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)

Mammalian 3 alpha-hydroxysteroid dehydrogenases (3 alpha-HSDs) regulate steroid hormone levels. cDNA cloning indicates that the rat and human liver isoforms display high sequence identity and that they belong to the aldo-keto reductase (AKR) superfamily. Of these the most extensively characterized is rat liver 3 alpha-HSD. The recently solved X-ray crystal structure shows that this enzyme adopts an (alpha/beta)8-barrel scaffold (Hoog et al. 1994). NAD(P)H binds in an extended anti-conformation and lies along the inner surface of the barrel. The nicotinamide ring is stabilized by interaction with Y216. The 4-pro(R)-hydrogen transferred in the reaction is in close proximity to Y55. K84, D50 and H117 which are implicated in catalysis. These residues are located at the base of a hydrophobic pocket which is presumed to be involved in binding steroid hormone. This catalytic tetrad is conserved in members of the AKR superfamily. Mutant enzymes support roles for Y55 in steroid binding and for K84 as the general acid involved in catalysis. The gene for rat 3 alpha-HSD has been cloned and is 47 kb in length and contains 9 exon-intron boundaries which are highly conserved in the human gene(s). The 5'-flanking regions of the rat and human genes contain consensus sequences for AP-1, Oct-1 and multiple copies of perfect and imperfect steroid hormone response elements (REs) (estrogen, glucocorticoid (GRE), and progesterone) which may comprise a steroid response unit (SRU) (Lin & Penning 1995). Constitutive and regulated expression of the rat 3 alpha-HSD gene has been studied by transiently transfecting reporter gene (chloramphenicol acetyltransferase, CAT) constructs into human hepatoma (HepG2) cells. With respect to the transcription start-site (+1), a proximal (-498 to -199bp) and distal (-20 to -4.0kb) enhancer, as well as a powerful silencer (-755 to -498 bp) were located in the promoter. Band-shift and supershift assays provide evidence that Oct-1 binds to the silencer. Tandem repeats of the imperfect proximal and distal GREs that reside in the SRU were inserted into tk-CAT vectors and transiently transfected. Stimulation of transfected cells with dexamethasone resulted in robust CAT activity. These data indicate that glucocorticoids may positively regulate transcription of the rat 3 alpha-HSD gene from the SRU.
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PMID:3 alpha-hydroxysteroid dehydrogenase: three dimensional structure and gene regulation. 894 1

Caffeic acid phenethyl ester (CAPE) is a phenolic antioxidant derived from the propolis of honeybee hives. CAPE was shown to inhibit the formation of intracellular hydrogen peroxide and oxidized bases in DNA of 12-O-tetradecanoylphorbol-13-acetate (TPA)-treated HeLa cells and was also found to induce a redox change that correlated with differential growth effects in transformed cells but not the nontumorigenic parental ones. Mediated via the electrophile or human antioxidant response element (hARE), induction of the expression of NAD(P)H quinone oxidoreductase (NQO1) and glutathione S-transferase Ya subunit genes by certain phenolic antioxidants has been correlated with the chemopreventive properties of these agents. Here, we determined by Northern analysis that CAPE treatment of hepatoma cells stimulates NQO1 gene expression in cultured human hepatoma cells (HepG2), and we characterized the effects of CAPE treatment on the expression of a reporter gene either containing or lacking the hARE or carrying a mutant version of this element in rodent hepatoma (Hepa-1) transfectants. A dose-dependent transactivation of human hARE-mediated chloramphenicol acetyltransferase (cat) gene expression was observed upon treatments of the Hepa-1 transfectants with TPA, a known inducer, as well as with CAPE. The combined treatments resulted in an apparent additive stimulation of the reporter expression. To learn whether this activation of cat gene expression was effected by protein kinase C in CAPE-treated cells, a comparison was made of cat gene activity after addition of calphostin, a protein kinase C inhibitor. Calphostin reduced the cat gene induction by TPA but not by CAPE, suggesting that stimulation of gene expression in this system by these agents proceeds via distinct mechanisms. Band-shift experiments to examine binding of transactivator proteins from nuclear extracts of treated and untreated cells to a hARE DNA probe showed that TPA exposure increased the binding level. In contrast, binding of factors to this probe was inhibited after either in vivo treatment of cells with CAPE or in vitro addition of this compound to the nuclear extract. In view of the clear stimulation by CAPE of gene expression mediated by hARE, possible explanations of this result are discussed.
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PMID:Caffeic acid phenethyl ester stimulates human antioxidant response element-mediated expression of the NAD(P)H:quinone oxidoreductase (NQO1) gene. 901 71

