Discriminating analysis of “in vitro” prostaglandin release by myometrial and luminal sides of the ewe endometrium

An original perifusion device which allows a discrimination between the 30 mn releases of prostaglandins F₂α and E₂ by the luminal and the myometrial faces of sheep endometrium is described. Tissue was sampled on day 4, 14, 16 or 17 of the cycle and on day 14 or 17 of pregnancy. Total prostaglandin (PG) release measured with this device was in good agreement with PG's concentrations in media of in vitro endometrium incubations already described.Discrimination analysis of the PGs release by each side of the endometrial tissue during the 30 mn perifusion time releaved that PGF₂α concentrations of the perifusion medium issued from the lumen compartment were higher than those of the myometrial compartment in all physiological status where corpus luteum is active (including early pregnancy). Therefore in the ewe, it seems that luteal structure maintenance during early pregnancy is not due, as in the giltm to a shift in PGF₂α secretion towards the uterine lumen.     

Effects of pregnancy, oxytocin, ovine trophoblast protein-1 and their interactions on endometrial production of prostaglandin F2 alpha in vitro in perifusion chambers

Pregnancy and intrauterine infusion of ovine trophoblast protein one (oTP-1) decrease oxytocin-induced secretion of prostaglandin F2 alpha (PGF) from the uterus. In the present study, effects of oTP-1 and pregnancy on endometrial secretion of PGF were examined in an in vitro perifusion system. In Experiment 1, endometrium from day 14 pregnant and cyclic ewes was perifused sequentially on both the lumenal and myometrial sides with Krebs Ringers Bicorbonate solution (KRB), KRB plus oxytocin (1 IU/ml) and KRB alone. Endometrium from pregnant ewes secreted more PGF from both lumenal and myometrial sides than endometrium from cyclic ewes (P less than 0.05). Oxytocin stimulated secretion of PGF from both sides of endometrium regardless of status. Secretion of PGF was greater from the lumenal surface of endometrium compared to myometrium (P less than 0.05) for pregnant and cyclic ewes. For Experiment 2, endometrium was collected from day 15 cyclic ewes and perifused sequentially with KRB, KRB plus 300 ng/ml of either Bovine Serum Albumin (BSA) or oTP-1, KRB with or without BSA or oTP-1 plus oxytocin (1 IU/ml) and then KRB alone. Oxytocin stimulated greater release of PGF from oTP-1-treated than BSA-treated endometrium. Pretreatment of endometrium with oTP-1 had the same effect on oxytocin-induced PGF secretion as cotreatment with oTP-1 and oxytocin. In Experiment 3, uterine horns of cyclic ewes were catheterized on day 10 of the estrous cycle, and infused with either oTP-1 or day 16 pregnant sheep serum proteins on days 12, 13 and 14. Endometrium was collected on day 15 and perifused sequentially with KRB, KRB plus oxytocin (1 IU/ml) and then KRB alone. Treatment of ewes with oTP-1 attenuated endometrial secretion of PGF in response to oxytocin. Results of this study indicate that: (1) pregnancy stimulates basal secretion of PGF from endometrium and has no effect on oxytocin-induced secretion of PGF in vitro; (2) short-term oTP-1 treatment enhances oxytocin-induced PGF secretion from day 15 cyclic endometrium and (3) long-term oTP-1 treatment in vivo inhibits oxytocin-induced PGF secretion in ewes.     

Oxytocin (OT) action in uterine tissues of cyclic and early pregnant gilts: OT receptors concentration, prostaglandin F(2)alpha secretion, and phosphoinositide hydrolysis

