Action of PGI2 analogs on the pulmonary and systemic circulations in the conscious newborn lamb

Previous work (Lock et al., J. Pharm . Exp. Ther. 215:156, 1980) has shown that conventional screening procedures for vasoactive PGI2 analogs were little value in predicting pulmonary vasodilator activity in the newborn lamb. To gain a better insight into the structural requirements for pulmonary vasoactivity and possibly identify useful compounds for the management of neonatal pulmonary hypertensive disorders, we have tested the following PGI2 analogs in normoxic and hypoxic newborn lambs: 15(S)-9-deoxy-15-methyl-9 alpha,6- nitrilo -PGF1 (analog I); 9-deoxy-9 alpha,5- nitrilo -PGF1 (analog II); (6S, 15S)-15-methyl-PGI2 (analog III); and ( 6R , 15S)-15-methyl-PGI1 (analog IV). A prostaglandin analog mimicking PGI2 (compound BW245C ; (+/-)-5-(6- carboxyhexyl )-1-(3-cyclohexyl-3-hydroxypropyl)hydantoin ) was tested as well. Compounds were injected into a branch pulmonary artery and any local pulmonary effect could be assessed from the change in the ratio of blood flow to the injected lung over total flow. None of the analogs tested proved to be a selective pulmonary dilator. BW245C was a potent peripheral vasodilator (threshold around 0.5 microgram/kg) and indirectly lowered pulmonary vascular resistance through its systemic effects. Analog I also dilated the systemic circulation, but only at the highest dose tested (100 micrograms/kg). The latter finding is surprising because it was previously shown that the parent, non-methylated compound is a fairly potent and selective pulmonary vasodilator. Analog II and IV were inactive at a dose up to, respectively, 30 and 20 micrograms/kg. Analog III, on the other hand, weakly constricted the systemic circulation at a dose of 10 micrograms/kg.(ABSTRACT TRUNCATED AT 250 WORDS)     

Selective antagonism of the systemic vasodepressor response to PGE1 by d,1–11, 15-bisdeoxy PGE1

Several bisdeoxy PGE₁ analogs are potent, competitive antagonists of PGE₁-induced colonic contractions in the gerbil. The efficacy of these analogs in antagonizing PGE₁-mediated systemic vasodepression has not been previously demonstrated. In this study, serial doses of PGs were administered before, during and after infusion of d,1–11, 15-bisdeoxy PGE₁. Bolus injections of PGE₁ (3.0 μk/kg), PGE₂ (3.0 μg/kg) and PGI₂ (0.3 μg/kg) were administered via the right external jugular vein to male Wistar rats. PGE₁, PGE₂ and PGI₂ decreased systemic arterial pressure 41%, 38% and 38%, respectively. The PGE₁ analog was infused (200 μg/kg/min) through the right common carotid artery. The analog itself had no effect on mean systemic arterial pressure, but maximum reversible inhibition (51%) of PGE₁-mediated vasodepression occurred following a 50 minute infusion. No significant effect of the PGE₁ analog was observed on PGE₂ or PGI₂-mediated vasodepression. These data demonstrate the ability to antagonize PGE₁-mediated vasodepression, and to differentiate the vascular responses to PGE₁ and PGE₂ or PGI₂.     

Effects of 13, 14-dehydroprostacyclin methyl ester on the feline intestinal vascular bed

The effects of a stable PGI₂ analog, 13, 14-dehydro-PGI₂ methyl ester and several vasoactive hormones were compared in the feline intestinal vascular bed under conditions of controlled blood flow so that changes in perfusion pressure directly reflect changes in vascular resistance. The PGI₂ analog decreased perfusion pressure in a dose-dependent fashion when injected in the range of dose of 0.03–3 μg and was quite similar to PGE₂ whereas isoproterenol was somewhat more potent as a vasodilator in the feline intestinal vascular bed. The present data show that 13, 14-dehydro-PGI₂ methyl ester has potent vasodilator activity in the intestinal vascular bed.     

