Inhibition of uterine prostaglandin-F2 alpha production by bovine conceptus secretory proteins

The effect of bovine conceptus secretory proteins (CSP) on uterine prostaglandin (PG)-F2 alpha production was evaluated in dairy cattle following injection of estradiol-17 beta. Intrauterine injections of dialyzed serum proteins (Control, n = 5) or CSP (n = 5) were administered from days 15 through 18 post-estrus. Following intrauterine treatments on day 18, all cows were injected with E2 (3 mg) to stimulate uterine PGF2 alpha production. Plasma concentrations of progesterone (P4) and 15-keto-13,14-dihydro-PGF2 alpha (PGFM) were determined by RIA. The PGFM responses following E2 challenge were decreased (p less than 0.01) for cows receiving CSP versus serum proteins into the uterine lumen. Individual PGFM, P4 and cycle length responses are discussed. Data suggest that proteins secreted by the bovine conceptus suppress uterine PGF2 alpha production during pregnancy recognition in the cow.     

Intracellular recording of cerebral cortical actions of prostaglandin F2α (PGF2α)

Our reported data on the cortical inhibitory actions of prostaglandin F_2α (PGF_2α) and the diversity of data in the literature on cerebral PG actions are examined here in the light of intracellular recording which provides the requisite membrane data for the first time. Thus, 1) intracellular recording from the cat cerebral cortex is obtained for the actions of PGF_2α and for norepinephrine (NE) and serotonin (5HT). 2) The parallel changes in firing and polarization and the simultaneous transmembrane conductance changes are qualitatively identical for PGF_2α, NE and 5HT. 3) The reduction in firing accompanied by hyperpolarization indicates that PGF_2α, NE and 5HT all inhibit these cells. 4) The ionic species responsible for this inhibition is such that it increased the transmembrane resistance, and this was true for all three. 5) The changes in membrane parameters, identical in direction for PGF_2α and NE, but stronger for the latter, constitute conditions that can lead to competitive inhibition and therefore conote, presumably, actions at the same or related receptors. Such competition with evoked cortical field potentials is shown in the preceding paper.     

Release of prostaglandin F2α during the bovine peripartal period

Progesterone, estrone and 15-keto-13,14-dihydro-PGF_2α levels were determined in the peripheral blood circulation during the peripartal period in 12 cows. Plasma concentrations of progesterone showed a gradual and continuous decrease during the last 60 days before parturition. This gradual decrease was followed by an abrupt decline in the progesterone concentration occurring 24–48 hours before delivery. The plasma levels of estrone started to increase about 30 days prior to parturition with high concentrations attained during the last days of pregnancy. After delivery the estrone content decreased to baseline levels. Increased levels of the PGF_2α metabolite were recorded 24–48 hours before parturition. These increased PGF_2α metabolite levels occurred before or in conjunction with prepartum luteolysis. Prostaglandin metabolite levels remained high during parturition and returned to baseline 10–20 days after delivery.     

Disappearance of prostaglandins F2α and 15-methyl F2α from amniotic fluid as measured by radioimmunoassay

A sensitive and relatively specific radioimmunoassay for 15 (S) 15 methyl prostaglandin F_2α was used to determine the levels of the drug in amniotic fluid after it had been injected intra-amniotically for termination of second trimester pregnancy. The disappearance of the free acid (tham salt) and methyl ester of the prostaglandin analogue were similar. The results of this preliminary study suggest that the drug rapidly equilibrates in the fluid and this is followed by a slow removal from the amniotic sac. A comparison with a similar study with PGF_2α, revealed that the analogue had a longer half-life in the amniotic fluid.     

Cross-Reactivity of Δ17-6-Keto-PGF1α with 6-Keto-PGF1α antibodies

The cross-reactivity of the PGI₃ metabolite, Δ17-6-keto-PGF_1α, with antibodies against 6-keto-PGF_1α for radioimmunoassays (RIA) has been investigated. Δ17-6-keto-PGF_1α was obtained either from commercial sources or after its purification from endothelial cells. In the latter case, primary cultured bovine aortic endothelial cells were incubated for 20 min at 37°C with 10 μM eicosapentaenoic acid (EPA) in the presence of 2 μM 13-hydroperoxy-octadecadienoic acid, an activator of the EPA cyclooxygenation, and the 6-keto-PGF_1α and Δ17-6keto-PGF_1α produced were separated by RP-HPLC. Then, cross-reactivities of the commercial and purified Δ17-6-keto-PGF_1α with 6-keto-PGF_1α antibodies were determined and found not to exceed 10%. In addition, the amounts of prostacyclin-related compounds detected by direct measurements in media of cells loaded with EPA were compared with those obtained after purification of 6-keto-PGF_1α. In accordance with the cross-reactivity data, we found that RIA in media mainly measured 6-keto-PGF_1α, the Δ17-6-keto-PGF_1α formed being undetected at 90%. It is concluded that 6-keto-PGF_1α antibodies generally used for RIA of 6-keto-PGF_1α are highly specific since they can discriminate a metabolite bearing an additional double bond such as the PGI₃ metabolite Δ17-6-keto-PGF_1α.     

