15-methylation augments the cardiovascular effects of prostaglandin F2α

Intramuscular injection of the 15-methyl analogue of prostaglandin F_2α (15-me-PGF_2α) is being used to initiate second trimester abortion. The natural prostaglandin F_2α (PGF_2α) is a known pulmonary pressor agent but there is little information about the cardiovascular effects of the analogue. Consequently, we compared the hemodynamic responses to the two forms in twenty-three anesthetized dogs. Given I.M. or I.V. 15-me-PGF_2α produced a greater and more sustained rise in pulmonary arterial pressure than PG F_2α. Intramuscular 15-me-PGF_2α also elicited a more prolonged increase in pulmonary vascular resistance than prostaglandin F_2α given I.M. or I.V. The methyl analogue (I.M. or I.V.) causes a greater initial fall in systemic arterial oxygen tension and cardiac output, and a greater increase in systemic resistance than I.M. PG F_2α Breathlessness seen during abortion induced by prostaglandin F_2α or its methyl analogue may be caused by acute pulmonary hypertension in addition to bronchoconstriction.     

The 8-isoprostaglandins: Evidence for eight compounds in human semen

Evidence is presented for the existence of a group of 8-iso prostaglandins in human semen, comprising 8-iso PG E₁^*, 8-iso PG E₂, 8-iso PG F_1α, 8-iso PG F_2α and the four corresponding 19-hydroxy prostaglandins. The E and F compounds have been positively identified by comparison of their mass spectra and chromatographic properties with those of authentic standards. Preliminary measurements of levels of these compounds in pooled semen are presented.     

Effect of endotoxin administration in pregnant camels

Intravenous administration of Escherichia coli endotoxin at a dose of 0.05 μg/kg bodyweight to pregnant camels resulted in abortion. The injection of endotoxin caused significant increases in the plasma concentration of 13,14-dihydro-15-prostaglandin F_2α, the metabolite of prostaglandin F_2α (PG F_2α) and cortisol and a significant decrease in the concentration of progesterone. It is suggested that endotoxin caused abortion in camels was a consequence of endotoxin induced PG F_2α secretion resulting in luteal regression and decreased progesterone concentration.     

Differential effects of detergents and dimethylsulfoxide on membrane prostaglandin E1 and F2α receptors

Pretreatment of membranes for 1 hr at 4° with up to 0.1% Triton X-100 (TX-100) and sodium desoxycholate (SDC), resulted in a greater loss of [³H] prostaglandin (PG)F₂α binding compared to E₁ binding. Lubrol WX (LWX) tended to cause a greater loss of [³H]PGF₂α than E₁ binding. However, the differential loss was not as marked as with TX-100 or SDC. Triton X-305 was relatively ineffective, but loss of [³H]PGE₁ binding was greater than for PGF₂α. Increasing concentrations of dimethylsulfoxide (DMSO) progressively inhibited PGF₂α binding without affecting PGE₁ binding. The detergent, but not DMSO, induced losses of membrane PG binding were due to solubilization of the receptors. Greater amounts of membrane protein and phospholipids were solubilized at detergent (TX-100 and SDC) concentrations that solubilized 100% of PGE₁ receptors compared to 100% solubilization of F₂α receptors. Neither the duration of preincubation nor the amount of membrane protein chosen were responsible for differential PGE₁ and F₂α receptor losses. These differential membrane PG receptor losses raise the possibility of differences in PGE₁ and F₂α receptors association with the membrane structure.     

Determination of prostaglandin F2α, E2, D2 and 6-keto-F1α in human cerebrospinal fluid

Prostaglandin (PG)F_2α, E₂, D₂ and 6-keto-F_1α were determined in human cerebrospinal fluid by a mass spectrometric technique. The samples were obtained from 12 patients with suspected intracranial disease. A 64 fold variation in PG levels was observed. The major PG was 6-keto-F_1α (0.12–15 ng/ml). PGF_2α and PGE₂ were present in lower concentrations PGD₂ was below the level of detection (0.05 ng/ml) except in one patient with extremely high total levels of PGs.     

