Influence of chronic antigen exposure on the metabolism of PGH2 by microsomal preparations of airway and parenchymal tissues in the sheep

The effects of repeated antigen exposure on the synthesis of mediators by lung tissues are not well understood. To investigate the influence of antigen challenge on the synthesis of prostaglandins by central airway and peripheral lung tissues, fourteen sensitive sheep underwent biweekly exposure to aerosolized $\text{Ascaris suu}$ antigen (7) or saline (7). Following the fifth exposure, microsomal and high speed supernatant fractions were prepared from trachealis muscle and lung parenchyma. Synthesis of thromboxane (TX) A₂, prostaglandin (PG) D₂ and PGI₂ from the PG endoperoxide intermediate, PGH₂, was assayed over a range of substrate concentrations from 3–200 uM. Synthesis of PGI₂ by trachealis microsomes was approximately 5-fold greater than that of TXA₂. PGI₂ and TXA₂ production was identical in tracheal preparations from Ascaris- and saline-exposed animals. In parenchymal tissues, where TXA₂ production predominated over PGI₂ by 9-fold, preparations from Ascaris- exposed animals synthesized 50% more TXA₂ than controls at PGH₂ concentrations of 25 uM and above, whereas synthesis of PGI₂ and PGD₂ were similar in preparations from both groups of animals. The density of pulmonary mast cells was decreased by 21% in the Ascaris group, whereas polymorphonuclear leukocyte density was unchanged. These results demonstrate the differential synthesis of TXA₂ and PGI₂ in central airways and peripheral lung regions of the sheep. They further indicate that repeated exposure of the airways to antigen selectively enhances TXA₂ synthesis in the lung periphery of sensitized animals. The site of this increased enzymatic activity, whether in resident cells or newly-infiltrated cells, has not been determined.     

Effect of taurine on arterial, uterine and cardiac PGI2 and TXA2 synthesis in the rat

The influence of taurine (in drinking water for 6 weeks) on PGI₂ and TXA₂ synthesis by some female rat organs was investigated using radioimmunoassay and platelet antiaggregatory bioassay. Taurine 100 and 200 mg/kg/day increased aortic PGI₂ release from 0.59 ± 0.04 (control) to 0.85 ± 0.05 and 1.01 ± 0.06 ng/mg, respectively and that by the myometrium from 0.24 ± 0.02 (control) to 0.38 ± 0.01 and 0.50 ± 0.04 ng/mg wet tissue, respectively (P < 0.05, n = 6). It did not affect PGI₂ and TXA₂ production in the heart or TXA₂ in the aorta. Taurine 200 mg/kg depressed uterine TXA₂ synthesis from 148.6 ± 9.8 (control) to 85.4 ± 6.8 pg/mg (P < 0.05, n = 6). Furthermore taurine 0.4 and 0.8 mM in vitro stimulated PGI₂ released by the myometrial and aortic tissues from pregnant rats. The stimulant effect of taurine on PGI₂ may be related to its antioxidant effect whereas its inhibitory effect on uterine TXA₂ may result from direction of synthesis towards PGI₂. It is concluded that endogenous taurine may participate in regulation of PGs synthesis and that prostanoids may contribute to its known actions. On broad basis, taurine-induced release of PGI₂ may prove of potential value in those ailments characterised by deficiency in PGI₂ release.     

The effects of thyroxine and methimazole treatment on the synthesis of prostacyclin (PGI2) in the rat

The effects of thyroxine (T4) and methimazole administration on plasma prostacyclin (PGI₂) levels $\text{in vivo}$ and on PGI₂ release by aortic rings incubated $\text{in vitro}$ were investigated in rats. Male rats were given single injection of T4 (200 μg/100 g body wt) ip every 24 h for either 3, 7 or 14 days for hyperthyroid rats. For hypothyroid rats, a group of rats were given methimazole (0.01 % in drinking water) for 14 days. PGI₂ concentrations were determined in plasma and also in the medium in which aortic rings were incubated. PGI₂ was measured as 6-keto-PGF1α by RIA. Plasma PGI₂ levels in T₄-treated groups were found to be significantly higher than those of control animals. Aortic rings obtained from rats given single injection of T₄ for 7 and 14 days showed significant increases in release of PGI₂ into the incubation medium. In contrast, rats given methimazole for 14 days showed a significant decrease in the production of PGI₂ by aortic rings without any significant changes in plasma levels. Direct addition of T₄ into the incubation medium did not cause any significant changes in PGI₂ release by aortic rings obtained from control rats.These results suggest the regulatory role of thyroid hormone in PGI₂ synthesis $\text{in vivo}$.     

