Inhibition of poly(ADP-ribose) polymerase increases (+/-)-anti-benzo [a]pyrene diolepoxide-induced micronuclei formation and p53 accumulation in isolated human peripheral blood lymphocytes.
In response to DNA damage, in particular DNA strand breaks, the proposed roles for normal tumour suppressor protein p53 are to increase the period of time available for DNA repair prior to replication, or to direct damaged cells into programmed cell-death. Since treatment of mammalian cells with (+/-)-anti-benzo[a]pyrene diolepoxide [(+/-)-anti-BPDE] --a mixture of metabolites comprising the most reactive (+)-anti-enantiomer of the full environmental carcinogen benzo[a]pyrene--has been shown to result in induction of DNA repair processes and consequently in DNA strand break formation, the aim of the present study was to investigate whether p53 accumulation is induced in (+/-)-anti-BPDE-treated phytohaemagglutinin-stimulated human peripheral blood lymphocytes (PBLs). Both immunocytochemical and immunoblot analysis indicated that treatment of PBLs with (+/-)-anti-BPDE results in p53 accumulation. Optimal accumulation was observed at 2.5 microM, while no increase of p53 levels was observed at concentrations < 2.5 microM and > 10 microM. Further, (+/-)-anti-BPDE-induced p53 accumulation in PBLs was found to be time-dependent with accumulation up to 24 h after the onset of treatment. Treatment of PBLs with 2.5 microM of (+/-)-anti-BPDE and 1 mM of 3-aminobenzamide, an inhibitor of the DNA strand break-dependent enzyme poly(ADP-ribose) polymerase, resulted in increased p53 levels, in comparison to cells treated with (+/-)-anti-BPDE alone. This combination also potentiated the frequency of (+/-)-anti-BPDE-induced micronuclei. These findings suggest that (+/-)-anti-BPDE-induced DNA strand break formation is responsible for the observed p53 accumulation. It is unlikely that poly(ADP-ribose) polymer formation is a prerequisite in the process of p53 accumulation, as triggered by DNA strand-break inducing agents like (+/-)-anti-BPDE. It is hypothesized that p53-dependent pathways may be activated in phytohaemagglutinin-stimulated human peripheral blood lymphocytes exposed ex vivo to (+/-)-anti-BPDE.
Stierum
R H
RH
Department of Health Risk Analysis and Toxicology, University of Limburg, Maastricht, The Netherlands.
van Herwijnen
M H
MH
Pasman
P C
PC
Hageman
G J
GJ
Kleinjans
J C
JC
van Agen
B
B
D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide / Q000633:toxicity*; D000328:Adult; D000818:Animals; D002273:Carcinogens / Q000633:toxicity*; D002478:Cells, Cultured; D004249:DNA Damage; D006801:Humans; D008214:Lymphocytes / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D008297:Male; D048629:Micronuclei, Chromosome-Defective / Q000187:drug effects*; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011065:Poly(ADP-ribose) Polymerases / Q000502:physiology*; D011817:Rabbits; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
D002273:Carcinogens; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D016159:Tumor Suppressor Protein p53; D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; D011065:Poly(ADP-ribose) Polymerases