PGF2α Concentrations Research Articles

Effect of α-linolenic acid on oocyte maturation and embryo development of prepubertal sheep oocytes

The purpose of this study was to evaluate the effect of omega-3 α-linolenic acid (ALA) added to the IVM medium on embryo development of prepubertal sheep oocytes. Experiment 1 investigated the effect of ALA at different concentrations (0 [control], 50, 100, and 200 μM) and DMSO (100 μM) in IVM media on cumulus cell expansion and oocyte nuclear maturation and on synthesis of prostaglandins (PGE2 and PGF2α). Experiment 2 investigated the effects of ALA at different concentrations in the IVM medium on oocyte fertilization, cleavage, and developmental potential to blastocyst stage and changes in estradiol and progesterone concentrations in the spent IVM media. IVM oocytes were fertilized with frozen-thawed spermatozoa capacitated in a serum-free sperm medium. Presumptive zygotes were cultured 8 days in synthetic oviductal fluid (SOF) medium without serum. Blastocyst quality was assessed by counting total cell number and the number of apoptotic cells using Hoechst and Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. Nuclear maturation of oocytes and the number of fully expanded cumulus cells were reduced after treatment with 200 μM of ALA compared with other groups (P ≤ 0.05). Supplementation with ALA increased both PGE2 and PGF2α concentrations in the spent media (P ≤ 0.05). No differences were observed in blastocyst development among control (12.2%) and 50, 100, and 200 μM ALA groups (6.9%, 11.5% and 14.0%, respectively). However, the total cell number (46.50 ± 5.85, 67.94 ± 6.71, 45.20 ± 6.37, and 59.80 ± 5.51, respectively; P ≤ 0.05) and apoptotic cell number (6.45 ± 0.89, 2.48 ± 0.81, 4.02 ± 1.15, and 3.67 ± 1.15, respectively; P ≤ 0.05) were significantly improved. After IVM, estradiol concentration was lower and progesterone concentration was higher in ALA groups compared with the control group (P ≤ 0.05). In conclusion, these results revealed that ALA affects prepubertal sheep embryo quality associated with alteration of releasing reproductive hormones.

Prostaglandin E1 or E2 inhibits an oxytocin-induced premature luteolysis in ewes when oxytocin is given early in the estrous cycle

The objective of this study was to determine whether PGE1 or PGE2 prevents a premature luteolysis when oxytocin is given on Days 1 to 6 of the ovine estrous cycle. Oxytocin given into the jugular vein every 8 hours on Days 1 to 6 postestrus in ewes decreased (P ≤ 0.05) luteal weights on Day 8 postestrus. Plasma progesterone differed (P ≤ 0.05) among the treatment groups; toward the end of the experimental period, concentrations of circulating progesterone in the oxytocin-only treatment group decreased (P ≤ 0.05) when compared with the other treatment groups. Plasma progesterone concentrations in ewes receiving PGE1 or PGE1 + oxytocin were greater (P ≤ 0.05) than in vehicle controls or in ewes receiving PGE2 or PGE2 + oxytocin and was greater (P ≤ 0.05) in all treatment groups receiving PGE1 or PGE2 than in ewes treated only with oxytocin. Chronic intrauterine treatment with PGE1 or PGE2 also prevented (P ≤ 0.05) oxytocin decreases in luteal unoccupied and occupied LH receptors on Day 8 postestrus. Oxytocin given alone on Days 1 to 6 postestrus in ewes advanced (P ≤ 0.05) increases in PGF2α in inferior vena cava or uterine venous blood. PGE1 or PGE2 given alone did not affect (P ≥ 0.05) concentrations of PGF2α in inferior vena cava and uterine venous blood when compared with vehicle controls or oxytocin-induced PGF2α increases (P ≤ 0.05) in inferior vena cava or uterine venous blood. We concluded that PGE1 or PGE2 prevented oxytocin-induced premature luteolysis by preventing a loss of luteal unoccupied and occupied LH receptors.

Calcium potentiates the effect of estradiol on PGF2α production in the bovine endometrium.

BackgroundEstradiol (E2) is required for luteolysis in cows and its injection stimulates prostaglandin F2α (PGF2α) release. The main goal of our study was to investigate the ability of endometrial explants and cells treated with E2 and the calcium ionophore (CI) A23187 to synthesize PGF2α.ResultsTreatment with E2in vivo resulted in a 48.4% increase of PGF2α production by endometrial explants treated in vitro with A23187. Production of PGF2α was better stimulated with A23187 at concentrations of 10-6 and 10-5 mol/L compared with other concentrations used. The concentration of PGF2α for untreated bovine endometrial cell cultures was 33.1 pg/mL, while for cultures treated with E2, A23187, or a combination of E2 and A23187, the PGF2α concentration was 32.5, 92.4 and 145.6 pg/mL, respectively.ConclusionsTreatment with A23187 tended to stimulate PGF2α production. In the presence of E2, A23187 significantly stimulated PGF2α synthesis. It appears that A23187 potentiates the effects of E2 with respect to synthesis of endometrial PGF2α in cattle.

