15-hydroxy-11-alpha-9-alpha-(epoxymethano)prosta-5-13-dienoic-acid and ridogrel

We previously showed that ramatroban (Baynastrade mark), a thromboxane A(2) (TxA(2)) antagonist, had inhibited prostaglandin D(2) (PGD(2))-stimulated human eosinophil migration mediated through activation of chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2). However, detailed pharmacological characterization of its inhibitory activity has not been described. In the present study, we showed that [(3)H]ramatroban bound to a single receptor site on CRTH2 transfectants with a similar K(d) value (7.2 nM) to a TxA(2) receptor (8.7 nM). We also demonstrated that ramatroban inhibited PGD(2)-, 15-deoxy-Delta(12, 14)-PGJ(2) (15d-PGJ(2))- and indomethacin-induced calcium responses on CRTH2 transfectants in a competitive manner with similar pA(2) values (8.5, 8.5, and 8.6, respectively). This is the first report showing the evidence for direct binding of ramatroban to CRTH2, revealing its competitive inhibitory effects and another interesting finding that PGD(2), indomethacin and 15d-PGJ(2) share the same binding site with ramatroban on CRTH2.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Anti-Inflammatory Agents, Non-Steroidal; Binding, Competitive; Bridged Bicyclo Compounds, Heterocyclic; Calcium; Carbazoles; Cell Line; Cell Movement; Cyclic AMP; Dose-Response Relationship, Drug; Fatty Acids, Unsaturated; Humans; Hydrazines; Indomethacin; Models, Biological; Pentanoic Acids; Prostaglandin D2; Pyridines; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides; Transfection; Tritium

This study compares the effects of threshold concentrations of endothelin-1 in isolated rat basilar arteries with those in mesenteric arterial branches and investigates the mechanisms of inhibitory and potentiating endothelin-1-effects. In basilar arteries, endothelin-1 reduces the contractions induced by 5-hydroxytryptamine (5-HT), by the thromboxane A2 agonist U46619, and by vasopressin. The inhibitory effect of endothelin-1 on the contraction induced by 5-HT is abolished by deendothelialization, by the endothelin ET(B) receptor antagonist RES 701-1, by indomethacin, or by glibenclamide. In mesenteric arteries, endothelin-1 potentiates the contractile effects of 5-HT, U46619, and vasopressin. The potentiation of the contractile effect induced by 5-HT is only somewhat modified by deendothelialization, but abolished by the thromboxane A2 receptor antagonists GR32191 and ridogrel. U46619 potentiates the 5-HT-effect in mesenteric arteries. Thus, though the contractile endothelin ET(A) receptors were not blocked, threshold concentrations of endothelin-1 inhibited contractile effects in the rat basilar artery via activation of endothelial ET(B) receptors. Prostaglandins and ATP-sensitive K+ channels are involved in this inhibitory action. In contrast, endothelin-1 potentiates contractile actions in mesenteric arteries via the release of endogeneous thromboxane A2 from non-endothelial cells. The study points out the completely different role of the endothelium in combined effects of endothelin-1 between cerebral and mesenteric arteries.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Basilar Artery; Biphenyl Compounds; Drug Synergism; Endothelin Receptor Antagonists; Endothelin-1; Glyburide; Heptanoic Acids; Indomethacin; Male; Mesenteric Arteries; Muscle Contraction; Muscle, Smooth, Vascular; Pentanoic Acids; Peptides, Cyclic; Pyridines; Rats; Rats, Sprague-Dawley; Serotonin; Vasoconstriction; Vasopressins

