Bronchopulmonary effects of prostaglandin F2α and three of its metabolites in the dog

The airway and lung dynamics of prostaglandin F_2α (PGF_2α) and three of its metabolites were examined in the spontaneously-ventilated, pentobarbital anesthetized dog. Changes in expiratory flow rate, tidal volume, respiration rate, lung resistance and dynamic lung compliance were evaluated and compared quantitatively. In a dose range of 0.3–3.0 μg/kg i.v., PGF_2α and its 13,14-dihydro metabolite were found to be exceptionally potent agents. This metabolite was approximately twice as potent as PGF_2α on most parameters studied. Two other metabolites, 15-keto-PGF_2α and 15-keto-13,14-dihydro-PGF_2α, were only slightly effective, even in a dose range of 1.0–30.0 μg/kg i.v. These latter two metabolites produced dose-response curves with significantly shallower slopes than PGF_2α and were shown to be at least thirty-five times less potent than the parent compound. Therefore, oxidation of PGF_2α at the carbon-15 position by 15-hydroxy prostaglandin dehydrogenase appears to produce compounds with minimal bronchopulmonary activity.     

Use of tritium labeled amino acid conjugates of prostaglandins and thromboxanes as labeled ligands in prostanoid radioimmunoassay

Conjugates of prostaglandins and thromboxanes with tritium labeled amino acids were prepared and employed as labeled ligands in porstaglandin and thromboxane radioimmunoassays. Assays for PGF_2α, 15-keto-13, 14-dihydro-PGF_2α, TXB₂ and 15-keto-13, 14-dihydro-TXB₂ were evaluated in comparative studies using either these heterologous ligands or the corresponding homologus tritiated eicosanoid as tracers. Binding properties for the respective antibodies were found to be similar using either tracer.Three biological studies were also conducted, viz. study of the release of TXB₂ during collagen induced platelet aggregation, of 15-keto-13, 14-dihydro-TXB₂ during guinea pig pulmonary anaphylaxis, and of PGF_2α (measured as 15-keto-13, 14-dihydro-PGF_2α in peripheral plasma) during bovine luteolysis. The analyses gave comparable results using either the heterologous or the homologous assay.Thus, this type of labeled prostanoid conjugates may serve as a convenient alternative to homologous tracers in radioimmunoassay. Heterologous tracers may even in certain cases provide the only simple solution to the problem of preparing a labeled ligand of high specific activity.     

Catabolism of prostaglandin F2α in rat ovary : Differences between ovarian and uterine tissues

The catabolism of prostaglandin F_2α(PGF_2α) in rat ovarian homogenate was studied comparing with uterine homogenate. Two kinds of metabolite were recognized by incubation of PGF_2α with ovarian or uterine homogenate; 13,14-dihydro-15-keto-PGF_2α(15KD-PGF_2α) and 13,14-dihydro-PGF_2α(13,14H₂-PGF_2α) in ovarian homogenate and 15-keto-PGF_2α(15K-PGF_2α) and 15KD-PGF_2α in uterine homogenate. Incubation of 15KD-PGF_2α with ovarian homogenate resulted in the formation of 13,14H₂-PGF_2α but incubation with uterine homogenate did not produce 13,14H₂-PGF_2α. 13,14H₂-PGF_2α was in accord with Rf value of a compound formed by reduction of 15KD-PGF_2α with sodium borohydride.     

Inhibition of uterine prostaglandin-F2α production by bovine conceptus secretory proteins

The effect of bovine conceptus secretory proteins (CSP) on uterine prostaglandin (PG)-F_2α production was evaluated in dairy cattle following injection of estradiol-17β. 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 PGF_2α production. Plasma concentrations of progesterone (P₄) and 15-keto-13,14-dihydro-PGF_2α (PGFM) were determined by RIA. The PGFM responses following E₂ challenge were decreased (p<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 PGF_2α production during pregnancy recognition in the cow.     

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.     

Measurement of 11-ketotetranor PGF metabolites: An approach for monitoring prostaglandin F2α release in the mare

The appearance and disappearance of 15-keto-13,14-dihydro-PGF_2α and 11-ketotetranor PGF metabolites have been measured in the blood and urine of the mare following i.v. injection of prostaglandin F_2α (PGF_2α). The basal plasma concentration of the 11-ketotetranor PGF metabolites was 10-fold greater than 15-keto-13,14-dihydro PGF_2α; after injection of PGF_2α, however, 15-keto-13,14-dihydro PGF_2α increased rapidly to concentrations exceeding those of the 11-ketotetranor PGF metabolites, which also increased but to a much lesser extent. The half-life for the disappearance of 15-keto-13,14-dihydro PGF_2α was about 30-fold shorter than that of the 11-ketotetranor PGF metabolites. Similar profiles were seen in the urine, except that the concentration of the 11-ketotetranor PGF metabolites was always greater than that of the 15-keto-13,14-dihydro PGF_2α. In the mare, the main plasma metabolite of PGF_2α appears to be 15-keto-13,14-dihydro PGF_2α, whereas the 11-ketotetranor PGF metabolites predominate in the urine.Similar patterns were seen for both types of metabolite in the plasma during luteolysis and early pregnancy. Because of the differences in rates of appearance and disappearance of these metabolites, measurement of both allows the detection of peaks of PGF_2α release in samples taken less frequently than is necessary when either type is measured alone.     