Oxytocin (OT) is involved in the regulation of luteolysis in pigs. However, it is still not clear if OT is responsible for initiation of luteal regression in this species. The objectives of the study were: (1) to compare OT receptors (OTr) concentrations in endometrium and myometrium of cyclic and early pregnant pigs, (2) to examine the effect of OT on plasma PGF(2)alpha secretion during the progressive luteal regression, (3) to ascertain the effect of OT on inositol phosphates (IPs) accumulation in endometrial and myometrial cells of cyclic and early pregnant pigs. Concentrations of OTr on the endometrium and myometrium of cyclic (n = 33) (days 2-4; 11-13; 14-16; 18-20; day 21) and early pregnant (n = 4) (days 14-16) gilts were determined and they ranged from 7 +/- 3 (days 11-13) to 377 +/- 113 fmol/mg protein (day 21) in the endometrium and from 33 +/- 11 (days 2-4) to 167 +/- 28 fmol/mg protein (days 18-20) in the myometrium. In both tissues, concentrations of OTr were low during the luteal phase and increased (P < 0.01) during the follicular phase. In contrast to myometrial OTr, endometrial OTr during pregnancy were undetectable. In next experiment, mature gilts (n = 12) were injected with OT (20IU; i.v.) for three consecutive days starting on days 14 and 15 of the oestrous cycle and plasma PGF(2)alpha metabolite-13,14-dihydro-16-keto PGF(2)alpha (PGFM) concentration was determined. On days 15-16 and 16-17, OT increased plasma PGFM level. This effect was not observed on days 14-15 of the estrous cycle. A negative correlation was noticed between plasma concentrations of PGFM and progesterone (r = -0.3; P < 0.05). In last experiment, OT (100 nM) augmented (P < 0.01) an accumulation of inositol phosphates (IPs) in isolated myometrial cells on days 14-16 (n = 4) and 18-20 (n = 3) of the estrous cycle and on days 14-16 (n = 4) of pregnancy. Oxytocin-stimulated accumulation of IPs was not observed in endometrial cells. In summary: (1) concentrations of OTr on both the endometrium and myometrium were the highest during perioestrus-period in pigs, (2) myometrium of early pregnant sows possessed functional OTr, (3) oxytocin increased plasma PGFM concentration after initiation of luteolysis; and (4) OT-stimulated accumulation of IPs in myometrial, but not in endometrial cells. In conclusion, OT appears to not be involved in the initiation of luteal regression in sows and functional OTr are still present in the myometrium during early pregnancy (days 14-16).     

Effect of porcine conceptus secretory proteins on in vitro secretion of prostaglandins-F2 alpha and -E2 from luminal and myometrial surfaces of endometrium from cyclic and pseudopregnant gilts

Uterine endometrium collected from pseudopregnant (PP) and cyclic gilts on day (D) 15 after estrus were perifused in vitro with 10 ug/ml of porcine conceptus secretory proteins (pCSP) or serum proteins (SP) in Krebs ringer bicarbonate (KRB) buffer. In Experiment 1, samples were collected from luminal and myometrial surfaces of endometrium and concentrations of prostaglandin F2 alpha (PGF) determined by radioimmunoassay (RIA). Secretion of PGF by endometrium from cyclic gilts was stimulated (P less than .05) by pCSP. In Experiment 2, endometrium from D 14 cyclic and PP gilts was perifused and concentrations of PGF and prostaglandin E2 (PGE) in perfusate were determined by RIA. Across both statuses, luminal surface secretion of PGF was stimulated (P less than .05) by pCSP. Treatment with pCSP decreased secretion of PGE from myometrial surface of endometrium from cyclic gilts and increased (P less than .01) secretion of PGE from the myometrial surface of endometrium from PP gilts. In Experiment 3, pCSP were separated into acidic and basic fractions by anion exchange chromatography and each fraction was perifused separately over the luminal surface of endometrium from cyclic and PP gilts. Perifusion with acidic pCSP suppressed secretion of PGF by endometrium from cyclic or PP gilts; while basic pCSP did not influence secretion of PGF. These results demonstrated that products secreted by Day 15 pig conceptuses stimulate release of PGF and PGE from porcine uterine endometrium.     

Effects of pregnancy, oxytocin, ovine trophoblast protein-1 and their interactions on endometrial production of prostaglandin F2α in vitro in perifusion chambers