Relaxation of isolated human pulmonary muscle preparations with prostacyclin (PGI2) and its analogs

The effects of PGI₂ and two analogs Iloprost and ZK 96480 were examined on isolated human pulmonary muscle preparations. High concentrations of these agents reduced the basal tone in all types of preparations. In addition, they relaxed tissues which had been maximally contracted with histamine (50 μM). PGI₂ was more potent on pulmonary arterial muscle preparations (pD₂ value : 6.33, n = 3) than on bronchial muscles. The relaxations induced by PGI₂ in bronchial preparations were quite variable, that is, some tissues relaxed while others did not. The analogs also relaxed arterial preparations and the pD₂ values were approximately the same (Iloprost : 7.42, n = 4 and ZK 96480 : 7.48, n = 4). The isolated human pulmonary vascular preparations were approximately 10-fold more sensitive to the analogs than bronchial muscle preparations. In bronchial tissues we noted that the PGI₂ relaxant effect was spontaneously reversed with time, an activity not observed with both analogs. A pretreatment of the bronchial tissues with indomethacin (1.7 μM) did not reduce the variations observed with PGI₂ nor modify the transient relaxation observed with this agent. These data demonstrate that vascular tissues from the human lung are considerably more sensitive to these relaxant agonists than bronchial preparations.     

Pharmacological profile of a novel carbacyclin derivative with high metabolic stability and oral activity in the rat

A novel carbacyclin derivative (16S)-13,14-dehydro-16,20-dimethyl-3-oxa-18,18,19,19-tetradehydro-6a-carbaprostaglandin-I₂ (3-oxa-analogue) has been synthesized in order to find chemically and metabolically stable prostacyclin-memetics with a potency equal or even superior to PGI₂.The 3-oxa-analogue was found to be stabilized against β-oxidation, a main metabolic degradation step also for chemically stable PGI₂-analogues. The compound is orally available and displays a long duration of 4,5 – 48 h of antiaggregatory and hypotensive action. The 3-oxa-analogue inhibits ADP-induced platelet aggregation with an IC₅₀ of 3.0 nM. Following intravenous application the 3-oxa-analogue lowers diastolic blood pressure in a dose dependent manner, the ED₂₀ being 0.1 – 0.2 μg/kg after injection and ≤ 0.05 μg/kg/min after infusion respectively. In vivo platelet aggregation is inhibited after i.v. infusion of the 3-oxa-analogue with an IC₅₀ of 0.037 μg/kg/min. As compared to Iloprost, the 3-oxa-analogue is 5 – 12 fold more potent with respect to in vivo hypotensive and anti-aggregatory effects.The results of the present studies indicate that the 3-oxa-analogue has a pharmacological profile comparable to prostacyclin (PGI₂) and Iloprost. Due to the fact that the 3-oxa-analogue is chemically and metabolically stable, long term oral treatment can be achieved in clinical conditions in which PGI₂ and Iloprost have already been shown to be therapeutically useful principles.     

The mechanism of enhancement by fatty acid hydroperoxides of anaphylactic mediator release

Fragments of chopped lung from indomethacin treated guinea-pigs had an anti-aggregating effect when added to human platelet rich plasma (PRP), probably due to the production of prostacyclin (PGI₂) since the effect was inhibited by 15-hydroperoxy arachidonic acid (15-HPAA, 10 μg ml⁻¹). Both 15-HPAA (1–20 μg ml⁻¹ min⁻¹) and 13-hydroperoxy linoleic acid (13-HPLA, 20 μg ml⁻¹ min⁻¹) caused a marked enhancement of the anaphylactic release of histamine, slow-reacting substance of anaphylaxis (SRS-A) and rabbit aorta contracting substance (RCS) from guinea-pig isolated perfused lungs. This enhancement was not reversed by the concomitant infusion of either PGI₂ (5 μg ml⁻¹ min⁻¹) or 6-oxo-prostaglandin F_1α (6-oxo-PGF_1α, 5 μg ml⁻¹ min⁻¹). Anaphylactic release of histamine and SRS-A from guinea-pig perfused lungs was not inhibited by PGI₂ (10 ng - 10 μg ml⁻¹ min⁻¹) but was inhibited by PGE₂ (5 and 10 μg ml⁻¹ min⁻¹). Antiserum raised to 5,6-dihydro prostacyclin (PGI₁) in rabbits, which also binds PGI₂, had no effect on the release of anaphylactic mediators. The fatty acid hydroperoxides may enhance mediator release either indirectly by augmenting thromboxane production or by a direct effect on sensitized cells. Further experiments to distinguish between these alternatives are described in the accompanying paper (27).     

