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Epoxide-metabolizing Enzymes in Mammary Gland and Liver from BALB/c Mice and Effects of Inducers on Enzyme Activity¹

State of California, Department of Food and Agriculture, Medical Toxicology Branch, Sacramento, California 95814 [M. H. S.] and Departments of Entomology and Environmental Toxicology, University of California, Davis, California 95616 [R. N. W., B. D. H]

Epoxide hydrolases (EC 3.3.2.3) (EH) are hydrolytic enzymes which may play an important role in the activation and detoxification of mammary carcinogens. In this study, microsomal, cytosolic, and cholesterol epoxide hydrolases along with glutathione S-transferase were characterized in liver and mammary gland from nulliparous and lactating BALB/c mice and from mice transplanted with preneoplastic hyperplastic out-growths. Clofibrate, butylated hydroxyanisole, and ß-naphthoflavone were used to induce EH. Significant epoxide hydrolysis was observed in microsomal and cytosolic subcellular fractions assayed with cis- and trans-stilbene oxide, benzo(a(pyrene-4,5-oxide, and cholesterol epoxide. The hydrolysis rates were significantly different for nulliparous and lactating animals, in both mammary gland and liver. Clofibrate increased the activity of all forms of EH in liver, but not mammary gland. Butylated hydroxyanisole and ß-naphthaflavone appeared to induce cytosolic glutathione S-transferase as well as some, but not all, forms of EH in liver and mammary gland regardless of hormonal stimuli. The inducers produced different effects in mammary gland as compared with liver. This may be due to either differing amounts of inducer reaching the target site or different regulation of the enzymes in mammary gland and liver. Hyperplastic outgrowths and liver from hyperplastic outgrowth-transplanted animals demonstrated significantly different EH and cytosolic glutathione S-transferase activities from those of nulliparous and lactating animals. This observation offers preliminary evidence that levels of epoxide-metabolizing enzymes are altered when mammary tissue is transformed. Mammary gland cytosolic EH was purified by affinity chromatography and compared to that from liver by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Western blotting, enzyme-linked immunosorbent assay, isoelectric focusing, and enzyme inhibition by 4-phenylchalcone oxide. Cytosolic EH from the mammary gland appears to be identical to the liver enzyme by all the above mentioned biochemical and biophysical parameters.

¹ This investigation was supported in part by NIH Grant ES02710-07. M. H. S. is a recipient of National Institute of Environmental Health Sciences Postdoctoral Fellowship 1-F32-ES05328-01 BI-2, R. N. W. is a recipient of the National Institute of Environmental Health Sciences Training Grant 5-T32-ES07059, and B. D. H. is the recipient of the Burroughs Wellcome Toxicology Scholar Award.

² To whom requests for reprints should be addressed, at the Department of Entomology, University of California, Davis, CA 95616.

Received 4/27/87. Revised 8/19/87. Revised 11/23/87. Accepted 12/ 1/87.