Éditeur(s) : HAL CCSD American Chemical Society
Résumé : International audience
Aromatic amines and structurally related heterocyclic aromatic amines (HAAs) are produced during the combustion of tobacco or during the high-temperature cooking of meat. Exposure to some of these chemicals may contribute to the etiology of several common types of human cancers. 2-Amino-9H-pyrido[2,3-b]indole (AαC) is the most abundant HAA formed in mainstream tobacco smoke: it arises in amounts that are 25-100 times greater than the levels of the arylamine, 4-aminobiphenyl (4-ABP), a human carcinogen. 2-Amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) is a prevalent HAA formed in cooked meats. AαC and MeIQx are rodent carcinogens; however, their carcinogenic potency in humans is unknown. A preliminary assessment of the carcinogenic potential of these HAAs in humans was conducted by examining the capacity of primary human hepatocytes to form DNA adducts of AαC and MeIQx, in comparison to 4-ABP, followed by the kinetics of DNA adduct removal by cellular enzyme repair systems. The principal DNA adducts formed were N-(deoxyguanosin-8-yl) (dG-C8) adducts. Comparable levels of DNA adducts were formed with AαC and 4-ABP, whereas adduct formation was ∼5-fold lower for MeIQx. dG-C8-AαC and dG-C8-4-ABP were formed at comparable levels in a concentration-dependent manner in human hepatocytes treated with procarcinogens over a 10,000-fold concentration range (1 nM-10 μM). Pretreatment of hepatocytes with furafylline, a selective inhibitor of cytochrome P450 1A2, resulted in a strong diminution of DNA adducts signifying that P450 1A2 is a major P450 isoform involved in bioactivation of these procarcinogens. The kinetics of adduct removal varied for each hepatocyte donor. Approximately half of the DNA adducts were removed within 24 h of treatment; however, the remaining lesions persisted over 5 days. The high levels of AαC present in tobacco smoke and its propensity to form persistent DNA adducts in human hepatocytes suggest that AαC can contribute to DNA damage and the risk of hepatocellular cancer in smokers.
ISSN: 0893-228X

inserm-00869907
http://www.hal.inserm.fr/inserm-00869907
DOI : 10.1021/tx4002226
PUBMED : 23898916

Éditeur(s) : HAL CCSD American Society for Biochemistry and Molecular Biology
Résumé : International audience
2-Amino-9H-pyrido[2,3-b]indole (AαC) is a carcinogenic heterocyclic aromatic amine (HAA) that arises in tobacco smoke. UDP-glucuronosyltransferases (UGTs) are important enzymes that detoxicate many procarcinogens, including HAAs. UGTs compete with P450 enzymes, which bioactivate HAAs by N-hydroxylation of the exocyclic amine group; the resultant N-hydroxy-HAA metabolites form covalent adducts with DNA. We have characterized the UGT-catalyzed metabolic products of AαC and the genotoxic metabolite 2-hydroxyamino-9H-pyrido[2,3- b]indole (HONH-AαC) formed with human liver microsomes, recombinant human UGT isoforms, and human hepatocytes. The structures of the metabolites were elucidated by 1H NMR and mass spectrometry. AαC and HONH-AαC underwent glucuronidation by UGTs to form, respectively, N 2-(β- D-glucosidurony1)-2-amino-9H-pyrido[2,3-b]indole (AαC-N 2-Gl) and N 2-(β-D-glucosidurony1)-2-hydroxyamino-9H-pyrido[2,3-b] indole (AαC-HON 2-Gl). HONH-AαC also underwent glucuronidation to form a novel O-linked glucuronide conjugate, O-(β-D-glucosidurony1)-2-hydroxyamino-9H-pyrido[2,3-b]indole (AαC-HN 2-O-Gl). AαC-HN 2-O-Gl is a biologically reactive metabolite and binds to calf thymus DNA (pH 5.0 or 7.0) to form the N-(deoxyguanosin-8-yl)-AαC adduct at 20-50-fold higher levels than the adduct levels formed with HONH-AαC. Major UGT isoforms were examined for their capacity to metabolize AαC and HONH-AαC. UGT1A4 was the most catalytically efficient enzyme (V max/K m) at forming AαC-N 2-Gl (0.67 μl*min -1*mg of protein -1), and UGT1A9 was most catalytically efficient at forming AαC-HN-O-Gl (77.1 μl*min -1*mg of protein -1), whereas UGT1A1 was most efficient at forming AαC-HON 2-Gl (5.0 μl*min -1*mg of protein -1). Human hepatocytes produced AαC-N 2-Gl and AαC-HN 2-O-Gl in abundant quantities, but AαC-HON 2-Gl was a minor product. Thus, UGTs, usually important enzymes in the detoxication of many procarcinogens, serve as a mechanism of bioactivation of HONH-AαC. © 2012 by The American Society for Biochemistry and Molecular Biology, Inc.
ISSN: 0021-9258

hal-00696860
https://hal-anses.archives-ouvertes.fr/hal-00696860
DOI : 10.1074/jbc.M111.320093
PUBMED : 22393056

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
: Previously, we demonstrated that eicosapentaenoic acid enhanced ethanol-induced oxidative stress and cell death in primary rat hepatocytes via an increase in membrane fluidity and lipid raft clustering. In this context, another n-3 polyunsaturated fatty acid, docosahexaenoic acid (DHA), was tested with a special emphasis on physical and chemical alteration of lipid rafts. Pretreatment of hepatocytes with DHA reduced significantly ethanol-induced oxidative stress and cell death. DHA protection could be related to an alteration of lipid rafts. Indeed, rafts exhibited a marked increase in membrane fluidity and packing defects leading to the exclusion of a raft protein marker, flotillin. Furthermore, DHA strongly inhibited disulfide bridge formation, even in control cells, thus suggesting a disruption of protein-protein interactions inside lipid rafts. This particular spatial organization of lipid rafts due to DHA subsequently prevented the ethanol-induced lipid raft clustering. Such a prevention was then responsible for the inhibition of phospholipase C-γ translocation into rafts, and consequently of both lysosome accumulation and elevation in cellular low-molecular-weight iron content, a prooxidant factor. In total, the present study suggests that DHA supplementation could represent a new preventive approach for patients with alcoholic liver disease based upon modulation of the membrane structures.
ISSN: 0278-6915

hal-00860153
https://hal-univ-rennes1.archives-ouvertes.fr/hal-00860153
https://hal-univ-rennes1.archives-ouvertes.fr/hal-00860153/document
https://hal-univ-rennes1.archives-ouvertes.fr/hal-00860153/file/Sergent-et-al-incl-figures-accepted.pdf
DOI : 10.1016/j.fct.2013.07.061