Hepatocellular carcinoma (HCC) remains a highly prevalent and deadly disease, being among the top causes of cancer-related deaths worldwide. Despite the fact that the liver is the major site of biotransformation, studies on drug metabolizing enzymes in HCC are scarce. It is known that malignant transformation of hepatocytes leads to a significant alteration of their metabolic functions and overall deregulation of gene expression. Advanced stages of the disease are thus frequently associated with liver failure, and severe alteration of drug metabolism. However, the impact of dysregulation of metabolic enzymes on therapeutic efficacy and toxicity in HCC patients is largely unknown. Here we demonstrate a significant down-regulation in European Caucasian patients of cytochromes P450 (CYPs), the major xenobiotic-metabolizing enzymes, in HCC tumour samples as compared to their surrounding non-cancerous (reference) tissue. Moreover, we report for the first time the association of the unique CYP profiles with specific transcriptome changes, and interesting correlations with expression levels of nuclear receptors and with the histological grade of the tumours. Integrated analysis has suggested certain co-expression profiles of CYPs with lncRNAs that need to be further characterized. Patients with large tumours with down-regulated CYPs could be more vulnerable to drug toxicity; on the other hand, such tumours would eliminate drugs more slowly and should be more sensitive to pharmacotherapy (except in the case of pro-drugs where activation is necessary).

• Klíčová slova
• CYP, Cytochrome P450, Drug metabolism, Gene expression, Hepatocellular carcinoma, Non-coding RNA,
• MeSH
• dospělí MeSH
• hepatocelulární karcinom enzymologie   patologie   MeSH
• hepatocyty metabolismus   MeSH
• játra metabolismus   MeSH
• kohortové studie MeSH
• lidé středního věku MeSH
• lidé MeSH
• metabolická inaktivace genetika   MeSH
• nádory jater enzymologie   patologie   MeSH
• receptory cytoplazmatické a nukleární genetika   metabolismus   MeSH
• regulace genové exprese enzymů * MeSH
• senioři MeSH
• stanovení celkové genové exprese MeSH
• stupeň nádoru MeSH
• systém (enzymů) cytochromů P-450 genetika   MeSH
• transkriptom * MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• receptory cytoplazmatické a nukleární MeSH
• systém (enzymů) cytochromů P-450 MeSH

UNLABELLED: Introduction and aim. Hepatocellular carcinoma (HCC) is the most common primary malignant liver tumor. It is primarily caused by hepatic cirrhosis or chronic viral hepatitis. Hepatic carcinogenesis is associated with increased oxidative stress. Thus, the aim of our study was to assess expression of the genes involved in the homeostasis of oxidative stress in patients with HCC. MATERIAL AND METHODS: The study was performed on 32 patients with primary HCC (verified by liver histology in 29 patients) and 27 control subjects (in 11 subjects, liver histology was available either with no or minimal changes in the liver tissue). Gene expressions of heme oxygenase 1 (HMOX1), biliverdin reductase A/B (BLVRA/B), NADPH oxidase 2 (NOX2) and p22phox were analyzed in the liver and peripheral blood leukocytes (PBL) in the subjects. RESULTS: Compared to controls, almost a 3 times higher mRNA level of BLVRA was detected in livers of HCC patients (p = 0.002); while those of BLVRB as well as HMOX1 were unchanged (p > 0.05). In accord with these results in the liver tissue, BLVRA mRNA levels in PBL were also significantly increased in HCC patients (p = 0.012). mRNA levels of NOX2 and p22phox in the liver tissue, although higher in HCC patients, did not differ significantly compared to control subjects (p > 0.05). Nevertheless, NOX2 mRNA level in PBL was significantly higher in HCC patients (p = 0.003). CONCLUSIONS: BLVRA mRNA levels in the liver as well as in PBL are significantly higher in HCC patients most likely as a feedback mechanism to control increased oxidative stress associated with HCC progression.

• MeSH
• hemoxygenasa-1 genetika   MeSH
• hepatocelulární karcinom krev   enzymologie   genetika   patologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• membránové glykoproteiny genetika   MeSH
• messenger RNA genetika   MeSH
• nádorové biomarkery krev   genetika   MeSH
• nádory jater krev   enzymologie   genetika   patologie   MeSH
• NADPH-oxidasa 2 MeSH
• NADPH-oxidasy genetika   MeSH
• oxidační stres genetika   MeSH
• oxidoreduktasy působící na CH-CH vazby krev   genetika   MeSH
• progrese nemoci MeSH
• regulace genové exprese enzymů MeSH
• regulace genové exprese u nádorů MeSH
• senioři MeSH
• signální transdukce MeSH
• studie případů a kontrol MeSH
• upregulace MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• biliverdin reductase MeSH Prohlížeč
• CYBA protein, human MeSH Prohlížeč
• CYBB protein, human MeSH Prohlížeč
• hemoxygenasa-1 MeSH
• HMOX1 protein, human MeSH Prohlížeč
• membránové glykoproteiny MeSH
• messenger RNA MeSH
• nádorové biomarkery MeSH
• NADPH-oxidasa 2 MeSH
• NADPH-oxidasy MeSH
• oxidoreduktasy působící na CH-CH vazby MeSH

