title: Effect of TCDD exposure on CYP1A1 and CYP1B1 expression in explant cultures of human endometrium.
abstract: Endometriosis is a debilitating disease estimated to affect 10% of reproductive-age women and characterized by the growth of endometrial tissue outside of the uterus. The present study characterizes a human endometrial explant culture model for studying the direct effects of TCDD exposure by assessing the expression of CYP1A1 and CYP1B1 mRNA (Northern blotting), protein (Western blotting), and activity (7-ethoxyresorufin-O-deethylase; EROD) in explants cultured with and without TCDD. Explants were obtained at laparoscopy or laparotomy from women undergoing surgery for tubal ligation, endometriosis, or pelvic pain unrelated to endometriosis. The explants were cultured with 10 nM estradiol (E(2)) or 1 nM E(2) plus 500 nM progesterone (P(4)) with or without TCDD (first 24 h). The expression of CYP1A1 and CYP1B1 mRNA was greatest with 10 nM TCDD and increased up to 72 h after initial exposure. EROD activity increased up to 120 h. Explants from a secretory phase biopsy became reorganized in culture and formed a new epithelial membrane, while maintaining basic endometrial morphology and viability for up to 120 h. At 24 h, TCDD significantly increased CYP1A1 and CYP1B1 mRNA, and at 72 h, TCDD significantly increased EROD activity and CYP1B1 protein compared to explants cultured without TCDD for similar times. CYP1B1 protein also exhibited substantial constitutive expression that was similar in uncultured biopsies, where CYP1B1 protein was immunolocalized in the cytoplasm of epithelial glands, with only occasional patches of protein in the surface epithelial membrane. In explants cultured with and without TCDD exposure, CYP1B1 protein was localized in the cytoplasm of the new surface epithelial membrane and glands closest to the surface. CYP1A1 protein was not detected in uncultured biopsies or explants. Both younger age (age 30 and under) and proliferative phase were associated with higher TCDD-induced EROD activity in specimens treated with E(2):P(4). No significant endometriosis-related differences were observed for any of the biomarkers, but the detection of disease-specific change was limited by small sample size and variability in tissue-cycle phase. The human endometrial explant culture model will be useful for future studies of the effects of dioxin-like compounds on human endometrium in relationship to cycle phase and hormonal exposure.
authors.affiliation: Department of Biotechnical and Clinical Laboratory Science, State University of New York at Buffalo, 3435 Main Street, Buffalo, NY 1421, USA. dbofinge@acsu.buffalo.edu
authors.forename: D P
authors.initials: DP
authors.lastname: Bofinger
authors.forename: L
authors.initials: L
authors.lastname: Feng
authors.forename: L H
authors.initials: LH
authors.lastname: Chi
authors.forename: J
authors.initials: J
authors.lastname: Love
authors.forename: F D
authors.initials: FD
authors.lastname: Stephen
authors.forename: T R
authors.initials: TR
authors.lastname: Sutter
authors.forename: K G
authors.initials: KG
authors.lastname: Osteen
authors.forename: T G
authors.initials: TG
authors.lastname: Costich
authors.forename: R E
authors.initials: RE
authors.lastname: Batt
authors.forename: S T
authors.initials: ST
authors.lastname: Koury
authors.forename: J R
authors.initials: JR
authors.lastname: Olson
mesh_terms: D001189:Aryl Hydrocarbon Hydroxylases; D051784:Aryl Hydrocarbon Receptor Nuclear Translocator; D001483:Base Sequence; D046508:Culture Techniques; D019363:Cytochrome P-450 CYP1A1 / Q000235:genetics*; D065633:Cytochrome P-450 CYP1B1; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers; D004268:DNA-Binding Proteins; D004717:Endometrium / Q000187:drug effects* / Q000201:enzymology; D005260:Female; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D006801:Humans; D007150:Immunohistochemistry; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology*; D018336:Receptors, Aryl Hydrocarbon / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics
chemical_list: C497396:ARNT protein, human; D017931:DNA Primers; D004268:DNA-Binding Proteins; D000072317:Polychlorinated Dibenzodioxins; D018336:Receptors, Aryl Hydrocarbon; D014157:Transcription Factors; D051784:Aryl Hydrocarbon Receptor Nuclear Translocator; D003577:Cytochrome P-450 Enzyme System; D001189:Aryl Hydrocarbon Hydroxylases; C585496:CYP1B1 protein, human; D019363:Cytochrome P-450 CYP1A1; D065633:Cytochrome P-450 CYP1B1