Ovarian steroid-dependent tumor necrosis factor-α production and its action on the equine endometrium in vitro

Tumor necrosis factor-α (TNF) is a cytokine that plays important roles in functions of the endometrium. The aims of this study were to determine whether (i) ovarian steroids modulate TNF production by endometrial cells (Experiment 1); (ii) TNF effects on prostaglandin (PG) production in cultured equine endometrial cells and tissue (Experiment 2). Epithelial and stromal cells were isolated from equine endometrium (Days 2–5 of the estrous cycle; n=20) and treated after passage 1. In Experiment 1, epithelial and stromal cells were exposed to progesterone (P4; 10−7M), 17-β estradiol (E2; 10−9M) or P4+E2 (10−7/10−9M) for 24h. Then, TNF mRNA transcription was determined using Real-time PCR. Additionally, TNF protein production was investigated in response to ovarian steroids for 24h using Enzyme-Linked Immunosorbent Spot (EliSpot). In Experiment 2, epithelial and stromal cells and endometrial explants (mid-luteal phase of the estrous cycle; n=5) were exposed in vitro to TNF (10ng/ml) and to oxytocin (OT; positive control; 10−7M) for 24h. The concentrations of PGE2 and PGF2α were determined using a direct enzyme immunoassay (EIA) method. The transcription of prostaglandin-endoperoxide synthase-2 (PTGS-2), prostaglandin E2 synthase (PGES) and PGF2α synthase (PGFS) mRNAs in the endometrial explants was determined using Real-time PCR. Results showed that TNF is produced by two types of equine endometrial cells and its production is up-regulated by ovarian steroids (P<0.05) in stromal cells and by P4 (P<0.05) and E2 (P<0.01) in epithelial cells. Epithelial and stromal cells can also produce PG in response to TNF. In endometrial explants, TNF stimulated PGE2 production to a large extent and PGF2α secretion to a lesser extent. These actions are mediated by up-regulation of PG synthases mRNA transcription. The study indicates that TNF production is closely related to ovarian steroid actions and that the interaction between TNF and PG regulates physiologic processes in the equine endometrium.

Response of pro-inflammatory prostaglandin contents in anti-inflammatory supplements from green mussel Perna viridis L. in a time-dependent accelerated shelf-life study

The effect of natural antioxidant additives on pro-inflammatory prostaglandins, PGE2 and PGF2αvis-à-vis C20n-6 fatty acid in the green mussel, Perna viridis extract (MSL) and those of MSL supplemented with additives (in GMET1 through GMET4) were investigated in an accelerated shelf-life study (50°C) of 90days. HPLC was employed for quantifying PGE2 and PGF2α and their chemical structures were assigned by signature peaks based on ESI-LC–MS and 1H NMR. The concentrations of PGE2 and PGF2α after an accelerated shelf-life study of 90days were significantly higher (P<0.05) in MSL on day 90 of the intervention than among GMET2 and GMET4. An inverse correlation between antioxidative effects of the additives and pro-inflammatory eicosanoids in GMET2 and GMET4 at baseline and at the end of the study was apparent. The results describe the utility of quantifying PGE2 and PGF2α as an index of antioxidant status and lipid peroxidation.

The modulation of prostaglandin production by coumestrol in the equine endometrium in vitro