Cardioprotective effect of prostacyclin and 7-oxo-PGI2 in rats against chronic isoproterenol damage

Isoproterenol (ISO) was injected in 5 mg/kg i.p. doses to rats, daily for two weeks. We evaluated the developed myocardial hypoxia and necrosis quantitatively by histological methods. To follow the time course of cardioprotection prostacyclin or 7-oxo-PGI₂ were injected daily, i.p. 5, 30 min and 1, 2, 3, 4 hours before or after the ISO to groups of ten rats, respectively. Cardioprotection was defined as the reduction of necrotized areas and was expressed as percentage change compared to the control (saline treated) group. 1 μg/kg PGI₂ and 50 μ/kg 7-oxo-PGI₂₊ showed nearly equipotent cardioprotection (37.3±7.9% and 38.3±6.8%, respectively). The peak effect of both compounds appeared when injected prior to ISO in the 120. min but the action of 7-oxo-PGI₂ was more prolonged. The different doses of prostacyclin analogs given after the ISO injection were ineffective with the exception of 50 μ/kg 7-oxo-PGI₂ (29.75± 5.2 %).     

Thromboxane and pulmonary hypertension following E. coli endotoxin infusion in sheep: Effect of an imidazole derivative

We assessed the effect of a specific thromboxane synthetase inhibitor (an imidazole derivative) on pulmonary hemodynamics and the concentrations of TxB₂ (TxA₂), 6-keto-PGF_1α (PGI₂), and PGF_2α in pulmonary lymph and transpulmonary blood samples following intravenous administration of E. coli endotoxin (1 μg/kg) in sheep. In control animals the rise in pulmonary artery pressure correlated with increases in plasma and lymph TxB₂ concentrations and large transpulmonary concentration gradients of this metabolite were measured. In imidazle treated animals both pulmonary hypertension as well as increases in plasma and lymph TxB₂ concentrations were substantially reduced. In contrast, peak concentrations of 6-keto-PGF_1α (PGI₂) and PGF_2α were severalfold higher than those measured in control animals. This suggests a shunting of endoperoxide metabolism towards prostacyclin and primary prostaglandins and documents the specificity of the thromboxane synthetase inhibitor. Out study provides evidence that endotoxin-induced pulmonary hypertension is mediated by pulmonary synthesis of TxA₂.     

The central nervous effect of prostaglandins I2, E2 and F2α on aconitine — Induced cardiac arrhythmia in rats

Intracerebroventricular administration of PGI₂ or PGE₂ reduced aconitine-induced cardiac arrhythmia in rats. PGF_2α had no antiarrhythmic effect under the same conditions. The ED₅₀ values of PGI₂ and E₂ were 0.25 μg/kg and 1.1 μg/kg, respectively. Central mechanisms may participate in the antiarrhythmic effect of these PGs.     

The effects of itnravenous ZK36-374, a stsable prostacyclin analogue, on normal volunteers

Prostacyclin (PGI₂) therapy has been evaluated in many vascular diseases. However, it is unstable and a potent vasodilator, able to lower blood pressure. Although such effects may be desirable in some situations, they are unwanted in others. ZK36-374 (Schering AG) is a carbacyclin derivatives with a similar action to PGI₂; however, it is chemically stable and has less of a hypotensive action.We evaluated the effects of a 4-hour I.V. infusion of ZK36-374 at a maximum dose of 2ng/Kg/min. in ten normal volunteers. Prior to the infusion and at 2 and 4 hours, blood was sampled for estimation of platelet aggregation in both platelet rich plasma and whole blood. β-thromboglobulin, 6-keto-PGF_1α and TXB₂ were measuerd by radioimmunoassay, as were other coagulation and rheological tests. The infusion was well tolerated with facial flushing, jaw trismus and some nausea at max dose. Blood pressure and pulse rate were not significantly altered. During infusion of ZK36-374, the rates of platelet aggregation to 2μm AdP and 2μg collagen in PRP were significantly decreased when compared to baseline, as was whole blood aggregation to 2μm ADP and 0.5 μg collagen. βTG also fell significantly, as did the levels of 6-keto-PGF_1α and TXB₂. Fibrinolysis, blood viscosity, and red cell deformability were unchanged.ZK36-374 is an effective anti-platelet agent without major toxic or hypotensive effects.     

