Lipid nanoparticle delivery of siRNA targeting Cyp2e1 gene attenuates subacute alcoholic liver injury in mice / 浙江大学学报·医学版

OBJECTIVES@#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.@*METHODS@#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.@*RESULTS@#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).@*CONCLUSIONS@#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.

Pharmacokinetics of isoniazid in Wistar rats exposed to ethanol

Abstract Tuberculosis treatment consists of a drug combination, where isoniazid is the core drug and alcoholism is a factor highly related to poor patient compliance with the therapy. CYP2E1 is an enzyme involved both in the metabolism of ethanol and in the formation of hepatotoxic compounds during the metabolism of isoniazid. The shared metabolism pathway accounts for the possibility of pharmacokinetic interaction in cases of concomitant alcohol use during tuberculosis treatment. The aim of this study was to evaluate the effect of repeated exposure of Wistar rats (males, 250 g, n=6) to ethanol on the pharmacokinetics of a single dose of isoniazid in combination with pyrazinamide and rifampicin (100 mg/kg, 350 mg/kg and 100 mg/kg, respectively). An animal group received the combination of drugs and ethanol and was compared to a control group, which received the combination of drugs without exposure to ethanol. The plasma concentrations of isoniazid were determined by a UHPLC/UV bioanalytical method that was previously validated. Biochemical markers of liver function were measured to assess potential damage. A lower elimination half-life of isoniazid was observed in the ethanol group than in the control group (t1/2 0.91 h versus 1.34 h). There was no evidence of hepatotoxicity through the biomarker enzymes evaluated. The results allow us to infer that although there are no biochemical changes related to liver damage, there is a slight influence of ethanol exposure on the pharmacokinetic profile of isoniazid. This change may have a relevant impact on the efficacy of isoniazid in the outcome of tuberculosis treatment.

Extracts of Poria cocos polysaccharides improves alcoholic liver disease in mice via CYP2E1 and NF-κB inflammatory pathways / 中国中药杂志

The present study investigated the effect of extract of Poria cocos polysaccharides(PCP) on cytochrome P450 2 E1(CYP2 E1) and nuclear factor κB(NF-κB) inflammatory signaling pathways in alcoholic liver disease(ALD) mice and explored its protective effect and mechanism. Sixty male C57 BL/6 N mice of SPF grade were randomly divided into a control group, a model group, a positive drug group(bifendate, 200 mg·kg~(-1)), and high-(200 mg·kg~(-1)) and low-dose(50 mg·kg~(-1)) PCP groups. Gao-binge mo-del was induced and the mice in each group were treated correspondingly. Liver morphological and pathological changes were observed and organ index was calculated. Serum levels of alanine aminotransferase(ALT) and aspartate aminotransferase(AST) were detected. Malondialdehyde(MDA) and superoxide dismutase(SOD) in liver tissues were detected by assay kits. The levels of interleukin-6(IL-6) and tumor necrosis factor-α(TNF-α) were detected by ELISA. The activation of macrophages was observed by immunofluorescence staining and protein expression of CYP2 E1, Toll-like receptor 4(TLR4), NF-κB p65, and phosphorylated NF-κB p65(p-NF-κB p65) were analyzed by Western blot. The ALD model was properly induced. Compared with the model group, the PCP groups significantly improved the pathological injury of liver tissues. Immunofluorescence staining revealed that compared with the model group, the groups with drug intervention showed decreased macrophages in liver tissues. Additionally, the PCP groups showed reduced ALT, AST, MDA, IL-6, and TNF-α(P<0.05), and potentiated activity of SOD(P<0.01). PCP extract has the protective effect against alcoholic liver injury in mice, and the underlying mechanism may be related to the regulation of the expression of CYP2 E1 and inhibition of TLR4/NF-κB inflammatory signaling pathway to reduce oxidative stress and inflammatory injury, thereby inhibiting the development of ALD.

Liver Tissue-specific Genes IGFALS,CYP3A4,SLC22A1 and CYP2E1 May be Associated with Poor Prognosis of Liver Cancer / 中国医学科学院学报

Objective To explore the function and mechanism of related genes in the occurrence and development of liver cancer, and the possibility of key genes as potential biomarkers and prognostic indicators for the treatment of liver cancer.Methods We selected 4 datasets(GSE57957, GSE121248, GSE36376 and GSE14520)from the GEO database.With

Biomarkers of susceptibility and effect in car painters exposed to organic solvents / Biomarcadores de susceptibilidad y efecto en pintores de carros expuestos a solventes orgánicos

