title: Benzo[a]pyrene-induced cell cycle arrest in HepG2 cells is associated with delayed induction of mitotic instability.
abstract: The environmental carcinogen benzo[a]pyrene (B[a]P) after being metabolised by cytochrome P450 enzymes forms DNA adducts. This abnormal situation induces changes in the cell cycle, DNA damage, chromosomal and mitotic aberrations, all of which may be related to carcinogenesis. In order to further investigate the mechanistic basis of these effects, HepG2 cells were treated with 3μM B[a]P for various time periods, followed by further incubation in the absence of B[a]P for up to 192h. B[a]P treatment led initially to S-phase arrest followed by recovery and subsequent induction of G2/M arrest, indicating activation of the corresponding DNA damage checkpoints. Immunofluorescence-based studies revealed accumulation of B[a]P-induced DNA adducts and chromosomal damage which persisted beyond mitosis and entry into a new cycle, thus giving rise to a new round of activation of the S-phase checkpoint. Prolonged further cultivation of the cells in the absence of B[a]P resulted in high frequencies of various abnormal mitotic events. Abrogation of the B[a]P-induced S-phase arrest by the Chk1 inhibitor UCN-01 triggered a strong apoptotic response but also dramatically decreased the frequency of mitotic abnormalities in the surviving cells, suggesting that events occurring during S-phase arrest contribute to the formation of delayed mitotic damage. Overall, our data demonstrate that, although S-phase arrest serves as a mechanism by which the cells reduce their load of genetic damage, its prolonged activation may also have a negative impact on the balance between cell death and heritable genetic damage.
authors.affiliation: Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research Foundation, Athens, Greece. Electronic address: dstellas@bioacademy.gr.
authors.forename: Dimitris
authors.initials: D
authors.lastname: Stellas
authors.affiliation: Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research Foundation, Athens, Greece.
authors.forename: Vassilis L
authors.initials: VL
authors.lastname: Souliotis
authors.affiliation: Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research Foundation, Athens, Greece.
authors.forename: Margarita
authors.initials: M
authors.lastname: Bekyrou
authors.affiliation: Hellenic Pasteur Institute, Athens, Greece.
authors.forename: Despina
authors.initials: D
authors.lastname: Smirlis
authors.affiliation: Laboratory for Cell Genetics,Vrije Universiteit Brussel, Brussels, Belgium.
authors.forename: Micheline
authors.initials: M
authors.lastname: Kirsch-Volders
authors.affiliation: Institute of Molecular Biology and Pathology C.N.R., Rome, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Degrassi
authors.affiliation: Laboratory for Cell Genetics,Vrije Universiteit Brussel, Brussels, Belgium; Institute of Molecular Biology and Pathology C.N.R., Rome, Italy.
authors.forename: Enrico
authors.initials: E
authors.lastname: Cundari
authors.affiliation: Institute of Biology, Medicinal Chemistry and Biotechnology, National Hellenic Research Foundation, Athens, Greece.
authors.forename: Soterios A
authors.initials: SA
authors.lastname: Kyrtopoulos
mesh_terms: D001564:Benzo(a)pyrene / Q000494:pharmacology*; D002274:Carcinogens, Environmental / Q000494:pharmacology*; D002453:Cell Cycle / Q000187:drug effects / Q000235:genetics; D059447:Cell Cycle Checkpoints / Q000187:drug effects*; D018736:DNA Adducts / Q000187:drug effects / Q000235:genetics; D004249:DNA Damage; D042822:Genomic Instability / Q000187:drug effects*; D056945:Hep G2 Cells; D006801:Humans; D008938:Mitosis / Q000187:drug effects / Q000235:genetics; D013997:Time Factors; D014407:Tumor Cells, Cultured
chemical_list: D002274:Carcinogens, Environmental; D018736:DNA Adducts; D001564:Benzo(a)pyrene
keywords: Benzo[a]pyrene; Carcinogenesis; Chk1; Mitotic abnormalities; S-phase arrest