The metabolism of acetaldehyde (ACA), benzaldehyde (BA), propionaldehyde (PA) and valeraldehyde (VA) has been studied in two hepatoma cell lines, the rat HTC and mouse Hepa 1c1c7 cells. The cytotoxicity of the four aldehydes to these two cell lines has been compared. The end-points for evaluating cytotoxicity were 1) total macromolecular content (TMC) of confluent cultures, and 2) colony forming ability of dividing cells. These two assay systems had different sensitivities for the toxicity of aldehydes, probably due to different numbers of target cells. The activities of aldehyde dehydrogenases (NAD- and NADP-dependent, ALDH), alcohol dehydrogenase and aldehyde reductase were markedly greater in the HTC cell line compared to the Hepa 1c1c7 cell line, especially with BA as substrate. The cytotoxicities of aldehydes were generally stronger in the HTC cell line than in the Hepa 1c1c7 cell line; with the CF test. Particularly, BA was highly toxic to the HTC cells, which possessed the highest ALDH levels. Moreover, the treatment with (diethylamino)benzaldehyde, an ALDH inhibitor, completely abolished the toxicity of BA. Taken together, all these findings suggest that several cell lines expressing different aldehyde metabolizing activities could be used especially in the pre-screening phase to distinguish the metabolism-dependent cytotoxic effects from the metabolism independent effects.
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PMID:Comparative evaluation of cytotoxicity and metabolism of four aldehydes in two hepatoma cell lines. 929 76

The genotoxicity of nitroaromatic compounds was examined in two cultured cell lines, namely, a human hepatoma cell line, HepG2, and a brown bullhead fibroblast cell line, BB. Furthermore, the role of the quinone-reducing enzyme DT diaphorase [NAD(P)H:(quinone acceptor) oxidoreductase] was examined with respect to its influence on the genotoxic effects of model nitroaromatic pollutants. The nitroreductive characteristics of these two cell lines were examined using an acetylated cytochrome c reduction assay for enzymatic nitroreductase activity. Subsequently, the influence of DT diaphorase on the genotoxicity of two model nitroaromatics, 4-nitroquinoline 1-oxide (4NQ) and nitrofurantoin (NF), revealed that DT diaphorase was the predominant 4NQ reductase in cytosols of both cell lines, but played a lesser role in NF reduction in both species. Despite these interspecific similarities, results revealed marked qualitative differences between the two species in terms of the influence of DT diaphorase on quinone-mediated genotoxicity. When pretreated with the DT diaphorase inhibitor dicoumarol, HepG2 cells exhibited an exacerbation of genotoxicity in the presence of 4NQ, indicating a protective influence of the enzyme. In contrast, 4NQ genotoxicity in BB cells was reduced in the presence of dicoumarol, indicating a deleterious effect of DT diaphorase activity. Conversely, dicoumarol pretreatment was moderately protective against NF-mediated genotoxicity in HepG2 cells but exacerbated NF toxicity in BB cells. This study illustrates the manner in which functionally analogous enzymes may have markedly distinct influences on xenobiotic toxicity in different cellular systems.
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PMID:Roles of DT diaphorase in the genotoxicity of nitroaromatic compounds in human and fish cell lines. 931 Jan 46