s / Journal of Equine Veterinary Science 34 (2014) 133 133 determined using Real-time PCR method. The concentrations of PGE2 and PGF2a in culture medium were determined using a direct enzyme immunoassay (EIA) method. In Experiment 2, the influence of Cou on the proliferation of epithelial and stromal cells was determined using MTT (3[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) method. 3. Results and Discussion In epithelial cells, E2 as well as Cou, augmented PGE2 and PGF2a secretion compared to control group (P<0.05; Figure 1A,B). In contrast, in stromal cells, only Cou increased PGE2 secretion compared to control group (P<0.001). However, E2 as well as Cou stimulated PGF2a secretion compared to control group in stromal cells (P<0.001). Both 17-b estradiol and Cou up-regulated PTGS2 mRNAs transcription in stromal and epithelial cells compared with the control group (P<0.05; Figure 1C,D). In epithelial cells, E2 and Cou up-regulated PGES mRNA transcription compared to the control group (P<0.05). In epithelial and stromal cells, E2 as well as Cou, up-regulated PGFS mRNAs transcription compared to the control group (P<0.05). Besides, after the 24h incubation, Cou increased Fig. 1. Effects of 17b-estradiol (E2) and coumestrol (Cou) on prostaglandin E2 secretion (A,B), expression of PTGS-2 mRNA (C,D) and cell proliferation (E,F) in the cultured equine epithelial (A,C,E) and stromal (B,D,F) endometrial cells. epithelial and stromal cells proliferation (p<0.05), while E2 only increased stromal cells proliferation (P<0.05; Figure 1E,F). The present data provide one of the possible explanations how alfalfa derived phytoestrogens and its metabolites act as endocrine disruptors, leading to disruption of the reproductive processes and to temporal infertility in mares. Cou mimics E2 action in equine endometrium. However, in contrast to E2, only Cou was able to stimulate PGE2 secretion by stromal cells. Additionally, we showed that Cou is a stronger stimulator of PG production than E2 itself. Thus, since PG play a crucial role in estrous cycle and pregnancy regulation, secretory disorders occurring locally in the uterus may have a tremendous impact on reproductive functions in the mare. Coumestrol disrupts the ratio of PGE2 to PGF2a which might lead to the non-physiological PG actions in equine reproductive tract during the estrous cycle.

Specialty Supplement Use and Biologic Measures of Oxidative Stress and DNA Damage

Oxidative stress and resulting cellular damage have been suggested to play a role in the etiology of several chronic diseases, including cancer and cardiovascular disease. Identifying factors associated with reduced oxidative stress and resulting damage may guide future disease-prevention strategies. In the VITamins And Lifestyle (VITAL) biomarker study of 209 persons living in the Seattle area, we examined the association between current use of several specialty supplements and oxidative stress, DNA damage, and DNA repair capacity. Use of glucosamine, chondroitin, fish oil, methylsulfonylmethane (MSM), coenzyme Q10 (CoQ10), ginseng, ginkgo, and saw palmetto was ascertained by a supplement inventory/interview, whereas the use of fiber supplements was ascertained by questionnaire. Supplements used by more than 30 persons (glucosamine and chondroitin) were evaluated as the trend across number of pills/week (non-use, <14 pills/week, 14+ pills/week), whereas less commonly used supplements were evaluated as use/non-use. Oxidative stress was measured by urinary 8-isoprostane and PGF2α concentrations using enzyme immunoassays (EIA), whereas lymphocyte DNA damage and DNA repair capacity were measured using the Comet assay. Multivariate-adjusted linear regression was used to model the associations between supplement use and oxidative stress/DNA damage. Use of glucosamine (Ptrend: 0.01), chondroitin (Ptrend: 0.003), and fiber supplements (P: 0.01) was associated with reduced PGF2α concentrations, whereas CoQ10 supplementation was associated with reduced baseline DNA damage (P: 0.003). Use of certain specialty supplements may be associated with reduced oxidative stress and DNA damage. Further research is needed to evaluate the association between specialty supplement use and markers of oxidative stress and DNA damage.

The involvement of peroxisome proliferator activated receptors (PPARs) in prostaglandin F2α production by porcine endometrium

In the present study, we investigated the in vitro effects of peroxisome proliferator activated receptor (PPAR) ligands on PGF2α secretion and mRNA expression of prostaglandin F synthase (PGFS) in porcine endometrial explants collected on days 10–12 and 14–16 of the estrous cycle or pregnancy. The explants were incubated for 6h with: PPARα ligands – WY-14643 (agonist) and MK 886 (antagonist); PPARβ ligands – l-165,041 (agonist) and GW 9662 (antagonist); PPARγ ligands – 15d-prostaglandin J2 (PGJ2, agonist), rosiglitazone (agonist) and T0070907 (antagonist). The expression of PGFS mRNA in the endometrium and the concentration of PGF2α in culture media were determined by real time RT-PCR and radioimmunoassay, respectively. During the estrous cycle (days 10–12 and 14–16), the agonists – WY-14643 (PPARα), l-165,041 (PPARβ), PGJ2 and rosiglitazone (PPARγ) – increased PGF2α secretion but did not affect PGFS mRNA abundance. During pregnancy (days 10–12 and 14–16), PPARα and PPARγ ligands did not change PGF2α release, whereas PPARβ agonist augmented PGF2α release on days 14–16 of pregnancy. In addition, WY-14643 and l-165,041 increased PGFS mRNA level in both examined periods of pregnancy. PPARγ agonist (PGJ2) and antagonist (T0070907) enhanced PGFS mRNA abundance in the endometrium on days 10–12 and 14–16 of pregnancy, respectively. The results indicate that PPARs are involved in the production of PGF2α by porcine endometrium, and that the sensitivity of the endometrium to PPAR ligands depends on reproductive status of animals.