Pregnancy and intrauterine infusion of ovine trophoblast protein one (oTP-1) decrease oxytocin-induced secretion of prostaglandin F2α (PGF) from the uterus. In the present study, effects of oTP-1 and pregnancy on endometrial secretion of PHF were examined in an in vitro perifusion system. In Experiment 1, endometrium from day 14 pregnant and cyclic ewes was perifused sequentially on both the lumenal and myometrial sides with Krebs Ringers Bicorbonate solution (KRB), KRB plus oxytocin (1 IU/ml) and KRB alone. Endormetrium pregnant ewes secreted more PGF fro both lumenal and myotrial sides than endometrium from cyclic ewes (P<0.05). Oxytocin stimulated secretion of PGF was greater from the lumenal surface of endometrium compared to myometrium was collected from day 15 cyclic ewes and perifused sequentially with KRB, KRB plus 300 ng/ml of either Bovine Serum Albumin (BSA) or oTP-1, KRB with or without BSA or oTP-1 plus oxytocin (1 IU/ml) and then KRB alon. Oxytocin stimulated greater release of PGF from oTP-1-treated than BSA-treated endometrium. Pretreament of endometrium with oTP-1 has the same effect on oxytocin-induced PGF section was cotreatment with oTP-1 and oxytocin. In Experiment 3, uterine horns of cyclic ewes were catheterized on day 10 of the estrous cycle, and infused with either oTP-1 or day 16 pregnant sheep serum proteins on days 12, 13 and 14. Endometrium was collected on day 15 and perifused sequentially with KRB, KRB plus oxytocin (1 IU/ml) and then KRB alone. Treatment of ewes with oTP-1 attenuated endometrial secretion of PGF in response to oxytocin. Results of this study indicate that: (1) preganancy stimulates basal secretion of PGF from endometrium and has no effect on oxytocin-induced secretion of PGF in vitro; (2) short-term oTP-1 treatment enhances oxytocin-induced PGF secretion from day 15 cyclic endometrium and (3) long-term oTP-1 treatment in vivo inhibits oxytocin-induced PGF secretion in ewes.     

Antiluteolytic strategies to improve fertility in cattle.

During early pregnancy, a "critical period" may be defined between Days 15 and 17. Embryonic mortality associated with this period causes significant economic losses to the cattle industry. During this period, the endometrium will follow a default program to release luteolytic pulses of PGF2alpha, unless the conceptus sends appropriate antiluteolytic signals to block PGF2alpha, production. Maintenance of pregnancy is dependent on a successful blockage of endometrial PGF2alpha production. Biology of the critical period is complex and multifactorial. Endocrine, cellular and molecular factors, both from maternal and conceptus origins act in concert to determine whether luteolysis or maintenance of pregnancy will prevail. Understanding the influences of such factors in the biology of the critical period allowed researchers to produce a series of strategies aiming to favor maintenance of pregnancy in lieu of luteolysis. Strategies include hormonal and nutritional manipulations to decrease plasma concentrations of estradiol 17beta (E2) while increasing those of progesterone (P4), and inhibiting the PGF2alpha-synthesizing enzymatic machinery in the endometrium during the critical period. Experimental results indicate that use of such strategies has improved pregnancy rates following artificial insemination and embryo transfer programs.     

The effect of progesterone and human interferon alpha-2 on the release of PGF2 alpha and PGE from epithelial cells of human proliferative endometrium.

Progesterone and interferon-like trophoblastic proteins modulate prostaglandin (PG) synthesis from endometrium in early ovine and bovine pregnancy. Enriched epithelial cells were prepared from human endometrium removed in the proliferative phase of menstrual cycle (n = 8). Progesterone at a concentration of 1 microM suppressed PGE release from the cells during the first 24 hours in culture. After 48 hours in culture progesterone at a dose of 100 nM and 1 microM suppressed both the release of PGF2 alpha and PGE from the cells and this suppression was maintained for a further two days. Addition of exogenous 30 microM arachidonic acid (AA) abolished this effect of progesterone on both PGF2 alpha and PGE release. Interferon alpha-2 did not suppress the basal release of PGF2 alpha nor PGE. In the presence of progesterone, interferon alpha-2 attenuated the progesterone mediated suppression of PGF2 alpha but not PGE release from endometrial cells. These findings suggest that progesterone suppresses the basal release of PGs from human endometrium, but unlike the sheep, interferon alpha-2 does not exert this action on human endometrium.     

