Growth kinetics in MCF-7 cells modulate benzo[a]pyrene-induced CYP1A1 up-regulation.
Pro-carcinogens, such as benzo[a]pyrene (B[a]P), that are exogenous ligands of the aromatic hydrocarbon receptor may influence the susceptibility of target-cell populations through the up-regulation of cytochrome P450 (CYP) mixed function oxidases. We examined whether the growth kinetics of MCF-7 cells might determine the level of up-regulation of CYP1A1, CYP1A2 or CYP1B1 by B[a]P, and whether this could then influence subsequent levels of DNA damage. Cell cultures manipulated to be G(0)/G(1)-phase concentrated, S-phase concentrated or G(2)/M-phase concentrated were treated with B[a]P and the expression levels of CYP1A1, CYP1A2, CYP1B1, cyclin-dependent kinase inhibitor 1A [CDKN1A (P21(WAF1/CIP1))], B-cell leukaemia/lymphoma-2 (BCL-2), and Bcl-2-associated X levels were determined. Levels of DNA damage were measured as DNA single-strand breaks (SSBs) by the alkaline single-cell gel electrophoresis (comet) assay or as DNA adducts by (32)P-postlabelling analysis. B[a]P-induced up-regulation of CYP1A1 was >100-fold in S-phase-concentrated cells, but in G(0)/G(1)-phase- or G(2)/M-phase-concentrated cultures up-regulation occurred to a significantly lower extent. Consistent with this, B[a]P-treated S-phase-concentrated cultures exhibited markedly up-regulated P21(WAF1/CIP1), higher levels of dose-related increases in DNA SSBs, and increased DNA adduct levels presumably as a result of CYP1A1-mediated activation of B[a]P to B[a]P-diol-epoxide compared with the cultures enriched for the other cell cycle phases. Growth kinetics in vitro may be an important predeterminant of susceptibility to an exogenous pro-carcinogen in short-term test systems and these findings have important implications when extrapolating such results to a particular target-cell population in vivo.
Jiao
Haiyan
H
Biomedical Sciences Unit, Department of Biological Sciences, Lancaster University, Lancaster LA1 4YQ, UK.
Allinson
Sarah L
SL
Walsh
Michael J
MJ
Hewitt
Rebecca
R
Cole
Kathy J
KJ
Phillips
David H
DH
Martin
Francis L
FL
D001564:Benzo(a)pyrene / Q000494:pharmacology*; D002453:Cell Cycle / Q000187:drug effects; D048708:Cell Growth Processes / Q000187:drug effects; D045744:Cell Line, Tumor; D020552:Comet Assay; D019363:Cytochrome P-450 CYP1A1 / Q000096:biosynthesis* / Q000235:genetics*; D018736:DNA Adducts / Q000187:drug effects; D004249:DNA Damage; D004790:Enzyme Induction / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007700:Kinetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects*
D018736:DNA Adducts; D012333:RNA, Messenger; D001564:Benzo(a)pyrene; D019363:Cytochrome P-450 CYP1A1