PGE2and PGF2α Concentrations in Human Endometrial Fluid as Biomarkers for Embryonic Implantation

Prostaglandin (PG) signaling has been implicated in embryonic implantation in several animal species including humans; however, this knowledge has not yet been clinically translated. The objective of this work is to investigate whether PGE2 and PGF2α in endometrial fluid (EF) can be used as biomarkers of human embryonic implantation. Lipidomic profile of human EF (n = 173) obtained through natural cycles, hormonal replacement therapy, controlled ovarian stimulation, and refractory endometrium induced by the insertion of an intrauterine device was analyzed by liquid chromatography and tandem mass spectrometry. Immunohistochemistry, Western blotting, immunolocalization of PG receptors on mouse embryos, embryo adhesion assay, pharmacological interventions, and statistical analysis were conducted. PGE2 and PGF2α concentrations increased significantly in the human EF during the window of implantation in natural cycles and assisted reproductive technologies patients undergoing in vitro fertilization and ovum donation. This profile was abrogated in the refractory endometrium. We also demonstrated that PGE2 and PGF2α synthases are located in the endometrial epithelium being hormonally regulated during the window of implantation, and PG receptors are expressed in the trophoectoderm and inner cell mass of mouse blastocysts. Using an in vitro model of embryo adhesion, we demonstrated that inhibition of PGE2 and PGF2α or PG receptors (EP2 and FP) prevents embryo adhesion, which can be overcome by adding these molecules back or using their agonists. Finally, in a pilot study, we demonstrated that PGE2 and PGF2α levels from EF 24 hours prior to embryo transfer could predict pregnancy outcome. Our results suggest that PGE2 and PGF2α concentrations 24 hours prior to embryo transfer are potential noninvasive biomarkers of endometrial receptivity.

Prostaglandin E1 or E2 (PGE1, PGE2) prevents premature luteolysis induced by progesterone given early in the estrous cycle in ewes

The objective of this study was to determine whether prostaglandin E1 (PGE1) or prostaglandin E2 (PGE2) prevents premature luteolysis in ewes when progesterone is given during the first 6 days of the estrous cycle. Progesterone (3 mg in oil, im) given twice daily from Days 1 to 6 (estrus = Day 0) in ewes decreased (P < 0.05) luteal weights on Day 10 postestrus. Plasma progesterone concentrations differed (P < 0.05) among the treatment groups; toward the end of the experimental period, concentrations in jugular venous blood decreased (P < 0.05) compared with the other treatment groups. Plasma progesterone concentrations in ewes receiving PGE1 or PGE1 + progesterone were greater (P < 0.05) than in vehicle controls or in ewes receiving PGE2 or PGE2 or PGE2 + progesterone. Chronic intrauterine treatment with PGE1 or PGE2 prevented (P < 0.05) decreases in plasma progesterone concentrations, luteal weights, and the proportion of luteal unoccupied and occupied LH receptors on Day 10 postestrus in ewes given exogenous progesterone, but did not affect (P > 0.05) concentrations of PGF2α in inferior vena cava blood. Progesterone given on Days 1 to 6 in ewes advanced (P < 0.05) increases in PGF2α in inferior vena cava blood. We concluded that PGE1 or PGE2 prevented progesterone-induced premature luteolysis by suppressing loss of luteal LH receptors (both unoccupied and occupied).

Prostaglandin F2α formation is associated with mortality in a Swedish community-based cohort of older males