Abstract Introduction: Car painters are routinely exposed to organic solvents classified as carcinogenic and mutagenic substances. Objective: To characterize the population susceptibility and evaluate the genotoxic effects of exposure to organic solvents. Methods: A cross-sectional study comparing a group of car painters exposed to organic solvents with a non-exposed group. CYP2E1 polymorphisms and the presence of micronuclei in lymphocytes were determined. Results: One hundred twenty-two workers participated in the study: 62 who worked in car paint shops and were exposed to solvents, and 60 who were not exposed. There were statistically significant differences between the two groups regarding micronucleated cells and nucleoplasmic bridges frequencies (p=0.042 and p=0.046, respectively; exact likelihood ratio). Significant differences were found at the interaction between the CYP2E1 genotype c1c1 and occupational exposure to solvents, with higher frequencies of micronuclei (p= 0.013) and micronucleated cells (p= 0.015). However, when the frequencies of micronuclei, micronucleated cells and nucleoplasmic bridges in the exposure group were compared between the c1c1 and c2c2/c1c2 allele groups of the CYP2E1 polymorphism, statistically significant differences were found. Conclusions: This study confirms that when workers with CYP2E1 polymorphisms, specifically the c1c1 genotype, are exposed to organic solvents, they are more likely to have somatic cell mutations, a condition associated with increased susceptibility to diseases such as cancer

Resumen Introducción: Los pintores de vehículos automotores están rutinariamente expuestos a agentes como los solventes orgánicos, capaces de producir efectos mutágenos y carcinógenos. Objetivo: Caracterizar la susceptibilidad poblacional y evaluar los efectos genotóxicos debidos a la exposición a solventes orgánicos. Métodos: Estudio de corte transversal que comparó a un grupo de pintores de carros expuestos a solven tes orgánicos con un grupo de personas no expuestas. Fueron determinados tanto los polimorfismos de CYP2E1 como la presencia de micronúcleos en linfocitos. Resultados: Participaron 122 personas, 62 trabajadores de talleres de pintura de autos expuestos a solventes y 60 personas no expuestas. Con relación al cuestionario Q 16, 32% de los expuestos refirieron síntomas sugestivos de neurotoxicidad. Las frecuencias de células micronucleadas y de puentes nucleoplásmicos fueron significativamente mayores en los expuestos que en los no expuestos: p= 0.042 y p= 0.046, respectivamente, Razón de verosimilitud exacta). Fueron halladas diferencias significativas en la interacción de CYP2E1 (c1c1) y la exposición ocupacional a solventes, con mayores frecuencias de micronúcleos (p= 0.013) y de células micronucleadas (p= 0.015). Conclusiones: Este estudio reafirma que los trabajadores expuestos a solventes orgánicos con polimorfismos de CYP2E1, específicamente con genotipo c1c1, tienen mayor probabilidad de presentar mutaciones en las células somáticas, condición asociada con una mayor susceptibilidad a enfermedades como el cáncer

Relationship of anti-tuberculosis drug-induced liver injury and genetic polymorphisms in CYP2E1 and GST

ABSTRACT Setting: Treatment of tuberculosis (TB) can result in Drug-Induced Liver Injury (DILI) since hepatotoxic metabolites are formed during the biotransformation of isoniazid (INH).DILI can be related to the genetic profile of the patient. Single nucleotide polymorphisms in the CYP2E1 gene and GSTM1 and GSTT1 deletion polymorphisms have been associated with adverse events caused by INH. Objective: To characterize the genetic polymorphisms of CYP2E1, GSTT1 and GSTM1 in TB carriers. Design: This is an observational prospective cohort study of 45 patients undergoing treatment of TB. PCR-RFLP and multiplex-PCR were used. Results: The distribution of genotypic frequency in the promoter region (CYP2E1 gene) was: 98% wild genotype and 2% heterozygous. Intronic region: 78% wild genotype; 20% heterozygous and 2% homozygous variant. GST enzyme genes: 24% Null GSTM1 and 22% Null GSTT1. Patients with any variant allele of the CYP2E1 gene were grouped in the statistical analyses. Conclusion: Patients with the CYP2E1 variant genotype or Null GSTT1 showed higher risk of presenting DILI (p = 0.09; OR: 4.57; 95% CI: 0.75-27.6). Individuals with both genotypes had no increased risk compared to individuals with one genotype.