Effect of the progesterone antagonist RU486 on human myometrial spontaneous contractility and PGI2 release

We studied the effect of antiprogesterone RU 486 on spontaneous uterine contractility and PGI₂ release with human myometrial strips superfused “in vitro”. A decrease of PGI₂ release into the superfusion medium was observed after 20 min superfusion. The inhibition was dose-dependent and reversible. After 20 min washing with tyrode medium without RU 486, the uterine strips recovered their initial rate of release. R5020, a progesterone agonist, did not affect PGI₂ release nor dexamethasone and testosterone. Parallel to the decrease of PGI₂ observed during RU 486 superfusion, the uterine spontaneous contraction frequency decreased, while the amplitude and duration of contractions increased. The alteration of uterine contractility was also rapid, dose-dependent and reversible. Modifications of uterine strip spontaneous contractility, similar to those induced by RU 486, were also observed with superfusions of R5020 at concentrations as low as 10⁻⁹M, dexamethasone (10⁻⁸M), but not with superfusions of testosterone. These observations are not in favour of a progesterone-receptor mediated effect of RU 486 in our model. The mechanism of action may be related to the antiprogesterone specific structure i.e. the bulky substituent at the C-11 position. The RU 486 effect on uterine strip contractility, mimicked by other steroids, could point to a non-specific lipid/membrane interaction. However, the fact that testosterone did not affect motility, may indicate a possible specificity of steroids having a 3 oxo pregnene structure.     

A comparison of the effects of PGI2, PGE2 and PGH2 on the cyclic nucleotide levels in rat anterior pituitary glands

Rat anterior pituitary explants were incubated with PGI₂, PGH₂ and PGE₂ in the presence of theophylline (1mM) and the production of cyclic AMP was measured. PGE₂ was found to be about 20 times more potent than PGI₂ while PGH₂ was slightly more effective than PGI₂. The results suggest that PGI₂ does not play a physiological role in cyclic AMP mediated events in the rat anterior pituitary.     

A comparison of the effects of PGI2, PGE2 and PGH2 on the cyclic nucleotide levels in rat anterior pituitary glands in vitro

Rat anterior pituitary explants were incubated with PGI₂, PGH₂ and PGE₂ in the presence of theophylline (1mM) and the production of cyclic AMP was measured. PGE₂ was found to be about 20 times more potent than PGI₂ while PGH₂ was slightly more effective than PGI₂. The results suggest that PGI₂ does not play a physiological role in cyclic AMP mediated events in the rat anterior pituitary.     

PGI2-specific antibodies administered in vivo suggest against a role for endogenous PGI2 as a circulating vasodepressor hormone in the normotensive and spontaneously hypertensive rat

Immunoglobulins raised against 5,6-dihydro PGI₂ crossreact with PGI₂. When infused $\text{in vivo}$ into the rat, these immunoglobulins are capable of I) neutralising the vasodepressor effects (bolus or continuous infusion) of exogenous PGI₂, 2) blocking the catabolism of exogenous ³H-PGI₂ and prolonging its life-time in the circulation (t$\text{1}\text{2}$ approx 60 min) while that of ³H-PGE₂ is unaffected, 3) trapping an endogenously produced substance which after extraction from blood and dissociation from the ligand-antibody complex, is immunoreactive with 6-keto PGF_1α-specific antiserum. Yet the anti-5,6-dihydro PGI₂ immunoglobulins have no effect on resting arterial blood pressure both in the normotensive and spontaneously hypertensive rat. These experiments indicate that endogenously produced PGI₂ does not play a significant $\text{systemic}$ role in blood pressure control although in combination with other vasodilators it could still participate in the regulation of vascular tone at a local level.     

The effect of prostaglandin I2 on the contractility of the term pregnant human myometrium

Small myometrial strips were dissected from the upper and lower segments of the term pregnant human uterus. The specimens were superfused in organ chambers and contractile activity was recorded isometrically.In strips from the upper segment, prostacyclin (PGI₂), induced an initial excitatory response followed in the majority of experiments by transient inhibition. In the lower segment the response was generally the same although direct inhibition without initial stimulation occurred in some cases.During the period of inhibition the specimens were refractory to iterated exposure to PGI₂. Furthermore, during this period of PGI₂-induced inhibition the muscle strip was also refractory to PGE₂ but responded to PGF_2α and oxytocin by stimulation.After inhibition of spontaneous contractile activity induced by indomethacin PGI₂ induced an excitatory response.The results do not indicate any critical change in the myometrial responsiveness of the upper uterine segment to PGI₂ during labor. In strips from the lower segment obtained before labor there tended to be a dominance of non-responders and inhibition only as compared to the results during labor. Nevertheless, whether or not PGI₂ under physiological or pharmacological conditions has any significant influence on the contractility of the term pregnant human uterus, still remains obscure.As judged from earlier reports from our laboratory and the present study it is evident that the uterine vessels are considerably more sensitive to the action of PGI₂ than the myometrium.     