An increasing number of clinical studies highlight the importance of the inflammatory mediator prostaglandin F2 α (PGF(2α)). Prostaglandin F2 α activity has been suggested to play pivotal roles in the development of cardiovascular diseases and cancer. However, whether systemic PGF(2α) concentrations may signal mortality is unknown. The aim was to evaluate in vivo PGF(2α) formation, by measuring urinary 15-keto-dihydro-PGF(2α), and mortality risk in a community setting. Urinary 15-keto-dihydro-PGF(2α) was measured in a Swedish population of 670 men (aged 77-78 years) and the participants were followed up for a median of 9.7 years (383 died, among them 156 of cardiovascular causes and 102 of cancer). In Cox regression models, urinary 15-keto-dihydro-PGF(2α) was significantly associated with cardiovascular mortality [multivariate hazard ratio (HR) for 1 SD increase of urinary 15-keto-dihydro-PGF(2α): 1.18; 95% CI:1.04-1.34; P = 0.01) independent of established cardiovascular risk factors including C-reactive protein. Urinary 15-keto-dihydro-PGF(2α) was also independently associated with total mortality (multivariate HR for 1 SD increase of urinary 15-keto-dihydro-PGF(2α): 1.11; 95% CI: 1.01-1.21; P = 0.03). The combination of 15-keto-dihydro-PGF(2α) concentrations above the median and high serum high-sensitive C-reactive protein (>3 mg/L) was independently associated with a two-fold increased risk of cancer and total mortality (P = 0.02 and P < 0.001, respectively). This is the first study to show that the inflammatory mediator PGF(2α) was independently associated with mortality and specifically cardiovascular mortality 10 years later. The results are in line with the emerging evidence of the importance of the inflammatory mediator PGF(2α) in fatal cardiovascular disease.

Clinical Relevance of Plasma Prostaglandin F2α Metabolite Concentrations in Patients with Idiopathic Pulmonary Fibrosis

BackgroundIdiopathic pulmonary fibrosis (IPF) is a devastating lung disease of unknown etiology with few current treatment options. Recently, we determined an important role of prostaglandin F2α (PGF2α) in pulmonary fibrosis by using a bleomycin-induced pulmonary fibrosis model and found an abundance of PGF2α in bronchoalveolar lavage fluid of IPF patients. We investigated the role of PGF2α in human IPF by assessing plasma concentrations of 15-keto-dihydro PGF2α, a stable metabolite of PGF2α.MethodsWe measured plasma concentrations of 15-keto-dihydro PGF2α in 91 IPF patients and compared these values with those of controls (n = 25). We further investigated the relationships of plasma 15-keto-dihydro PGF2α concentrations with disease severity and mortality.ResultsPlasma concentrations of 15-keto-dihydro PGF2α were significantly higher in IPF patients than controls (p<0.001). Plasma concentrations of this metabolite were significantly correlated with forced expiratory volume in 1 second (Rs [correlation coefficient] = −0.34, p = 0.004), forced vital capacity (Rs = −0.33, p = 0.005), diffusing capacity for carbon monoxide (Rs = −0.36, p = 0.003), the composite physiologic index (Rs = 0.40, p = 0.001), 6-minute walk distance (Rs = −0.24, p = 0.04) and end-exercise oxygen saturation (Rs = −0.25, p = 0.04) when patients with emphysema were excluded. Multivariate analysis using stepwise Cox proportional hazards model showed that a higher composite physiologic index (relative risk = 1.049, p = 0.002) and plasma 15-keto-dihydro PGF2α concentrations (relative risk = 1.005, p = 0.002) were independently associated with an increased risk of mortality.ConclusionsWe demonstrated significant associations of plasma concentrations of PGF2α metabolites with disease severity and prognosis, which support a potential pathogenic role for PGF2α in human IPF.

Oxytocin stimulated release of PGF2α and its inhibition by a cyclooxygenase inhibitor and an oxytocin receptor antagonist from equine endometrial cultures

Uterine inflammation results in a poor uterine environment and early embryonic loss in the mare due to an inhibition of maternal recognition of pregnancy caused from increased prostaglandin F2α (PGF2α). Oxytocin binds to endometrial cell receptors to activate prostaglandin synthesis. An oxytocin receptor antagonist (Atosiban) and a cyclooxygenase inhibitor (indomethacin) both decrease PGF2α production. The aim of this study was to evaluate the in vitro effects of Atosiban and indomethacin on equine uterine prostaglandin secretion. Equine endometrial explants were harvested on day two of behavioral estrus. Endometrial explant cultures were challenged with oxytocin (250nM) and PGF2α concentrations were measured over time. Explants were also cultured with Atosiban and indomethacin for 6h to determine the influence on PGF2α secretion. When endometrial explants were challenged with oxytocin, PGF2α concentrations were greater (P<0.0001) at each time point over the 24h of culture as compared to controls. Oxytocin failed (P<0.001) to elicit PGF2α release in explants cultured with either Atosiban or indomethacin. These findings show equine endometrial explants can be stimulated with oxytocin to increase secretion of PGF2α and this secretion can be inhibited through an oxytocin receptor antagonist and a Cox inhibitor, suggesting that this response to oxytocin involves an oxytocin receptor mediated event that activates the prostaglandin synthesis cascade through cyclooxygenase. Furthermore, this data suggests a role for the use of these inhibitors in vivo to decrease uterine PGF2α secretion and prevent early luteal regression and embryonic loss.