Platelet aggregation at sites of vascular injury releases both peptide growth factors and vasoactive compounds. Although significant attention has been focused on peptide growth factors, very little is known about the mitogenic effect of vasoactive compounds. We evaluated the effect of serotonin (5-HT) and thromboxane A2 (TXA2) mimetic U46619 alone and in combination on aortic endothelial cells. Stimulation of endothelial cells by 5-HT resulted in an increase in tritiated thymidine uptake and an increase in cell number, whereas U46619 did not have any significant effect. However, when endothelial cells were exposed to both compounds, U46619 potentiated the mitogenic effect of 5-HT on endothelial cells. When endothelial cells were preincubated with LY281067 (a 5-HT2 receptor antagonist) or ridogrel (a combined TXA2 synthase inhibitor and receptor antagonist), LY281067 blocked the mitogenic effect of 5-HT and ridogrel blocked the potentiating effect of U46619 on 5-HT2-induced tritiated thymidine incorporation. When endothelial cells were preincubated with both antagonists, the effects of both 5-HT and U46619 were blocked. Recent studies have indicated that regenerating endothelial cells at sites of vascular injury may release growth factors for vascular smooth muscle cells, leading to smooth muscle cell proliferation and development of neointima. This study suggests that the combined use of 5-HT and TXA2 receptor antagonists may inhibit the growth of endothelial cells at sites of vascular injury and attenuate the formation of neointima.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Cell Division; Cells, Cultured; Disease Models, Animal; Endothelium, Vascular; Lysergic Acid; Male; Muscle, Smooth, Vascular; Pentanoic Acids; Platelet Aggregation; Pyridines; Rabbits; Serotonin; Serotonin Antagonists; Thromboxane A2; Thromboxane-A Synthase; Thymidine; Vasoconstrictor Agents

To investigate the nature of the arachidonic acid metabolite involved in the altered reactivity of microvessels of two-kidney, one-clip hypertensive rats and the possible contribution of this product to the elevated blood pressure levels found in two-kidney, one-clip hypertension, mesenteric arterioles either perfused in vitro or studied in vivo were used along with blood pressure determinations. The decreased response to acetylcholine observed was normalized by ridogrel, a thromboxane A2 receptor antagonist, and dazoxiben, a thromboxane A2 synthase inhibitor. The smooth muscle response to nitric oxide, tested with sodium nitroprusside, was unaltered in two-kidney, one-clip hypertensive microvessels. Neither ridogrel nor dazoxiben modified the response to this vasodilator. In contrast, the potentiated response to noradrenaline was corrected by ridogrel and dazoxiben in vitro but not in vivo. Noradrenaline and acetylcholine increased the release of thromboxane A2 from the mesenteric microvessels of two-kidney, one-clip hypertensive rats. Ridogrel and dazoxiben decreased but did not normalize the elevated blood pressure of hypertensive rats. Based on these results, we concluded that: 1) the decreased responsiveness of smooth muscle to acetylcholine resulted from an increase in thromboxane A2 formation rather than a decrease in sensitivity to nitric oxide; 2) thromboxane A2 contributes to the increased noradrenaline response in mesenteric microvessels perfused in vitro while in in vivo other blood borne vasoactive agents may also be involved since the potentiated noradrenaline response was not corrected by inhibiting thromboxane A2 synthesis or receptors; 3) in addition to thromboxane A2, another as yet unidentified factor, may contribute to the elevated blood pressure in two-kidney, one-clip hypertension.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Arterioles; Enzyme Inhibitors; Hypertension, Renovascular; Imidazoles; Male; Mesentery; Microcirculation; Nitric Oxide; Nitroprusside; Pentanoic Acids; Perfusion; Pyridines; Rats; Rats, Wistar; Receptors, Prostaglandin; Receptors, Thromboxane; Receptors, Thromboxane A2, Prostaglandin H2; Renal Circulation; Thromboxane A2; Thromboxane-A Synthase; Vascular Resistance; Vasoconstrictor Agents

