Changes in the cytochrome P450 monooxygenase system were investigated in Hep G2 cells treated for 24 hours with various concentrations of benomyl. Decreases in both benzo[a]pyrene hydroxylase (AHH) and ethoxyresorufin deethylase (EROD), markers of the P4501A1 isoenzyme, were noted. Ethoxycoumarin deethylase (ECOD), a marker of the P4502B1 isoenzyme, showed a dose-dependent increase. Characterisation by SDS-polyacryl-amide gel electrophoresis of Hep G2 cell microsomal proteins revealed a decrease in the polypeptide bands at 55.5kD (P4501A1) and 48kD (P4501A1 and P4502B1) and an increase in the polypeptide band at 52kD (P4502B1). Benomyl induced a decrease in cytochrome P4501A1 and an increase in cytochrome P4502B1 in Hep G2 cells, as indicated by variations in AHH, EROD and ECOD activity, and by characterisation of microsomal proteins by SDS-polyacryl-amide gel electrophoresis.
MarinovichM., GuizzettiM., and GalliC.L. (1994). Mixtures of benomyl, pirimiphos-methyl, dimethoate, diazinon and azinphos-methyl affect protein synthesis in HL-60 cells differently.Toxicology94, 173–185.
2.
PiattiE., MarabiniL., and ChiesaraE. (1994). Increase of micronucleus frequency in cultured rat hepatocytes treated in vitro with benomyl and pirimiphos-methyl separately and in mixture.Mutation Research324, 59–64.
3.
ZemanF.J., HoogenboomE.R., KavlockR.J., and SempleJ.L. (1986). Effect on the fetus of maternal benomyl exposure in the protein-deprived rat.Journal of Toxicology and Environmental Health127, 405–413.
4.
CarterS.D., HeinJ.F., RehnbergG.L., and LaskeyJ.W. (1984). Effect of benomyl on the reproductive development of male rats.Journal of Toxicology and Environmental Health13, 53–61.
5.
DouchP.G.C. (1973). The metabolism of benomyl in mammals.Xenobiotica3, 364–380.
6.
DalviR.R. (1992). Effect of the fungicide benomyl on xenobiotic metabolism in rats.Toxicology71, 63–68.
7.
RoeA.L., SnawderJ.E., BensonR.W., RobertsD.W., and CascianoD.A. (1993). Hep G2 cells: an in vitro model for P450 dependent metabolism of acetaminophen.Biochemical and Biophysical Research Communications190, 15–19.
8.
ZhangS.Z., LipskyM.M., TrumpB.F., and HsuI.C. (1990). Neutral red (NR) assay for cell viability and xenobiotic-induced cytotoxicity in primary cultures of human and rat hepatocytes.Cell Biology and Toxicology6, 219–234.
9.
NebertD.V., and GelboinH.V. (1968). Substrate inducible microsomal aryl hydrocarbon hydroxylase in mammalian cell culture.The Journal of Biological Chemistry234, 6242–6249.
10.
BurkeM.D., ThompsonS., ElcombeC.R., HalpertJ., HaaparantaT., and MayerR.T. (1985). Ethoxy-pentoxy- and benzyloxyphenoxazones and homologues: a series of substrates to distinguish between different induced cytochromes P450.Biochemical Pharmacology34, 3337–3345.
11.
EdwardsA.M., GlistakM.L., and LucasCM. (1984). 7-Ethoxycoumarin deethylase activity as a convenient measure of liver drug metabolizing enzymes, regulation in cultured rat hepatocytes.Biochemical Parmacology33, 1537–1546.
12.
LowryO.M., RosenbroughN.J., FarrA.L., and RandallR.J. (1951). Protein measurements with the Folin phenol reagent.The Journal of Biological Chemistry193, 265–275.
13.
MaizelJ.B. (1979). Polyacrylamide gel electrophoresis of viral proteins. In Methods in Virology, Vol. 5 (ed. MarasmoroschK., KopprowskiN.), pp. 355–375. New York: Academic Press.
14.
DonatoM.T., BassiA.M., GomezM.J., PencoS., HerreroE., AdamD., CastellJ.V., and FerroM. (1994). Evaluation of the xenobiotic biotransformation capability of six rodent hepatoma cell lines in comparison with rat hepatocytes.In Vitro Cellular and Developmental Biology304, 574–580.
15.
GrantM.H., DuthieS.J., GrayA.G., and BurkeM.D. (1988). Mixed function oxidase and UDP-glucuronyltransferase activities in the human Hep G2 hepatoma cell line.Pharmacology37, 4111–4116.
16.
GonzalesF.J. (1993). Cytochrome P450 in humans. In Handbook of Experimental Pharmacology (ed. SchenkmanJ.B., GreimH.), pp. 239–257. Berlin, Heidelberg, New York.Springer-Verlag.
17.
RadiceS., CipellettiB., ChiesaraE., and MarabiniL. (1996). Effect of benomyl on cytochrome P450 and on mixed function oxidases in primary cultured rat hepatocytes.Pharmacological Research, in press.
18.
DiveC., WorkmanP., and WatsonJ.W. (1987). Novel dynamic flow cytoenzymological determination of intracellular esterase inhibition by BCNU and related isocyanates.Biochemical Pharmacology36, 3731–3738.
19.
HellmanB., and LaryeaD. (1990). Inhibitory effects of benomyl and carbendazim on the [3H]thymidine incorporation in various organs of the mouse: evidence for a more pronounced action of benomyl.Toxicology61, 161–169.
20.
JochheimC.M., and BaillieT.A. (1994). Selective and irreversible inhibition of glutathione reductase in vitro by carbamate thioester conjugates of methyl-isocyanate.Biochemical Pharmacology47, 1197–1206.
21.
IshizakiH., BradyJ.F., NingS.M., and YangC.S. (1990). Effect of phenethylisothiocyanate on microsomal N-nitrosodimethylamine metabolism and other monooxygenase activities.Xenobiotica20, 255–264.
22.
SmithT.J., GuoZ., LiC., NingS.M., ThomasP.E., and YangC.S. (1993). Mechanisms of inhibition of 4(methylnitrosoamino)-l-(3-pyridyl)-l-butanone: bioactivation in mouse by dietary phenethylisothiocyanate.Cancer Research53, 3276–3282.
23.
KocarekT.A., SchuetzE.G., and GuzelianP.S. (1991). Selective induction of cytochrome P450 by Kepone (chlordecone) in primary cultures of adult rat hepatocytes.Molecular Pharmacology40, 203–210.
24.
BradyJ.F., LiD., IshizakiH., and YangC.S. (1988). Effect of diallyl sulphide on rat liver microsomal nitrosamine metabolism and other monooxygenase activities.Cancer Research48, 5937–5940.
25.
BradyJ.F., LeeM.J., IshizakiH., and YangC.S. (1987). Diethylether as a substrate for acetone/ethanol-inducible cytochrome P450 and as an inducer for cytochrome(s) P450.Molecular Pharmacology33, 148–154.
26.
YamazoeY., ShimadaM., MarayamaN., and KatoR. (1987). Suppression of levels of phenobarbital-inducible rat liver cytochrome P450 by pituitary hormone.The Journal of Biological Chemistry262, 7423–7428.
