Endothelial vasodilator production by ovine uterine and systemic arteries:: ovarian steroid and pregnancy control of ERα and ERβ levels

Pregnancy and the follicular phase are physiological states of elevated oestrogen levels and rises in uterine blood flow (UBF). The dramatic increase in utero-placental blood flow during gestation is required for normal fetal growth and development. Oestrogen exerts its vasodilatory effect by binding to its specific oestrogen receptors (ER) in target cells, resulting in increased expression and activity of endothelial nitric oxide synthase (eNOS) to relax vascular smooth muscle (VSM). However, the regulation of endothelial versus VSM ER alpha and ER beta expression in uterine arteries (UAs) during the ovarian cycle, pregnancy and with exogenous hormone replacement therapy (HRT) are currently unknown. ER mRNA and protein localization was determined by in situ hybridization (ISH) using S-35-labelled riboprobes and immunohistochemistry (IHC), respectively. UA endothelial (UAendo), UA VSM, omental artery endothelium (OA endo), and OAVSM proteins were isolated and ER alpha and ER beta protein expression was determined by Western analysis. We observed by ISH and IHC that ER alpha and ER beta mRNA and protein were localized in both UAendo and UA VSM. Immunoblot data demonstrated ovarian hormone specific regulation of ER alpha and ER beta protein in UAendo and UA VSM. Compared to luteal phase sheep, both ER beta and ER beta levels in UAendo were elevated in follicular phase sheep. Whereas ER beta was elevated by pregnancy in UAendo and UA VSM, ER alpha was not appreciably altered. eNOS was increased in UAendo from follicular and pregnant sheep. Ovariectomized ewes (OVEX) had substantially reduced UAendo ER beta, but not UAendo ER alpha or OAendo ER alpha and ER beta. In contrast, OVEX increased UA VSM ER alpha and ER beta and decreased OA VSM ER alpha and ER beta. Treatment with oestradiol-17 beta (E2 beta), but not progesterone or their combination, increased UAendo ER alpha levels. The reduced ER beta in UAendo from OVEX ewes was reversed by E-2 beta and progesterone treatment. While ER alpha and eNOS were not elevated in any other reproductive or non-reproductive endothelia tested, ER beta was augmented by pregnancy in uterine, mammary, placenta, and coronary artery endothelia. ER alpha and ER beta mRNA and protein are expressed in UA endothelium with expression levels depending on the endocrine status of the animal, indicating UA endothelium is a target for oestrogen action in vivo, and that the two receptors appear to be differentially regulated in a spatial and temporal fashion with regard to the reproductive status or HRT.