Responsiveness of the lamb ductus arteriosus to prostaglandins and their metabolites

The relative potencies of the prostaglandins A₁, A₂, E₁, E₂, F_2α and their 15-keto-, 15-keto-13,14-dihydro-, and 13,14-dihydro-metabolites were investigated on isolated lamb ductus arteriosus preparations contracted by exposure to elevated PO₂. All the prostaglandins (except PGF_2α and its 15-keto-metabolites) relaxed the tissue. However, only PGE₁, E₂, and their 13,14-dihydro-metabolites, were effective at concentrations below 10⁻⁸ M. Therefore, events that alter metabolism of circulating PGs in the perinatal period may have significant effects on the relative patency or closure of the ductus arteriosus.     

Determination of 15-keto-13,14-dihydro-metabolites of PGE2 and PGF2α in plasma using high performance liquid chromatography and gas chromatography-mass spectrometry

A method is described for the measurement of 15-keto-13,14-dihydrometabolites of PGE₂ and PGF_2α in peripheral human plasma. This involves purification by high performance liquid chromatography followed by determination of levels by combined gas chromatography-mass spectrometry using tetradeuterated analogs of the metabolites as internal standards. The levels of these metabolites in plasma are considered to be a more reasonable index of the entry of PGE₂ and PGF_2α into peripheral blood than are the levels of the corresponding primary prostaglandins. The endogenous levels of 15-keto-13,14-dihydro-PGE₂ and 15-keto-13,14-dihydro-PGF_2α found in peripheral plasma are 33 ± 10 pg/ml (SD; n=6) and 40 ± 16 pg/ml (SD; n=6), respectively.     

The simultaneous use of two prostaglandin radioimmunoassays employing two antisera of differing specificity: II. Relative Stability of Prostaglandins E1, E2 and F1α in Cell Cultures of BALB/c 3T3 and SV3T3 Mouse Fibroblasts.

Progesterone and the main plasma metabolite of PGF_2α, 15-keto-13,14-dihydro-PGF_2α, were determined at hourly intervals in the peripheral circulation during luteolysis in two heifers. The prostaglandin release was found to occur during 2–3 days as rapid pulses with a duration of 1–5 hours prior to and during luteolysis, which was indicated by decreasing levels of progesterone.     

Separation of prostaglandins E2 and F2α from their pulmonary metabolites by thin-layer chromatography

A simple thin-layer chromatographic system has been developed to separate PGE₂ and PGF_2α from 13,14-dihydro-PGF_2α and other pulmonary metabolites of prostaglandins. The separation is achieved on Kieselgel-60 F-254 plates impregnated with both silver nitrate and boric acid. The system should be of use in the purification and tentative identification of primary prostaglandins.     

Identification and quantitative determination of prostaglandins by high pressure liquid chromatography

High pressure liquid chromatography (HPLC) using 4′ × 1/8″ columns of a pellicular silica support (Corasil-II) allows identification of prostaglandins diastereomers based on their characteristic retention relative to a standard, PGE₂ in this study. Surprisingly this simple method allows separation of PGE₂, PGE₁, and PGE_o (dihydro-PGE₁) or PGF₂α, PGF₁α, and PGF_oα without resort to silver nitrate impregnated stationary phases. Even more subtle distinctions such as that between 13,14-dihydro-PGF₂α, PGF₁α and 5,6- -PGF₂α are possible by HPLC. The differential refractometer detector used throughout can also be used for quantitation. This is illustrated by a study of the relative rates of degradation of natural PGF₂α (an oil at the temperature employed, 41°C) and racemic PGF₂α (mp. 63°) on exposure to air.     