The effect of progesterone on oxytocin-stimulated intracellular mobilization of Ca2+ and prostaglandin E2 and F2alpha secretion from porcine myometrial cells

Our past studies have shown that porcine myometrium produce prostaglandins (PG) during luteolysis and early pregnancy and that oxytocin (OT) and its receptor (OTr) support myometrial secretion of prostaglandins E2 and F2alpha (PGE2 and PGF2alpha) during luteolysis. This study investigates the role of intracellular Ca2+ [Ca2+]i as a mediator of OT effects on PG secretion from isolated myometrial cells in the presence or absence of progesterone (P4). Basal [Ca2+]i was similar in myometrial cells from cyclic and pregnant pigs (days 14-16). OT (10(-7)M) increased [Ca2+]i in myometrial cells of cyclic and pregnant pigs, although this effect was delayed in myometrium from pregnant females. After pre-incubation of the myocytes with P4 (10(-5)M) the influence of OT on [Ca2+]i)was delayed during luteolysis and inhibited during pregnancy. Myometrial cells in culture produce more PGE2 than PGF2alpha regardless of reproductive state of the female. OT (10(-7)M) increased PGE2 secretion after 6 and 12 h incubation for the tissue harvested during luteolysis and after 12 h incubation when myometrium from gravid females was used. In the presence of P4 (10(-5)M), the stimulatory effect of OT on PG secretion was diminished. In conclusion: (1) porcine myometrial cells in culture secrete PG preferentially during early pregnancy and produce more PGE2 than PGF2alpha, (2) OT controls myometrial PGF2alpha secretion during luteolysis, (3) release of [Ca2+]i is associated with the influence of OT on PG secretion, and (4) the effects of OT on PG secretion and Ca2+ accumulation are delayed by P4 during luteolysis and completely inhibited by P4 during pregnancy.     

Oxytocin-stimulated production of prostaglandin F2alpha by isolated endometrium of rabbit: modulation by ovarian steroids

To test the hypothesis that ovarian steroid hormones modulate oxytocin-induced release of prostaglandin F2alpha (PGF2alpha) from uterine endometrium, 2 ovariectomized rabbits were pretreated with progesterone (5 mg/day for 10 days), 2 with estradiol-17 beta (25 microgram/day for 10 days), 2 with both steroids, and one with sesame oil only. On the last day of treatment, endometrial fragments were excised and incubated in vitro with or without oxytocin (100 muU/ml). Although endometrium from rabbits pretreated with combined steroids released more PGF2alpha immediately after excision than did tissue from animals pretreated with either steroid by itself, endometrium from animals pretreated with estradiol-17 beta alone released the most PGF2alpha during sustained incubation in vitro. Moreover, only this tissue exhibited significant oxytocin-dependent release of PGF2alpha. At the dosages used, progesterone completely antagonized both of these effects of estradiol-17 beta. The results support the hypothesis that ovarian steroid hormones regulate oxytocin-dependent release of PGF2alpha from endometrial cells. A posible mechanism of action is suggested.     

Prostaglandin secretion by perifused bovine endometrium: secretion towards the myometrial and luminal sides at day 17 post-estrus as altered by pregnancy

Bilateral perifusion devices were utilized to measure prostaglandin secretion towards luminal and myometrial sides of bovine endometria. Tissues were collected at Day 17 post-estrus from cyclic (n = 4), pregnant (n = 5) and bred but subsequently non-pregnant (n = 6) cows. Tissue from each cow was placed into two perifusion devices, perifused with Krebs-Ringer Bicarbonate solution (3 ml/10 min) for 2.5 h and fractions collected every 10 min. Oxytocin (1 IU/ml) was perifused during fractions 7-12 to the luminal side of one device and to the myometrial side of the other device. Regardless of status, prostaglandin secretion rates (PGF and PGE2) were higher (P less than 0.01) from the luminal side than the myometrial side. Secretion rates of PGF were lower (P less than 0.01) for endometria from pregnant cows than for endometria from cyclic or bred/non-pregnant cows, whereas secretion rates of PGE2 were not affected by pregnancy status. Regardless of the side of perifusion, secretion rates of PGF and PGE2 from endometria of cyclic and bred/non-pregnant cows were elevated (P less than 0.01) throughout the period of oxytocin treatment, whereas prostaglandin secretion by endometria from pregnant cows was not stimulated by oxytocin. Decreased secretion of PGF from endometria of pregnant cows suggests that the corpus luteum and pregnancy are maintained because of an inhibition of endometrial prostaglandin synthesis or an inability to respond to stimulators of prostaglandin synthesis (i.e. oxytocin).     