Environmental exposure to benzene: evaluation of urinary S-PMmorphism (CYP2E1-129A and poly3G>C and NQO1 609C> T) in Campos Elíseos residents, Duque de Caxias, Rio de Janeiro State, Brazil / Exposição ambiental ao benzeno: avaliação do S-PMA urinário e de polimorfismos (CYP2E1-1293G>C e NQO1 609C> T) em residentes de Campos Elíseos, Duque de Caxias, Rio de Janeiro, Brasil / Exposición ambiental al benceno: análisis del S-PMA en la orina y polimorfismo (CYP2E1-1293G>C y NQO1 609C> T) en residentes de Campos Elíseos, Duque de Caxias, Río de Janeiro, Brasil

Benzene is one of the most important substances for assessment, due to its significant use, the environmental contamination resulting from its emission and the effects on human health. It is classified by the International Agency for Research on Cancer (IARC) as a known carcinogen to humans (group 1) and associated with the development of leukemia. In general, the population is exposed to this substance by inhaling contaminated air, which varies according to the location and intensity of its potential sources. The petrochemical industry is one of the most important sources of this compound. The municipality of Duque de Caxias, specifically the Campos Elíseos district, in Rio de Janeiro State, Brazil, houses the Industrial Complex of Campos Elíseos (PICE), a grouping of over 25 industries, which includes the second largest oil refinery in Brazil. Environmental contamination from the PICE has been recognized, but there is a lack of studies concerning its impact on the health of the surrounding population. S-phenylmercapturic acid (S-PMA) concentrations ranging from 0.80 to 8.01μg.g-1 creatinine were observed in the local population, apparently related to hematological changes also observed in exposed population. The quantifiable presence of urinary S-PMA from the benzene metabolism is associated with the fact that 60% of the participants present specific hematological changes, which may be due to the environmental benzene exposure. The allele and genotype frequencies of the CYP2E1 and NQO1 enzymes observed in the study population were similar to those reported in other studies. The presence of the variant allele in the NQO1 genotype may be a risk factor for the observed hematological changes.

O benzeno é uma das substâncias mais importantes para a biomonitorização, em função do uso disseminado, da contaminação ambiental que resulta da emissão e dos efeitos sobre a saúde humana. O benzeno é classificado pela Agência Internacional de Pesquisa em Câncer (IARC) como carcinógeno conhecido em seres humanos (grupo 1) e está associado ao desenvolvimento de leucemias. Em geral, a população fica exposta a essa substância através da inalação do ar contaminado, que varia de acordo com a localização e a intensidade das fontes potenciais. A indústria petroquímica é uma das fontes mais importantes desse composto. O Município de Duque de Caxias, especificamente o Distrito de Campos Elíseos, no Estado do Rio de Janeiro, Brasil, é sede do Polo Industrial de Campos Elíseos (PICE), um conjunto de mais de 25 indústrias que inclui a segunda maior refinaria de petróleo no Brasil. A contaminação ambiental produzida pelo PICE já é conhecida, mas faltam estudos sobre o impacto na saúde da população local. Foram observadas concentrações de ácido S-fenilmercaptúrico (S-PMA) entre 0,80 e 8,01μg.g-1 creatinina na população local, aparentemente implicadas nas alterações hematológicas também observadas na população exposta. A presença quantificável do S-PMA urinário do metabolismo do benzeno está associada ao fato de 60% dos participantes apresentarem alterações hematológicas específicas, o que pode ser devido à exposição ambiental ao benzeno. As frequências alélicas e genotípicas das enzimas CYP2E1 e NQO1, observadas na população do estudo, foram semelhantes àquelas relatadas em outros estudos. A presença da variante alélica do genótipo NQO1 pode ser um fator de risco para as alterações hematológicas observadas.

El benceno es una de las sustancias más importantes susceptibles de estudio, debido a su uso significativo, la contaminación ambiental resultante de sus emisiones y sus efectos sobre la salud humana. Está clasificado por el Centro Internacional de Investigaciones sobre el Cáncer (IARC) como un conocido carcinógeno para los humanos (grupo 1) y está asociado con el desarrollo de leucemias. En general, la población está expuesta a esta sustancia por inhalación de aire contaminado, que varía según el lugar y la intensidad de las emisiones. La industria petroquímica es un de las fuentes emisoras más importantes de este compuesto. La municipalidad de Duque de Caxias, específicamente el distrito de Campos Elíseos, en Río de Janeiro, Brasil, alberga el Complejo Industrial de Campos Elíseos (PICE), un conglomerado de más de 25 industrias, que incluye la segunda mayor refinería de petróleo en Brasil. La contaminación ambiental procedente del PICE ya ha sido reconocida, pero es notable la falta de estudios respecto a su impacto en la salud de la población circundante. Se observaron en la población local concentraciones de ácido s-fenilmercaptúrico (SPMA por sus siglas en inglés) que oscilan entre los 0,80 a 8,01μg.g-1 creatinina, aparentemente relacionadas con cambios hematológicos también hallados en la población expuesta. La presencia cuantificable de SPMA en la orina, procedente del metabolismo del benceno, está asociada con el hecho de que un 60% de los participantes presenta cambios específicos hematológicos, los cuales tal vez se deben a la exposición ambiental al benceno. Las frecuencias alélicas y genotípicas del CYP2E1 y enzimas NQO1 observadas en el estudio fueron similares a las reportadas en otros estudios. La presencia de la variante alélica en el genotipo NQO1 podría ser un factor de riesgo para los cambios hematológicos observados.