It has been shown in previous studies that liver HEP-G2 cells (human hepatocellular carcinoma) lose their ability to express active alcohol dehydrogenase (ADH) and cytochrome P450 2E1 (CYP2E1). Although both are ethanol-inducible enzymes, short-term exposure to ethanol does not cause any changes in expression or activity in cultured HEP-G2 cells. Therefore, we tested the effect of long-term exposure to ethanol on the expression and activity of both ADH and CYP2E1 in these cells. The expression of ADH and CYP2E1 was assessed at the mRNA and/or protein level using real-time PCR and Western blot analysis. Specific colorimetric assays were used for the measurement of ADH and CYP2E1 enzymatic activities. Caco-2 cells (active CYP2E1 and inactive ADH) were used as control cells. Significantly increased protein expression of ADH (about 2.5-fold) as well as CYP2E1 (about 1.6-fold) was found in HEP-G2 cells after long-term (12 mo) exposure to ethanol. The activity of ADH and CYP2E1 was also significantly increased from 12 ± 3 and 6 ± 1 nmol/h/mg of total protein to 191 ± 9 and 57 ± 9 nmol/h/mg of total protein, respectively. We suggest that the loss of activity of ethanol-metabolizing enzymes in cultured HEP-G2 cells is reversible and can be induced by prolonged exposure to ethanol. We are therefore able to reactivate HEP-G2 cells metabolic functions concerning ethanol oxidation just by modification of in vitro culture conditions without necessity of transfection with its side effect - enzyme overexpression.

• MeSH
• alkoholdehydrogenasa biosyntéza   genetika   MeSH
• apoptóza účinky léků   MeSH
• buňky Hep G2 MeSH
• cytochrom P-450 CYP2E1 biosyntéza   genetika   MeSH
• enzymová indukce účinky léků   MeSH
• ethanol farmakologie   MeSH
• hepatocelulární karcinom enzymologie   patologie   MeSH
• játra účinky léků   enzymologie   MeSH
• lidé MeSH
• nádory jater enzymologie   patologie   MeSH
• oxidace-redukce účinky léků   MeSH
• regulace genové exprese u nádorů účinky léků   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• alkoholdehydrogenasa MeSH
• cytochrom P-450 CYP2E1 MeSH
• ethanol MeSH

Aryl hydrocarbon receptor (AhR) and glucocorticoid receptor (GR) play crucial role in the regulation of drug metabolizing enzymes and in many essential physiological processes. Cellular signaling by these receptors shares several functional and regulatory features. Here we investigated regulatory cross-talk between these two receptors. Human hepatoma cells (HepG2) were the model of choice. We analyzed the effects of dexamethasone (DEX) and dioxin (TCDD) on i) expression of AhR and GRalpha mRNAs; ii) levels of AhR and GR proteins; iii) transcriptional activities of AhR and GR in reporter assays; iv) 7-ethoxyresorufin-O-deethylase activity (EROD). We found that both DEX and TCDD affected AhR and GR mRNAs expression, proteins levels and transcriptional activities in HepG2 cells. These effects on cellular signaling by AhR and GR comprised up-/down-regulation of gene expression and ligand-dependent protein degradation. We conclude that interactive regulatory cross-talk between GR and AhR receptors in HepG2 cells defines possible implications in physiology and drug metabolism. Future research should be focused on the investigation of AhR-GR cross-talk in various normal human cells and tissues both in vitro and in vivo.

