Mutagenic response of mouse lymphoma cells after activation of benzidine and 2-aminofluorene with purified prostaglandin H synthase.
The mutagenic response of L5178Y mouse lymphoma cells to the model aromatic amine carcinogens, benzidine (BNZ) and 2-aminofluorene (2-AF) in the presence of the mammalian peroxidase prostaglandin H synthase (PHS) was examined. Standard incubation conditions for mouse lymphoma cells and the PHS system were developed. The cells were exposed to BNZ and 2-AF with purified PHS in the presence or absence of a peroxide, 5-phenyl-4-pentenyl hydroperoxide (PPHP) which is required for PHS-dependent amine oxidation. Incubations were carried out in a medium consisting of Hanks' balanced salt solution with calcium and magnesium and 0.1% pluronic F-68. BNZ by itself or in the presence of PPHP induced a weak mutagenic response in mouse lymphoma cells, but the addition of PHS or PHS and its co-factor PPHP increased the mutagenic response approximately 5-fold over that observed in the absence of PHS. A maximal mutagenic response for BNZ was observed after incubation with the complete activating system, PHS and PPHP. These data are in agreement with the fact that BNZ is an excellent substrate for PHS. When 2-AF was incubated with mouse lymphoma cells, only a minimal mutagenic response was observed. Incubation of 2-AF with either PPHP or PPHP and PHS (complete peroxidase system produced a significant enhancement in mutagenic response. Thus, the mutagenic response of the mouse lymphoma cells to 2-AF was dependent on the peroxide, PPHP but not the enzyme PHS. These data suggest that 2-AF, which is a poor PHS substrate, is oxidized by a different catalyst than PHS. This work demonstrates that BNZ and 2-AF are converted by peroxide-dependent mechanisms to mutagens that can be detected in mammalian cells.
Sarkar
F H
FH
Oxford Biomedical Research, Inc., Rochester, MI 48309.
Radcliff
G
G
Callewaert
D M
DM
Marnett
L J
LJ
Eling
T
T
Daston
D S
DS
Caspary
W J
WJ
D000475:Alkenes; D000818:Animals; D001560:Benzidines / Q000378:metabolism / Q000633:toxicity*; D001711:Biotransformation; D005449:Fluorenes / Q000378:metabolism / Q000633:toxicity*; D007940:Leukemia L5178; D051379:Mice; D009152:Mutagenicity Tests / Q000379:methods*; D009153:Mutagens; D010545:Peroxides; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism*; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured / Q000187:drug effects
D000475:Alkenes; D001560:Benzidines; D005449:Fluorenes; D009153:Mutagens; D010545:Peroxides; C029876:benzidine; C012177:2-aminofluorene; C047254:5-phenylpent-4-enyl-1-hydroperoxide; D011451:Prostaglandin-Endoperoxide Synthases