Rdh13 deficiency weakens carbon tetrachloride-induced liver injury by regulating Spot14 and Cyp2e1 expression levels / 医学前沿

Mitochondrion-localized retinol dehydrogenase 13 (Rdh13) is a short-chain dehydrogenase/reductase involved in vitamin A metabolism in both humans and mice. We previously generated Rdh13 knockout mice and showed that Rdh13 deficiency causes severe acute retinal light damage. In this study, considering that Rdh13 is highly expressed in mouse liver, we further evaluated the potential effect of Rdh13 on liver injury induced by carbon tetrachloride (CCl). Although Rdh13 deficiency showed no significant effect on liver histology and physiological functions under regular culture, the Rdh13 mice displayed an attenuated response to CCl-induced liver injury. Their livers also exhibited less histological changes and contained lower levels of liver-related metabolism enzymes compared with the livers of wild-type (WT) mice. Furthermore, the Rdh13 mice had Rdh13 deficiency and thus their liver cells were protected from apoptosis, and the quantity of their proliferative cells became lower than that in WTafter CCl exposure. The ablation of Rdh13 gene decreased the expression levels of thyroid hormone-inducible nuclear protein 14 (Spot14) and cytochrome P450 (Cyp2e1) in the liver, especially after CCl treatment for 48 h. These data suggested that the alleviated liver damage induced by CCl in Rdh13 mice was caused by Cyp2e1 enzymes, which promoted reductive CCl metabolism by altering the status of thyroxine metabolism. This result further implicated Rdh13 as a potential drug target in preventing chemically induced liver injury.

Comparison of toxic responses to acetaminophen challenge in ICR mice originating from different sources / 한국실험동물학회지

Acetaminophen (APAP) is the most common antipyretic analgesic worldwide. However, APAP overdose causes severe liver injury, especially centrilobular necrosis, in humans and experimental animals. At therapeutic dosage, APAP is mainly metabolized by sulfation and glucuronidation, and partly by cytochrome P450–mediated oxidation. However, APAP overdose results in production of excess reactive metabolite, N-acetyl-p-benzoquinone imine (NAPQI), by cytochromes P450; NAPQI overwhelms the level of glutathione (GSH), which could otherwise detoxify it. NAPQI binds covalently to proteins, leading to cell death. A number of studies aimed at the prevention and treatment of APAP-induced toxicity are underway. Rats are more resistant than mice to APAP hepatotoxicity, and thus mouse models are mainly used. In the present study, we compared the toxic responses induced by APAP overdose in the liver of ICR mice obtained from three different sources and evaluated the usability of the Korl:ICR stock established by the National Institute of Food and Drug Safety Evaluation in Korea. Administration of APAP (300 mg/kg) by intraperitoneal injection into male ICR mice enhanced CYP2E1 protein expression and depleted hepatic GSH level 2 h after treatment accompanied with significantly increased level of hepatic malondialdehyde, a product of lipid peroxidation. Regardless of the source of the mice, hepatotoxicity, as evidenced by activity of serum alanine aminotransferase, increased from 8 h and peaked at 24 h after APAP treatment. In summary, hepatotoxicity was induced after the onset of oxidative stress by overdose of APAP, and the response was the same over time among mice of different origins.

Polimorfismos en los genes de la enzima alcohol deshidrogenasa (ADH1) y la citocromo P450 2E1 (CYP2E1) en pacientes con cirrosis y carcinoma hepatocelular / Polymorphisms in alcohol dehydrogenase (ADH1) and cytochrome p450 2E1 (CYP2E1) genes in patients with cirrhosis and/or hepatocellular carcinoma