The effect of prostacyclin on intestinal ion transport in the rat

The actions of PGI₂ and PGE₂ on electrically monitored ion transport in rat jejunum and colon have been determined both $\text{in vivo}$ and $\text{in vitro}$. Whilst PGE₂ was shown to induce a marked change in ion transport PGI₂ was relatively ineffective. The ability of the prostanoids to influence ion transport is related to their capacity to change mucosal cyclic AMP levels since in isolated small intestinal enterocytes PGE₂ caused a marked stimulation in cyclic AMP levels whilst PGI₂ had little effect. In colonic mucosal scrapes PGE₂ was more effective than PGI₂ in stimulating changes in cyclic AMP levels. It appears doubtful that PGI₂ plays a direct role in the regulation of intestinal ion transport.     

The antagonism by anti-inflammatory analgesics of prostaglandin F2α-induced contractions of human and rabbit myometrium in vitro

The anti-inflammatory analgesic drugs, aspirin, indomethacin, phenylbutazone, mefenamic acid, ibuprofen and flurbiprofen are shown to inhibit in a dose-dependent manner the force of contraction of isolated human pregnant myometrial strips which have been stimulated to contract by adding prostaglandin (PG) F_2α to the tissue bath. These drugs and also flufenamic acid and salicin show a similar antagonism of the action of PGF_2α with isolated rabbit non-pregnant myometrium. The ratio of the inhibitory concentration $\text{in vitro}$ to the maximum plasma level after a normal dose $\text{in vivo}$ suggests that phenylbutazone and possibly ibuprofen may be capable of inhibiting human uterine contractions $\text{in vivo}$. Patients who were treated with aspirin during induction of abortion using PGF_2α during the second trimester of pregnancy showed no significant change in the induction-abortion interval compared with patients not taking aspirin.     

Modulation of platelet thromboxane A2 and arterial prostacyclin by dietary vitamin E

Platelets from vitamin E-deficient and vitamin E-supplemented rats generate the same amount fo thromboxane A₂ (TxA₂) when they are incubated with unesterified arachidonic acid. Platelets from vitamin E-deficient rats produce more TxA₂ than platelets from vitamin E-supplemented rats when the platelets are challenged with collagen. Arterial tissue from vitamin E-deficient rats generates less prostacyclin (PGI₂) than arterial tissue from vitamin E-supplemented rats. The vitamin E effect with arterial tissue is observed when the tissue is incubated with and without added unesterified arachidonic acid. These data show that arterial prostacyclin synthesis is diminished in vitamin E-deficient rats. Vitamin E, $\text{in}$$\text{vivo}$, inhibits platelet aggregation both by lowering platelet TxA₂ and by raising arterial PGI₂.     

The effects of prostacyclin (PGI2) and 6-keto-PGF1α on bovine plasma progesterone and LH concentrations

Injections of 1 mg PGI₂ directly into the bovine corpus luteum significantly increased peripheral plasma progesterone concentrations within 5 min. Concentrations were higher in the PGI₂-treated heifers than in saline-injected controls between 5 and 150 min and at 3.5, 4, 5, and 7 h post-treatment. Levels tended to remain elevated through 14 h. Saline and 6-keto-PGF_1α were without effect on plasma progesterone levels. The luteotrophic effect of PGI₂ was not due to alterations in circulating LH concentrations. An $\text{in vitro}$ experiment assessed the effects of either PGI₂ alone or in combination with LH on progesterone production by dispersed luteal cells. Progesterone accumulation over 2 h for control, 5 ng LH, 1 μg PGI₂, 10 μg PGI₂, and 10 μg PGI₂ plus 5 ng LH averaged 99 ± 42, 353 ± 70, 152 ± 35, 252 ± 45, and 287 ± 66 ng/ml (n=4), respectively. Thus PGI₂ has luteotrophic effects on the bovine CL both $\text{in vivo}$ and $\text{in vitro}$.     