Effects of prostaglandin endoperoxide analogs on contractile elements in lung and gastrointestinal tract

Prostaglandin endoperoxides are formed in the lung as intermediate compounds in the biosynthesis of prostaglandins and thromboxanes. The effects of different doses of two analogs of prostaglandin endoperoxide PGH₂ were compared with those of PGF_2α and PGE₂ on superfused preparations of isolated trachea, bronchiole, peripheral lung, pulmonary artery and gastrointestinal smooth-muscle tissues. Endoperoxide analogs induced contraction of all smooth-muscle structures in the lung and airways. Compared to PGF_2α, analog I was approximately 71 times as potent on guinea-pig trachea, 214 times as potent on guinea-pig lung, and 57 times as potent on guinea-pig polmunary artery. Analog II was moderately less potent on all tissues than analog I. On gastrointestinal smooth-muscle organs, the PGH₂ analogs were generally closer in activity to PGF_2α and PGE₂, or even weaker. The findings show that PG endoperoxide vessels, and suggest that the naturally occurring compounds may participate in the mediation of bronchoconstriction and pulmonary vasoconstriction in disease states.     

The effects of a 5-lipoxygenase inhibitor, AA-861, on PGI2-like substance production in guinea-pig lungs

To determine the effects of AA-861 on PGI₂ production in guinea-pig lungs, 3 g of guinea-pig lung was chopped in 4 ml of buffer (control group), in buffer with 4 μg/ml indomethacin (indomethacin group) and in buffer with 2.5 × 10⁻⁵M AA-861 (AA-861 group). The chopped lungs were incubated for 30 min. 250 μl of incubation medium from each group was assessed before and after 3, 5, 10, 15, 20, 25 and 30 min of incubation. The incubation medium was centrifuged and the supernatant was tested for a PGI₂-like substance (PGI₂) by platelet aggregation inhibition. PGI₂ was produced mainly during the initial 3–5 min of incubation and was decreased thereafter. PGI₂ production was almost completely inhibited in the indomethacin group at all of the incubation times and was partially inhibited in the AA-861 group during the initial 3–5 minutes. Endogenous 5-lipoxygenase products generated in the early stages of incubation seem to be involved in PGI₂ production in guinea-pig lungs.     

12-Hydroxyeicosatetraenoic acid reduces prostacyclin production by endothelial cells

12-Hydroxyeicosatetraenoic acid (12-HETE), a lipoxygenase product released by activated platelets and macrophages, reduced prostacyclin (PGI₂) formation in bovine aortic endothelial cultures by as much as 70%. Maximal inhibition required 1 to 2 h to occur and after 2 hr, a concentration of 1 μM 12-HETE produced 80% of the maximum inhibitory effect. 5-HETE and 15-HETE also inhibited PGI₂ formation. The inhibition was not specific for PGI₂; 12-HETE reduced the formation of all of the radioactive eicosanoids synthesized from [1-¹⁴C]arachidonic acid by human umbilical vein endothelial cultures. Inhibition occurred in the human cultures when PGI₂ formation was elicited with arachidonic acid, ionophore A23187 or thrombin. These findings suggest that prolonged exposure to HETEs may compromise the antithrombotic and vasodilator properties of the endothelium by reducing its capacity to produce eicosanoids, including PGI₂.     

Prostacyclin contributes to inhibition of hypoxic pulmonary vasoconstriction by alkalosis

The mechanism by which extracellular alkalosis inhibits hypoxic pulmonary vasoconstriction is unknown. We investigated whether the inhibition was due to intrapulmonary production of a vasodilator prostaglandin such as prostacyclin (PGI₂). Hypoxic vasoconstriction in isolated salt-solution-perfused rat lungs was blunted by both hypocapnic and NaHCO₃_induced alkalosis (perfusate pH increased from 7.3 to 7.7). The NaHCO₃-induced alkalosis was accompanied by a significant increase in the perfusate level of 6-keto-prostaglandin F_1α (6-keto-PGF_1α), an hydrolysis product of PGI₁. Meclofenamate, an inhibitor of cyclooxygenase, counteracted both the blunting of hypoxic vasoconstriction and the increased level of 6-keto-PGF_1α. In intact anesthetized dogs, hypocapnic alkalosis (blood pH increased from 7.4 to 7.5) blunted hypoxic pulmonary vasoconstriction before but not after administration of meclofenamate. In separate cultures of bovine pulmonary artery endothelial and smooth muscle cells stimulated by bradykinin, the incubation medium levels of 6-keto-PGF_1α were increased by both hypocapnia and NaHCO₃-induced alkalosis (medium pH increased from 7.4 to 7.7). These results suggest that inhibition of hypoxic pulmonary vasoconstriction by alkalosis is mediated at least partly by PGI_2.     