Introducción. Uno de los principales factores de riesgo del carcinoma hepatocelular es el consumo crónico de alcohol. En estudios en diferentes poblaciones, se sugiere que las variantes genéticas de las enzimas que participan en el metabolismo del alcohol, como la alcohol deshidrogenasa (ADH) y la citocromo P450 (CYP2E1), estarían asociadas con riesgo de enfermedades hepáticas terminales. Objetivo. Identificar y caracterizar las variantes alélicas de los genes ADH1B, ADH1C y CYP2E1 en pacientes colombianos con diagnóstico de cirrosis y carcinoma hepatocelular. Materiales y métodos. Se incluyeron muestras de pacientes atendidos entre el 2005 y el 2007, y entre el 2014 y el 2016, en la unidad de hepatología de un hospital de Medellín. La genotipificación de las muestras se hizo mediante reacción en cadena de la polimerasa (Polymerase Chain Reaction, PCR) con análisis de los polimorfismos en la longitud de los fragmentos de restricción (Restriction Fragment Length Polymorphism, RFLP). Los resultados se compararon con los de dos grupos de control y con lo reportado en la base de datos del 1000 Genomes Project. Resultados. Se recolectaron 97 muestras de pacientes con diagnóstico de cirrosis y carcinoma hepatocelular. Los dos factores de riesgo más frecuentes fueron el consumo crónico de alcohol (18,6 %) y las colangiopatías (17,5 %). Los genotipos más frecuentes en la población de estudio fueron el ADH1B*1/1 (82 %), el ADH1C*1/1 (59 %) y el CYP2E1*C/C (84 %). Conclusiones. En este primer estudio de los polimorfismos en pacientes colombianos con diagnóstico de cirrosis y carcinoma hepatocelular, los genotipos más frecuentes fueron el ADH1B*1/1, el ADH1C*1/1 y el CYP2E1*C/C. No se observaron diferencias estadísticamente significativas en la frecuencia de los genotipos entre los casos y los controles. Se requieren estudios adicionales en población colombiana para evaluar el riesgo de la enfermedad hepática terminal por consumo crónico de alcohol y la asociación con los polimorfismos.

Introduction: One of the most important risk factors for hepatocellular carcinoma (HCC) is alcohol consumption: Studies in different populations suggest that the risk of liver disease could be associated with genetic variants of the enzymes involved in alcohol metabolism, such as alcohol dehydrogenase (ADH) and cytochrome P450 CYP2E1. Objective: To identify and characterize the allelic variants of ADH1B, ADH1C and CYP2E1 genes in Colombian patients with cirrhosis and/or HCC. Materials and methods: We included samples from patients attending the hepatology unit between 2005-2007 and 2014-2016 of a hospital in Medellin. Samples were genotyped using PCR-RFLP. We compared the results with two control groups and the 1000 Genomes Project database. Results: We collected 97 samples from patients with a diagnosis of cirrhosis and/or HCC. The two main risk factors were chronic alcohol consumption (18.6%) and cholangiopathies (17.5%). The most frequent genotypes in the study population were ADH1B*1/1 (82%), ADH1C*1/1 (59%), and CYP2E1*C/C (84%). Conclusions: This first study of polymorphisms in Colombian patients diagnosed with cirrhosis and/or HCC showed genotypes ADH1B*1/1, ADH1C*1/1 and CYP2E1*C/C as the most frequent. We found no significant differences in the genotype frequency between cases and controls. Further studies are necessary to explore the association between polymorphisms and the risk of end-stage liver disease from alcohol consumption.

Hepatoprotective effect of Alhagi sparsifolia against Alcoholic Liver injury in mice

Overconsumption of alcohol leads to alcoholic liver disease (ALD). Natural compounds have been investigated previously for their hepatoprotective activities against liver injury. This study investigated the protective effect of Alhagi sparsifolia on ALD. Alcohol was administered to mice for three consecutive days; either alone or in combination with Alhagi sparsifolia extract (150, 300, 600 mg/kg). Serum aspartate aminotransferase and alanine transaminase as biomarkers of liver injury, the content of malonaldehyde, hydrogen peroxide (H2O2) and glutathione which indicated the redox status of liver and the antioxidant enzyme activity of super oxide dismutase were detected, respectively. Moreover, the expression of protein cytochrome P450 2E1 (CYP2E1) the key enzyme of alcohol metabolism, and also tested by western blot experiment. Subsequently, the mRNA levels of inflammatory factors including TNF- α and TLR4 was determined real-time PCR. Results showed that Alhagi sparsifolia significantly alleviated alcohol-induced liver injury by reducing serum ALT and AST, inhibiting MDA and H2O2 content, increasing SOD, and GSH level in the liver (P< 0.05). In addition, the Alhagi sparsifolia treatment inhibited the expression of CYP2E1 (P< 0.05). The results suggest that Alhagi sparsifolia could be a promising natural substance for ameliorating acute alcohol-induced oxidative stress and hepatic injury

Hepaprotective Effect of Standardized Ecklonia stolonifera Formulation on CCl₄-Induced Liver Injury in Sprague-Dawley Rats

