Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019204 (hepatocellular carcinoma)
 71,386 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The oxidation of O6-benzylguanine, an inactivator of O6-alkylguanine-DNA alkyltransferase, was examined using human liver cytosol, microsomes, and several P450 isoforms. Incubation of O6-benzylguanine with human liver cytosol resulted in the formation of O6-benzyl-8-oxoguanine, which was inhibited by menadione, a potent inhibitor of aldehyde oxidase. Inhibition by allopurinol, a xanthine oxidase inhibitor, was less dramatic. Oxidation of O6-benzylguanine also occurred with pooled human liver microsomes and was inhibited by both furafylline and troleandomycin, selective inhibitors of CYP1A2 and CYP3A4, respectively. Human P450s CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2E1, and CYP3A4 expressed in Hep G2 hepatoma cells using vaccinia virus vectors were incubated with 10 or 200 microM O6-benzylguanine. At 10 microM, O6-benzylguanine was oxidized primarily by CYP1A2 and to a lesser extent by CYP3A4. However, an appreciable increase in CYP3A4 contribution was noted at 200 microM. CYP1A2 exhibited a more than 200-fold higher relative catalytic activity (Vmax/Km) compared with CYP3A4. Therefore, at therapeutically relevant concentrations of O6-benzylguanine, CYP1A2 could be primarily involved in its oxidation since it shows a much lower Km value (1.3 microM) than CYP3A4 (52.2 microM) and cytosol (81.5 microM). However, one would expect interindividual variation in the extent of oxidation of O6-benzylguanine depending on the levels of aldehyde oxidase, CYP1A2, and CYP3A4.
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PMID:Human liver oxidative metabolism of O6-benzylguanine. 750 88

Experiments were carried out to stably and constitutively express the coding sequence of the human cytochrome P4502E1 in HepG2, a human-hepatoma-derived cell line, by recombinant retroviral expression. Southern blot analysis showed a successful integration of a single copy of unaltered viral DNA into the genome of each transduced clone tested. Northern blot analysis showed that the transduced clones produced an RNA species which hybridized to the CYP2E1 cDNA probe. Western blot analysis using anti-human P4502E1 IgG indicated that the transduced clones produced a protein band with molecular weight of 54 000. Microsomes from transduced clones were catalytically active with p-nitrophenol, dimethylnitrosamine, aniline, and ethanol as substrates; little or no activity was found with control clones. Oxidation of p-nitrophenol was inhibited by anti-human P4502E1 IgG, diethyl dithiocarbamate, 4-methylpyrazole, and ethanol. ESR spectroscopy showed that microsomes from clone MV2E1-9 produced superoxide radical. Rates were an order of magnitude higher than that for control microsomes, most likely reflecting the loose coupling associated with P4502E1. The rate of H2O2 production by microsomes from MV2E1-9 was 2-fold greater than that of control clones. The elevated rate of H2O2 production in clone MV2E1-9 is about half the rate of superoxide radical production, suggesting that this H2O2 is largely derived from superoxide radical dismutation. Microsomal lipid peroxidation was determined using ferric-ATP as the iron catalyst. When the concentration of iron was "high" (0.025 mM), rates of production of thiobarbituric acid reactive components were identical for microsomes from MV2E1-9 and control clones. However, when the concentration of iron was lowered to 0.005 mM, control clones did not display lipid peroxidation, whereas microsomes from MV2E1-9 were reactive. This peroxidation was sensitive to antioxidants such as trolox, propyl gallate, and glutathione but not to catalase or superoxide dismutase. Rates of superoxide and H2O2 production and of lipid peroxidation were 7-20-fold higher on a per nanomole of P450 basis with clone MV2E1-9 compared to human liver microsomes, indicating that the human P4502E1 is especially reactive in production of reactive oxygen intermediates and in catalysis of lipid peroxidation.
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PMID:Stable expression of human cytochrome P4502E1 in HepG2 cells: characterization of catalytic activities and production of reactive oxygen intermediates. 768 64

