17A cells, which are recombinant, ethanol-oxidizing HepG2 cells that express both alcohol dehydrogenase (ADH) and cytochrome P450 2E1 (CYP2E1). We exposed ethanol-non-oxidizing HepG2 cells (ADH-/CYP2E1-) or ethanol-oxidizing VL-17A (ADH+/CYP2E1+) to varying levels of ethanol for 24 h or 72 h. After these treatments, we stained cells for aggresomes (detected microscopically) and quantified their numbers and sizes. We also conducted flow cytometric analyses to confirm our microscopic findings. Additionally, aggresome content in liver cells of patients with alcohol-induced hepatitis was quantified. After we exposed VL-17A cells to increasing doses of ethanol for 24 h or 72 h, 20S proteasome activity declined in response to rising ethanol concentrations. After 24 h of ethanol exposure, aggresome numbers in VL-17A cells were 1.8-fold higher than their untreated controls at all ethanol concentrations employed. After 72 h of ethanol exposure, mean aggresome numbers were 2.5-fold higher than unexposed control cells. The mean aggregate size in all ethanol-exposed VL-17A cells was significantly higher than in unexposed control cells but was unaffected by the duration of ethanol exposure. Co-exposure of cells to EtOH and rapamycin, the latter an autophagy activator, completely prevented EtOH-induced aggresome formation. In the livers of patients with alcohol-induced hepatitis (AH), the staining intensity of aggresomes was 2.2-fold higher than in the livers of patients without alcohol use disorder (AUD). We conclude that ethanol-induced proteasome inhibition in ethanol-metabolizing VL-17A hepatoma cells causes accumulation of protein aggregates. Notably, autophagy activation removes such aggregates. The significance of these findings is discussed.

Regulation of NOX/p38 MAPK/PPARα pathways and miR-155 expression by boswellic acids reduces hepatic injury in experimentally-induced alcoholic liver disease mouse model: novel mechanistic insight

Alcoholic liver disease (ALD) refers to hepatic ailments induced by excessive alcohol intake. The pathogenesis of ALD comprises a complex interplay between various mechanistic pathways, among which inflammation and oxidative stress are key players. Boswellic acids (BAs), found in Boswellia serrata, have shown hepatoprotective effects owing to their antioxidant and anti-inflammatory activities, nevertheless, their therapeutic potential against ALD has not been previously investigated. Hence, this study was performed to depict the possible protective effect of BAs and detect their underlying mechanism of action in an experimentally-induced ALD mouse model. Male BALB/c mice were equally categorized into six groups: control, BAs-treated, ALD, and ALD that received BAs at three-dose levels (125, 250, and 500 mg/kg) by oral gavage for 14 days. Results showed that the high dose of BAs had the most protective impact against ALD according to histopathology examination, blood alcohol concentration (BAC), and liver function enzymes. Mechanistic investigations revealed that BAs (500 mg/kg) caused a significant decrease in cytochrome P450 2E1(CYP2E1), nicotine adenine dinucleotide phosphate oxidase (NOX) 1/2/4, p38 mitogen-activated protein kinase (MAPK), and sterol regulatory element-binding protein-1c (SREBP-1c) levels, and the expression of miR-155, yet increased peroxisome proliferator-activated receptor alpha (PPARα) levels. This led to an improvement in lipid profile and reduced hepatic inflammation, oxidative stress, and apoptosis indices. In summary, our study concludes that BAs can protect against ethanol-induced hepatic injury, via modulating NOX/p38 MAPK/PPARα pathways and miR-155 expression.

Hepatoprotective Effect of Albumin Peptide Fractions from Corn Germ Meal against Alcohol-Induced Acute Liver Injury in Mice.