Interferon γ stimulates prostaglandin E2 production by mouse Kupffer cells

When mononuclear phagocytes, including Kupffer cells, are activated by various agents, they synthesize and release arachidonic acid metabolites, prostaglandins (PGs) and leukotrienes (LTs). In this study, we examined the effect of in vitro Kupffer cell activation with recombinant murine IFN gamma on PGE2 and LTB4 secretion. IFN gamma enhanced PGE2 secretion, and this effect of IFN gamma was stronger than that of IL-1 or TNF. Moreover, IFN gamma promoted LTB4 release especially in the absence of PGs. On the other hand, dexamethasone and indomethacin inhibited and, EGTA and TMB-8, which reduce intracellular Ca++ Levels, blocked IFN gamma induced PGE2 production, which suggested that the activation of phospholipase A2 and cyclooxygenase in Kupffer cells requires the elevation of intracellular Ca++ levels.     

Cardiovascular actions of prostaglandins F2α; 15-me-F2α; F1α; F2β and F1β in the cat

Prostaglandin F_2α (5μg/kg, i.v.) causes an increase in pulmonary arterial pressure, decrease in systemic arterial pressure, and reflex bradycardia in the anesthetized cat. The same dose of the 15-methyl analogue of PGF_2α produces the same triad of effects but of greater magnitude and duration. Although prostaglandins F_1α, F_2β and F_1β also cause the same cardiovascular effects as F_2α, there is a decrease in potency for all parameters measured, with PGF_2α>PGF_1α>PGF_2β>PGF_1β. When compared to the actions of PGF_2α in producing an increase in pulmonary arterial pressure, PGs F_1α, F_2β and F_1β were less potent by approximately 10, 100, and 1000 fold respectively.     

Stimulation by prostaglandin F2α of acidic glycosaminoglycan production in cultured fibroblasts

The effect of PGF2α on the synthesis of hexosamine-containing substances (acidic glycosaminoglycans and glycoproteins) was studied in cultured fibroblasts derived from a rat carrageenin granuloma. Treatment with PGF2α ranging from 0.01 μg/ml to 20 μg/ml resulted in a significant increase of the production of these macromolecules by the cells. The stimulatory effect was found significant even at the low concentration of 10 ng/ml, and could be seen as early as 3h after exposure to PGF2α. The hexosamine-containing substances increased by PGF2α revealed that 80% of the increase was due to acidic glycosaminoglycans and the rest was due to glycoproteins.     

PGE2 and PGF2α release by human peritoneal macrophages in endometriosis

Objective: To test for differences in the amount and activity of peritoneal macrophages present in the peritoneal fluid of women with, and without endometriosis using prostaglandin release by macrophages in culture as a marker.Patients: Women of reproductive age undergoing laparoscopy for infertility or chronic pelvic pain with postoperative diagnosis of endometriosis and women undergoing laparoscopy for sterilization.Methods: Peritoneal fluid was aspirated during laparoscopy, volume was recorded, macrophages were isolated via a Ficoll Paque gradient and kept in primary culture. PGE₂ and PGF_2α release of the cells were measured before and after stimulation with zymosan.Results: Women with endometriosis had significantly more peritoneal macrophages than controls. Peritoneal macrophages of women with endometriosis released significantly more PGE₂ than those of the control group: 8.4 ± 2.0 versus 1.4 ± 0.4 ng/ml/10⁶cells (mean ± SEM, p=0.0005) and PGF_2α : 10 ± 4.3 (endometriosis) versus 1.8 ± 0.4 (control) ng/ml/10⁶cells (mean ± SEM, p = 0.045).Conclusion: There is a significant increase in the amount of prostaglandins released by peritoneal macrophages from women with endometriosis. These prostaglandins might alter uterine and tubal contractility, thereby affecting fertility.     

Vasoactive intestinal polypeptide (VIP) inhibits prostaglandin-F2α-induced activity of the rabbit myometrium

Seven female rabbits had myometrial autografts transplanted into ear-chambers. The electrical activity of the graft was recorded and mechanical activity observed. Vasoactive intestinal polypeptide (VIP) dose-dependently inhibited myometrial activity induced by prostaglandin-F_2α. This evidence and the occurrence of “VIPergic” nerve fibres, which seem to innervate the myometrial cells, suggest that VIP may play a physiological role in the local control of the myometrial activity.     

