Estrogen receptor-beta messenger ribonucleic acid ontogeny in the prostate of normal and neonatally estrogenized rats.

Abstract

Neonatal exposure to estrogens permanently alters rat prostate growth and epithelial differentiation leading to prostatic dysplasia on aging. The effects are lobe-specific, with the greatest response observed in the ventral lobe. Recently, a novel estrogen receptor (ER) complementary DNA was cloned from the rat prostate and termed ER-beta (ER beta) due to its high homology with the classical ER alpha. The protein possesses high affinity for 17beta-estradiol, indicating that ER beta is an alternate molecule for mediating estrogenic effects. Importantly, ER beta messenger RNA (mRNA) was localized to rat prostatic epithelial cells, which contrasts with the stromal localization of ER alpha in the rat prostate. The present study was undertaken to determine the ontogeny of ER beta mRNA expression in the rat prostate lobes and to examine the effects of early estrogen exposure on prostatic ER beta expression. Male rat pups were given 25 microg estradiol or oil on days 1, 3, and 5; were killed on day 1, 3 (oils only), 6, 10, 30, or 90; and prostate lobes were frozen. Longitudinal sections were processed for in situ hybridization using an 35S-labeled antisense mRNA probe corresponding to a 400-bp EcoRI-AccI fragment in the 5' untranslated region of rat ER beta complementary DNA. Image analysis was used to quantitate silver grains. In addition, total RNA was isolated from the ventral prostate (VP) and used for semiquantitative RT-PCR. Results from in situ hybridization revealed that at birth, ER beta was equivalently expressed at low levels in both mesenchymal and epithelial cells in oil-treated rats. From day 1 onwards, expression in all stromal cells slowly and significantly declined, so that in the control adult prostate, stromal ER beta mRNA was slightly above background. In the oil-treated control rats, epithelial ER beta mRNA increased to moderate levels between days 6-10 in the VP and days 10-15 in the dorsal and lateral lobes as cells began differentiation and ducts lumenized. A further significant increase in ER beta message was observed at day 30, which indicates that full epithelial ER beta expression may require the completion of functional differentiation. By day 90, expression levels were maximal and similar between the lobes. RT-PCR substantiated this developmental increase in ER beta between days 1-90. Neonatal exposure to estrogens did not have an immediate effect on prostatic ER beta mRNA levels as determined by in situ hybridization and RT-PCR. However, the marked increase in epithelial cell expression at day 30 observed in the control VP was dampened in the VP of animals exposed neonatally to estrogens. By day 90, the VP of estrogenized rats possessed low ER beta message levels compared with the high expression in oil controls. In contrast, the dorsal and lateral lobes of neonatally estrogenized rats possessed high levels of ER beta mRNA at day 90, equivalent to controls. The present data demonstrate that ER beta mRNA expression in the rat prostate is developmentally regulated, and that neonatal estrogen can affect this expression in the adult VP. Because the effect of neonatal estrogens was not immediate, the data imply that early estrogen exposure may not directly autoregulate ER beta expression, and suggests that the adult effects on ER beta mRNA expression may be indirect. The differences in ER beta mRNA imprinting in the separate lobes may account for or reflect the lobe-specific neonatal estrogen imprints previously observed in the rat prostate.