Effects of prostaglandin PGE2 on the synthesis of placental proteins and human placental lactogen (HPL)

The biosynthesis of placental proteins and placental lactogen (HPL) was studied $\text{in vitro}$ in 10–12 week, 16–18 week and term human placenta in the presence and absence of PGE_2α. The highest ¹⁴C-leucine incorporation was detected in 10 to 12 weeks old placentas. Addition of PGE_2α to the induction medium depressed the rate of incorporation of ¹⁴C-leucine into placental proteins on a dose dependent manner. Placentas most sensitive to this action of PGE_2α were those obtained at 18 weeks gestation followed by placentas at term. $\text{In vivo}$ application of PGE_2α for tharapeutic induction of abortions resulted in the marked inhibition of placental protein synthesis $\text{in vitro}$.     

Potent bronchoconstrictor effects of aerosolized prostaglandin D2 in dogs

Previously, we demonstrated that prostaglandin D₂ (PGD₂), a natural product of the endoperoxide PGH₂, evoked bronchoconstriction when given I.V. to dogs (PROSTAGLANDINS $\text{13}$:255–269, 1977). The present investigation in anesthetized dogs demonstrated that aerosols of PGD₂ (0.001–0.1%) produced concentration-dependent increases in pulmonary resistance (R_L) and decreases in dynamic lung compliance (C_DYN) which were short-lived and equipotent to PGF_2α. These alterations in pulmonary mechanics were partially, yet significantly, inhibited by atropine, thereby suggesting that at least a portion of the bronchoconstriction may be cholinergically mediated. Concomitant cardiovascular depressant effects of both PGD₂ and PGF_2α aerosols were much less and more variable than their bronchopulmonary effects.These results demonstrate a potent bronchoconstrictor effect of aerosolized PGD₂ in dogs. PGD₂ warrants further attention as a possible mediator of the bronchospasm seen in acute, reversible airways obstruction.     

Prostaglandin and thromboxane production by fibroblasts and vascular endothelial cells

We have compared the production of prostaglandins in fibroblast-like cells and endothelial cells in culture. Of the fibroblasts studied $\text{10T}\text{1}\text{2}$, SHE, BP6T and KD produce significant amounts of PGI₂, PGE₂ and PGF_2F2 under optimal culture conditions, but only 3T3 and BHK produce TxA₂ in addition to PGI₂. The adult bovine aortic endothelial cells (ABAE) and fetal bovine heart endothelium (FBHE) synthesise PGI₂ but not TxA₂, either from endogenous or exogenous substrates. Both cultured endothelial cells and fibroblasts apparently lack 15-hydroxyprostaglandin dehydrogenase pathway and the ability to convert 6-Keto PGF_1α into 6-Keto PGE₁. PGI₂ production by ABAE was 3–5 times that of FBHE, about twice that of SHE cells and 6–8 times that of $\text{10T}\text{1}\text{2}$ or BP6T cells. Supernatants or media obtained from these cells inhibited aggregation of human platelet-rich plasma, a known biological effect of PGI₂. This effect was abolished when cell monolayers were preincubated with indomethacin or tranylcypromine. RIA and chromatographic data of 6-Keto PGF_1α from these experiments confirmed that the inhibition of platelet aggregation was due to the formation of PGI₂. The production of all prostanoids by endothelial cells or fibroflasts was significantly higher during the exponential phase of growth as compared to confluent monolayers. We propose that fibroblasts $\text{10T}\text{1}\text{2}$ or SHE can serve as useful experimental models for the study of metabolism and transport of PGI₂ and/or TxA₂ in cells of nonendothelial nature.     

The action of prostacyclin (PGI2) on the contractility of the isolated circular and longitudinal muscle layers of the human oviduct

Small strips from the circular and longitudinal muscle layers of the ampullary-isthmic portion of the human oviduct were mounted in organ chambers for recording of their spontaneous contractility. Concentrations in the order of 1–300 ng/ml of PGI₂ were tested and compared with similar concentrations of PGE₂ and PGF_2α. It was found that PGI₂ contracted the longitudinal muscle layer in the same manner as did PGE₂. The spontaneous activity of the circular layer was markedly suppressed by PGE₂ but only moderately inhibited by PGI₂ even at high concentrations.     

