Radiation induced alterations in rabbit aortic prostacyclin formation

Earlier studies have demonstrated very severe vascular lesions occuring after irradiation in human and experimental animals as well. We examined the rabbit aortic PGI2-formation using the platelet bioassay technique in one year aged rabbits after different irradiation doses and a time course. The findings demonstrate a significant increase in prostacyclin formation, which might be due to the damage of endothelial cells. This stage is followed by a long lasting severe depression. This behaviour in addition demonstrates a dose dependent manner, The findings of a long lasting decrease in vascular PGI2-formation might contribute to the understanding of the high incidence of vascular lesions being found at the site of irradiation.     

The effect of radiation on prostacycliln (PG2) productnio by cultured endothelial cells

The effect of ionizing irradiation on the synthesis of prostacyclin (PGI₂) by cult8erd bovine aortic endothelial cells was detemined. PGI₂ was measuured in the culture medium by a radioimmunoassay for 6-Kto PGF_1α. Two phenomena were observed following irradiation: a) Cells which suffered an immediate radiation damage (1000–5000 rads) released high quantities of PGI₂ to the culture medium. This was due to a de novo synthesis of PGI₂ stimulated by radiation induced cellular damage, since pretreatment with aspirin of the endothelial cell monolayers resulted in a marked inhibition of PGI₂ release following irradiation. b) Metabolically active cells which remained confluent and firmly attaached to the culture dish following single, low and intermediate doses (200–1200 rads) radiation, exhibited a marked decreased in their capacity to synthesize PGI₂ upon exposure to various stimuli of the archidonic acid cascade (arachidonic acid, melittin, ionophore A23187 and PGH₂. Similar results were observed with cells treated with fractionated radiation.The quantities of PGI₂ produced by the endothelial cells decreased as a function of the dose of radiation and time interval between irradiation and subsequent stimulation. Radiation had a minimal effect on the nonthrombogenic properties of the endothelial cells, as evidenced by the small increase in the platelet adherence to the endothelial cells. The effect of radiation on PGI₂ pruction by the vascular endothelium may be relevant to the development of radiation induced capillary occlusions, and the enhancement of atherosclerotic lesions in large vesses.     

The hyperalgesic effects of prostacyclin and prostaglandin E2

Hyperalgesia induced in rat paws or dog knee joints by prostacyclin (PGI₂) and prostaglandin E₂ was measured by a modification of the Randall-Selitto method (1) of by the degree of incapacitation (2). In both species PGI₂ induced an immediate hyperalgesic effect but the effect of PGE₂ had a longer latency. Low doses of PGI₂ caused a short lasting effect but PGE₂, large doses of PGI₂ or successive administration of small doses of PGI₂ caused a long lasting effect.It is suggested that prostacyclin mediates rat paw hyperalgesia induced by carrageenin. The long lasting hyperalgesic effect of PGE₂ and high doses of PGI₂ is possibly an indirect effect caused by stimulation of a sensory nerve sensitising mechanism.     

Prostacyclin increases portal venous flow

Prostacyclin (PGI₂) is a potent vasolidator and is a potential therapeutic agent to increase blood flow during several disease states. PGI₂ is alos elevated in plasma during sepsis or pancreatitis. The hemodynamic effect of PGI₂ has not been investigated with regard to the portal venous system. In five anesthetized swine, cardiac output (CO), central venous pressure (CVP), femoral artery pressure (FAP), heart rate (HR), pulmonary artery pressure (PAP), portal venous flow (PoVF), and portal venous pressure (PoVP) were measured before and after increasing doses of PGI₂. The infusions were then repeated after atropine administration. The previously reported effects on the peripheral and pulmonary vascular systems were confirmed. after an injection of 0.5 to 5.0 ug/kg of PGI₂ into the left atrium, a significant decline in CO, FAP, and PAP occured. Atropinization further depressed CO. The most marked effecr of PGI₂, however, was an increase in PoVF without a change in PoVP. This effect was more pronounced when atropine was administered. In anethetized swine, PGI₂ is a potent vasodilator in all vascular beds, including the portal venous system. These hemodynamic changes should be realized when exogenous PGI₂ is considered as a therapeutic agent or when endogenous PGI₂ might increase in association with disease states like pancreatitis or sepsis.     

Prostacyclin is not a circulating hormone

Gas chromatography with electron-capture detection of the extensively purified pentafluorobenzyl derivative of 6-oxo-PGF_1α was used to determine prostacyclin in blood. Neither human peripheral plasma or whole blood, nor blood drawn directly from the human heart (blood from the right and left atrium which is comparable to pulmonary artery and vein blood), contained any detectable prostacyclin (< 20 pg/ml). Even hyperventilation did not result in detectable PGI₂-formation. During intravenous infusion of PGI₂ into one arm, large amounts were found in blood drawn from the other arm. Increased levels were also found during severe infection and in endotoxin shock. These results lend no support to theories based on the concept that prostacyclin is a circulating hormone under normal conditions.     