The rate of formation of styrene glycol from styrene was compared in human, rat, and mouse liver microsomes. At a low styrene concentration (0.085 mM), the rates decreased in the order, mouse (2.43 +/- 0.29 nmol/(mg of protein.min)) > rat (1.07 +/- 0.20) > human (0.73 +/- 0.45); at a high concentration (1.85 mM), the order was rat (4.21 +/- 0.72) > mouse (2.72 +/- 0.11) > human (1.91 +/- 0.84). Kinetic analysis indicated the presence of at least two forms of styrene-metabolizing cytochrome P450s with different Km values in human liver microsomes. Styrene was also metabolized in human lung microsomes: the rate of styrene glycol formation was higher in the lung microsomes from smokers than in those from current nonsmokers. The P450 forms responsible for transforming styrene to styrene glycol were determined by analyzing cDNA-expressed individual P450 forms produced in cultured hepatoma G2 cells by recombinant vaccinia viruses. Of the 12 human P450 forms studied, CYP2B6 and CYP2E1 existing in human liver and/or lungs and CYP2F1 in human lungs were the most active in the forming of styrene glycol, followed by CYP1A2 and CYP2C8. Human CYP3A3, CYP3A4, CYP3A5, and CYP4B1 also catalyzed the metabolism but were much less active. CYP2A6, CYP2C9, and CYP2D6 had only a little detectable activity. CYP1A2, CYP2B6, CYP2C8, CYP2E1, and CYP3A4/3A3 were expressed in human liver microsomes, and CYP2C8 was expressed in human lung microsomes, although the expression of CYP2F1 and CYP4B1 could not be investigated. These data indicate that several human hepatic and/or pulmonary P450 forms are capable of metabolizing styrene, albeit at different rates.
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PMID:Styrene metabolism by cDNA-expressed human hepatic and pulmonary cytochromes P450. 769 48

Acetaminophen (APAP) when administered in excess can cause severe hepatic necrosis in vivo. To study the mechanism of APAP toxicity and the role of cytochrome P450, a previously established human hepatoma HepG2 subline, MVh2E1-9, that constitutively expresses human CYP2E1 was used as a model. At high concentrations (above 5 mM) and when intracellular reduced glutathione (GSH) was depleted, APAP caused severe cytotoxicity in MVh2E1-9, but not in MV-5 cells which lack CYP2E1. The APAP cytotoxicity was dependent on the concentration of APAP and time of exposure, and could be blocked by 4-methylpyrazole, ethanol, diallyl sulfide, N-acetylcysteine and N-t-butyl-alpha-phenylnitrone, but not by propylgallate, an inhibitor of lipid peroxidation. Significantly more 14C-labeled APAP protein adduct was detected in MVh2E1-9 cells than MV-5 cells, especially after depletion of GSH. The formation of the APAP adducts could be inhibited by the same agents which prevent APAP cytotoxicity. At a lower concentration (1-2 mM), APAP inhibited proliferation in both MVh2E1-9 and the control MV-5 cells to similar extents. This antiproliferative action of APAP did not require depletion of GSH as did the cytotoxic action of APAP. These data suggest that APAP has a dual toxic effect on MVh2E1-9 cells: a P450-independent antiproliferative effect and the CYP2E1-dependent cytotoxic effect. These results demonstrate the ability of human CYP2E1 to activate APAP to reactive metabolites which form covalent protein adducts and cause toxicity to a hepatoma cell line.
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PMID:Cytotoxicity of acetaminophen in human cytochrome P4502E1-transfected HepG2 cells. 779 Nov 25

