Embryonic exposure to the fungicide vinclozolin causes virilization of females and alteration of progesterone receptor expression in vivo: an experimental study in mice.
BACKGROUND
Vinclozolin is a fungicide that has been reported to have anti-androgenic effects in rats. We have found that in utero exposure to natural or synthetic progesterones can induce hypospadias in mice, and that the synthetic progesterone medroxyprogesterone acetate (MPA) feminizes male and virilizes female genital tubercles. In the current work, we selected a relatively low dose of vinclozolin to examine its in utero effects on the development of the genital tubercle, both at the morphological and molecular levels.


METHODS
We gave pregnant dams vinclozolin by oral gavage from gestational days 13 through 17. We assessed the fetal genital tubercles from exposed fetuses at E19 to determine location of the urethral opening. After determination of gonadal sex, either genital tubercles were harvested for mRNA quantitation, or urethras were injected with a plastic resin for casting. We analyzed quantified mRNA levels between treated and untreated animals for mRNA levels of estrogen receptors alpha and beta, progesterone receptor, and androgen receptor using nonparametric tests or ANOVA. To determine effects on urethral length (males have long urethras compared to females), we measured the lengths of the casts and performed ANOVA analysis on these data.


RESULTS
Our morphological results indicated that vinclozolin has morphological effects similar to those of MPA, feminizing males (hypospadias) and masculinizing females (longer urethras). Because these results reflected our MPA results, we investigated the effects of in utero vinclozolin exposure on the mRNA expression levels of androgen, estrogen alpha and beta, and progesterone receptors. At the molecular level, vinclozolin down-regulated estrogen receptor alpha mRNA in females and up-regulated progesterone receptor mRNA. Vinclozolin-exposed males exhibited up-regulated estrogen receptor alpha and progesterone receptor mRNA, effects we have also seen with exposure to the synthetic estrogen, ethinyl estradiol.


CONCLUSIONS
The results suggest that vinclozolin virilizes females and directly or indirectly affects progesterone receptor expression. It also affects estrogen receptor expression in a sex-based manner. We found no in vivo effect of vinclozolin on androgen receptor expression. We propose that vinclozolin, which has been designated an anti-androgen, may also exert its effects by involving additional steroid-signaling pathways.
Buckley
Jill
J
Institute for the Study and Treatment of Hypospadias, Department of Urology, School of Medicine, University of California, HSW 1434, San Francisco, CA 94143, USA. jbuckley@urology.ucsf.edu
Willingham
Emily
E
Agras
Koray
K
Baskin
Laurence S
LS
D000704:Analysis of Variance; D000726:Androgen Antagonists / Q000633:toxicity*; D000818:Animals; D005260:Female; D005333:Fetus / Q000187:drug effects*; D005659:Fungicides, Industrial / Q000008:administration & dosage / Q000633:toxicity*; D005865:Gestational Age; D007021:Hypospadias / Q000139:chemically induced; D008297:Male; D051379:Mice; D010080:Oxazoles / Q000008:administration & dosage / Q000633:toxicity*; D011247:Pregnancy; D012333:RNA, Messenger / Q000032:analysis / Q000187:drug effects; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018709:Statistics, Nonparametric; D014521:Urethra / Q000187:drug effects; D014770:Virilism / Q000139:chemically induced*
D000726:Androgen Antagonists; D005659:Fungicides, Industrial; D010080:Oxazoles; D012333:RNA, Messenger; C025643:vinclozolin