The liver is an essential organ for the detoxification of exogenous xenobiotics, drugs and toxic substances. The incidence rate of non-alcoholic liver injury increases due to dietary habit change and drug use increase. Our previous study demonstrated that Ecklonia stolonifera (ES) formulation has hepatoprotective effect against alcohol-induced liver injury in rat and tacrine-induced hepatotoxicity in HepG2 cells. This present study was designated to elucidate hepatoprotective effects of ES formulation against carbon tetrachloride (CCl₄)-induced liver injury in Sprague Dawley rat. Sixty rats were randomly divided into six groups. The rats were treated orally with ES formulation and silymarin (served as positive control, only 100 mg/kg/day) at a dose of 50, 100, or 200 mg/kg/day for 21 days. Seven days after treatment, liver injury was induced by intraperitoneal injection of CCl₄ (1.5 ml/kg, twice a week for 14 days). The administration of CCl₄ exhibited significant elevation of hepatic enzymes (like AST and ALT), and decrease of antioxidant related enzymes (superoxide dismutase, glutathione peroxidase and catalase) and glutathione. Then, it leaded to DNA damages (8-oxo-2′-deoxyguanosine) and lipid peroxidation (malondialdehyde). Administration of ES formulation inhibited imbalance of above factors compared to CCl₄ induced rat in a dose dependent manner. Real time PCR analysis indicates that CYP2E1 was upregulated in CCl₄ induced rat. However, increased gene expression was compromised by ES formulation treatment. These findings suggests that ES formulation could protect hepatotoxicity caused by CCl₄ via two pathways: elevation of antioxidant enzymes and normalization of CYP2E1 enzyme.

A non-hepatotropic parasite infection increases mortality in the acetaminophen-induced acute liver failure murine model: possible roles for IL-5 and IL-6

We evaluated the effects of a non-hepatotropic parasite infection (Taenia crassiceps) on the outcome of acetaminophen-induced acute liver failure in mice. Uninfected and T. crassiceps infected mice orally received either 300 mg/kg acetaminophen or water as vehicle (n = 5 per group). Survival analysis, hepatocyte necrosis, alanine aminotransferase (ALT) levels, CYP2E1 protein, interleukin (IL-) 5, and IL-6 were assessed for all groups. All infected mice died within 16 h after exposure to acetaminophen (Tc+APAP group), whereas only one-third of uninfected animals exposed to acetaminophen (APAP group) died. Uninfected (Control group) and infected (Tc group) mice that received the vehicle showed no liver damage. Tc+APAP mice exhibited massive liver necrosis characterised by marked balloning degeneration of hepatocytes and higher serum ALT compared to Control, Tc, and APAP animals. Liver tissue from Tc+APAP mice also displayed increased expression of CYP2E1 protein and higher mRNA and protein levels of IL-5 and IL-6 compared to the other groups. These findings suggest that non-hepatotropic parasite infections may increase mortality following acute liver failure by promoting hepatocyte necrosis via IL-5 and IL-6-dependent CYP2E1 overproduction. This study identifies new potential risk factors associated with severe acute liver failure in patients.

Alcohol, cirrosis y predisposición genética / Alcohol, Cirrhosis, and Genetic Predisposition

La cirrosis hepática es la tercera causa de muerte alrededor del mundo que es atribuible al consumo de alcohol. Más del 80% de los consumidores crónicos de alcohol desarrollan esteatosis y entre el 20% al 40% presentan otras complicaciones como fibrosis, hepatitis alcohólica y cirrosis; sin embargo, no todos los individuos con consumo crónico de alcohol desarrollan cirrosis, en parte debido al componente genético de cada individuo. El grado de actividad de las enzimas que metabolizan el alcohol está influenciado por polimorfismos presentes en los genes que codifican para estas enzimas, y corresponde a uno de los factores determinantes para el desarrollo de una hepatopatía terminal en respuesta al consumo de alcohol. Entre las enzimas implicadas en el metabolismo del alcohol están la alcohol deshidrogenasa (ADH), el citocromo P450 2E1 (CYP2E1) y la acetaldehído deshidrogenasa (ALDH), de las cuales se ha reportado que la mayor actividad de ADH y CYP2E1 y la menor actividad de ALDH pueden conferir riesgo en algunas poblaciones por la acumulación de acetaldehído, el cual es tóxico para el organismo. Se realizó una revisión en la literatura de los principales aspectos del metabolismo del alcohol y polimorfismos (genotipos) de enzimas que intervienen en el metabolismo del alcohol como factor de riesgo. Esto se hizo mediante la búsqueda de material bibliográfico a través de la base de datos PubMed desde 1990 hasta el 2013 utilizando las palabras claves alcohol liver disease, ADH, ALDH, CYP2E1 y polymorphism.