Expression of enzymes of cyclooxygenase pathway and secretion of prostaglandin E2 and F2alpha by porcine myometrium during luteolysis and early pregnancy

Past studies of uterine prostaglandin (PGs) and pig reproduction have focused on endometrial rather than myometrial PGs. This study documents the synthesis and secretion of myometrial prostaglandins (PGs) in pigs and the involvement of oxytocin (OT) in these processes. Cyclooxygenase-2 (COX-2) expression was similar in myometrial explants from cyclic and pregnant pigs (days 14-16) and OT (10(-7) M) in vitro significantly increased COX-2 protein regardless of reproductive state. Basal expression of prostaglandin E2 synthase (PGES) was higher during pregnancy than during luteolysis. Conversely, prostaglandin F synthase (PGFS) was highest during luteolysis and lower in myometrium from gravid animals. OT had no influence on the expression of PGES and PGFS. In another tissue culture experiment, myometrial slices produced more PGE2 than PGF2alpha regardless of reproductive state of the female. OT stimulated PGE2 production in myometrium harvested during luteolysis and increased PGF2alpha production in all tissues examined. Progesterone (P4; 10(-5) M) blocked stimulatory effect of OT on myometrial PG release. Myometrial OTr mRNA was higher (P=0.03) during luteolysis than during pregnancy. In conclusion: (1) oxytocin increases myometrial COX-2 expression, but does not influence the expression of terminal enzymes of PGs synthesis (PGES and PGFS); (2) porcine myometrium preferentially produces PGs during early pregnancy and secretes more PGE2 than PGF2alpha; (3) myometrial OT and OTr support secretion of PGs from myometrium during luteolysis.     

Endocrine secretion of prostaglandin F2alpha in cyclic gilts is decreased by intrauterine administration of exogenous oxytocin

When administered systemically, oxytocin (OT) stimulates secretion of uterine prostaglandin F2alpha (PGF2alpha) in swine, but the role of endometrially-derived OT in control of PGF2alpha release is not clear. This study determined the effect of exogenous OT, administered into the uterine lumen of intact cyclic gilts, on PGF2alpha secretion during late diestrus. Intrauterine infusion of 40USP units OT (in 30 ml 0.9% saline) was performed for 30 min (1 ml/min) into each uterine horn between 7:00 and 9:00 h on days 10, 12, 14 and 16 after estrus. Beginning 20 min before infusion, samples of jugular venous blood were drawn at 5-10-min intervals for 140 min for quantification of 13,14-dihydro-15-keto-PGF2alpha (PGFM), the major stable metabolite of PGF2alpha. Progesterone was analyzed in samples collected 0, 60 and 120 min after initiation of OT infusion. Treatment with OT did not alter plasma concentrations of PGFM on days 10 or 12 but decreased (P<0.001) PGFM concentrations for 40 min after onset of infusion on day 16. Concentrations of PGFM also were reduced in the pre-treatment samples on day 14 (P=0.05) and day 16 (P<0.001) in OT-infused gilts. Plasma progesterone declined (P<0.01) between days 10 and 16 in control-infused gilts but did not decline until after day 14 (P<0.001) in gilts infused with OT. These results indicate that when OT is administered into the uterine lumen of pigs during late diestrus, it has an anti-luteolytic effect to reduce endocrine secretion of PGF2alpha and delay the decline in progesterone that occurs during luteolysis.     

