Studying endocrine disrupter action and toxicology in ER knock-out mice. Discussion

Estrogen receptors (ER) are thought to play a crucial role in development, reproduction, carcinogenesis and normal physiology. Using gene targeting techniques, we have produced lines of transgenic mice homozygous for the disrupted ERa gene (aERKO) and ERβ genes (βERKO). Comparable levels of ER-β mRNA were found in tissues of aERKO mice suggesting that ER-β expression is not dependent on ERα. aERKO mice were totally unresponsive to uterotropic assays with estradiol, Genistein, hydroxy TAM, DES or EGF treatment. Progesterone receptor (PR) mRNA was detected in aERKO mice, but not stimulated by estrogen in the uterus, mammary gland and ovary. Pituitary hormones are elevated in aERKO females but same as WT in βERKO mice indicating that ERa plays a major role in peptide hormone regulation. Genistien was equally effective in aERKO as WT mice in suppressing serum LH levels suggesting its action in this estrogen target response involved a non-ER mediated mechanism compared to the uterine genistein activity Analysis of the mammary glands of adult aERKO mice, showed a primitive ductal rudiment while the βERKO had the fully developed ductal tree seen in WT siblings. The influence of ERa activity on mammary tumorgenicity was evaluated by crossing aERKO with an oncogenic WNT-1 transgenic mouse line having an increased incidence of mammary tumors. Mice with WT ER levels and WNT-1 transgene show 98% tumor incidence at 27 weeks of age. αERKO/WNT-1 mice showed a delayed onset (54 weeks). These results indicate that mammary tumors can develop from ER negative tissues, but with a delayed onset. A second cross with erB2 (Neu) transgenics shows that WT and hetero Neu mice have a tumor incidence at 54 wks of age, aERKO/Neu have an extremely low incidence past 90 wks. Compared to the WNT/ ERKO the ERKO/Neu have a poorer tumor rate suggesting that ERa plays a more prominent role in this oncogenic tumor induction. Since the initial description of DES carcinogenesis in both humans and mice, a major question was what aspect of the DES action was ER mediated or non-receptor mediated. Perinatal DES treatment to WT mice showed numerous phenotypic toxicological and carcinogenic effects including, persistent vaginal cornification, uterine hyperplasia, oviductal hypertrophy in females. In similarly treated males, DES induced seminal vesicle atrophy and prostatic hyperplasia. aERKO treated males and females showed none of the observed toxic effects indicating that ERa was involved in mediating these DES effects and not ERβ. Further characterization of the mice and comparison of the DES effects in βERKO mice and double ER gene KO mice will be needed to more fully the specific roles of the two different forms of ER in estrogen hormone toxicology and hormonal carcinogenesis.