Sulforaphane and quercetin modulate PhIP-DNA adduct formation in human HepG2 cells and hepatocytes

Bacon, James R., Williamson, Gary, Garner, R. Colin , Lappin, Graham, Langouet, Sophie and Bao, Yongping (2003) Sulforaphane and quercetin modulate PhIP-DNA adduct formation in human HepG2 cells and hepatocytes. Carcinogenesis, 24 (12). pp. 1903-1911. ISSN 0143-3334

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The formation of DNA adducts in human HepG2 cells and human hepatocytes exposed to 14C-labelled 2-amino-1-methyl-6-phenylimidazo4,5-bpyridine (PhIP) was examined using Accelerator Mass Spectrometry (AMS). PhIP generated DNA adducts in a linear dose-dependent manner between 100 pM and 20 μM. Co-treatment with the dietary isothiocyanate, sulforaphane (SFN, 1-10 μM), or the flavonoid, quercetin (5-20 μM), significantly reduced the level of PhIP-DNA adducts in a dose-dependent manner. The degree of protection was dependent on PhIP concentration, i.e. after 100 pM PhIP exposure, SFN or quercetin reduced adduct levels to below the limit of detection (0.15 amol PhIP/μg DNA) but at higher PhIP exposure (10 nM and 1 μM), the protection was 60 and 10%, respectively. The involvement of phase I, phase II and DNA repair enzymes in this protection against PhIP-DNA adduct formation was investigated using real-time RT-PCR and enzyme activity assays. In intact HepG2 cells, quercetin inhibited cytochrome P450 (CYP)1A2, the main phase I enzyme responsible for PhIP bioactivation. In contrast, SFN induced phase II detoxification enzymes, UDP-glucuronosyltransferase 1A1 and glutathione S-transferase A1 mRNA expression. SFN and quercetin showed no effect on DNA repair, neither in terms of the level of PhIP-DNA adducts, when cells were treated with phytochemicals after the carcinogen exposure, nor the regulation of mRNA expression of two DNA repair enzymes, apurinic endonuclease and DNA polymerase β. This study indicates that dietary isothiocyanates and flavonoids modulate phase I and phase II enzyme expression, hence increasing the rate of detoxification of the dietary carcinogen PhIP in human HepG2 cells but do not affect the rate of PhIP-DNA adduct repair. The formation of PhIP-DNA adducts in human hepatocytes was also dose-dependent with PhIP-concentration and the levels of protection by SFN or quercetin were up to 60% after 10 nM PhIP treatment, but showed large inter-individual variation with no observed protection in some individuals.

Additional Information:First published online August 29th 2003
Keywords:2 amino 1 methyl 6 phenylimidazo4,5 bpyridine, carbon 14, carcinogen, cytochrome P450 1A2, DNA (apurinic or apyrimidinic site) lyase, DNA directed DNA polymerase beta, flavonoid, glucuronosyltransferase, glutathione transferase, isothiocyanic acid, messenger RNA, polydeoxyribonucleotide synthase, quercetin, sulforaphane, article, cell protection, cell strain HepG2, concentration response, controlled study, detoxification, dietary intake, DNA adduct, DNA repair, enzyme activity, enzyme assay, enzyme induction, enzyme inhibition, exposure, human, human cell, isotope labeling, linear system, liver cell, mass spectrometer, nucleotide sequence, phytochemistry, priority journal, protein expression, regulatory mechanism, reverse transcription polymerase chain reaction, statistical significance, variance, Anticarcinogenic Agents, Biological Markers, Carcinogens, Cell Line, Cell Line, Tumor, Cytochrome P-450 CYP1A2, DNA Adducts, DNA Polymerase beta, Dose-Response Relationship, Drug, Hepatocytes, Humans, Imidazoles, Isothiocyanates, Mass Spectrometry, Reverse Transcriptase Polymerase Chain Reaction, RNA, RNA, Messenger, Thiocyanates, Time Factors
Subjects:C Biological Sciences > C130 Cell Biology
Divisions:College of Science > School of Pharmacy
ID Code:8241
Deposited On:23 Mar 2013 19:32

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