• MeSH
• cytochrom P-450 CYP1A1 metabolismus   MeSH
• dexamethason farmakologie   MeSH
• genetická transkripce účinky léků   MeSH
• glukokortikoidy farmakologie   MeSH
• hepatocelulární karcinom enzymologie   genetika   metabolismus   MeSH
• interakce mezi receptory a ligandy * MeSH
• lidé MeSH
• messenger RNA metabolismus   MeSH
• nádorové buněčné linie MeSH
• nádory jater enzymologie   genetika   metabolismus   MeSH
• pilotní projekty MeSH
• polychlorované dibenzodioxiny farmakologie   MeSH
• receptory aromatických uhlovodíků agonisté   genetika   metabolismus   MeSH
• receptory glukokortikoidů agonisté   genetika   metabolismus   MeSH
• regulace genové exprese enzymů účinky léků   MeSH
• signální transdukce účinky léků   MeSH
• transfekce MeSH
• transkripční faktory bHLH MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• AHR protein, human MeSH Prohlížeč
• cytochrom P-450 CYP1A1 MeSH
• dexamethason MeSH
• glucocorticoid receptor alpha MeSH Prohlížeč
• glukokortikoidy MeSH
• messenger RNA MeSH
• NR3C1 protein, human MeSH Prohlížeč
• polychlorované dibenzodioxiny MeSH
• receptory aromatických uhlovodíků MeSH
• receptory glukokortikoidů MeSH
• transkripční faktory bHLH MeSH

Cytochrome P450 (CYP) 1A1 attracts attention mainly because of its role in production of carcinogenic reactive metabolites from polycyclic aromatic hydrocarbons such as benzo[a]pyrene, but recent developments indicate its apparent role in cell cycle progression. Expression of the enzyme is subject to regulation by aryl hydrocarbon receptor (AhR). It has been shown that induction of CYP 1A1 in HepG2 cells and primary rat hepatocytes by tetrachloro-p-dibenzodioxin (TCDD) is diminished by colchicine and nocodazole. Both compounds decrease CYP1A1 mRNA, protein, and activity levels in HepG2 cells and mRNA level in primary rat hepatocytes. Neither compound significantly affected [(3)H]-TCDD binding to AhR, thus their effect on AhR transcriptional activity proceeds via indirect means. For colchicine and nocodazole are well-known microtubule interfering agents, we also assessed their effect on microtubule integrity in both cell types under investigation. Both compounds disrupt cytoskeleton integrity with differential potency depending on cell type. The observed suppression of AhR transcriptional activity by colchicine and nocodazole can be associated with G2/M cell cycle arrest in HepG2 cells, as demonstrated by Myt1 protein hyperphosphorylation and FACS analysis. However, in primary rat hepatocytes, cytoskeleton disruption is independent of cell cycle while displaying the same influence on AhR-dependent gene transcription. In our view, this is evidence in favor of modulatory role of cytoskeleton in AhR-dependent expression.

• MeSH
• buněčné dělení účinky léků   MeSH
• cytochrom P-450 CYP1A1 biosyntéza   genetika   metabolismus   MeSH
• cytoskelet fyziologie   MeSH
• G2 fáze účinky léků   MeSH
• hepatocelulární karcinom enzymologie   MeSH
• hepatocyty enzymologie   MeSH
• kolchicin farmakologie   MeSH
• krysa rodu Rattus MeSH
• kultivované buňky MeSH
• lidé MeSH
• messenger RNA biosyntéza   genetika   MeSH
• mikrotubuly účinky léků   ultrastruktura   MeSH
• nádorové buněčné linie MeSH
• nádory jater enzymologie   MeSH
• nokodazol farmakologie   MeSH
• polychlorované dibenzodioxiny farmakologie   MeSH
• průtoková cytometrie MeSH
• receptory aromatických uhlovodíků fyziologie   MeSH
• teratogeny farmakologie   MeSH
• viabilita buněk účinky léků   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• cytochrom P-450 CYP1A1 MeSH
• kolchicin MeSH
• messenger RNA MeSH
• nokodazol MeSH
• polychlorované dibenzodioxiny MeSH
• receptory aromatických uhlovodíků MeSH
• teratogeny MeSH

The flavonolignan silybin and its derivative dehydrosilybin have been proposed as candidate UV-protective agents in skin care products. This study addressed the effect of silybin and dehydrosilybin on the activity of cytochrome P450 isoform CYP1A1 in human keratinocytes (HaCaT) and human hepatoma cells (HepG2). CYP1A1 catalytic activity was assessed as O-deethylation of 7-ethoxyresorufin using fluorescence detection. Silybin and dehydrosylibin inhibited basal and dioxin-inducible CYP1A1 catalytic activity in both cell lines used. The inhibitory effect of tested compounds was more pronounced in HaCaT cells than in HepG2 cells, and dehydrosilybin was a much stronger inhibitor than silybin. Analyses on CYP1A1 human recombinant protein yielded IC(50) values of 22.9 +/- 4.7 micromol/L and 0.43 +/- 0.04 micromol/L for silybin and dehydrosilybin, respectively. Since CYP1A enzymes are some of the most prominent actors in the process of chemically induced carcinogenesis, the inhibitory activity of the flavonolignans tested against CYP1A1 favors their use as cytoprotective agents in terms of skin and hepatic metabolism. In addition, the capability of dehydrosilybin to inhibit CYP1A1 in submicromolar concentrations makes this compound a potential biological probe in CYP1A1 analyses.