Anti-platelet aggregating and disaggregating activities of 6,9-methano PGI2

Anti-platelet aggregating and disaggregating activities of the chemically stable 6,9-methano prostaglandin I₂ (6,9-methano PGI₂) were investigated. 6,9-Methano PGI₂ inhibited ADP-induced platelet aggregation in PRP from humans, rabbits and rats. 6,9-Methano PGI₂ also inhibited rabbit platelet aggregation induced by ADP, collagen, thrombin, arachidonic acid and 11,9-epoxy-methano PGH₂. Antiaggregating activities of 6,9-methano PGI₂ were 0.3 to 2.0 times greater than those of PGE₁. 6,9-Methano PGI₂ facilitated platelet disaggregation in a dose related manner. Antiaggregating and disaggregating activities of 6,9-methano PGI₂ were markedly enhanced by incubation with the phosphodiesterase inhibitor, theophylline.     

Effects of hydrocortisone on the hypercalcemia and plasma levels of 13,14-dihydro-15-keto-prostaglandin E2 in mice bearing the HSDM1 fibrosarcoma

In mice bearing the prostaglandin-producing HSDM₁ fibrosarcoma, the plasma concentration of 13,14-dihydro-15-keto-PGE₂ was elevated before the development of hypercalcemia, and the magnitude of the rise was greater than that of PGE₂. When hydrocortisone, which inhibits synthesis of PGE₂ by HSDM₁ cells in culture, was administered to tumor-bearing mice, the steroid hormone prevented the rises in plasma PGE₂ metabolite and calcium concentrations. At the dose levels used, hydrocortisone did not inhibit the calcium-mobilizing action of parathyroid hormone $\text{in vivo}$ or the bone resorption-stimulating activity of $\text{PGE}{}_2\text{in vitro}$. These findings are consistent with our hypothesis that the hypercalcemic syndrome in HSDM₁ tumor-bearing mice is due to the secretion of PGE₂ by the tumor.     

The interaction of ethanol and exogenous arachidonic acid in the formation of leukotrienes and prostaglandin D2 in mastocytoma cells

The present studies were undertaken to examine the hypothesis that ethanol could effect cellular biosynthesis in the murine mastocytoma cell of prostaglandins and leukotrienes, oxidative metabolites of arachidonic acid, at concentrations that could be encountered $\text{in vivo}$ as well as during $\text{in vitro}$ experiments. The effects of ethanol which encompass these concentration ranges (200–1000 mg%) can be summarized as follows: first in the absence of exogenous arachidonic acid, ethanol caused a dose dependent decrease in the production of leukotrienes which was statistically significant at 200 mg%. At 1000 mg%, ethanol caused a 20–50% decrease in leukotrienes and a 21% decrease in the amount of prostaglandins D₂ (PGD₂) formed in these cells. Secondly, when cells were incubated with exogenous arachidonic acid (14 μg/ml), large increases in both PGD₂ and leukotrienes occurred. Under these conditions, ethanol caused a further increase in the amount of leukotrienes and a small increase in the amount of PGD₂ formed. This stimulatory effect was specific for ethanol since neither t-butanol nor n-butanol caused the enhanced production of leukotrienes with exogenous arachidonic acid. Thus, these experiments sugsests that ethanol affects metabolsim of arachidonic acid at reasonably low doses (200–400 mg%) of ethanol in a manner dependent on the free arachidonic acid in the tissue. Also, $\text{in vitro}$ experiments in which ethanol is used as a solvent for arachidonic acid could be greatly affected by high levels of ethanol (500–1000 mg%) which are frequently utilized.     

Prostaglandins and myogenic control of tension in lower esophageal sphincter in vitro

Active tension is produced by the lower esophageal sphincter (LES) of North American opossum $\text{in vitro}$ by a myogenic mechanism. Strips of LES, but not those from the esophageal body, contracted to prostaglandin (PG)F_2α, stable expoxymethano derivatives of PGH₂ and to thromboxane B₂. Stable endoperoxides were more than 500 times more potent than PGF_2α. PGI₂ and 6-keto PGF_1α were weak relaxants of LES strips. LES strips transformed arachidonic acid into contractile substances. This transformation was prevented by agents which interfere with PG synthesis by inhibiting cyclo-oxygenase [indomethacin (IDM), 5,8,11,14-eicosatetraynoic acid (ETA) or thromboxane synthetase [imidazole]. Tranylcypromine 500 μg/ml also inhibited contractions to arachidonic acid. These agents also reduced muscle tone, so that endogenous PG formation may contribute to active tension in the LES. ETA and IDM increased tone before inhibiting it, and this effect was prevented by prior treatment with ETA or imidazole. There may also be an endogenous PG which inhibits LES tone. The possibility that this may be PGI₂ is discussed.