Effects of the precalving administration of omega-3 fatty acids alone or in combination with acetylsalicylic acid in periparturient dairy cows1

This study investigated the effects of the administration of long chain omega-3 fatty acids (ω-3 FA) and acetylsalicylic acid (ASA) on inflammation, performance, and fertility in periparturient dairy cows. Five weeks before calving, 26 multiparous dairy cows were randomly assigned to 1 of 3 treatments: ω-3 FA (n = 9; OME), ω-3 FA and ASA (n = 9; OMAS), or palm oil (n = 8; CTR). During the last 3 wk of pregnancy, OME and OMAS groups received daily 12.0 g of fish-derived ω-3 FA, whereas CTR cows received only SFA. In addition, OMAS cows received daily 6.0 mg ASA/kg BW starting at 7 d before calving. Only a few cows had health problems after calving, but those in OMAS were most affected (n = 3 vs. 1 in CTR). Inflammatory status around calving did not improve in OME cows, as confirmed by the patterns of concentration of acute-phase proteins (APP), which were similar to CTR. Compared with CTR and OME, the increase of the positive APP and the decrease of the negative APP (e.g., albumin; P < 0.01) observed in OMAS cows suggested a severe inflammatory status after calving. Compared with OMAS, postcalving energy metabolism was better in OME cows as shown by a lower degree of lipomobilization (smaller BCS drop, greater glucose) and milder ketogenesis (less β-hydroxybutyrate; P < 0.01). Cows in CTR had optimal fertility indices, whereas OMAS was the worst group. The severe inflammation and the more negative energy balance likely contributed to the poor fertility parameters in those cows. It is known that ASA exerts an inhibitory effect on cyclooxygenases, causing a possible decrease in the synthesis of PGF2α. A decreased concentration of PGF2α is connected with alterations in the physiologic processes related to labor and to uterine motility. Cows in OMAS had a longer pregnancy (P < 0.10 vs.OME) and a greater frequency of retained placenta, which may be attributed to decreased synthesis of PGF2α. The administration of ω-3 FA alone did not delay calving or the expulsion of fetal membranes. In conclusion, long-chain ω-3 FA improved the physiological status of cows, partly through better energy balance. The administration of ASA before calving (even at a low dose) in combination with ω-3 FA did not exert any synergistic positive effect on inflammation and performance.

Effects of conjugated linoleic acids on prostaglandin secretion by bovine endometrial epithelial cells in vitro

To determine the effects of 2 conjugated linoleic acid (CLA) isomers (cis-9, trans-11 and trans-10, cis-12) on synthesis of prostaglandin (PG) E(2) and F(2α) and expression of prostaglandin H synthase-2 (PGHS-2) of adult and fetal bovine endometrial epithelial cells in vitro. Primary cultures of endometrial epithelial cells obtained from 4 adult cows and 4 fetal bovine carcasses. Cells were exposed to 0, 50, 100, or 200μM cis-9, trans-11 or trans-10, cis-12 CLA isomers for 24 hours. Culture media collected before and after 6 hours of stimulation of cells with phorbol 12-myristate 13-acetate were assayed to detect PGE(2) and PGF(2α) via ELISA. After stimulation, cells were collected for western blot analysis to quantify PGHS-2. Concentrations of PGF(2α) and PGE(2) were significantly lower in culture media of adult and fetal endometrial epithelial cells exposed to any concentration of either CLA than they were in media of cells not exposed to CLAs. The trans-10, cis-12 CLA isomer seemed to decrease PG production more markedly than did the cis-9, trans-11 CLA isomer. Most concentrations of both CLAs significantly reduced culture media PGE(2):PGF(2α) concentration ratios of cells. Exposure of cells to CLAs did not affect expression of PGHS-2 protein. Results of this study indicated CLAs significantly decreased PGF(2α) and PGE(2) concentrations and PGE(2):PGF(2α) concentration ratios for cultures of adult and fetal endometrial epithelial cells with no apparent effect on PGHS-2 expression. Similar effects in cows could have effects on maternal recognition of pregnancy and immune function.