Prostaglandin secretion by perifused bovine endometrium: Secretion towards the myometrial and luminal sides at day 17 post-estrus as altered by pregnancy

Bilateral perifusion devices were utilized to measure prostaglandin secretion towards luminal and myometrial sides of bovine endometria. Tissues were collected at Day 17 post-estrus from cyclic (n=4), pregnant (n=5) and bred but subsequently non-pregnant (n=6) cows. Tissue from each cow was placed into two perifusion devices, perifused with Krebs-Ringer Bicarbonate solution (3 ml/10 min) for 2.5h and fractions collected every 10 min. Oxytocin (1 IU/ml) was perifused during fractions 7–12 to the luminal side of one device and to the myometrial side of the other device. Regardless of stratus, prostaglandin secretion rates (PGF and PGE₂) were higher (P< 0.01) from the luminal side than the myometrial side. Secretion rates of PGF were lower (P< 0.01) for endometria from pregnant cows than for endometria from cyclic or bred/non-pregnant cows, whereas secretion rates of PGE₂ were not affected by pregnancy status. Regardless of the side of perifusion, secretion rates of PGF and PGE₂ from endometria of cyclic and bred/non-pregnant cows were elevated (P< 0.01) throughout the period of oxytocin treatment, whereas prostaglandin secretion by endometria from pregnant cows wasnot stimulated by oxytocin. Decreased secretion of PGF from endometria of pregnant cows suggests that the corpus luteum and pregnancy are maintained because of an inhibition of endometrial prostaglandin synthesis or an inability to responsd to stimulators of prostaglandin synthesis (i.e. oxytocin).     

Effects of superovulation, arachidonic acid or endometrium on release of prostaglandins and synthesis of proteins by bovine trophoblast

This study was conducted in vitro to examine factors that may regulate prostaglandin release by bovine trophoblast and endometrial slices. Trophoblastic tissues and endometrial slices were recovered from superovulating and normally-ovulating cattle on day 16 or 20 of pregnancy and incubated for 24 h. Release of PGF2 alpha and 13,14-dihydro-15-keto-PGF2 alpha (PGFM), and incorporation of [14C]-leucine into proteins were quantified and expressed per microgram DNA, which gives a measure of cellular activity. Activity of trophoblastic tissue for synthesizing protein was decreased (P less than .05) and for releasing PGFM was increased (P less than .05) on day 20 compared to day 16 of pregnancy. Neither superovulation nor day of pregnancy altered trophoblastic activity for releasing PGF2 alpha. Superovulation increased (P less than .05) endometrial release of PGF2 alpha. Endometrial release of PGF2 alpha was less (P less than .05) on day 20 than on day 16 of pregnancy. When arachidonic acid (0, 100, 200 or 400 micrograms) was added at the start of incubation, trophoblastic release of PGF2 alpha changed (P less than .05) quadratically with dose of arachidonic acid. When arachidonic acid was added 8 h after the start of incubation, trophoblastic release of PGF2 alpha increased linearly (P less than .01) with dose of arachidonic acid. Adding arachidonic acid to incubation medium did not affect trophoblastic or endometrial protein synthesis. Endometrial slices suppressed (P less than .05) trophoblastic protein synthesis and release of PGF2 alpha. Apparently, endometrium can modulate trophoblastic release of prostaglandins and synthesis of proteins in vitro, and trophoblastic tissue from superovulated cattle 16 or 20 days pregnant can be used to study trophoblastic synthesis of prostaglandins and proteins.     

Effects of tumor necrosis factor-alpha on secretion of prostaglandins E2 and F2alpha in bovine endometrium throughout the estrous cycle

To determine the physiological significance of tumor necrosis factor-alpha (TNFalpha) in the regulation of endometrial prostaglandin (PG) release in cattle, we investigated the effects of TNFalpha on the secretion of PGE2 and PGF2alpha by bovine endometrium during the estrous cycle. Bovine uteri were classified into six stages (estrus: Day 0, early luteal 1: Days 2 to 3, early luteal 11: Days 5 to 6, mid-luteal: Days 8 to 12, late luteal: Days 15 to 17 and follicular: Days 19 to 21). After 1 h of pre-incubation, endometrial tissues (20 to 30 mg) were exposed to 0 or 0.6 nM TNFalpha for 4 h. The PGE2 concentrations in the medium were higher in the luteal stages than in the follicular stage and in estrus. In contrast, PGF2alpha concentrations were higher in the follicular stage and in estrus than in the luteal stages. The ratio of the basal concentrations of PGE2 and PGF2alpha (PGE2/PGF2alpha ratio) was higher in the luteal stages than in the follicular stage and in estrus. Although TNFalpha stimulated both PGE2 and PGF2alpha secretion during the entire period of the estrous cycle, the level of stimulation of TNFalpha on PGE2 output by the bovine endometrium does not show the same cyclical changes as that shown on PGF2alpha output. The stimulation of TNFalpha resulted in a decrease in the PGE2/PGF2alpha ratio only in the late luteal stage. Furthermore, TNFalpha stimulated PGE2 secretion in stromal, but not epithelial cells. The overall results suggest that TNFalpha is a potent regulator of endometrial PGE2 secretion as well as PGF2alpha secretion during the entire period of estrous cycle, and that TNFalpha plays different roles in the regulation of secretory function of bovine endometrium at different phases of the estrous cycle.     