Effect of prostaglandins E1, E2 and F2α on neutrophil aggregation

Recently we have found that chemotactic factors stimulate neutrophils in suspension to aggregate. Because of an obvious analogy to platelet aggregation, we examined the influence of three prostaglandins on this process. Prostaglandins E₁, E₂ and F_2α alone did not cause aggregation of the neutrophils but were able to partially inhibit the aggregation response induced by the synthetic chemotactic tripeptide, formly-methionyl-leucyl-phenylalanine. The minimal inhibitory concentrations for prostaglandins E₁, E₂ and F_2α were 10⁻⁷, 10⁻⁶ and 10⁻⁵M, respectively. These results are similar to those found for the prostaglandin-induced inhibition of platelet aggregation. It may be, therefore, that neutrophil aggregation, like platelet aggregation, is modulated by intracellular prostaglandins and other products of arachidonic acid metabolism.     

Stereospecificity of enzymatic reduction of prostaglandin E2 to F2α

Antibodies directed toward PGF_2β were prepared in rabbits. The serologic specificity of the immune reaction was determined by inhibition of sodium borohydride-reduced (³H) PGE₂ anti-PGF_2β binding by several prostaglandins. The antibodies to PGF_2β recognize the β-hydroxyl configuration in the cyclopentane ring of PGF_2β. With the use of both anti-PGF_2α and anti-PGF_2β, the product of PGE₂ reduction by 9-ketoreductase purified from chicken heart was identified as PGF_2α. Guinea pig liver and kidney homogenates were examined for PGE 9-ketoreductase activity. Although enzyme activity was present, no evidence of PGF_2β production was found.     

The enzymatic conversion of prostaglandin D2 to prostaglandin F2α

Prostaglandins E₂ and D₂ were both converted to prostaglandin F_2α (9α, 11α) by an enzyme present in sheep blood. Neither the 9β, 11α epimer nor the 9α, 11β epimer was produced from PGE₂ or PGD₂ respectively. The rate of reduction was measured using isotope dilution (D₄ PGF_2α) and multiple-ion detection gas chromatography-mass spectrometry.     

Inhibition of adipose differentiation by 9α, 11β-prostaglandin F2α

Prostaglandin F2α (PGF2α) is a potent adipose differentiation inhibitor for the adipogenic cell line 1246 and for adipocyte precursors in primary culture with an ED50 of 3×10⁻⁸ M. In this paper, we examined the effect of several prostaglandins which have structural similarities with PGF2α on the differentiation of 1246 cells and of adipocyte precursors in primary culture. The results show that only 9α,11β-PGF2α is as potent as PGF2α to inhibit differentiation of adipocyte precursors in primary culture and of the adipogenic cell line 1246. In the presence of 9α,11β-PGF2α, the cells remained fibroblast-like, typical of undifferentiated adipocyte precursors. Triglyceride accumulation and increase of specific activity for glycerol-3-phosphate dehydrogenase were inhibited. In addition, mRNA expression of early markers of differentiation such as lipoprotein lipase (LPL) and fatty acid binding protein (FAB) was decreased. The isomer 9β,11α-PGF2α and other PGF2α derivatives were inactive. These results provide new information on the biological activity of 9α,11β-PGF2α as an inhibitor of adipose differentiation and about the structural characteristics of prostaglandins required for maintenance of a high adipose differentiation inhibitory effect.     

Induction of second trimester abortion: Comparison between vaginal 15-methyl-PGF2α methyl ester and intra-amniotic PGF2α

The efficiency and acceptability of a single-dose, long-acting vaginal suppository containing 3.0 mg of 15-methyl PGF_2α methyl ester was compared with intra-amniotic administration of 50 mg of PGF_2α in 100 patients with a second trimester pregnancy termination. Within 24 hours, 78 per cent of the patients in the vaginal group and 92 per cent in the intra-amniotic group had aborted. The mean induction-abortion interval was 17.9 hours in the vaginal group and 15.8 hours in the intra-amniotic group.Gastrointestinal side-effects were more frequent, but the procedure was less painful, with vaginal 15-methyl PGF_2α methyl ester than with intra-amniotic PGF_2α.The vaginal route is technically simple for adaptation to large-scale use, but the high frequency of gastrointestinal side-effects still limits the acceptability of 15-methyl PGF_2α methyl ester in vaginal administration.     