Increased platelet activity after termination of prostacyclin infusion into man

Infusion of PGI₂ at a dose of 5 or 10 ng/kg/min during 72 hours into patients with peripheral vascular disease was followed by increased susceptibility of platelets to proaggregatory action of ADP and collagen but not that of arachidonate. The above effects were observed 24 hours after termination of infusion of PGI₂. A tendency to an increased formation of TXA₂ in PRP aggregated by arachidonate was also noticed. Infusion of PGI₂ at a dose of 2 mg/kg/min during 72 hours into the patients caused the decreased platelt aggregability to ADP and arachidonate but not to collagen, and a decreased tendency of production of TXA₂ in PRP aggregated by arachidonate. The existence of a “rebound effect” in platelets after a long term PGI₂ therapy is suggested.     

Histamine stimulates prostacyclin synthesis in cultured human umbilical vein endothelial cells

Human venous endothelial cells synthesize prostacyclin (PGI₂) in response to treatment with histamine. The amount of PGI₂ produced is proportional to the histamine concentration over the range of 10^?7 to 10^?5 M, with a maximal response at 2–5 × 10^?6 M. PGI₂ synthesis occurs as a burst lasting less than 3 minutes after histamine addition. The H1 histamine receptor antagonist pyrilamine causes an 87% inhibition of PGI₂ synthesis, whereas the H2 antagonist cimetidine gives no significant inhibition, suggesting that PGI₂ synthesis in response to histamine is mediated by an H1 receptor.     

The effects of prostacyclin on glycemia and insulin release in man

Prostacyclin /PGI₂/ administered intra-arterially or intravenously to patients with peripheral vascular disease exerted a hyperglycemic effect. In normoglycemic patients receiving PGI₂ at a dose of 5 ng/kg/min these effects were barely detectable, but they became unmasked by a rapid glucose injection. In diabetic patients the same PGI₂ dose led to distinct elevation in blood glucose. Prostacyclin at a dose of 10 ng/kg/min raised blood glucose levels both at rest and after stimulation with glucose, and opposed effectively hypoglycemic action of tolbutamide in non-diabetic patients. PGI₂ repressed glucose-induced insulin release in some normoglycemic patients but in others it either increased it or did not affect it. While hyperglycemic effects are reversible when PGI₂ infusion is stopped, and do not interfere with the usual therapeutic administration of prostacyclin for a few days they, nevertheless, might constitute a risk in a patient with poorly controlled diabetes.     

Enhanced formation of PGI2, a potent hypotensive substance, by aortic rings and homogenates of the spontaneously hypertensive rat

Intact rings and homogenates of aorta from spontaneously hypertensive rats (SHR) contain enhanced capacity over normal rats (NR) to convert arachidonic acid into PGI₂. The PGI₂ synthetic system in SHR is stimulated to a greater extent than NR by norepinephrine. Indomethacin blocks this stimulation. PGE₂ and PGF_2α were detected in much smaller amounts in homogenates (undetected in rings) but their formation was not enhanced by the hypertensive tissue. The identity of PGI₂ was based on 1) direct pharmacological assay on the rat blood pressure. In this system identical vasodepressor responses to PGI₂ are observed after intracarotid and intrajugular administration 2) indirectly as 6-keto PGF_1α isolated after incubation of aortic homogenates with tritiated arachidonic acid and 3) indirectly by GC-MS assay of PGE₂, PGF_2α and 6-keto PGF_1α formed during incubation of aortic homogenates with excess unlabeled arachidonic acid. These results provide additional support to our recent hypothesis that PGI₂, of aortic origin, might actively participate in the regulation of systemic blood pressure. Its enhanced formation by intact hypertensive vascular tissue reflects an increase in the number of enzyme molecules immediately available to the substrate. This could probably be an adaptive response to the elevated levels of catecholamines in the circulation.     

Metabolism of PGI2 by 15-hydroxyprostaglandin-dehydrogenase and Δ13-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.     

Prostacyclin does not affect insulin secretion in humans

The effects of Prostacyclin (PGI₂) infusion on insulin secretion and glucose tolerance were investigated in 7 healthy subjects. PGI₂ infusion caused no statistically significant changes of either glucose or insulin concentration, over the range 2.5–20 ng/Kg/min. A constant PGI₂ infusion (10 ng/Kg/min) did not inhibit acute insulin responses to a glucose (20 g i.v.) pulse (response before PGI₂ = 612±307%; during PGI₂ = 515±468%, mean ± SD, mean change 3–5 min insulin, % basal; P=NS). Glucose disappearance rates were similar after the first and second glucose pulse.Thus, in contrast to PGE₂, PGI₂ does not affect insulin secretion nor glucose disposal at doses producing platelet and vascular changes. It is hypothesized that an altered PGI₂/PGE₂ balance in diabetes may represent a link between vascular, platelet and metabolic changes.     