Reversal of indomethacin-induced inhibition of tubuloglomerular feedback by prostaglandin infusion

Experiments were performed in rats to study the effect of infusion of PGI₂, PGE₂, and PGF_2α on tubuloglomerular feedback responses (i.e. the change of SNGFR in response to a change of loop of Henle flow rate) in the presence and absence of simultaneous inhibition of endogenous PG synthesis with indomethacin. Infusion of PGI₂ or PGE₂ at rates that did not alter arterial blood pressure did not significantly modify the magnitude of feedback responses (PGI₂) 8.5 μg/hr, PGE₂ 85 μg/hr). Some inhibition of feedback responses was seen when PGI₂ and PGE₂ were administered at higher rates were associated with a reduction of blood pressure (PGI₂ 20 μg/hr, PGE₂ 200 μg/hr). PGI₂ (8.5 μg/hr) and PGE₂ (85 μg/hr) largely prevented feedback inhibition induced by indomethacin. When given subsequent to indomethacin PGI₂ and PGE₂ restored feedback responsiveness almost to normal. In contrast, PGF_2α did not influence feedback inhibition caused by indomethacin. Infusion of PGI₂ induced partial restoration of feedback responses in DOCA-salt treated animals in which the feedback system is virtually completely inactive. Our results indicate that availability of PGI₂ or PGE₂ is necessary for the normal operation of the tubuloglomerular feedback mechanism for control of nephron filtration rate.     

Effects of prostaglandins E2, I2 and F2α, arachidonic acid and indomethacin on pressor responses to norepinephrine in conscious rats

The effects of prostaglandins E₂ (PGE₂), I₂ (PGI₂) and F₂α (PGF₂α), arachidonic acid and indomethacin on pressor responses to norepinephrine were examined in conscious rats. Intravenously infused PGE₂ (0.3, 1.25 μg/kg/min), PGI₂ (50, 100 ng/kg/min), PGF₂α (1.8, 5.4 μg/kg/min) and arachidonic acid (0.7, 1.4 mg/kg/min) did not change the basal blood pressure. Both PGE₂ and PGI₂ significantly attenuated pressor responses to norepinephrine, whereas PGF₂α significantly potentiated them. Arachidonic acid, a precursor of the prostaglandins (PGs), significantly attenuated pressor responses to norepinephrine. Since the attenuating effect of arachidonic acid was completely abolished by the pretreatment with indomethacin (5 mg/kg), arachidonic acid is thought to exert an effect through its conversion to PGs. On the contrary, intravenously injected indomethacin (0.2–5.0 mg/kg) facilitated pressor responses to norepinephrine in a dose-related manner without any direct effect on the basal blood pressure. These results suggest that endogenous PGs may participate in the regulation of blood pressure by modulating pressor responses to norepinephrine in conscious rats.     

Prostacyclin inhibits 5-hydroxytryptamine release but stimulates thromboxane synthesis during cardiopulmonary bypass

The antiaggregating agent prostacyclin (PGI₂) was infused into ten dogs during cardiopulmonary bypass (CPB) to minimize thrombocytopenia and platelet dysfunction. The animals were anesthetized, placed on mechanical ventilation and underwent thoracotomy. After heparinization with 300 u/kg, animals were assigned to control (n=5) or PGI₂ treated groups (n=5). Thoracotomy and then CPB decreased platelet numbers to below 30, 000/mm³ (p < 0.05) and fibrinogen to less than 150 mg/dl (p < 0.05). PGI₂ at 100 ng/kg·min was infused for the 2 h period of CPB. PGI₂ infusion did not prevent these changes, but did prevent platelet serotonin release. In the control group after CPB, platelet serotonin fell from the baseline value of 1.11 μg/10⁹ to 0.35 μg/10⁹ platelets (p < 0.05). In contrast, PGI₂ treatment resulted in a serotonin increase to 2.27 μg/10⁹ platelets (p < 0.05). Thromboxane B₂ concentrations of platelets and plasma rose during CPB (p < 0.05). Surprisingly, PGI₂ infusion accentuated this rise in platelet and plasma thromboxane B₂ (p < 0.05). These data indicate that during CPB, an infusion of PGI₂: 1) does not prevent thrombocytopenia; 2) increases platelet serotonin uptake despite, 3) an associated rise in platelet and plasma thromboxane B₂.     

