Characteristics of aryl hydrocarbon hydroxylase activity in rat mammary epithlial cells grown in primary culture.
Rat mammary epithelial cells grown in primary culture contain the microsomal enzyme, aryl hydrocarbon (benzo[a]pyrene) hydroxylase (AHH), which catalyses the oxidative conversion of polycyclic aromatic hydrocarbons (PAH) to more polar derivatives. Constitutive AHH activity, measured with an established fluorometric method, was 46 pmol/mg protein/h in homogenates of rat mammary epithelial cells after 5 days in culture. The addition of dimethylbenz[a]anthracene (DMBA), benz[a]anthracene (BA), or 3-methylcholanthrene (3-MC) to the cell culture medium increased AHH activity 5.3-, 4.7- and 2.4-fold, respectively. Kinetic studies revealed that maximal hydroxylase induction occurred 16 h after 1 micro M DMBA was added to the culture medium. The decay of the DMBA-induced hydroxylase was biphasic: one component had a t1/2 of 15--30 min and another a t1/2 of 4 h. Norepinephrine, 17 beta-estradiol and 5,6-benzoflavone also increased AHH activity in mammary epithelial cells in vitro, however, sodium phenobarbital had no effect. Fetal bovine serum (FBS), previously shown to be a potent in vitro inducer of AHH activity, had no effect on either constitutive or DMBA-induced mammary epithelial hydroxylase activities following treatment with 1% activated charcoal. Metyrapone and 7,8-benzoflavone, inhibitors of microsomal mixed function oxidase activity, reduced both constitutive and DMBA-induced AHH activities when added to homogenates of untreated and DMBA-treated mammary epithelial cells. The addition of 7,8-benzoflavone reduced both constitutive and DMBA-induced hydroxylase activities by approx. 80%, whereas metyrapone addition inhibited these activities by 20%. The study demonstrates several in vitro factors which alter AHH activity in primary cultures of rat mammary epithelial cells.
D015127:9,10-Dimethyl-1,2-benzanthracene / Q000494:pharmacology; D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000378:metabolism*; D001551:Benz(a)Anthracenes / Q000494:pharmacology; D001571:Benzoflavones / Q000494:pharmacology; D001769:Blood; D002478:Cells, Cultured; D003470:Culture Media; D004790:Enzyme Induction / Q000187:drug effects; D005260:Female; D008321:Mammary Glands, Animal / Q000201:enzymology*; D008748:Methylcholanthrene / Q000494:pharmacology; D008797:Metyrapone / Q000494:pharmacology; D051381:Rats
D001551:Benz(a)Anthracenes; D001571:Benzoflavones; D003470:Culture Media; D008748:Methylcholanthrene; D015127:9,10-Dimethyl-1,2-benzanthracene; D001189:Aryl Hydrocarbon Hydroxylases; D008797:Metyrapone