Isothiocyanates occur in many edible plants and are consumed in substantial quantities by humans. A number of isothiocyanates block chemical carcinogenesis in a variety of animal models by inhibiting Phase 1 enzymes involved in carcinogen activation and by inducing Phase 2 enzymes that accelerate the inactivation of carcinogens. There are large but unexplained potency differences among individual isothiocyanates. When murine hepatoma (Hepa 1c1c7) and several other cell lines were exposed to low concentrations (1-5 microM) of certain isothiocyanates, the intracellular isothiocyanate/dithiocarbamate concentrations (measured by cyclocondensation with 1,2-benzenedithiol) rose rapidly (30 min at 37 degrees C) to very high levels (e.g., 800-900 microM). The intracellular accumulation of isothiocyanates/dithiocarbamates was temperature, structure, and glutathione dependent and could not be saturated under experimentally achievable conditions. When murine hepatoma cells were exposed to nine isothiocyanates (5 microM for 24 h at 37 degrees C) that differed considerably in structure and Phase 2 enzyme inducer potencies, the intracellular concentrations (area under curve) correlated closely and linearly with their potencies as inducers of the Phase 2 enzymes: NAD(P)H:quinone reductase and glutathione S-transferases. Isothiocyanates that did not accumulate to high levels were not inducers. These observations suggest strongly that induction of Phase 2 enzymes depends on intracellular levels of isothiocyanates/dithiocarbamates. Depletion of glutathione by treatment of Hepa cells with buthionine sulfoximine increased the inducer potencies of several isothiocyanates but could not be directly related to changes in intracellular isothiocyanate/dithiocarbamate concentrations, suggesting that glutathione may play several roles in the induction process.
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PMID:Mechanism of differential potencies of isothiocyanates as inducers of anticarcinogenic Phase 2 enzymes. 978 15

Induction of phase 2 enzymes (e.g., glutathione transferases, NAD(P)H:quinone reductase, glucuronosyltransferases, epoxide hydrolase) is a major strategy for reducing the susceptibility of animal cells to neoplasia and other forms of electrophile toxicity. In a search for new chemoprotective enzyme inducers, a structure-activity analysis was carried out on two types of naturally occurring and synthetic substituted phenylpropenoids: (a) Ar-CH=CH-CO-R, where R is OH, OCH3, CH3, or Ar, including cinnamic, coumaric, ferulic, and sinapic acid derivatives, their ketone analogues, and chalcones; and (b) bis(benzylidene)cycloalkanones, Ar-CH=C(CH2)n(CO)C=CH-Ar, where n = 5, 6, or 7. The potencies of these compounds in inducing NAD(P)H:quinone reductase activity in murine hepatoma cells paralleled their Michael reaction acceptor activity (Talalay, P.; De Long, M. J.; Prochaska, H. J. Proc. Natl. Acad. Sci. U.S.A. 85, 1988, 8261-8265). Unexpectedly, the bis(benzylidene)cycloalkanones also powerfully quenched the lucigenin-derived chemiluminescence evoked by superoxide radicals. Introduction of o-hydroxyl groups on the aromatic rings of these phenylpropenoids dramatically enhanced their potencies not only as inducers for quinone reductase but also as quenchers of superoxide. These potentiating o-hydroxyl groups are hydrogen-bonded, as shown by moderate downfield shift of their proton NMR resonances and their sensitivities to the solvent environment. The finding that the potencies of a series of bis(benzylidene)cycloalkanones in inducing quinone reductase appear to be correlated with their ability to quench superoxide radicals suggests that the regulation of phase 2 enzymes may involve both Michael reaction reactivity and radical quenching mechanisms.
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PMID:Chemoprotective properties of phenylpropenoids, bis(benzylidene)cycloalkanones, and related Michael reaction acceptors: correlation of potencies as phase 2 enzyme inducers and radical scavengers. 985 96

This study applies biophysical methods like light absorption spectrophotometry of cytochromes, determination of NAD(P)H-dependent superoxide anion (O2-) formation and localisation of hydroxyl radicals (*OH) by 3-dimensional (3D) confocal laser scanning microscopy to reveal in human cells putative members of the oxygen sensing signal pathway leading to enhanced gene expression under hypoxia. A cell membrane localised non-mitochondrial cytochrome b558 seems to be involved as an oxygen sensor in the hepatoma cell line HepG2 in cooperation with the mitochondrial cytochrome b563 probably probing additionally metabolic changes. *OH the putative second messenger of the oxygen sensing pathway generated by a Fenton reaction could be visualized in the perinuclear space of the three human cell lines used. Substances like cobalt or the iron chelator desferrioxamine, which have been applied in HepG2 cells to mimic hypoxia induced gene expression, interact on various sides of the oxygen sensing pathway confirming the importance of b-type cytochromes and the Fenton reaction.
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PMID:Cytochromes and oxygen radicals as putative members of the oxygen sensing pathway. 985 48