Acute alcoholic liver disease can cause serious liver damage. This study reports on the hepatoprotective effect of albumin peptide fractions from corn germ meal (MW < 1 kDa) (APF4) on acute alcohol hepatic damage in mice. In the mice model, the results indicated that APF4 at a dose of 800 mg/kg/bw could markedly boost alcohol metabolism, which was shown in the reduced duration of the loss of the righting reflex; the reduced level of blood alcohol concentration (BAC), cytochrome P450 2E1 (CYP2E1), alanine aminotransferase (ALT), aminotransferase (AST), triglycerides (TG), and malondialdehyde (MDA) (p < 0.01); the enhanced activity of aldehyde dehydrogenase (ALDH); and the superoxide dismutase (SOD) and glutathione (GSH) levels being increased by up to 84.02% and 193.22% (p < 0.01) compared to the control group. The antioxidant capability and lipid peroxidation inhibition activity of APF4 may be responsible for its protective effect against liver damage induced by alcohol. The findings suggested that APF4 had the hepatoprotective property against liver damage induced by alcohol.

Полиморфные варианты генов ферментов системы детоксикации ксенобиотиков CYP2E1, GSTM1, GSTT1, EPHX1 как биомаркеры чувствительности к экспозиции побочными продуктами водоподготовки (на примере хлороформа)

Chloroform accumulation in the body and the increase in its steady-state concentrations in blood of exposed people have been established to be associated with polymorphisms of enzyme genes in a genotype involved in metabolism of water disinfection byproducts (A415G of EPHX1 gene, C1091T of CYP2E1 gene, zero mutations of GSTT1 and GSTM1 genes) (р &lt; 0.000001). These polymorphisms in a genotype correlate with higher chloroform levels in blood of people consuming chlorinated drinking water: by 43.8 % and higher for GSTM1 gene polymorphism; by 68.2 % and higher for GSTT1; by 80.4 % and higher for EPHX1 (р &lt; 0.01). EPHX1 genetic polymorphism makes chloroform accumulation much more probable (levels in blood ≥ Р75), which is the most pronounced when combined with GSTТ1 genetic polymorphism. The study results allow us to consider hetero- and homozygous polymorphic genotypes AG/GG for the EPHX1 gene, CT/TT for the CYP2E1 gene, and the null allele in the GSTT1 and GSTM1 genes as genetic predisposition factors for chloroform accumulation in the body. This increases the probability of health outcomes associated with chronic exposure to this disinfection byproduct. The A415G polymorphism of the EPHX1 gene and null alleles of GSTT1 gene, their combinations including the combination with the null allele of the GSTM1 gene and/or the C1091T polymorphism of the CYP2E1 gene can be used as the most informative biomarkers of sensitivity when assessing risks associated with exposure to trihalomethanes (chloroform) at levels not exceeding MPC in water.

Polymorphisms of xenobiotic metabolism enzyme genes cyp2e1, gstm1, gstt1, ephx1 as biomarkers of sensitivity to exposure to water disinfection byproducts (using chloroform as an example)

Chloroform accumulation in the body and the increase in its steady-state concentrations in blood of exposed people have been established to be associated with polymorphisms of enzyme genes in a genotype involved in metabolism of water disinfection byproducts (A415G of EPHX1 gene, C1091T of CYP2E1 gene, zero mutations of GSTT1 and GSTM1 genes) (р &lt; 0.000001). These polymorphisms in a genotype correlate with higher chloroform levels in blood of people consuming chlorinated drinking water: by 43.8 % and higher for GSTM1 gene polymorphism; by 68.2 % and higher for GSTT1; by 80.4 % and higher for EPHX1 (р &lt; 0.01). EPHX1 genetic polymorphism makes chloroform accumulation much more probable (levels in blood ≥ Р75), which is the most pronounced when combined with GSTТ1 genetic polymorphism. The study results allow us to consider hetero- and homozygous polymorphic genotypes AG/GG for the EPHX1 gene, CT/TT for the CYP2E1 gene, and the null allele in the GSTT1 and GSTM1 genes as genetic predisposition factors for chloroform accumulation in the body. This increases the probability of health outcomes associated with chronic exposure to this disinfection byproduct. The A415G polymorphism of the EPHX1 gene and null alleles of GSTT1 gene, their combinations including the combination with the null allele of the GSTM1 gene and/or the C1091T polymorphism of the CYP2E1 gene can be used as the most informative biomarkers of sensitivity when assessing risks associated with exposure to trihalomethanes (chloroform) at levels not exceeding MPC in water.