Liver cirrhosis is the third most common cause of death attributable to alcohol consumption throughout the world. More than 80% of chronic drinkers develop steatosis, and 20% to 40% develop other complications such as fibrosis, alcoholic hepatitis and cirrhosis. However, not everyone who chronically consumes alcohol develops cirrhosis. This is partly because of the genetic component of each individual. The level of activity of the enzymes that metabolize alcohol is influenced by polymorphisms of the genes that coding for these enzymes. This is one of the determining factors in the development of terminal liver disease in response to alcohol consumption. Among the enzymes involved in alcohol metabolism are alcohol dehydrogenase (ADH), cytochrome P450 2E1 (CYP2E1) and acetaldehyde dehydrogenase (ALDH). It has been reported that higher levels of activity of ADH and CYP2E1 and lower levels of activity of ALDH may be risk factors in some populations for accumulation of acetaldehyde which is toxic for the organism. This literature review covers the most important aspects of alcohol metabolism including polymorphisms (genotypes) of enzymes involved in the metabolism of alcohol as a risk factor. A search through the PubMed database from 1990 to be held 2013 was conducted using the keywords alcoholic liver disease, ADH, ALDH, CYP2E1, and polymorphism.

Maternal protein restriction during lactation modulated the expression and activity of rat offspring hepatic CYP1A1, CYP1A2, CYP2B1, CYP2B2, and CYP2E1 during development

Early nutrition plays a long-term role in the predisposition to chronic diseases and influences the metabolism of several drugs. This may happen through cytochromes P450 (CYPs) regulation, which are the main enzymes responsible for the metabolism of xenobiotics. Here, we analyzed the effects of maternal protein restriction (MPR) on the expression and activity of hepatic offspring’s CYPs during 90 days after birth, using Wistar rats as a mammal model. Hepatic CYP1A1, CYP1A2, CYP2B1, CYP2B2 and CYP2E1 mRNA and protein expression, and associated catalytic activities (ECOD, EROD, MROD, BROD, PROD and PNPH) were evaluated in 15-, 30-, 60-, and 90-day-old offspring from dams fed with either a 0% protein (MPR groups) or a standard diet (C groups) during the 10 first days of lactation. Results showed that most CYP genes were induced in 60- and 90-day-old MPR offspring. The inductions detected in MPR60 and MPR90 were of 5.0- and 2.0-fold (CYP1A2), 3.7- and 2.0-fold (CYP2B2) and 9.8- and 5.8– fold (CYP2E1), respectively, and a 3.8-fold increase of CYP2B1 in MPR90. No major alterations were detected in CYP protein expression. The most relevant CYP catalytic activities’ alterations were observed in EROD, BROD and PNPH. Nevertheless, they did not follow the same pattern observed for mRNA expression, except for an induction of EROD in MPR90 (3.5-fold) and of PNPH in MPR60 (2.2-fold). Together, these results suggest that MPR during lactation was capable of altering the expression and activity of the hepatic CYP enzymes evaluated in the offspring along development.

Induction of rat hepatic CYP2E1 expression by arecoline in vivo / 药学学报

The regulation mechanism of arecoline on rat hepatic CYP2E1 was studied in vivo. After oral administration of arecoline hydrobromide (AH; 4, 20 and 100 mg x kg(-1) x d(-1)) to rats for one week, the hepatic CYP2E1 mRNA level remained unchanged, but the hepatic CYP2E1 protein content was dose-dependently increased. Additionally, although the hepatic CYP2E1 activity was induced by AH treatment, the induction was attenuated with the increase in dosage. The results indicate that the effect of arecoline on rat hepaticdoes not involve transcriptional activation of the gene, but largely involves the stabilization of CYP2E1 protein against degradation or increased efficiency of CYP2E1 mRNA translation, and additionally involve the post- ranslational modification of CYP2E1 protein. Furthermore, the CYP2E1 response is fairly equal among the different species, the induction of rat hepatic CYP2E1 by arecoline suggests that there is a risk of metabolic interaction among the substrate drugs of CYP2E1 in betel-quid use human.

A Fruit Extract of Paeonia anomala Attenuates Chronic Alcohol-induced Liver Damage in Rats

Prolonged alcohol consumption causes alcoholic liver damage due to the generation of reactive oxygen species, the accumulation of fatty acids, and an increase in inflammatory cytokines in the liver. In this study, the protective effect of a fruit extract of Paeonia anomala (FEPA) against chronic alcohol-induced liver damage was evaluated in Sprague-Dawley rats fed an ethanol or a control Lieber-DeCarli diet for 5 weeks to induce alcoholic liver damage. FEPA (50, 25, and 10 mg/kg body weight/day) as well as the reference control silymarin (25 mg/kg body weight/day) were administered along with the ethanol diet. FEPA protected against increases in alanine aminotransferase and aspartate aminotransferase in serum and attenuated alcohol-induced increases in triglycerides, tumor necrosis factor alpha, thiobarbituric acid-reactive substances, and cytochrome P450 2E1 enzyme activity in the liver compared with the group treated with ethanol only. Anti-oxidative defenses such as the total glutathione level and glutathione peroxidase activity were increased by FEPA treatment. These results suggest that FEPA exerts protective effects against chronic alcohol-induced liver damage by attenuating hepatosteatosis and pro-inflammatory cytokine production and enhancing anti-oxidative defense mechanisms in the liver.