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.     

Prostaglandin synthesis by fetal rat bone in vitro: Evidence for a role of prostacyclin

Prostaglandin synthesis by fetal rat bones was examined by thin-layer chromatography of culture media after preincubation with labeled arachidonic acid. Cultures in rabbit complement (non-heat inactivated serum) were compared with cultures in heat-inactivated serum or cultures treated with indomethacin. The major complement-dependent products were PGE₂, PGF_2α and 6-keto-PGF_1α, the metabolite of prostacyclin (PGI₂). Since PGI₂ had not been previously identified in bone its ability to stimulate bone resorption was tested. Repeated addition of PGI₂ stimulated release of previously incorporated ⁴⁵Ca from fetal rat long bones in both short-term and long-term cultures at concentrations of 10⁻⁵ to 10⁻⁹M. Because of the short half life of PGI₂ in solution at neutral pH, we tested a sulfur analog, thiaprostacyclin (S-PGI₂) which was found to be a stimulator of bone resorption at concentrations of 10⁻⁵ to 10⁻⁶M. These studies suggest that endogenous PGI₂ production may play a role in bone metabolism. Since vessels produce PGI₂ it is possible that PGI₂ release may be responsible for the frequent association between vascular invasion and resorption of bone or calcified cartilage in physiologic remodeling and pathologic osteolysis.     

Contractile responses to prostacyclin (PGI2) of isolated human saphenous and rat venous tissue

Prostacyclin (PGI₂), in a wide concentration range, produced neither contraction nor relaxation of isolated human saphenous vein. Isolated portal veins and vena cava from normal and spontaneously hypertensive rats (SHR) responded only with an increase in contractile tension when exposed to PGI₂. This constrictor effect was absent in a calcium-free buffer. PGI₂ failed to relax KCI contracted vena cava. The constrictor effect of PGI₂ on portal vein was attenuated in a glucose-free, oxygen deficient buffer. No tachyphylaxis or tolerance to the constrictor effect of PGI₂ was noted. Results emphasize that PGI₂ may produce differing effects on vascular smooth muscle tension depending on species and type of blood vessel studied.     

Fish oil feeding lowers thromboxane- and prostacyclin production by rat platelets and aorta and does not result in the formation of prostaglandin I3

It is demonstrated that feeding cod-liver oil to rats leads to a considerable reduction in the formation of platelet T×A₂ and of vascular PGI₂. No appreciable formation off T×A₃ and PGI₃ is observed, although arterial thrombosis is depressed and bleeding time is prolonged. These findings contradict the suggested role of prostaglandins of the 3-series in thromboregulation.     

Alterations of vascular prostacyclin and thromboxane A2 in Dahl genetical strain susceptible to salt-induced hypertension

To assess the implications of vascular eicosanoids system in the hypertension of Dahl salt-sensitive (Dahl S) strain, we investigated the production of vascular vasodepressor and vasoconstrictor eicosanoids in Dahl S rats. 14-week-old Dahl S rats on a 0.11% NaCl diet (normotension) or a 0.3% NaCl diet (borderline hypertension) had a significantly lowered generation of vascular prostacyclin (PGI₂), compared with Dahl salt-resistant (Dahl R) rats. The impairment of vascular PGI₂ in Dahl S rats was restored to the normal level of Dahl R rats with the elevation of blood pressure induced by a high salt diet (4% NaCl). The production of vascular PGI₂ was closely related to the height of blood pressure. The deterioration of vascular PGI₂ was also found in 4-week-old Dahl S rats with normotension. Conversely, vascular thromboxane A₂ (TXA₂) was significantly enhanced in 14-week-old Dahl S rats in all of the feeding groups. Thus, it seems possible that the proved alterations of the vasodepressor and vasoconstrictor eicosanoids partially contribute to the genesis of salt hypertension. Although the exact mechanisms remain obscure, the adaptation of vascular PGI₂ on a high salt diet may be suitable to compete with the high blood pressure and to protect against the vascular damage.     