Prostaglandin biosynthesis and its regulation in the bovine endometrium: A comparison between nonpregnant and pregnant status

Prostaglandin F(2alpha) (PGF(2alpha)), an arachidonic acid metabolism product of the prostaglandin synthetase pathway, is synthesized and released from the endometrium during luteolysis in nonpregnant animals. When proper conception occurs, the synthesis and release pattern is changed to maintain the corpus luteum (CL) function. The biosynthesis of prostaglandins in the bovine endometrium was highest in the microsomes but of low order. In nonpregnancy, the formation of prostaglandins from labelled precursor acid was higher than in pregnancy. Besides the prostaglandin synthetase, an inhibiting activity on the conversion of arachidonic acid to prostaglandins was found in both the nonpregnant and pregnant endometrium. During luteolysis (Day 17), a low inhibiting capacity was seen in comparison with other days of the estrous cycle (Days 1, 4 and 14). The inhibitory capacity was very high on Days 16 to 20, 25, and 31 of pregnancy. In the nonpregnant endometrium at Day 17, a very low inhibitor potency, calculated as IC(50) values, was found both in the cytoplasma and in the microsomes, whereas during early pregnancy (Days 17, 18, and 20) both cytoplasma and microsomes possessed very high inhibitor potency. This finding indicates that the bovine endometrium contains both prostaglandin synthetase and an unknown potent inhibitor of prostaglandin biosynthesis that regulates prostaglandin biosynthesis both during the estrous cycle and early pregnancy.     

Uterine blood supply as a main factor involved in the regulation of the estrous cycle--a new theory

The paper presents a new theory on the physiological mechanism of initiation of luteolysis, function of endometrial cells and protection of corpus luteum. This theory is based on previous studies published by the authors and their coworkers on the retrograde transfer of PGF2alpha in the uterine broad ligament vasculature during the estrous cycle, early pregnancy and pseudopregnancy. The studies were focused on cyclic changes in uterine blood supply and the apoptosis of endometrial cells. Moreover, the results of many other authors are cited. The statements of the theory are as follows: 1. The initiation of luteolysis is a consequence of regressive changes in the endometrium which are due to the reduction of the uterine blood supply below the level necessary to provide for the extended needs of active endometrium. 2. During the luteal phase, both a considerable increase in uterine weight and a decrease in blood flow through the uterine artery, resulting from increasing progesterone concentration, reduce the uterine blood supply. In comparison to the volume of blood flowing to the porcine uterus during the estrus period, only 30-40% of the blood volume is determined on day 12 of the estrous cycle. The uterine weight at that time is 40-60% larger than that in the early luteal phase. Thus, due to the considerable constriction of uterine blood vessels, there is a discrepancy between the requirement for oxygen and other factors transported by blood and the possibility of supplying the uterus with these substances. After reaching the threshold of uterine blood supply level, which in pigs takes place around day 12 of the estrous cycle, regressive changes and PGF2alpha release from endometrial cells occurs. 3. Estrogens and progesterone are the major factors affecting blood flow in vessels supplying the uterus. The factors that modulate, complement and support vasodilation and vasoconstriction are: PGE2, LH, oxytocin, cytokines, neurotransmitters and other local blood flow regulators. In some animal species these modulators, especially those of embryonic origin, may be crucial for the status of uterine vasculature. 4. During early pregnancy, the action of embryo signals (estrogens, cytokines), endometrial PGE2 as well as LH results in the relaxation of the uterine artery (pigs: day 12) and, consequently, in an increase in uterine blood supply. This reaction of the maternal recognition of pregnancy effectively prevents regressive changes in well developed endometrial cells to occur. 5. Local uptake and retrograde transfer of PGF2alpha into the uterine lumen during early pregnancy protects corpus luteum from PGF2alpha luteolytic action. 6. During the period of regressive changes resulting from the limited uterine blood supply, endometrial cells restrain PGF2alpha synthesis. They are, however, still capable of releasing prostaglandin when uterine blood supply is improved after the embryo appears in the uterus. This potential capability for PGF2alpha synthesis was demonstrated in in vitro studies when endometrial cells collected during its regressive phase were incubated in medium and stimulated by LH and oxytocin. 7. Prostaglandin F2alpha pulses in venous blood flowing from the uterus do not confirm pulsatile secretion of PGF2alpha. The pulses may result from the pulsatile excretion of PGF2alpha with venous blood according to the rhythmic uterine contractions associated with oxytocin secretion. 8. The results supporting this concept are presented and discussed in due course. The critique of Bazer and Thatcher's theory on exocrine versus endocrine secretion of prostaglandin F2alpha during the estrous cycle is also depicted.     

