title: Expression and activity of cytochromes P450 2E1, 2A, and 2B in the mouse ovary: the effect of 4-vinylcyclohexene and its diepoxide metabolite.
abstract: 4-Vinylcyclohexene (VCH), an occupational chemical, causes destruction of small preantral follicles (F1) in mice. Previous studies suggested that VCH is bioactivated via cytochromes P450 (CYP450) to the ovotoxic, diepoxide metabolite, VCD. Whereas hepatic CYP450 isoforms 2E1, 2A, and 2B can metabolize VCH, the role of ovarian metabolism is unknown. This study investigated expression of these isoforms in isolated ovarian fractions (F1, 25-100 microm; F2, 100-250 microm; F3, >250 microm; interstitial cells, Int) from B6C3F1 mice dosed daily (15 days; ip) with vehicle, VCH (7.4 mmol/kg/day) or VCD (0.57 mmol/kg/day). Ovaries were removed and either isolated into specific ovarian compartments for mRNA analysis, fixed for immunohistochemistry, or prepared for enzymatic assays. mRNA and protein for all isoforms were expressed/distributed in all ovarian fractions from vehicle-treated mice. In the targeted F1 follicles, VCH or VCD dosing increased (p < 0.05) mRNA encoding CYP2E1 (645 +/- 14% VCH; 582 +/- 16% VCD), CYP2A (689 +/- 8% VCH; 730 +/- 22% VCD), and CYP2B (246 +/- 7% VCH) above control. VCH dosing altered (p < 0.05) mRNA encoding CYP2E1 in nontargeted F3 follicles (168 +/- 7%) and CYP2A in Int (207 +/- 19%) above control. Immunohistochemical analysis revealed the greatest staining intensity for all CYP isoforms in the Int. VCH dosing altered (p < 0.05) staining intensity in Int for CYP2E1 (19 +/- 2.4% below control) and CYP2A (39 +/- 5% above control). Staining intensity for CYP2B was increased (p < 0.05) above control in granulosa cells of small preantral (187 +/- 42%) and antral (63 +/- 8%) follicles. Catalytic assays in ovarian homogenates revealed that CYP2E1 and CYP2B were functional. Only CYP2E1 activity was increased (149 +/- 12% above control; p < 0.05) by VCH dosing. The results demonstrate that mRNA and protein for CYP isoforms known to bioactivate VCH are expressed in the mouse ovary and are modulated by in vivo exposure to VCH and VCD. Interestingly, there is high expression of these isoforms in the Int. Thus, the ovary may contribute to ovotoxicity by promoting bioactivation of VCH to the toxic metabolite, VCD.
authors.affiliation: University of Arizona, Department of Pharmacology and Toxicology, Tucson, Arizona 85721, USA.
authors.forename: Ellen A
authors.initials: EA
authors.lastname: Cannady
authors.forename: Cheryl A
authors.initials: CA
authors.lastname: Dyer
authors.forename: Patricia J
authors.initials: PJ
authors.lastname: Christian
authors.forename: I Glenn
authors.initials: IG
authors.lastname: Sipes
authors.forename: Patricia B
authors.initials: PB
authors.lastname: Hoyer
mesh_terms: D000818:Animals; D003510:Cyclohexanes / Q000008:administration & dosage / Q000633:toxicity*; D053138:Cyclohexenes; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D005260:Female; D066298:In Vitro Techniques; D007274:Injections, Intraperitoneal; D007527:Isoenzymes; D051379:Mice; D008815:Mice, Inbred Strains; D018613:Microscopy, Confocal; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology; D010053:Ovary / Q000187:drug effects* / Q000201:enzymology; D011506:Proteins / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014020:Tissue Extracts / Q000378:metabolism; D014753:Vinyl Compounds / Q000008:administration & dosage / Q000633:toxicity*
chemical_list: D003510:Cyclohexanes; D053138:Cyclohexenes; D007527:Isoenzymes; D011506:Proteins; D012333:RNA, Messenger; D014020:Tissue Extracts; D014753:Vinyl Compounds; C013733:4-vinylcyclohexene; C012606:4-vinyl-1-cyclohexene dioxide; D003577:Cytochrome P-450 Enzyme System