Effect of conceptus on expression of prostaglandin F2α receptor in the porcine endometrium

Increased synthesis of prostaglandin F2α (PGF2α) in the endometrium and conceptus during the implantation period results in elevated concentration of PGF2α in the uterine lumen in pregnant gilts. PGF2α exerts its effects through PGF2α receptor (PTGFR), a G–protein-coupled receptor. However, besides studies concerning the function of PTGFR in endometrial abnormalities, the role of PTGFR in the endometrium during early pregnancy has not been elucidated. Therefore, the aim of this study was: (1) to evaluate the profile of PTGFR gene and protein expression in the porcine endometrium during early pregnancy and the estrous cycle; (2) to determine if the effect of conceptus on PTGFR expression is dependent on type of endometrial cells—luminal epithelial (LE) or stromal (ST) cells; and (3) to elucidate if the putative effect of conceptus on endometrial PTGFR expression is mediated by estrogen receptor. We evaluated the expression pattern of PTGFR gene and protein in the endometrium during day 9, 11, 12, 15, and 18 of the estrous cycle and pregnancy (N = 4–6 per group). The expression of PTGFR mRNA was greater on day 18 of pregnancy and the estrous cycle (vs. days 9–15 of the estrous cycle and pregnancy, P < 0.05). Expression of PTGFR protein was approximately 10-fold upregulated in the endometrium on day 15 of pregnancy when compared with day 15 of the estrous cycle (P < 0.01). Endometrial expression of PTGFR protein increased from day 12 to 18 of pregnancy (P < 0.05). PTGFR mRNA was expressed in LE and ST cells. In a subsequent experiment, we used a coculture model in which LE cells were cultured on collagen-coated inserts placed in wells plated with ST cells. Day 11 or 15 conceptus-exposed medium (CEM) elevated expression of PTGFR mRNA (2- and 1.5-fold, respectively, P < 0.05) in LE cells cocultured with ST cells. CEM did not have an effect on PTGFR mRNA expression in ST cells. The 11-day CEM-induced increase of PTGFR mRNA was abolished by incubation of LE cells in the presence of the estrogen receptor antagonist (ICI-182,780; P < 0.01). Summarizing, the conceptus upregulated expression of PTGFR in the endometrium during the implantation period. Moreover, this study indicates that expression of PTGFR gene was elevated in LE cells of endometrium by embryonic signal of estradiol. Our results suggest para- and autocrine effects of PGF2α through its receptor PTGFR in the porcine endometrium, especially in luminal epithelium which is in direct contact with the conceptus during the implantation period.

Effect of Prostaglandin F2α on Growth of Staphylococcus Aureus Associated with Bovine Mastitis

Staphylococcus aureus is a major infectious organism causing mastitis. Certain fatty acids have been shown to inhibit the growth of pathogens including Staphylococcus aureus. An in vitro experiment was conducted to determine the effects of prostaglandin F2α (PGF2α) on growth of S. aureus. Flasks containing tryptic soy broth (TSB) were inoculated with S. aureus Novel, and subsequently treated with PGF 2α (dinoprost tromethamine) at concentrations of 0, 0.3, 0.6, 1.2, and 2.4 mg/ml. Cultures were incubated at 37°C for 24 h and sampled every 3 h. The entire experiment was conducted three times in duplicates. Bacterial growth was assessed by counting colony forming units (CFU) in triplicates. Data were analyzed by repeated measures analysis of variance and reduced and full dummy variable regression models to determine the effect of PGF2α concentrations on growth patterns of S. aureus over time. There was an effect of treatment and treatment by time interaction (P<0.05) on mean log CFU, indicating that bacterial growth over 24 h was different across treatments. At time of inoculation (0 h), mean log CFU values were not different among treatments; however at 24 h, mean log CFU for each PGF 2α treatment was different from control and decreased (P<0.05) with increasing concentrations of PGF 2α . The predicted growth curve for each treatment was different (P<0.05) when compared with the control, and the rate of bacterial growth was less for 1.2 mg/ml PGF 2α when compared with the control and 0.6 mg/ml. The bacterial growth was completely inhibited during the 24 h period in 2.4 mg/ml PGF 2α treatment. Furthermore, a non-linear regression model was employed to estimate the dose response at the 24 h, and revealed that 1.2 mg/ml was the minimum inhibitory dose. These results provide evidence, for the first time, that PGF 2α , in the form of dinoprost tromethamine, has bacteriostatic and bactericidal effects on S. aureus in vitro.