Kinetic changes in rat renal 15-hydroxy-prostaglandin dehydrogenase induced by chronic ethanol exposure

Several lines of investigation have suggested that exposure to ethanol may lead to alterations in both the synthesis and degradation of the E and F series of prostaglandins (PG). It has been suggested that these changes in PG metabolism underlie cartain of the pathophysiological consequences of chronic alcoholism but few data re availalbe as to the mechanism resonsible for these changes. We now report that chronic exposure to ethanol in moderate doses (17% total dietary calories as ethanol) and high doses (35% total dietary calories as ethanol) results in a concentration dependent loss of renal 15-hydroxy-prostaglandin dehydrogenase for the NAD mediated reactioins. Soluble fractions of kidney homogenates in teh presnet of > 10 K_m concentratons of NAd exhibited dose dependent loss of specific and total organ PG dehydrogenase activity toward PGE₂ and F_2α. A similar dose dependent decrease in the V_max of the NAD mediated reaction was measured for the oxidation of PGE₁, E₂, and F_2α. Moderate doses of ethanol resulted in an increase in the K_m for PGE₂ and F_2α. K_m values for the NADP mediated reactions were not significantly influenced by exposure to high doses of ethanol other than for PGE₁. These data suggest that chronic ethanol consumption results in a dose dependent, selective inhibition of the metabolism of PGs of the “2” series by the renal PGDH enzyme which utilizes NAD.     

Maternal and fetal prostaglandin concentrations during late gestation in dairy cattle

A study was conducted to measure the blood plasma concentrations of prostaglandin F_2α (PGF_2α), 13,14-dihydro-15-keto-prostaglandin F_2α (PGFM), prostaglandin E₂ (PGE₂) and 13,14-dihydro-15-keto-prostaglandin E₂ (PGEM) in the jugular vein, umbilical vein and artery and uterine vein of 18 Holstein Friesian cows during late gestation. A caesarean section was performed on all cows before term in order to obtain blood samples from the different sources. Plasma PG concentrations in the uterine or fetal circulation were significantly higher than in jugular vein plasma. Correlations between peripheral PG metabolite concentrations and primary PG concentrations in the various sources of the uterus or fetus were not significant (r = .17 − .47) and demonstrated that prostaglandin values based upon peripheral blood alone are of limited value.     

Effect of prostaglandins F2α and E2 on milk ejection, blood pressure and blood flow through the mammary artery in the cow

The influence of prostaglandins (PG) F₂α and E₂ on milk ejection, mammary artery blood flow and arterial blood pressure was studied in lactating cows. Injections of both PG in the jugular vein or the carotid artery induced milk ejection after a relatively long latency period. The minimal effective dose amounted to 1 to 5 μg and to 100 to 300 μg for PGF₂α and PGE₂ respectively. In several cases with PGF₂α and once with PGE₂ milk ejection was accompanied with a simultaneous increase in blood flow through the mammary artery whereas arterial blood pressure remained unchanged. Both routes of administration showed the same response. It was suggested that the effect of the PG on the bovine myoepithelium is indirect, possibly secondary to a release of oxytocin from the neurohypophysis.     

Prostaglandin profiles in nervous tissue and blood vessels of the brain of various animals

The endogenous formation of prostaglandin (PG) D₂, E₂, F_2α, and 6-keto-PGF_1α was determined in homogenates of mouse, rat, and rabbit brain, and of rat cerebral blood vessels, using gas chromatography mass spectrometry. In all species tested, 6-keto-PGF_1α could be identified in the brain homogenates, but was a minor component in relation to other PGs. In contrast 6-keto-PGF_1α was the most abundant PG in the blood vessels, being present in about 40-fold higher levels than in the brain tissue. PGD₂ was the most abundant PG in rat and mouse brains, but was below detection limits in the analyzed blood vessels. These studies indicating differential metabolism of PG endoperoxides in nervous and vascular tissue, provide a biochemical basis for further studies on the role of the PGs in brain circulation and neuronal activity.     

