title: Evaluation of ovotoxicity induced by 7, 12-dimethylbenz[a]anthracene and its 3,4-diol metabolite utilizing a rat in vitro ovarian culture system.
abstract: The polycyclic aromatic hydrocarbon 7, 12-dimethylbenz[a]anthracene, (DMBA), targets and destroys all follicle types in rat and mouse ovaries. DMBA requires bioactivation to DMBA-3,4-diol-1,2-epoxide for ovotoxicity via formation of the intermediate, DMBA-3,4-diol (catalyzed by microsomal epoxide hydrolase; mEH). mEH was shown to be involved in DMBA bioactivation for ovotoxicity induction in B6C3F(1) mouse ovaries. The current study compared DMBA and DMBA-3,4-diol mediated ovotoxicity, and investigated mEH involvement in DMBA-3,4-diol bioactivation in Fischer 344 (F344) rat ovary. F344 postnatal day (PND) 4 rat ovaries were cultured in vehicle control or media containing 1) DMBA or DMBA-3,4-diol (12.5 nM - 1 muM; 15 days); 2) DMBA (1 muM; 6 h - 15 days); and 3) DMBA (1 muM) or DMBA-3,4-diol (75 nM)+/-the mEH activity inhibitor cyclohexene oxide (CHO; 2 mM; 4 days). Ovaries were histologically evaluated and mEH mRNA and protein were measured by reverse transcriptase PCR or Western blotting, respectively. Ovotoxicity following 15 days of culture occurred (P<0.05) at lower concentrations of DMBA-3,4-diol (12.5 nM - primordial; 75 nM - primary) than DMBA (75 nM - primordial; 375 nM - primary). The temporal pattern of mEH expression following DMBA exposure showed mRNA up-regulation (P<0.05) on day 2, with increased protein (P<0.05) on day 4, the earliest time of observed follicle loss (P<0.05). mEH inhibition prevented DMBA-induced, but not DMBA-3,4-diol-induced ovotoxicity. These results demonstrate a conserved response in mice and rats for ovarian mEH involvement in DMBA bioactivation to its ovotoxic, 3,4-diol-1,2-epoxide form.
authors.affiliation: Department of Physiology, University of Arizona, Tucson, AZ 85745, USA.
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Igawa
authors.forename: Aileen F
authors.initials: AF
authors.lastname: Keating
authors.forename: Kathila S
authors.initials: KS
authors.lastname: Rajapaksa
authors.forename: I Glenn
authors.initials: IG
authors.lastname: Sipes
authors.forename: Patricia B
authors.initials: PB
authors.lastname: Hoyer
mesh_terms: D015127:9,10-Dimethyl-1,2-benzanthracene / Q000378:metabolism / Q000633:toxicity*; D000818:Animals; D000831:Animals, Newborn; D001551:Benz(a)Anthracenes / Q000378:metabolism / Q000633:toxicity*; D001711:Biotransformation; D053138:Cyclohexenes / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004790:Enzyme Induction; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004851:Epoxide Hydrolases / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics; D005260:Female; D006080:Ovarian Follicle / Q000187:drug effects / Q000201:enzymology; D010053:Ovary / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D011916:Rats, Inbred F344; D013997:Time Factors; D046509:Tissue Culture Techniques
chemical_list: D001551:Benz(a)Anthracenes; D053138:Cyclohexenes; D004791:Enzyme Inhibitors; D012333:RNA, Messenger; C007193:cyclohexene oxide; D015127:9,10-Dimethyl-1,2-benzanthracene; C026832:3,4-dihydroxy-1,2-epoxy-1,2,3,4-tetrahydrobenz(a)anthracene; D004851:Epoxide Hydrolases