A panel of four novel human hepatoma cell lines was isolated from a single tumor from a male individual. BC1, B16 and B16A2 lines were well differentiated, while cells of the B9 line were only poorly differentiated, being essentially negative for the functions analyzed. These cell lines have been surveyed for expression of a large set of plasma proteins, accumulation of liver-specific mRNAs and DNA-binding activity of ubiquitous and liver-enriched transcription factors. BC1 cells expressed the highest levels of albumin mRNA, whereas B16 and B16A2 cells accumulated the largest amounts of haptoglobin mRNA. In addition, B16 and B16A2 cells were unique in that they expressed CYP2E1 mRNA, a species absent from the available human liver cells, including HepG2 hepatoma cells, and 3-methylcholanthrene-inducible CYP1A2 mRNA. The activities of genes encoding transcription factors were evidenced in all four cell lines which expressed mRNAs for nuclear factor interleukin 6 and hepatocyte nuclear factor 1 (HNF) together with the DNA-binding activity of NFY and AP1 nuclear proteins. Strikingly, HNF-1 and HNF-4-like DNA-binding activities were restricted to BC1, B16 and B16A2 cells, supporting the idea of the potential role of these (or closely related) factors in the maintenance and/or in the establishment of the differentiated phenotype. B9 cells contained variant HNF1-like DNA-binding activity, similar to dedifferentiated rat hepatoma cells of the H5 line. CCAAT/enhancer-binding protein and HNF-3-like activities were found in all cell lines, although at a lower level and/or activity in B9 cells. Finally, transfection experiments of plasmids containing the whole hepatitis-B virus genome demonstrated that B16 cells, but not B9 cells, were able to support hepatitis-B virus replication and virion production, in agreement with the notion that HNF-1 activity is necessary for viral replication. We believe that the specific complement of transcription factors expressed in the differentiated BC1, B16 and B16A2 cells, and in the poorly differentiated B9 cells, will allow studies on the regulation of hepatic gene expression in these human lines, and will also aid the analysis of xenobiotic metabolism and the biology of hepatitis-B virus replication.
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PMID:trans-Acting factors, detoxication enzymes and hepatitis B virus replication in a novel set of human hepatoma cell lines. 868 51

Increased risk of environmentally induced cancer is associated with various types of exposures and host factors, including differences in carcinogen metabolism. Since many carcinogenic compounds require metabolic activation to enable them to react with cellular macromolecules, individual features of carcinogen metabolism may play an essential role in the development of environmental cancer. In this context, cigarette smoking has often been the main type of carcinogenic exposure examined in human studies. Increasing attention has recently been paid to the dose level at which individual susceptibility may be observed. Present studies on increased risk of smoking-related lung cancer associated with phenotypic or genotypic variation of the genes encoding for CYP1A1 or CYP2D6 enzymes are summarized. Similarly, higher risks of lung or bladder cancer seen at various levels of smoking in association with polymorphism of the glutathione S-transferase gene GSTM1 or NAT1 and NAT2 genes involved in N-acetylation are reviewed. Finally, the influence of CYP2E1, GSTM1, or the combined at-risk genotype on the risk of hepatocellular carcinoma in smokers is briefly discussed.
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PMID:Interaction between dose and susceptibility to environmental cancer: a short review. 925 56

Induction of CYP2E1 (cytochrome P450 2E1) by ethanol appears to be one of the central pathways by which ethanol generates a state of oxidative stress. CYP2E1 is a loosely coupled enzyme; formation of reactive oxygen species occurs even in the absence of added substrate. GSH is critical for preserving the proper cellular redox balance and for its role as a cellular protectant. Since cells must maintain optimal GSH levels to cope with a variety of stresses, the goal of this study was to characterize the GSH homeostasis in human hepatocarcinoma cells (HepG2) that overexpress CYP2E1. This study was prompted by the finding that toxicity in CYP2E1-overexpressing cells was markedly enhanced after GSH depletion by buthionine sulfoximine treatment. CYP2E1-overexpressing cells showed a 40-50% increase in intracellular H(2)O(2); a 30% increase in total GSH levels; a 50% increase in the GSH synthesis rate; and a 2-fold increase in gamma-glutamylcysteine synthetase heavy subunit (GCS-HS) mRNA, the rate-limiting enzyme in GSH synthesis. This GCS-HS mRNA increase was due to increased synthesis since nuclear run-on assays showed increased transcription in CYP2E1-expressing cells, and the GCS-HS mRNA decay after actinomycin D treatment was similar in CYP2E1-expressing cells and empty vector-transfected cells. The facts that treatment with GSH ethyl ester almost completely prevented the increase in GCS-HS mRNA and decreased H(2)O(2) levels and that transient transfection with catalase (but not manganese-superoxide dismutase) produced a decrease in GCS-HS mRNA only in CYP2E1-expressing cells suggest a possible role for H(2)O(2) in the induction of GCS-HS gene transcription. In contrast to results with HepG2 cells expressing CYP2E1, no increase in GCS-HS mRNA was found with a HepG2 cell line engineered to express human cytochrome P450 3A4. In summary, CYP2E1 overexpression in HepG2 cells up-regulates the levels of reduced GSH by transcriptional activation of GCS-HS; this may reflect an adaptive mechanism to remove CYP2E1-derived oxidants such as H(2)O(2).
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PMID:CYP2E1 overexpression in HepG2 cells induces glutathione synthesis by transcriptional activation of gamma-glutamylcysteine synthetase. 1074 80

