title: Methoxychlor directly affects ovarian antral follicle growth and atresia through Bcl-2- and Bax-mediated pathways.
abstract: Methoxychlor (MXC) is an organochlorine pesticide and reproductive toxicant. While in vivo studies indicate that MXC exposure increases antral follicle atresia, in part by altering apoptotic regulators (Bcl-2 and Bax), they do not distinguish whether MXC does so via direct or indirect mechanisms. Therefore, we utilized an in vitro follicle culture system to test the hypothesis that MXC is directly toxic to antral follicles, and that overexpression of anti-apoptotic Bcl-2, or deletion of pro-apoptotic Bax, protects antral follicles from MXC-induced toxicity. Antral follicles were isolated from wild-type (WT), Bcl-2 overexpressing (Bcl-2 OE), or Bax deficient (BaxKO) mice, and exposed to dimethylsulfoxide (control) or MXC (1-100 microg/ml) for 96 h. Follicle diameters were measured every 24 h to assess growth. After 96 h, follicles were histologically evaluated for atresia or collected for quantitative PCR analysis of Bcl-2 and Bax mRNA levels. MXC (10-100 microg/ml) significantly inhibited antral follicle growth at 72 and 96 h, and increased atresia (100 microg/ml) compared to controls at 96 h. Furthermore, MXC increased Bax mRNA levels between 48-96 h and decreased Bcl-2 mRNA levels at 96 h. While MXC inhibited growth of WT antral follicles beginning at 72 h, it did not inhibit growth of Bcl-2 OE or BaxKO follicles until 96 h. MXC also increased atresia of small and large WT and BaxKO antral follicles over controls, but it did not increase atresia of large Bcl-2 OE antral follicles over controls. These data suggest that MXC directly inhibits follicle growth partly by Bcl-2 and Bax pathways, and increases atresia partly through Bcl-2 pathways.
authors.affiliation: Program in Toxicology and Department of Epidemiology and Preventive Medicine, University of Maryland, Baltimore, Maryland 21201, USA.
authors.forename: Kimberly P
authors.initials: KP
authors.lastname: Miller
authors.forename: Rupesh K
authors.initials: RK
authors.lastname: Gupta
authors.forename: Chuck R
authors.initials: CR
authors.lastname: Greenfeld
authors.forename: Janice K
authors.initials: JK
authors.lastname: Babus
authors.forename: Jodi A
authors.initials: JA
authors.lastname: Flaws
mesh_terms: D000818:Animals; D004305:Dose-Response Relationship, Drug; D005260:Female; D005496:Follicular Atresia / Q000187:drug effects* / Q000378:metabolism; D007306:Insecticides / Q000633:toxicity*; D008731:Methoxychlor / Q000633:toxicity*; D051379:Mice; D018345:Mice, Knockout; D009924:Organ Culture Techniques; D006080:Ovarian Follicle / Q000187:drug effects* / Q000254:growth & development / Q000473:pathology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051028:bcl-2-Associated X Protein
chemical_list: C494827:Bax protein, mouse; D007306:Insecticides; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D051028:bcl-2-Associated X Protein; D008731:Methoxychlor