Potentiation of anti-aggregating activity of PGI2 by 7-ethoxycarbonyl-6,8-dimethyl-4-hydroxymethyl-1(2H)-phthalazinone (EG-626) in rabbit platelets in vitro

Anti-aggregating activity of 7-ethoxycarbonyl-6,8-dimethyl-4-hydroxymethyl-1(2H)-phthalazinone (EG-626) was tested using rabbit platelets in vitro. EG-626 alone, when added before, prevented platelet aggregation induced by ADP, as did PGI₂, papaverine and dipyridamole. Spontaneous disaggregation was also accelerated when EG-626 was added after the maximal aggregation induced by ADP. EG-626 alone also inhibited platelet aggregation induced by collagen and arachidonic acid. ID₅₀s of these agents in ADP-induced aggregation were 7–9 nM for PGI₂, 223 μM for EG-626, 266 μM for papaverine and 957 μM for dipyridamole. When EG-626 was used in combination with PGI₂, a threshold dose (50 μM) of EG-626 potentiated the anti-aggregating effect of subthreshold dose (3 nM) of PGI₂ upto 100% inhibition in collagen-induced platelet aggregation. The marked potentiating effect of EG-626 was accompanied by an accumulation of cyclic AMP in the platelets. These effects might be due to inhibition of phosphodiesterase. Papaverine and dipyridamole, other phosphodiesterase inhibitors, also potentiated the anti-aggregating activity of PGI₂. The activity of papaverine, however, was one eighth of EG-626 and that of dipyridamole was much less. The most effective combination of PGI₂ and EG-626 to induce 50% inhibition was obtained with 20% of ID₅₀ of each agent, whereas that of PGI₂ and papaverine or dipyridamole was 39 or 41%, respectively.     

Prostacyclin (PGI2) effets on anterior pituitary hormones in the rat in vivo

Intravenous injection of 600 μg PGE₂ or PGI₂ significantly increased serum LH and prolactin levels in estradiol treated ovariectomized rats. There was no effect on serum FSH concentration. PGE₂ and PGI₂ stimulated LH release in a non-dose dependent manner, while prolactin levels were positively correlated with the dose administered following PGI₂ treatment. 6-keto-PGF_1α at a comparable dose had no effect on pituitary hormone levels. Subcutaneous administration of 1 mg/kg or 60 mg/kg PGI₂ for seven days significantly depressed serum LH level both in male and female rats. These doses had no effect on serum FSH or prolactin levels.     

Prostacyclin stimulation of dog arterial cyclic AMP levels

Prostacyclin (PGI₂) dose-dependently increases the adenosine 3′,5′-cyclic monophosphate (cyclic AMP) levels in canine femoral, carotid, and canine and bovine coronary arteries. The prostacyclin-stimulation is enhanced by phosphodiesterase inhibitors, and is readily measurable after 60 sec incubation. The prostaglandin endoperoxide PGH₂, but not PGH₁, also elevates cAMP levels in femoral arteries. Inhibition of arterial prostacyclin synthetase with 28 μM 9,11-azoprosta-5,13-dienoic acid (azo analog I) blocks the PGH₂-stimulation of cAMP accumulation. Azo analog I does not attenuate a direct PGI₂ stimulation, indicating that the PGH₂ dependent elevation of cAMP is due to conversion of PGH₂ to PGI₂ by the artery. PGI₂ and PGE₁ increase cyclic AMP levels and relax dog femoral and bovine coronary arteries, while PGE₂, which actually contracts bovine coronary arteries, has no effect on arterial cyclic AMP levels. The significance of the PGI₂-stimulation of arterial cyclic AMP is not known, but it is probably related to relaxation of arterial strips.