Study on inhibitory effect of water extract of Polygonum multiflorum on CYP1A2 and CYP2E1 enzymatic activities and mRNA expressions in rat liver / 中国中药杂志

Rats were continuously given different doses of water extract of Polygonum multiflorum (1, 10 g x kg(-1)) for 7 days to prepare liver microsomes. Cocktail in vitro incubation approach and Real-time quantitative PCR technology were used to observe the effect of water extract of P. multiflorum on CYP450 enzymatic activities and mRNA expressions in rat liver. Compared with the blank control group, both 1, 10 g x kg(-1) water extract of P. multiflorum treated groups showed significant inhibitions in CYP2E1 enzymatic activities and mRNA expressions (enzymatic activities of CYP2E1, P < 0.01; mRNA expression of CYP2E1, P < 0.05 in 1 g x kg(-1) group, P < 0.01 in 10 g x kg(-1) group). They revealed a significant increase in the enzymatic activity of CYP3A1 (P < 0.01), but without significant change in mRNA expressions. The 10 g x kg(-1) group showed a significant inhibition in CYP1A2 enzymatic activities and mRNA expressions in rat livers (P < 0.01).

Association of single nucleotide polymorphisms of cytochrome P450 gene with susceptibility to gout in ethnic Han males from coastal regions of Shandong province / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To assess the association of cytochrome P450 gene single nucleotide polymorphisms (SNPs) with susceptibility to gout in ethnic Han males from coastal regions of Shandong province.</p><p><b>METHODS</b>Four hundred and eighty male patients with gout and 480 healthy male controls were included. Genotyping was carried out with a custom Illumina GoldenGate Genotyping assay to detect SNP rs2275620 of CYP2C8 gene, SNP rs2070676 of CYP2E1 gene, SNP rs837395 of CYP4B1 gene, and SNP rs194150 of TBXAS1 gene. The association was assessed with chi-square test.</p><p><b>RESULTS</b>No significant difference has been found between the two groups in regard to the genotypic and allelic frequencies of the TT, AT, AA genotypes and A, T alleles of the SNP rs2275620 of the CYP2C8 gene (P=0.88; P=0.97), the CC, CG, GG genotypes and C,G alleles of SNP rs2070676 of the CYP2E1 gene (P=0.24; P=0.09), the TT, AT, AA genotypes and A, T alleles of SNP rs837395 of the CYP4B1 (P=0.88; P=0.97), and TT, AT, AA genotypes and the A,T alleles of SNP rs194150 of TBXAS1 gene (P=0.15; P=0.06).</p><p><b>CONCLUSION</b>This study has identified no association of SNP loci rs2275620(A/T) of CYP2C8, rs2070676(C/G) of CYP2E1, rs837395(A/T) of CYP4B1 and rs194150(A/T) of TBXAS1 with gout in ethnic Han males from coastal regions in Shandong province. However, our result needs to be replicated in larger sets of patients collected from other regions and populations.</p>

Inhibiting effects of three components of Astragalus membranaceus on oxidative stress in Chang Liver cells / 中国中药杂志

The main objective of this research is to investigate the effects of astragaloside IV, calycosin separately glucoside, formononetin on oxidative stress in Chang Liver cells induced by H2O2. In the experiments, Chang Liver cells (a kind of normal human hepatocytes) were used as the research object, bifendate which has a clear hepatoprotective effect was used as the positive control drug, then the oxidative damage model of Chang Liver cells were established by H2O2. Cells were divided into six groups: blank control group, oxidative stress group, astragaloside IV group, calycosin separately glucoside group, formononetin group and positive control group. Then endogenous antioxidant system related indexes were detected by micro plate and colorimetric method; intracellular reactive oxygen species (ROS) were detected by DCFH-DA fluorescent probe; and the expressions of CYP2E1 were evaluated by liver microsomes, mRNA, and protein, respectively with spectrophotometry, Real-time PCR method, and Western blot technique. Results showed that H2O2 decreased antioxidant activity, and increased ROS level and expression of CYP2E1. The above oxidative stress status had been changed with protections of the three components of Astragalus membranaceus (compared with oxidative stress group, P < 0.05, P < 0.01), which taken as a whole had equivalent effects as the drug of positive control group( bifendate). Taken together, three Astragalus membranaceus ingredients all had significant or extremely significant inhibiting effects on oxidative damaged Chang Liver cells which were induced by H2O2, and the oxidative damage of Chang Liver cells had been relieved.