Prostacyclin induces a surprising long-lasting motility in quiescent uterine strips (indomethacin-treated) isolated from ovariectomized rats

Dose-response curves for several prostaglandins (PGI₂; PGD₂; PGF₂ and PGE₂); BaCl₂ or prostaglandin metabolites (15-keto-PGF_2α; 13, 14-diOH-15-keto-PGF_2α; 6-keto-PGF_1α and 6-keto-PGE₁ in quiescent (indomethacin-treated) uterine strips from ovariectomized rats, were constructed. All PGs tested as well as BaCl₂, triggered at different concentrations, evident phasic contractions. Within the range of concentrations tested the portion of the curves for the metabolites of PGF_2α was shifted to the right of that for PGF_2α itself; the curve for 6-keto-PGF_1α was displaced to the right of the curve for PGI₂ and that for 6-keto-PGE₁ to the left.It was also demonstrated that the uterine motility elicited by 10⁻⁵ M PGF_2α and its metabolites was long lasting (more than 3 hours) and so it was the activity evoked by PGI₂; 6-keto-PGF_1α and BaCl₂, but not the contractions following 6-keto-PGE₁, which disappeared much earlier. The contractile tension after PGF_2α; 15-keto-PGF_2α; 13, 14-diOH-15-keto-PGF_2α and PGI₂, increased as time progressed whilst that evoked by 6-keto-PGF_2α or BaCl₂ fluctuated during the same period around more constant levels.The surprising sustained and gradually increasing contractile activity after a single dose of an unstable prostaglandin such as PGI₂, on the isolated rat uterus rendered quiescent by indomethacin, is discussed in terms of an effect associated to its transformation into more stable metabolites (6-keto-PGF_1α, or another not tested) or as a consequence of a factor which might protects prostacyclin from inactivation.     

Prostacyclin synthase and phospholipases in the vascular wall of experimental hypertensive rats

To reveal the role of enzymes involved in PGI₂ synthesis for vascular PGI₂ generation in experimental hypertensive models, we defined PGI₂ synthase and phospholipases activities in the aortic wall of two different experimental hypertensive rats, e.g. spontaneously hypertensive rats (SHR) and desoxycorticosterone acetate (DOCA)-salt hypertensive rats. In the stage of established hypertension both of the hypertensive models had a significantly large capacity of the vascular wall to produce PGI₂, as compared to respective control rats. PGI₂ synthase activities in the vascular wall were significantly increased by 27% for SHR and by 80% for DOCA-salt hypertensive rats. Moreover, the enzymatic activities were closely related to the blood pressure values for both of the models. On the other hand, phospholipase C or phospholipase A₂ activities were increased or unchanged in SHR, respectively, whereas both of the phospholipases were significantly decreased in DOCA-salt hypertensive rats. Thus, it is indicated that PGI₂ synthase is partly responsible for the increased PGI₂ generation in the vasclar wall of SHR and DOCA-salt hypertensive rats, and that vascular phospholipase C is playing a more important role in providing arachidonate for PGI₂ synthesis in SHR.     

Platelet products and vascular PGI2 production

The effects of products synthesized and/or secreted by activated platelets on production of PGI₂ by human, rat and rabbit vascular rings were investigated. Of the platelet dense body constituents, 5HT stimulated PGI₂ production by vascular tissue of all three species whereas ADP was active only on rat tissue. Of the lipids produced during platelet activation, PAF stimulated PGI₂ production by vascular tissue of all three species, Lysophosphatidate was less active than PAF on rabbit and human tissue and inactive in rat tissue, and the TxA₂-mimetic, U46619, was inactive on vascular tissue of all three species. It is concluded that there are species variations in the effects of agonists on vascular PGI₂ production and that platelet-derived products other than platelet-derived growth factor and β-thromboglobulin could modulate PGI₂ production to regulate platelet activation in vivo.     

On the multiplicity of platelet prostaglandin receptors: II. The use of N-0164 for distinguishing the loct of action for PGI2, PGD2, PGE2 and hydantoin analogs

The ω-chain variant analogs of prostacyclin (PGI₂) and PGD₂ in which the n-amyl side-chain has been replaced by a cyclohexyl group have been prepared and their cardiovascular activities have been compared to those of BW-245C(Fig. 1)(1) a potent anti-aggregatory vasodilator bearing a cyclohexyl-terminated side-chain on a hydantoin skeleton. The cyclohexyl group has little effect on PGI₂, but converts PGD₂ to a long lasting hypotensive agent and increases the platelet anti-aggregatory potency of PGD₂ by a factor of 8. The prostaglandin antagonist N-0164 selectively blocks the anti-aggregatory actions of PGD₂, cyclohexyl-PGD₂, and BW-245C; with essentially no effect on PGI₂, cyclohexyl-PGI₂ and PGE₂ at comparably effective doses. The latter observation is contrary to an earlier report by MacIntyre (2,3), but supports the view that the anti-aggregatory effect of high doses of PGE₂ ($\text{EC}{}_{50}\text{=50μ}\text{M}$) is mediated by the PGI₂ receptor (4). The hydantoin acts at the platelet PGD₂ receptor.