Oxytocin stimulates prostaglandin F2alpha secretion and prostaglandin F synthase protein expression in porcine myometrial tissue

The possibility of PGF(2)alpha production and presence of prostaglandin F synthase (PGFS; PGD(2) 11-ketoreductase) was studied in control and oxytocin (OT)-stimulated myometrial slices isolated from cyclic (Days 14-16) and early pregnant (Days 14-16) sows. Oxytocin (10(-7) M) stimulated (p<0.01) PGF(2)alpha production in both cycling and early pregnant myometrial slices. Prostaglandin F(2)alpha release was higher (p<0.01) in control as well as OT-treated myometrium of early pregnant sows in comparison to cycling myometrium. Prostaglandin F synthase expression at protein level was evident in myometrial slices of cyclic as well as early pregnant sows. The signals of PGFS was stronger (p<0.05) in cycling myometrium exposed to OT compared to that of control. There were no significant differences (p>0.05) in PGFS protein expression between control and OT-stimulated myometrial tissue of early-pregnant sows. The results of this study indicate the local PGF(2)alpha synthesis and the presence of PGFS in porcine cycling and early pregnant myometrial tissue. In addition, OT increased PGD(2) 11-ketoreductase protein expression in myometrium harvested during the porcine estrous cycle. However, the OT-stimulated PGF(2)alpha myometrial secretion was observed in both, cycling and pregnant gilts.     

Influence of a prostaglandin synthesis inhibitor administered at embryo transfer on pregnancy rates of recipient cows

Elevated uterine luminal concentrations of prostaglandin F(2alpha) (PGF(2alpha)) have been negatively associated with embryo quality and pregnancy rates. Two studies were performed in cows to determine PGF(2alpha) release from uterine endometrium following embryo transfer and to investigate administration of flunixin meglumine (FM), a prostaglandin synthesis inhibitor, on pregnancy rates following embryo transfer. In Experiment 1, blood samples were collected prior to and after embryo transfer from the posterior vena cava via saphenous vein cannulation. Serum profiles of PGF(2alpha) indicated that manipulation of the reproductive tract during embryo transfer was followed by increased release of PGF(2alpha) from the uterine endometrium. In Experiment 2, estrus (day=0) was synchronized in recipient animals and a single embryo transferred 7 days after estrus. At the time of non-surgical embryo transfer, animals were randomly assigned to receive either FM (FM; n=1300) or remain untreated (control (CON); n=797). Data collected at transfer included stage of embryo development, embryo quality, technician, and transfer quality score. Overall pregnancy rates of cows receiving FM (65%) were higher than control cows (60%; P<0.02). Pregnancy rates following transfer of quality 1 (good) embryos did not differ (P>0.05) between treatments. However, pregnancy rates of quality 2 (fair) embryos were higher in animals receiving FM than in CON (P<0.01). Moreover, pregnancy rates of transferred morula- and blastocyst-stage embryos were higher in FM-treated than in controls (P<0.06 and P<0.04, respectively). In conclusion, uterine release of PGF(2alpha) is elevated following embryo transfer and administration of a PGF(2alpha) synthesis inhibitor at the time of embryo transfer improved pregnancy rates in cows.