PLATFORM SESSION 13. TOXICOLOGY
Monday, August 2, 2004
2:00 PM–4:00 PM
Chair: Leon Earl Gray, Jr.
Co-Chair: Nigel Noriega
(317) METHOXYCHLOR INDUCES ANTRAL FOLLICLE ATRESIA IN VITRO BY ALTERING Bcl-2 FAMILY MEMBERS.
Gupta, Rupesh1, Miller, Kimberly1, Jones, Jenny2, Flaws, Jodi1, 1 University of Maryland-School of Medicine, Baltimore, MD2 University of Maryland-School of Medicine, Baltimore, MD
ABSTRACT- Females are born with a finite number of primordial follicles, of which a small fraction survive and reach the penultimate antral stage, while others die via apoptosis. Antral follicles are responsible for releasing an egg for fertilization and synthesizing steroid hormones, thus maintaining cyclicity. Previous in vivo studies with the organochlorine pesticide methoxychlor (MXC) showed that antral follicles were primary targets of MXC with no effect on other follicle types. Specifically, MXC exposure decreased antral follicle numbers and increased the percentage of atretic antral follicles. In addition, studies using in vitro follicle culture demonstrated that MXC exposure inhibited follicle growth compared to controls at 72 and 96 hrs and increased follicular atresia compared to controls at 96 hrs. While there are different pathways leading to toxicant induced cell death, the Bcl-2 family of proteins is a major regulator of cell death, and has been shown to be involved in regulating follicular atresia. Thus, this work tested the hypothesis that MXC induces atresia of antral follicles by altering the expression of Bcl-2 family members in a dose and time dependent manner. To test this hypothesis, antral follicles were isolated from 39 day old CD-1 mouse ovaries and cultured in supplemented -minimum essential media. Follicles were exposed to vehicle control (dimethylsulfoxide) or MXC (1-100g/mL) for 24, 48, 72 and 96 hrs. After culture, real time polymerase chain reaction was performed on the follicles to measure mRNA levels of selected Bcl-2 family members including pro-apoptotic Bax, and anti-apoptotic Bcl-2 and Bcl-XL. MXC (100g/mL) significantly increased Bax mRNA levels compared to control at 48 hrs (control = 1.35 ± 0.10 genomic equivalents (ge); MXC = 2.27 ± 0.03 ge; n = 3; p≤0.04) and at 96 hrs (control = 0.65 ± 0.07 ge; MXC = 1.16 ± 0.08 ge; n = 3; p≤0.01). In contrast, MXC significantly reduced Bcl-XL mRNA levels compared to control by 96 hrs (control = 1.92 ± 0.23 ge; MXC = 0.99 ± 0.11 ge; n = 3; p≤0.01). There were no significant differences in Bcl-2 mRNA expression between MXC-treated and control follicles at the selected time points. These data suggest that MXC alters Bcl-2 family signaling pathways to induce antral follicle toxicity in the mouse ovary. Supported by NIH HD38955 and T32 ES07263.
KEY WORDS: follicles, methoxychlor, bcl-2 family