Effects of PGF2α and PGF2α, 1–15 lactone on the corpus luteum and on early pregnancy in the rhesus monkey

The effects of prostaglandin (PG)F_2α and PGF_2α, 1–15 lactone were compared in luteal phase, non-pregnant and in early pregnant rhesus monkeys. Animals treated with either PG after pretreatment with human chorionic gonadotropin (hCG) had peripheral plasma progesterone concentrations that were not statistically different from those in animals treated with hCG and vehicle. However, menstrual cycle lengths in monkeys treated with PGF_2α, 1–15 lactone were significantly (P <0.02) shorter than those in vehicle treated animals. In the absence of hCG pretreatment, plasma progesterone concentrations were significantly (P <0.008) lower by the second day after the initial treatment with either PGF_2α or PGF_2α, 1–15 lactone than in vehicle treated monkeys. Menstrual cycle lengths in monkeys treated with either PG were significantly (P <0.04) shorter than those in animals treated with vehicle. There were no changes in plasma progesterone concentrations in early pregnant monkeys treated with PGF_2α, and pregnancy was not interrupted. In contrast, plasma progesterone declined and pregnancy was terminated in 5 of 6 early pregnant monkeys treated with PGF_2α, 1–15 lactone. These data indicate that PGF_2α, 1–15 lactone decreases menstrual cycle lengths in non-pregnant rhesus monkeys. More importantly, PGF_2α, 1–15 lactone terminates early pregnancy in the monkey at a dose which is less than an ineffective dose of PGF_2α.     

Activity of prostaglandin E, F, A and B on sphincter, dilator and ciliary muscle preparations of the cat eye

Both sphincter and dilator muscle preparations of the cat iris contract to prostaglandins; F_2α and E₂ are the most potent and A₁ and B₁ the least. Ciliary muscle strips relax to PG's provided that the strips are precontracted. E₁, E₂ and often F_2α are more potent relaxants than the remaining PG's. The effects of PG's are not altered by α or β blockade nor by atropine; however, propranolol blocks the PG induced relaxation of the ciliary muscle. The effects of PG's on the sphincter are antagonized by catecholamines; but the latter act synergistically in contracting the dilator and in relaxing the ciliary muscle. Indomethacin markedly potentiates the effects of PG's on all three muscle preparations.     

Determination of 9α,11β-prostaglandin F2 by stereospecific antibody in various rat tissues

In view of the recent finding that prostaglandin D₂ is stereospecifically converted to 9α,11β-prostaglandin F₂, an isomer of prostaglandin F₂α, a highly specific and sensitive radioimmunoassay for 9α,11β-prostaglandin F₂ was developed and applied to determine the content of this prostaglandin in various rat tissues. Antisera against 9α-11β-prostaglandin F₂ were raised in rabbits immunized with the bovine serum albumin conjugate, and [³H]9α,11β-prostaglandin F₂ was enzymatically prepared from [³H]prostaglandin D₂. The assay detected 9α,11β-prostaglandin F₂ over the range of 20 pg to 1 ng, and the antiserum showed less than 0.04% cross-section with prostaglandin F₂α, prostaglandin F₂β and 9β,11β-prostaglandin F₂. To avoid postmortem changes, tissues were frozen in liquid nitrogen immediately after removal. The basal level of 9α,11β-prostaglandin F₂ was hardly detectable in various tissues of the rat examined, including spleen, lung, liver and brain; although it was found to be 0.31 ± 0.06 ng/g wet weight in the small intestine. During convulsion induced by pentylenetetrazole, enormous amounts of prostaglandin D₂ (ca. 180 ng/g wet weight) and prostaglandin F₂α (ca. 70 ng/g) were produced in the brain; however, 9α,11β-prostaglandin F₂ was detected neither there nor in the blood. This result demonstrates that the conversion to 9α,11β-prostaglandin F₂ is a minor pathway, if one at all, of prostaglandin D₂ metabolism in the rat brain.     

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.     