• MeSH
• buněčné linie MeSH
• časové faktory MeSH
• cytochrom P-450 CYP1A1 antagonisté a inhibitory   metabolismus   MeSH
• dioxiny MeSH
• enzymová indukce MeSH
• hepatocelulární karcinom enzymologie   MeSH
• keratinocyty enzymologie   MeSH
• lidé MeSH
• molekulární struktura MeSH
• nádorové buněčné linie MeSH
• nádory jater enzymologie   MeSH
• silibinin MeSH
• silymarin farmakologie   MeSH
• viabilita buněk MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• cytochrom P-450 CYP1A1 MeSH
• dehydrosilybin MeSH Prohlížeč
• dioxiny MeSH
• silibinin MeSH
• silymarin MeSH

Quaternary benzo[c]phenanthridine alkaloids (QBA) sanguinarine and chelerythrine exhibit a wide spectrum of biological activities whence they are used in dental care products. Recent studies indicated that cytochrome P450 CYP1A attenuates sanguinarine toxicity both in vivo [Williams, M.K., Dalvi, S., Dalvi, R.R., 2000. Influence of 3-methylcholanthrene pretreatment on sanguinarine toxicity in mice. Vet. Hum. Toxicol. 42, 196-198] and in vitro [Vrba, J., Kosina, P., Ulrichová, J., Modrianský, M., 2004. Involvement of cytochrome P450 1A in sanguinarine detoxication. Toxicol. Lett. 151, 375-387]. However, CYP1A converts sanguinarine to the products that form DNA adducts [Stiborová, M., Simánek, V., Frei, E., Hobza, P., Ulrichová, J., 2002. DNA adduct formation from quaternary benzo[c]phenanthridine alkaloids sanguinarine and chelerythrine as revealed by the 32P-postlabeling technique. Chem. Biol. Interact. 140, 231-242]. In our work we examined the effects of sanguinarine and chelerythrine on CYP1A1 expression and catalytic activity in human hepatoma cells-HepG2. Sanguinarine and chelerythrine did not affect basal and dioxin-inducible expression of CYP1A1 mRNA and protein in HepG2 cells. The enzymatic activity of CYP1A1 was assessed by the fluorescent measurement of 7-ethyxoresorufin-O-deethylase (EROD) activity. We observed a slight decrease of dioxin-induced EROD activity in HepG2 cells by sanguinarine and chelerythrine. This decrease was attributed to the inhibition of CYP1A1 catalytic activity, as revealed by enzyme kinetic studies on recombinant CYP1A1 protein. The IC50 values for the inhibition of CYP1A1 by sanguinarine and chelerythrine were 2.1 and 1.9muM, respectively. In conclusion, albeit the CYP1A modulates QBA cytotoxicity and genotoxicity, the QBA themselves do not affect CYP1A1 expression. The data indicate that studied alkaloids do not have specific cellular target and their biological effects are rather pleiotropic.

• MeSH
• alkaloidy farmakologie   MeSH
• benzofenantridiny MeSH
• cytochrom P-450 CYP1A1 antagonisté a inhibitory   biosyntéza   MeSH
• cytochrom P-450 CYP1A2 biosyntéza   MeSH
• fenantridiny farmakologie   MeSH
• hepatocelulární karcinom enzymologie   MeSH
• inhibitory cytochromu P450 CYP1A2 MeSH
• inhibitory enzymů farmakologie   MeSH
• isochinoliny MeSH
• katalýza MeSH
• lidé MeSH
• messenger RNA biosyntéza   MeSH
• nádorové buněčné linie MeSH
• nádory jater enzymologie   MeSH
• rekombinantní proteiny chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• alkaloidy MeSH
• benzofenantridiny MeSH
• chelerythrine MeSH Prohlížeč
• cytochrom P-450 CYP1A1 MeSH
• cytochrom P-450 CYP1A2 MeSH
• fenantridiny MeSH
• inhibitory cytochromu P450 CYP1A2 MeSH
• inhibitory enzymů MeSH
• isochinoliny MeSH
• messenger RNA MeSH
• rekombinantní proteiny MeSH
• sanguinarine MeSH Prohlížeč