Nitric oxide (NO) production is essential to facilitate rises in uterine blood flow (UBF) during pregnancy. It has been proposed that the metabolites of E2β, 2-hydroxyestradiol (2-OHE2), 4-hydroxyestradiol (4-OHE2), 2-methoxyestradiol (2-ME2), and 4-methoxyestradiol (4-ME2) play a role in mediating vasodilation and rises in UBF during pregnancy. We previously showed that the E2β metabolites stimulate prostacyclin production in pregnancy-derived ovine uterine artery endothelial cells (P-UAECs); however, it is unknown whether the E2β metabolites also induce NO production. Herein, UAECs derived from nonpregnant and pregnant ewes were used to test the hypothesis that E2β metabolites stimulate NO production in a pregnancy-specific manner. Specific estrogen receptor (ER) and adrenergic receptor (AR) antagonists were used to determine the roles of ERs or ARs in E2β metabolite-induced NO production. E2β and its metabolites increased total nitric oxide metabolites (NOx) levels (NO2 + NO3) in P-UAECs, but not in NP-UAECs. Pretreatment with combined 1 µmol/L 1,3-bis(4-hydroxyphenyl)-4-methyl-5-[4-(2-piperidinylethoxy)phenol]-1H-pyrazole dihydrochloride (MPP; ER-α antagonist) and 1 µmol/L 4-[2-phenyl-5,7-bis(trifluoromethyl)pyrazolo[1,5-a]pyrimidin-3-yl]phenol (PHTPP; ER-β antagonist) inhibited the rises in NOx levels stimulated by E2β and 2-ME2, but had no effect on 2-OHE2-, 4-OHE2-, or 4-ME2-stimulated rises in NOx levels. Pretreatment with yohimbine (α2-AR antagonist) and propranolol (β2,3-AR antagonist) inhibited the rises in NOx levels stimulated by 2-OHE2, but not by E2β, 4-OHE2, 2-ME2, or 4-ME2. These data demonstrate that E2β metabolites stimulate NO synthesis via ERs or ARs in UAECs in a pregnancy-specific manner, suggesting that these metabolites contribute to rises in vasodilation and UBF during pregnancy.
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