Comparative studies of prostaglandins F2α and E2 in late cyclic and early pregnant sheep: synthesis by endometrium and conceptus effects of indomethacin treatment on establishment of pregnancy

Endometrial concentrations of prostaglandins F₂α (PGF₂α) and E₂ (PGE₂) were measured by specific radioimmunoassay in sheep, on day 14 of estrous cycle or pregnancy, during luteolysis (Day 16 of the cycle), and after implantation (Day 23 of pregnancy) : concentrations observed on day 14 of cycle and pregnancy were similar. During luteolysis, on day 16 of cycle, a consistent drop was noticed. If luteal regression did not occur, as a consequence of the presence of an embryo, endometrial concentrations of PGF₂α on day 23, were twice those of day 14, and PGE₂ remained unchanged. 2 hour incubations of endometrial caruncular tissue from 14 days cyclic or pregnant ewes resulted in de novo synthesis of PG which could be increased by Arachidonic Acid and inhibited by Indomethacin; during the first 30 min of incubation, the PGF₂α synthesis was comparable for both endometrial tissues, whereas PGE₂ synthesis was twice as great in pregnant endometrium. Fourteen and 23 day conceptuses had high PGF₂α and PGE₂ concentrations which were not due to maternal PG sequestration : PG synthesis which could be inhibited by Indomethacin was observed in incubated 14 day old embryos. Treatment of pregnant ewes from day 7 to day 22 after mating, either with Indomethacin (300 mg s.c. daily) or with Acetylsalicylic Acid (1 g I.V. daily) resulted in a sharp diminution of endometrial PG concentration and release, with no apparent effect on the establishment of pregnancy. These results tend to ascribe a less important role to PG during early pregnancy in sheep as compared with rodents, in terms of embryonic growth and implantation.     

Prostaglandins and oxytocin: Their effects on uterine smooth muscle

A subcellular fraction was isolated from uteri of non-pregnant and pregnant cows. ATP-dependent calcium binding was shown to take place in this fraction. This calcium binding was inhibited in a dose related fashion when increasing amounts of prostaglandin (PG) E₂ or F_2α were added to the in vitro experimental medium. The physiologically inactive PGF_1β had no inhibitory effect. Oxytocin caused inhibition of calcium binding in preparations from both pregnant and non-pregnant cows. The response to PGE₂ and PGF_2α was somewhat greater in preparations from pregnant uteri than from non-pregnant uteri. The response to oxytocin was very much greater in pregnant uteri. Because of the high PG sensitivity of calcium binding in preparations from the non-pregnant uterus, it is concluded that the PGs may be the more suitable agent in the control of reproduction.     

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.     

Surface behavior of prostaglandins (PGs)

This study establishes some correlations between molecular structure and surface function of six prostaglandins in a model membrane system. Using spread films at the air/water interface, we determined surface pressure and surface potential of PGs A₁, A₂, E₁, E₂, F_1α and F_2α. All the PGs formed films with low pressure (0 to 9 dyne/cm) and relatively low surface potentials (ΔV = 10 to 250 mV). On 0.15 M NaCl, the π and ΔV values were in the order E₁ > F_1α > A₁ > F_1α > A₁ > F₂α = A₂ > E₂ and F_1α > E₁ > A₁ > A₂ > F_2α > E₂ respectively. Clearly, the cis unsaturation in the carboxylic chain of the PG₂ series conferred greater instability to the films, as indicated by the lowest π and ΔV values. Also, members of the PG₁ series penetrated films of dipalmitoyl phosphatidyl choline (DPPC) better than PG₂ did, the ablest being E₁ (Δπ = 12 dyne/cm) and the poorest F₂α (Δπ = 2 dyne/cm); penetration of E₁ and F₂α was independent of the initial pressure (π_i) of the DPPC film, whereas with A₁ and F_1α Δπ decreased as π_i increased. The PGs expressed marked discriminating capacities for the electrolyte, as indicated by differences in their π and ΔV responses to Na⁺ and Ca⁺⁺ as well as for the lipid, as indicated by different penetration (Δπ values) into DPPC films.     