Kinetic and metabolic studies of Prostaglandin F2-alpha administered intra-amniotically for induction of abortion

The metabolism of intra-amniotically administered tritium labelled PGF_2α was studied in five patients admitted to the hospital for legal abortion. Three metabolites (15-keto-PGF_2α, 15-keto-13, 14-dihydro-PGF_2α and 13, 14-dihydro-PGF_2α) were identified in the amniotic fluid. The kinetics for the elimination of PGF_2α and the appearance of metabolites were studied. No correlation could be seen between the half life time of PGF_2α and the induction-abortion interval. PGF_2α, 15-keto-13, 14-dihydro-PGF_2α and 13, 14-dihydro-PGF_2α were also isolated from extracts of the placenta. Variable metabolite patterns were found in foetal tissues. These data indicate that both the foetus and the placenta are able to metabolize PGF_2α.     

Metabolism of PGI2 by 15-hydroxyprostaglandin-dehydrogenase and Δ-reductase in different vascular beds of the cat in vivo

The metabolism of endogenous PGI₂ (released by angiotensin II or bradykinin) and exogenous PGI₂ by 15-hydroxy-PG-dehydrogenase and Δ¹³-reductase was studied in five different vascular beds of the anaesthetized cat. Plasma concentrations of 6-keto-PGF_1α (the product of spontaneous hydrolysis of PGI₂) and 6,15-diketo-13,14-dihydro-PGF_1α (the metabolite formed from PGI₂ by 15-hydroxy-PG-dehydrogenase and Δ¹³-reductase) were determined in the efferent vessels of the respective vascular beds by specific radioimmunoassays.No major metabolism of PGI₂ by 15-hydroxy-PG-dehydrogenase and Δ¹³-reductase was detected in the head and the hindlimbs of the cat. In the lung exogenous (circulating) PGI₂ was not metabolized, whereas PGI₂ synthetized in the lung itself was converted to 6,15-diketo-13,14-dihydor-PGF_1α. No significant amounts of 6,15-diketo-13,14-dihydro-PGF_1α-immunoreactivity were detected in hepatic venous blood after infusion of PGI₂ into the portal vein. However as also no 6-keto-PGF_1α was found, the liver seems to efficiently extract PGI₂ from the circulation. The cat kidney had the highest capacity of all vascular beds investigated to release endogenous and exogenous PGI₂ as 6-15-diketo-13,14-dihydro-PGF_1α. In other organs (vascular beds) investigated PGI₂ is either metabolized less efficiently by the 15-hydroxy-PG-dehydrogenase or further transformed to other metabolites.     

The influence of prostaglandin metabolites on the uterine response to PGF2alpha. A clinical and pharmacokinetic study

Prostaglandin F_2α (PGF_2α) is rapidly metabolized in the human body following exogenous administration. Three main metabolites have been isolated and identified: 15-keto-PGF_2α, 15-keto-13, 14-dihydro-PGF_2α and 13, 14-dihydro-PGF_2α. One of these metabolites, 13, 14-dihydro-PGF_2α, stimulated human uterine contractility during midpregnancy both following intravenous and intra-amniotic administration. The activity approaches that of the parent compound, PGF_2α. Since the concentration of the metabolites in peripheral serum during continuous intravenous infusion of PGF_2α is only slightly lower than that found for PGF_2α, it is likely that 13, 14-dihydro-PGF_2α is of importance for the stimulatory effect of PGF_2α on human uterine activity in vivo. Intravenous injection of high doses of 15-keto-13, 14-dihydro-PGF_2α (2–4 mg) slightly stimulated uterine contractility but most likely this effect was not due to the compound itself but to the formation of its metabolite, 13, 14-dihydro-PGF_2α.     

Chemotactic activity of thromboxane B2, prostaglandins and their metabolites for polymorphonuclear leucocytes

(1) The chemotactic activities of thromboxane B₂ (TxB₂, PGE₂, PGF_2α, the 15-oxo, 15-oxo-13,14-dihydro and 13,14-dihydro metabolites of PGE₂, PGF_2α, and a metabolite of TxB₂ for polymorphonuclear leucocytes (PMN) have been investigated.(2) Thromboxane B₂ increased the directional migrationm of rat peritoneal PMN at a concentration of 2.0 μg/ml and of human peripheral neutrophils at a concentration of 0.5 μg/ml.(3) Neither PGE₂, PGF_2α nor their metabolites showed chemotactic activity for rat peritoneal PMN.(4) PGF_2α and 15-oxo-13,14-dihydro-thromboxane B₂ showed no chemotactic activity for human peripheral PMN.(5) The possible role of thromboxane B₂ in inflammation is discussed.     

