Aryl Hydrocarbon Hydroxylase and Epoxide Hydratase Activities: Age Effects in Mammary Epithelial Cells of Sprague-Dawley Rats23

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Abstract
By use of techniques developed for rat mammary epithelial cell isolation, studies were conducted to characterize aryl hydrocarbon hydroxylase (AHH) and epoxide hydratase (EH) activities and to determine whether changes in the enzyme activities might contribute to the age-dependent changes in polycyclic aromatic hydrocarbon (PAH)-induced tumorigenesis in the rat mammary gland. Constitutive AHH and EH activities in rat mammary epithelial cell homogenates were substantially lower than those measured in liver (9,500×g) supernatant. The administration of 3-methylcholanthrene (MCA) or 7,12-dimethylbenz[a]anthracene increased mammary epithelial AHH activity 93- and 44-fold, respectively, whereas hepatic AHH levels rose 17- and 6-fold. Neither inducer had any effect on constitutive EH activity in either tissue. The addition (0.1 mM) of 7,8-benzoflavone (7,8-BF), SKF 525-A, or metyrapone to homogenates of mammary epithelial cells from MCA-treated rats inhibited AHH activity 93, 48, and 22%, respectively. In contrast, 7,8-BF addition decreased rat liver AHH activity 83%, whereas SKF 525-A and metyrapone had little effect. EH activities in rat liver and mammary epithelial cells were reduced with addition of 1,1,1-trichloro-2-propene oxide and cyclohexene oxide; however, trans-stilbene oxide had no effect in either tissue preparation. Additional studies determined the age-dependent changes in rat mammary epithelial AHH and EH activities. Constitutive AHH activity remained unchanged in rats between 40 and 100 days of age. In contrast, mammary epithelial EH activity was highest in 40-day-old Sprague-Dawley rats, declined in rats between 40 and 60 days of age, and remained constant in rats between 60 and 100 days of age. The level of MCA-induced AHH activity in rat mammary epithelial cell homogenates was age-dependent. MCA administration induced constitutive AHH activity 85- and 74-fold in mammary epithelial cells isolated from 50- and 60-day-old Sprague-Dawley females. The magnitude of mammary epithelial AHH induction was significantly lower in both younger and older animals. The results demonstrated PAH-metabolizing enzymes in rat mammary epithelial cells and suggested that the age-dependent incidence of mammary tumorigenesis may be attributed in part to the age-dependent inducibility of AHH activity.