Cytochrome b-type NAD(P)H oxidoreductases are involved in many physiological processes, including iron uptake in yeast, the respiratory burst, and perhaps oxygen sensing in mammals. We have identified a cytosolic cytochrome b-type NAD(P)H oxidoreductase in mammals, a flavohemoprotein (b5+b5R) containing cytochrome b5 (b5) and b5 reductase (b5R) domains. A genetic approach, using BLAST searches against DBEST for FAD-, NAD(P)H-binding sequences followed by reverse transcription-PCR, was used to clone the complete cDNA sequence of human b5+b5R from the hepatoma cell line Hep 3B. Compared with the classical single-domain b5 and b5R proteins localized on endoplasmic reticulum membrane, b5+b5R also has binding motifs for heme, FAD, and NAD(P)H prosthetic groups but no membrane anchor. The human b5+b5R transcript was expressed at similar levels in all tissues and cell lines that were tested. The two functional domains b5* and b5R* are linked by an approximately 100-aa-long hinge bearing no sequence homology to any known proteins. When human b5+b5R was expressed as c-myc adduct in COS-7 cells, confocal microscopy revealed a cytosolic localization at the perinuclear space. The recombinant b5+b5R protein can be reduced by NAD(P)H, generating spectrum typical of reduced cytochrome b with alpha, beta, and Soret peaks at 557, 527, and 425 nm, respectively. Human b5+b5R flavohemoprotein is a NAD(P)H oxidoreductase, demonstrated by superoxide production in the presence of air and excess NAD(P)H and by cytochrome c reduction in vitro. The properties of this protein make it a plausible candidate oxygen sensor.
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PMID:Identification of a cytochrome b-type NAD(P)H oxidoreductase ubiquitously expressed in human cells. 1061 Dec 83

Isothiocyanates (ITCs) are abundant in the human diet. Many potently inhibit tumorigenesis induced by a wide variety of chemical carcinogens in rodents. Recently, we observed that several ITCs accumulated to very high concentrations in cultured cells and that their accumulated levels were closely related to their potencies in inducing phase II enzymes [NAD(P)H:quinone reductase and glutathione transferases] that detoxify carcinogens. To elucidate the molecular mechanism responsible for this accumulation, the intracellular chemical identities of two ITCs, sulforaphane [SF, 1-isothiocyanato-(4R,S)-(methylsulfinyl)butane] and benzyl-ITC, were investigated in murine hepatoma cells. Both ITCs accumulated very rapidly to high intracellular concentrations, but, remarkably, most of the intracellular forms of the ITCs were dithiocarbamates resulting from conjugation with reduced glutathione (GSH). For example, the intracellular concentration reached 6.4 mM when cells were exposed to 100 microM SF for 30 min at 37 degrees C and 95% of the accumulated product was the GSH conjugate. Cellular accumulation of each ITC was accompanied by a profound reduction in cellular GSH levels. These findings, together with our previous observation that accumulation of ITCs depended on cellular GSH levels, strongly suggest that intracellular conjugation of ITCs with GSH is mainly responsible for ITC accumulation. Surprisingly, rapid accumulation to high concentrations also occurred when cells were exposed to the GSH-ITC conjugates. However, these conjugates were apparently not absorbed intact, but were hydrolyzed extracellularly to free ITCs that were taken up by the cells. This conclusion is supported by the finding that suppression of dissociation of the conjugates by excess GSH or other thiols blocks accumulation of the conjugates.
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PMID:Role of glutathione in the accumulation of anticarcinogenic isothiocyanates and their glutathione conjugates by murine hepatoma cells. 1083 7

An extensive body of evidence supports the conclusion that by catalyzing obligatory two-electron reductions of quinones to hydroquinones, NAD(P)H:quinone reductase (QR1) protects cells against the deleterious effects of redox cycling of quinones, their ability to deplete glutathione, and to produce neoplasia. The effects of elevation of QR1 levels by various enzyme inducers, inhibition of the enzyme by dicumarol, and genetic deletion of the enzyme (knockout mouse) are all consistent with the proposed protective functions. Measurement of QR1 activity in murine hepatoma cells grown in 96-well microtiter plates has provided a rapid and quantitative method for detecting inducer activity and determining inducer potency. This constitutes a strategy for the identification of potential chemoprotectors against cancer. Epidemiological studies show that humans who are genetically deficient in QR1 are more susceptible to the hematological toxicity and carcinogenicity of benzene exposure, and may be more susceptible to the development of a number of malignant tumors.
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PMID:Persuasive evidence that quinone reductase type 1 (DT diaphorase) protects cells against the toxicity of electrophiles and reactive forms of oxygen. 1103 51


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