Determination of prostaglandin F in blood plasma using chemically modified bacteriophage technique

The levels of prostaglandin F found in human and rabbit plasma were determined by the chemically modified bacteriophage assay.Prostaglandin F₂α was coupled covalently to bacteriophage T4 using carbodiimide as cross linking agent and the conjugated phage could be inactivated by anti-prostaglandin F₂α antibodies. Prostaglandins specifically inhibited the modified phage inactivation caused by antiserum and as little as 200 picograms of prostaglandin F₂α could be detected by this system. Anti-prostaglandin F₂α antibodies had a high specificity toward prostaglandin F₂α and exhibited a very low degree of cross reaction to the other prostaglandin derivatives. The concentration of the extracted prostaglandin F₂α from the plasma containing exogenous prostaglandin F₂α could be determined with a high recovery.In normal human males and in male rabbits, 0.300.82 and 0.421.22 nanograms of prostaglandin F per ml of plasma were found, respectively. These levels of prostaglandin F in plasma agree with those determined by the radioimmunoassay.     

Differential pattern of activity of some prostaglandins in diverse superfused tissues

Dose-response curves to prostaglandins (PG) E₁, E₂, F_1α, F_2α, A₁ and A₂ were carried out in the rat stomach fundus, rat colon, chick rectum and rabbit aortic strip superfused with Krebs solution containing adrenergic, cholinergic, serotonergic and histaminergic blockers, and arranged in a cascade fashion. In the stomach and rectum, the order of potency was PGE's > PGF's > PGA's. In the aorta and colon, PG₂'s were more active than their corresponding PG₁'s. By determining the colon/stomach and aorta/colon activity ratios, it was possible to define differential patterns of activity for PGE₁, PGE₂, PGF_1α, PGF_2α and PGA's. The present results indicate the importance of determining complete dose-response curves in the cascade for the characterization of PG-like activity.     

Protein Kinase C-α Interaction with F0F1-ATPase Promotes F0F1-ATPase Activity and Reduces Energy Deficits in Injured Renal Cells

We showed previously that active PKC-α maintains F₀F₁-ATPase activity, whereas inactive PKC-α mutant (dnPKC-α) blocks recovery of F₀F₁-ATPase activity after injury in renal proximal tubules (RPTC). This study tested whether mitochondrial PKC-α interacts with and phosphorylates F₀F₁-ATPase. Wild-type PKC-α (wtPKC-α) and dnPKC-α were overexpressed in RPTC to increase their mitochondrial levels, and RPTC were exposed to oxidant or hypoxia. Mitochondrial levels of the γ-subunit, but not the α- and β-subunits, were decreased by injury, an event associated with 54% inhibition of F₀F₁-ATPase activity. Overexpressing wtPKC-α blocked decreases in γ-subunit levels, maintained F₀F₁-ATPase activity, and improved ATP levels after injury. Deletion of PKC-α decreased levels of α-, β-, and γ-subunits, decreased F₀F₁-ATPase activity, and hindered the recovery of ATP content after RPTC injury. Mitochondrial PKC-α co-immunoprecipitated with α-, β-, and γ-subunits of F₀F₁-ATPase. The association of PKC-α with these subunits decreased in injured RPTC overexpressing dnPKC-α. Immunocapture of F₀F₁-ATPase and immunoblotting with phospho(Ser) PKC substrate antibody identified phosphorylation of serine in the PKC consensus site on the α- or β- and γ-subunits. Overexpressing wtPKC-α increased phosphorylation and protein levels, whereas deletion of PKC-α decreased protein levels of α-, β-, and γ-subunits of F₀F₁-ATPase in RPTC. Phosphoproteomics revealed phosphorylation of Ser¹⁴⁶ on the γ subunit in response to wtPKC-α overexpression. We concluded that active PKC-α 1) prevents injury-induced decreases in levels of γ subunit of F₀F₁-ATPase, 2) interacts with α-, β-, and γ-subunits leading to increases in their phosphorylation, and 3) promotes the recovery of F₀F₁-ATPase activity and ATP content after injury in RPTC.