Synthesis and metabolism of prostaglandins E2, F2α and D2 by the rat gastrointestinal tract. Stimulation by a hypertonic environment in vitro

Whole cell preparations of rat stomach corpus, jejunum, and colon were incubated and the released prostaglandin E₂ (PGE₂), PGF_2α, PGD₂, 15 keto-13,14 dihydro PGE₂, and 15 keto-13,14 dihydro PGF_2α were measured by combined gas chromatography-mass spectrometry. All regions made PGD₂ and possessed a high capacity for producing 15 keto-13,14 dihydro derivatives of both PGE₂ and PGF_2α. Hypertonic sucrose solutions resulted in concentration-dependent increases in prostaglandin release, particularly of PGE₂ and its metabolite. It is suggested that PG's may play a role in the local effects of luminal hyperosomolarity on digestive tract functions.     

The identification of two novel prostaglandins and a thromboxane

6,15-Dioxo-PGF_1α, 6-15-dioxo-13,14-dihydro-PGF_1α and 15-oxo-thromboxane B₂ have been identified in incubates of ram seminal vesicle homogenates with added arachidonic acid or in the perfusate from sensitised challenged guinea pig lungs. These compounds are probably related to 6-oxo-PGF_1α and thromboxane B₂. The structures have been determined by gas chromatography mass spectrometry, following suitable chemical derivatisation.     

Metabolism of 17-phenyl-18,19,20-trinor-prostaglandin F2α in the cynomolgus monkey and the human female

[9β-³H]-17-Phenyl-18,19,20-trinor-PGF₂α was injected subcutaneously into female Cynomolgus monkeys and the structures of six products appearing in the urine were determined. The main urinary metabolites were the dinor- and tetranor-derivatives of 15-keto-13,14-dihydro-17-phenyl-18,19,20-trinor-PGF₂α. Unchanged 17-phenyl-18,19,20-trinor-PGF₂α was also identified among the urinary products, as well as its dinor- and tetranor-derivatives. Finally, the dinor-derivative of 13,14-dihydro-17-phenyl-18,19,20-trinor-PGF₂α was also found in urine. The same six products were also found in urine from human female subjects that had received 17-phenyl-18,19,20-trinor-PGF₂α either subcutaneously or intravenously.Studies on the half-life of the compound in the circulation were also performed in human females. Two less polar metabolites in plasma were identified, viz. 13,14-dihydro-17-phenyl-18,19,20-trinor-PGF₂α and 15-keto-13,14-dihydro-17-phenyl-18,19,20-trinor-PGF₂α.     

Enhanced lipid peroxidation and inflammation during heat exposure in rats of different ages: Role of α-tocopherol

To investigate early events possibly related to the development of heat shock, we examined whether inflammatory-(interleukin-6, tumor necrosis factor α and 15-keto-13,14-dihydro-PGF_2α) and peroxidative-(8-iso-PGF_2α and malondialdehyde) markers are altered during acute heat exposure and aging. We also studied the relationships between inflammatory and peroxidative markers in these settings. In order to prevent these reactions developed as a consequence of the conditions mentioned above, we tested the effects of α-tocopherol. Our results demonstrated that 15-keto-13,14-dihydro-PGF_2α and malondialdehyde in the liver were altered during acute heat exposure in the young and middle-aged rats and could be predicted by changes in the levels of circulatory cytokines. Regardless of age, the supplementation with α-tocopherol prevented changes in the plasma cytokine levels and 15-keto-13,14-dihydro-PGF_2α and malondialdehyde levels in the liver, during acute heat exposure. This study notably emphasized the ability of α-tocopherol to prevent different heat induced mechanisms, involved in induction of inflammatory or peroxidative reactions.     

Prostaglandins and prostaglandin metabolites in human gastric juice

Human gastric juice contains higher concentrations of PG metabolites than of unmetabolized PG indicating that local metabolism might play a role in limiting the biological activity of PG in gastric mucosa and has to be considered when investigating endogenous gastric PG. A major fraction of the 15-keto-13,14-dihydro-PGE₂ (KH₂PGE₂) formed in gastric mucosa and released into the gastric lumen seems to be rapidly dehydrated to a compound co-chromatographing with KH₂PGA₂, while the amounts of the bicyclic degradation product 11-deoxy-13,14-dihydro-15-keto-11,16-cyclo-PGE₂ (11-deoxy-KH₂-cyclo-PGE₂), as measured by radioimmunoassay, in freshly extracted gastric juice are negligible. Stimulation of secretion with pentagastrin does not influence significantly the concentrations of PG and PG metabolites in human gastric juice, but total output tends to increase parallel to the increase in secretion volume. Levels of immunoreactive 6-keto-PGF_1α in human gastric juice are much lower than those of PGE₂. Since human gastric mucosa synthesizes considerable amounts of PGI₂ and 6-keto-PGF_1α in vitro, the low levels of 6-keto-PGF_1α in gastric juice might indicate that PGI₂ formed by gastric mucosa in vivo is, like PGE₂ and PGF_2α, rapidly metabolized and/or removed preferentially via the blood stream.