A test using cultured cells with induced mixed-function oxygenase in the unscheduled DNA synthesis assay for detecting promutagens/procarcinogens

Abstract

The human FL cell line contains very low levels of constitutive AHH activity, but it could be greatly induced by NE, β-NF and 3-MC, and induced slightly by PB. When two different types of inducer, for example, 3-MC and PB or 3-MC and NE were given in combination, an additive inductive effect was not observed. Both the constitutive and induced AHH in FL cells have characteristics of MFO, namely, NADPH-dependence and CO-sensitivity. The fact that the constitutive and induced AHH in FL cells could be inhibited by a known hydroxylase inhibitor 7,8-BF indicated that the AHH in FL cells belongs to the cytochrome P-448 dependent MFO type. After removal of inducer from the medium, the induced AHH activity remained at a high level for at least 24–36 h. By using AHH-induced FL cells in the UDS assay system for the detection of promutagens/procarcinogens, we found that AFB 1 and 3-MC did not induce a UDS reaction in uninduced FL cells, while in β-NF induced cells,10 −6−10 −4 M AFB 1 and 10 −7−10 −6 M 3-MC elicited a very significant UDS reaction, which was concordant with the results obtained in the UDS assay system using HeLa cells or FL cells supplemented with liver microsomes or using primary cultured hepatocytes as indicator cells. B(a)P elicited the UDS reaction at concentrations of 10 −6−10 −3 M in β-NF induced cells, whereas 10 −4−10 −3 M was required in uninduced cells. The results above indicate that this new design is feasible, but further study is needed to assure its accuracy.