Long-term reproductive tract alterations in female mice treated neonatally with coumestrol.

The neonatal mouse has proven to be an excellent model to ascertain the deleterious effects of estrogenic compounds on the developing reproductive tract. This system has been used to determine the morphological consequences after neonatal exposure to coumestrol (a plant estrogen). The role of estrogens in carcinogenesis (as a cocarcinogen or a tumor promotor) is unresolved at this time. Many studies suggest that estrogens are capable of functioning in this manner and recent evidence in cellular and molecular oncology revealed that estrogens act by genetic and epigenetic mechanisms in cancer cells. This review discusses the long-term morphological alterations in the reproductive tract of mice exposed neonatally to coumestrol.

Reproductive abnormalities in female mice exposed neonatally to various doses of coumestrol.

Female C57BL/Crgl mice were neonatally exposed to various doses of coumestrol to determine the threshold doses required for the occurrence of reproductive tract abnormalities. Newborn mice received daily subcutaneous injections of 10(-3), 10(-2), 8 X 10(-2), 10(-1), 1, 5, 25, 50, and 100 micrograms coumestrol in 0.005 ml dimethyl sulfoxide (DMSO) or DMSO alone, or received no treatment for the first 5 d of life. Some of the animals were ovariectomized at 40 d of age. Mice were killed at 20-22 mo of age. All neonatal doses of coumestrol advanced vaginal opening before that of controls. At 2 and 20-22 mo of age, doses greater than or equal to 25 micrograms consistently resulted in ovary-independent persistent vaginal cornification as judged by vaginal smears. Intact untreated and DMSO-treated control mice exhibited aging changes in the genital tract, some cervical adenosis and early cervicovaginal pegs and downgrowths, uterine cystic glandular hyperplasia, corpora lutea, and scattered areas of ovarian ceroid deposition. Intact mice receiving neonatal coumestrol exhibited cervicovaginal pegs and downgrowths (at all doses with the exception of 25 and 50 micrograms), cervical adenosis (at doses greater than or equal to 8 X 10(-2) micrograms), uterine squamous metaplasia (significant at doses greater than or equal to 50 micrograms), and a decrease in uterine cystic glandular hyperplasia (significant at doses greater than or equal to 25 micrograms). The levels of 10(-1), 5, and 100 micrograms neonatal coumestrol daily resulted in hemorrhagic follicles. An increase in ovarian ceroid deposition (significant at doses greater than or equal to 5 micrograms) was observed. At 40 d and 20-22 mo of age, corpora lutea were consistently absent from the 100-micrograms-treated animals. Most of the ovariectomized untreated and DMSO-treated control animals showed typical castrate-like morphology of the genital tract, with the majority of the control mice exhibiting uterine cystic glandular hyperplasia. Ovariectomized mice receiving coumestrol neonatally exhibited various degrees of cervicovaginal alterations: pegs and downgrowths (significant at all doses with the exception of 10(-1) micrograms), endometrial collagen deposition (significant at greater than or equal to 25 micrograms), and reduced or absent uterine glands (significant at 10(-3), and 10(-11), and at all doses greater than or equal to 5 micrograms).

Long-term genital tract changes in female mice treated neonatally with coumestrol.

The neonatal mouse model has proven to be an effective system to examine long-term reproductive tract abnormalities resulting from early exposure to estrogens. Newborn C57BL/Crgl mice received 8 x 10(-2) micrograms diethylstilbestrol (DES) or 100 micrograms coumestrol (a plant estrogen) in 0.005 mL dimethyl sulfoxide (DMSO) or DMSO alone or received no treatment for the first 5 days of life. Half of the animals were ovariectomized at 40 days of age. Vaginal lavages were examined for 15 consecutive days before termination at 13 months of age, at which time genital tracts and mammary glands were removed for histological examination. Diethylstilbestrol- and coumestrol-treated animals exhibited ovary-independent persistent vaginal cornification as well as cervico-vaginal pegs and downgrowths, uterine squamous metaplasia, and an enhancement of age-related changes in the ovary including hemorrhagic follicles. In general, neonatal exposure to the naturally occurring plant estrogen, coumestrol, has long-term effects similar to those seen following exposure to natural and synthetic estrogens.

Reproductive alterations in female C57BL/Crgl mice exposed neonatally to zearalenone, an estrogenic mycotoxin.

Newborn female mice were injected daily for 5 days with 1 microgram zearalenone (Z, a weakly estrogenic mycotoxin present in cereal grains), resulting in ovary-dependent reproductive tract alterations at 8 months of age. Corpora lutea were absent from 25 of 34 (74%) Z-treated mice, indicating ovarian dysfunction. Fifty-six percent of Z-treated mice had dense collagen deposition in the uterine stroma and lacked uterine glands. Squamous metaplasia of the uterine luminal epithelium was found in 59% of Z-treated mice, and altered vaginal epithelium was found in 32% (2 mice had dysplastic lesions). Ovariectomized Z-treated mice were indistinguishable from ovariectomized controls.

Prolonged vaginal cornification and other changes in mice treated neonatally with coumestrol, a plant estrogen.

This study used the neonatal mouse model to determine if early exposure of female mice to coumestrol, a plant estrogen, would result in reproductive-tract alterations similar to those seen after neonatal exposure to diethylstilbestrol (DES). Newborn female C57BL/Crgl mice were given daily subcutaneous injections of 0.08 micrograms DES or 100 micrograms coumestrol in 0.005 ml dimethyl sulfoxide (DMSO), or DMSO alone, or were untreated, for the first 5 d of life. The doses chosen were equivalent in biological activity based on published uterine bioassay data (using young adult mice). Observations were made twice daily for 1.5 mo to determine the times of eye and complete vaginal opening. Half of the animals were ovariectomized at 40 d of age. Vaginal lavages were examined for 30 consecutive d beginning both at 2 and at 5 mo of age. DES and coumestrol significantly advanced the time of complete vaginal opening and induced a comparable degree of ovary-independent persistent vaginal cornification. In addition, coumestrol resulted in the occurrence of hemorrhagic ovarian follicles.