Formation and repair of benzo[a]pyrene-DNA adducts in cultured hamster tracheal epithelium determined by 32P-postlabeling analysis and unscheduled DNA synthesis.
Hamster tracheal organ cultures were used to investigate the relationship between DNA adduct formation measured directly by the 32P-postlabeling assay, and the DNA damage measured indirectly by the unscheduled DNA synthesis (UDS) assay. Hamster tracheas were treated with three concentrations of benzo[a]pyrene (B[a]P) for 2 days. Postlabeling and UDS assays were also carried out a few days after removal of the B[a]P. Furthermore, the types of B[a]P-DNA adducts formed in the in vitro organ culture were qualitatively compared with those formed in vivo after intratracheal intubation of B[a]P attached to Fe2O3 particles. In vivo only one adduct was detected by 32P-postlabeling. This adduct cochromatographed with the trans-addition produce of dG and (+)-anti-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE). In vitro, a clear B[a]P-DNA adduct pattern was also found with the 32P-postlabeling assay. Four different adducts were found. The main adduct spot migrated to the same position on the thin-layer chromatogram as the in vivo adduct. B[a]P-DNA adduct formation was both time- and dose-dependent. During the first day after removal of B[a]P the adduct levels still increased, thereafter they decreased at all B[a]P concentrations. A time- and dose-dependent increase in UDS was observed in the tracheal epithelial cells treated with B[a]P in vitro. After removal of the B[a]P, UDS decreased immediately, in contrast to the formation of DNA adducts. The results of the present study show that B[a]P induces time- and dose-dependently both DNA adducts and UDS in hamster tracheal organ culture. Moreover, the main DNA adduct formed in vitro, dG-(+)-anti-BPDE, was the same as that found in vivo.
Wolterbeek
A P
AP
TNO Toxicology and Nutrition Institute, Department of Biological Toxicology, Zeist, The Netherlands.
Roggeband
R
R
Steenwinkel
M J
MJ
Baan
R A
RA
Rutten
A A
AA
D000818:Animals; D001564:Benzo(a)pyrene / Q000378:metabolism*; D002478:Cells, Cultured; D006224:Cricetinae; D004247:DNA / Q000096:biosynthesis* / Q000378:metabolism*; D018736:DNA Adducts; D004260:DNA Repair; D004848:Epithelium / Q000378:metabolism; D008647:Mesocricetus; D010761:Phosphorus Radioisotopes; D014132:Trachea / Q000378:metabolism*
D018736:DNA Adducts; D010761:Phosphorus Radioisotopes; C056612:benzo(a)pyrene-DNA adduct; D001564:Benzo(a)pyrene; D004247:DNA