Endocrine and Molecular Control of Luteal and Placental Function in Dogs: A Review

In the domestic dog (Canis familiaris), the corpus luteum (CL) is the only source of progesterone (P4) in non-pregnant and pregnant animals. The progesterone secretion profiles are almost identical in both conditions until the last third of the luteal phase when the gradual P4 decline turns into a steep drop in pregnant bitches, indicating the onset of parturition. Consequently, the length of the CL-phase in non-pregnant dogs exceeds the luteal lifespan in pregnant animals. The canine CL-function is regulated by many species-specific regulatory mechanisms, the most intriguing of which is the reported independence of gonadotropic support during the first third of dioestrus. Recently, PGE2 has been proposed as one of the most important luteotropic factors acting locally during this time, but afterwards prolactin (PRL) appears to be the main luteotropic factor. Luteal regression/luteolysis occurs, however, in spite of an increased gonadotropic support. Lately, by demonstrating the expression of PRL-receptor (PRLr), a new insight into possible regulatory mechanisms has indicated that the supply of P4 could be controlled upstream of the steroidogenic machinery at the level of PRLr expression and/or function, subsequently leading to the functional suppression of the steroidogenic machinery. An endogenous source of a luteolytic agent is apparently lacking, implicating the luteal regression in non-pregnant bitches as a passive, degenerative process even if the PGF2α-receptor is constitutively expressed in canine CL. This is in contrast to pregnant dogs in which prepartum luteolysis seems to be an active process of CL destruction by PGF2α of utero/placental origin targeting the luteal PGF2α-receptor.

Endocrine and inflammatory factors and endometriosis-associated infertility in assisted reproduction techniques

Our research aimed to evaluate the effect of endometriosis on folliculogenesis and pregnancy, and to assess the involvement of inflammatory factors (IL1b, PGE2, PGF2α, and TGFβ2) in follicular fluid. A total of 65 follicular fluid aspirates were collected. Concentrations of inflammatory factors (IL1b, PGE2, PGF2α, and TGFβ2) and steroid hormones (E2, progesterone, FSH, and LH) within follicular fluid as well as serum E2 and LH concentrations were measured. The mRNA expression of IL1b, Ptgs2, aromatase, and PPARγ in granulosa cells was determined. The outcome of ART was monitored and recorded. The oocyte retrieval, rate of metaphase II oocyte, cleavage rate, effective embryo rate, and pregnancy rates of patients with endometriosis were all significantly lower than those of the control patients. In those with endometriosis, serum E2 concentrations were lower than those observed in controls. Aromatase levels in the granulosa cells of the endometriosis group were lower while concentrations of PGE2 in follicular fluid were higher than in the control group. Concentrations of PGE2, PGF2α, TGFβ2, and IL1b were significantly correlated with each other. These results suggest that the outcomes of ART, in relation to serum E2 concentration, were adversely affected by the presence of endometriosis. Furthermore, the results supported that, among the endocrine and inflammatory factors, PGE2 within the follicular fluid impairs the number and quality of oocytes.

Effects of oxytocin, lipopolysaccharide (LPS), and polyunsaturated fatty acids on prostaglandin secretion and gene expression in equine endometrial explant cultures

Increased secretion of prostaglandin F2α (PGF2α) within the uterus because of uterine inflammation can cause luteolysis and result in early embryonic loss. Supplementation with polyunsaturated fatty acids (PUFAs) has been shown to influence PG production in many species, although the effects on the mare remain unknown. The present study aimed to determine fatty acid uptake in equine endometrial explants and evaluate their influence on PG secretion and expression of enzymes involved in PG synthesis in vitro. Equine endometrial explants were treated with 100 μM arachidonic acid, eicosapentaenoic acid, or docosahexaenoic acid and then challenged with oxytocin (250 nM) or lipopolysaccharide (LPS; 1 μg/mL). Production of PGF2α and PG E2 (PGE2) was measured, and mRNA expression of enzymes involved in PG synthesis was determined with quantitative real-time PCR. Media concentrations of PGF2α and PGE2 were higher (P < 0.0001) from endometrial explants challenged with oxytocin or LPS compared with controls despite which fatty acid was added. Only DHA lowered (P < 0.0001) media concentrations of PGF2α and PGE2 from explants. Endometrial explants stimulated with oxytocin had increased expression of PG-endoperoxide synthase 1 (PTGS1; P < 0.02), PG-endoperoxide synthase 2 (PTGS2; P < 0.001), PG F2α synthase (PGFS; P < 0.01), PG E2 synthase (PGES; P < 0.01), and phospholipase A2 (PLA2; P < 0.005) compared with controls and regardless of fatty acid treatment; whereas stimulation with LPS increased expression of PTGS2 (P < 0.004), PGFS (P < 0.03), PGES (P < 0.01), and PLA2 (P < 0.01) compared with controls and regardless of fatty acid treatment. Treatment with PUFAs, specifically DHA, can influence PG secretion in vitro through mechanisms other than enzyme expression.