Much progress has been made in the understanding of the pathogenesis of alcoholic liver disease, resulting in improvement of treatment. Therapy must include correction of nutritional deficiencies, while taking into account changes of nutritional requirements. Methionine is normally activated to S-adenosylmethionine (SAMe). However, in liver disease, the corresponding enzyme is depressed. The resulting deficiencies can be attenuated by the administration of SAMe but not by methionine. Similarly, phosphatidylethanolamine methyltransferase activity is depressed, but the lacking phosphatidylcholine (PC) can be administrated as polyenylphosphatidylcholine (PPC). Chronic ethanol consumption increases CYP2E1, resulting in increased generation of toxic acetaldehyde and free radicals, tolerance to ethanol and other drugs, and multiple ethanol-drug interactions. Experimentally, PPC opposes CYP2E1 induction and fibrosis. Alcoholism and hepatitis C infection commonly co-exist, with acceleration of fibrosis, cirrhosis, and hepatocellular carcinoma. PPC is being tested clinically as a corresponding antifibrotic agent. Available antiviral agents are contraindicated in the alcoholic. Anti-inflammatory agents, such as steroids, may be selectively useful. Finally, anticraving agents, such as naltrexone or acamprosate, should be part of therapy.
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PMID:Liver diseases by alcohol and hepatitis C: early detection and new insights in pathogenesis lead to improved treatment. 1126 19

This article represents the proceedings of a symposium at the 2000 ISBRA Meeting in Yokohama, Japan. The chairs were Samuel W. French and R. J. Mayer. The presentations were (1) The ubiquitin-proteasome 26s pathway in liver cell protein turnover: Effect of alcohol and drugs, by Samuel W. French and F. Bardag-Gorce; (2) The role of CYP2E1 phosphorylation and degradation pathway in the induction of the enzyme, by Magnus Ingelman-Sundberg; (3) Role of proteasome in the proteolysis of oxidized proteins in experimental chronic alcoholism, by Helen Rouach; (4) Alcohol, proteolysis and liver cancer, by R. J. Mayer; (5) Effect of ethanol feeding on the ATP-ubiquitin-proteasome pathway in the liver cell, by F. Bardag-Gorce; (6) Novel mechanisms and targets for intracellular transport of CYP2E1, by E. Neve; and (7) Gankyrin, an oncoprotein commonly over expressed in hepatoma, by H. Higashitsuji.
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PMID:The ubiquitin-proteasome 26s pathway in liver cell protein turnover: effect of ethanol and drugs. 1139 Oct 75

This article represents the proceedings of a symposium at the 2000 ISBRA Meeting in Yokohama, Japan. The chairs were Terrence M. Donohue, Jr, and Dahn L. Clemens. The presentations were (1) Characterization of single and double recombinant hepatoma cells that express ethanol-metabolizing enzymes, by Terrence M. Donohue, Jr; (2) Inhibition of cell growth by ethanol metabolism, by Dahn L. Clemens; (3) Use of transfected HeLa cells to study the genesis of alcoholic fatty liver, by Andrea Galli and David Crabb; (4) CYP2E1-mediated oxidative stress induces COL1A2 mRNA in hepatic stellate cells and in a coculture system of HepG2 and stellate cells, by Natalia Nieto; (5) Transforming growth factor-alpha secreted from ethanol-exposed hepatocytes contributes to development of alcoholic hepatic fibrosis, by Junji Kato; and (6) Effect of ethanol on Fas-dependent caspase-3 activation and apoptosis in CD4+ T cells, by Shirish S. Barve.
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PMID:Use of cultured cells in assessing ethanol toxicity and ethanol-related metabolism. 1141 62


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