Prostaglandin E1 and F2α specific binding in bovine corpora lutea: Comparison with luteolytic effects

Preliminary characterization indicated the presence of separate prostaglandin (PG)E₁ and (PG)F_2α binding sites in membrane fractions prepared from bovine corpora lutea. These differ in the rate and temperature dependence of the specific binding. Equilibrium binding data indicate the apparent dissociation constants as 1.32 × 10⁻⁹M and 2.1 × 10⁻⁸M for PGE₁ and PGF_2α, respectively. Competition of several natural prostaglandins for the PGE₁ and PGF_2α bovine luteal specific binding sites indicates specificity for the 9-keto or 9α-hydroxyl moiety, respectively. Differences in relative ability to inhibit ³H-PG binding were found due to sensitivity to the absence or presence of the 5,6-cis-double bond as well.Bovine luteal function was affected following treatment of heifers with 25 mg PGF_2α as measured by reduced estrous cycle length, decreased corpus luteum size and significantly decreased plasma progesterone levels. In contrast, treatment with 25 mg PGE₁ resulted in cycle lengths comparable to those of non-treated herdmates with no apparent modification in corpus luteum size. However, plasma progesterone levels were increased significantly following PGE₁ treatment compared to pretreatment values. In so far as data obtained on PGF_2α relative binding affinity to the bovine CL can be compared to data obtained independently on PGF_2α induced luteolysis in the bovine, PGF_2α relative binding to the CL and luteolysis appeared to be associated. By similar reasoning, there was no apparent relationship between PGE₁ relative binding affinity in the luteal fractions and luteolysis in estrous cyclic cattle.     

Cardiac arrhythmias induced by prostaglandin F2α in cats

Intravenous administration of prostaglandin F_2α results in transient episodes of sinus bradycardia in anesthetized cats. In addition, ventricular bigeminy was observed in approximately 40% of those cats anesthetized with pentobarbital (36 mg/kg) and 58% of those anesthetized with chloralose (70 mg/kg). This arrhythmogenic effect of PGF_2α is abolished following bilateral vagotomy, indicating that the arrhythmias are most likely due to a marked stimulation of vagal tone in this species.     

Prostaglandin E1 and F2α specific binding in bovine corpora lutea: Comparison with luteolytic effects

Preliminary characterization indicated the presence of separate prostaglandin (PG)E₁ and (PG)F_2α binding sites in membrane fractions prepared from bovine corpora lutea. These differ in the rate and temperature dependence of the specific binding. Equilibrium binding data indicate the apparent dissociation constants as 1.32 × 10⁻⁹M and 2.1 × 10⁻⁸M for PGE₁ and PGF_2α, respectively. Competition of several natural prostaglandins for the PGE₁ and PGF_2α bovine luteal specific binding sites indicates specificity for the 9-keto or 9α-hydroxyl moiety, respectively. Differences in relative ability to inhibit ³H-PG binding were found due to sensitivity to the absence or presence of the 5,6-cis-double bond as well.Bovine luteal function was affected following treatment of heifers with 25 mg PGF_2α as measured by reduced estrous cycle length, decreased corpus luteum size and significantly decreased plasma progesterone levels. In contrast, treatment with 25 mg PGE₁ resulted in cycle lengths comparable to those of non-treated herdmates with no apparent modification in corpus luteum size. However, plasma progesterone levels were increased significantly following PGE₁ treatment compared to pretreatment values. In so far as data obtained in vitro on PGF_2α relative binding affinity to the bovine CL can be compared to data obtained independently in vitro on PGF_2α induced luteolysis in the bovine, PGF_2α relative binding to the CL and luteolysis appeared to be associated. By similar reasoning, there was no apparent relationship between PGE₁ relative binding affinity in the luteal fractions and luteolysis in estrous cyclic cattle.     

Radioimmunoassays of prostaglandin F2α and prostaglandin F2α-main urinary metabolite with prostaglandin-I-tyrosine methylester amide

Radioimmunoassays for measuring prostaglandin F_2α (PGF_2α) and 5α, 7α-dihydroxy-11-keto tetranorprosta-1,16-dioic acid, PGF_2α-main urinary metabolite (PGF_2α-MUM), with ¹²⁵I-tyrosine methylester amide (TMA) of PGF_2α and PGF_2α-MUM were developed.Antibody to PGF_2α was produced in rabbits immunized with conjugates of PGF_2α coupled to bovine serum albumine. Antibody to PGF_2α-MUM was also produced in rabbits immunized with conjugates of PGF_2α-MUM coupled to bovine serum albumin.PGF_2α-¹²⁵I-TMA had an affinity to antiserum to PGF_2α. PGF_2α-MUM-¹²⁵I-TMA also responded to antiserum to PGF_2α-MUM.