An in vitro investigation of the actions of reproductive hormones on the cervix of the ewe in the follicular stage: The effects of 17β-estradiol, oxytocin, FSH, and arachidonic acid on the cervical pathway for the synthesis of prostaglandin E2

Abstract

During the periovulatory period, the cervix of the ewe relaxes and this mechanism is thought to be mediated by oxytocin and prostaglandin E2 (PGE2) in response to increased concentrations of 17β-estradiol and perhaps FSH. The aim of the study was to determine the in vitro effects of 17β-estradiol, FSH, oxytocin, and arachidonic acid (AA) on the synthesis of PGE2 and on the expression of oxytocin receptor (OTR), cytoplasmic phospholipase A2 (cPLA2), and cyclooxygenase 2 (COX-2) in explants of cervical tissue collected from ewes in the periovulatory phase of the estrous cycle. Cervical minces from ewes in the follicular phase of the estrous cycle were cultured in supplemented Eagle's Minimum Essential Medium for 48 hours with 17β-estradiol, FSH, oxytocin, or AA. After incubation, the tissue was stored at −80 °C and the media at −20 °C. Western immunoblotting was used to determine relative levels of OTR, cPLA2, and COX-2 in cervical tissue, and the media was analyzed by RIA, to determine the concentration of PGE2. The addition of 17β-estradiol increased the concentration of PGE2 in the media (P = 0.001), the levels of COX-2 (P = 0.02) and OTR (P = 0.006) but not those of cPLA2 (P = 0.15). The addition of FSH increased the levels of COX-2 (P = 0.01) but, it had no effect on the concentration of PGE2 (P = 0.08) or on the levels of OTR (P = 0.07) and cPLA2 (P = 0.15). Oxytocin did not increase the levels of COX-2 (P = 0.38) but increased those of OTR (P = 0.001) and cPLA2 (P = 0.01) but not on the concentration of PGE2 in the media. Arachidonic acid increased the levels of cPLA2 (P = 0.01) and those of COX-2 (P = 0.02) but not the concentration of PGE2 in the media. Our findings suggest that the PGE2-mediated mechanisms of cervical relaxation in the ewe during the follicular phase are stimulated by FSH, 17β-estradiol, oxytocin, and AA. They all appear to act by inducing receptors and enzymes along the synthetic pathway for PGE2.