To investigate dilator effects of endothelins (ETs) on the pulmonary circulation and possible changes induced by chronic hypoxia, we examined vascular responses to ET-1 and ET-3 as well as ET binding to receptor subtypes ETA and ETB in the lungs from rats exposed to either room air (controls), hypoxia (10% O2) for 3 wk (3 WH), or 3 WH followed by recovery to room air (3 WH+R). In controls, both ETA and ETB receptor binding was present in smooth muscle of airways and vessels. Infusion of ET-1 or ET-3 (3-100 pM) to isolated perfused lungs preconstricted by U-46619 produced dose-dependent vasodilation with a greater potency of ET-3 (P < 0.01). The vasodilator responses to ET-1 and ET-3 were potentiated by the cyclooxygenase blocker meclofenamate (3 x 10(-6) M) or by the thromboxane synthetase inhibitor R-68070. In meclofenamate-treated lungs, the vasodilator responses to ET-1 and ET-3 remained unaffected by the inhibitor of nitric oxide synthesis, NG-monomethyl-L-arginine (5 x 10(-4) M) or by the guanylate cyclase inhibitor, methylene blue (10(-4) M). Conversely, the K+ channel blockers glibenclamide (10(-4) M) and tetraethylammonium (10(-4) M) attenuated the vasodilator responses to both ET-1 and ET-3. The selective ETA receptor antagonist BQ-123 did not alter ET-induced vasodilation, whereas it attenuated ET-induced vasoconstriction. Vasodilation to both ET-1 and ET-3 was abolished in lungs from 3 WH rats (P < 0.01) but was fully restored in lungs from 3 WH+R rats. Pulmonary vasodilation induced by the K+ channel opener pinacidil, which was suppressed by glibenclamide, did not differ between controls and 3 WH rat lungs. We found no change in ETA and ETB receptor binding from pulmonary vessels in H rat lungs compared with controls. In conclusion, endothelin-induced pulmonary vasodilation which may involve activation of K+ channels is abolished during chronic hypoxia. This abolition does not appear to be related to alterations in ET-receptor subtypes or to unresponsiveness of K+ channels in the pulmonary circulation.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Arginine; Autoradiography; Dose-Response Relationship, Drug; Endothelins; Glyburide; Guanidines; Hypoxia; In Vitro Techniques; Iodine Radioisotopes; Lung; Male; Meclofenamic Acid; Nitroarginine; Pentanoic Acids; Pinacidil; Potassium Channel Blockers; Prostaglandin Endoperoxides, Synthetic; Pulmonary Circulation; Pyridines; Rats; Rats, Wistar; Receptors, Endothelin; Tetraethylammonium; Tetraethylammonium Compounds; Thromboxane A2; Thromboxane-A Synthase; Vasoconstrictor Agents; Vasodilation; Vasodilator Agents

When injected i.v. to guinea pigs, the granulocyte secretagog N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP) induces bronchoconstriction (BC), lung platelet sequestration and increased transendothelial albumin exchanges in lungs. We evaluated BC and the variations of the lung contents in radiolabeled platelets, erythrocytes and extravascular albumin, as measurements of platelet lung entrapment, reduction of lung blood volume and increase of transendothelial albumin exchanges, respectively. Trimetoquinol, a thromboxane A2 (TXA2)-endoperoxide receptor antagonist, inhibited BC and platelet entrapment by lungs induced by fMLP, but protection was nonspecific because it also suppressed BC by histamine. The specific TXA2 synthetase inhibitor/endoperoxide receptor antagonist ridogrel suppressed BC and reduced lung platelet entrapment, but failed to prevent the increase of extravascular albumin and the decrease of erythrocyte lung contents due to fMLP. Consequently, the fMLP-induced increase of vascular albumin exchanges and reduction of lung blood volume are TXA2-independent. Aspirin prevented BC, but failed to suppress lung platelet entrapment by fMLP, indicating that in vivo platelet activation is not TXA2-dependent, even though the levels of circulating TXB2, the stable metabolite of TXA2, were increased after fMLP concomitantly with that of 6-keto-prostaglandin (PG)F1 alpha, the stable metabolite of PGI2. The ridogrel-treated animals showed reduced blood level of TXB2 and increased levels of 6-keto-PGF1 alpha after fMLP challenge. Blocking the cyclooxygenase pathway with aspirin prevented ridogrel-induced protection against lung platelet sequestration after fMLP, supporting the concept that rechanneling of arachidonate metabolism toward protective prostaglandins accounts for protection by ridogrel.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; 6-Ketoprostaglandin F1 alpha; Animals; Arachidonic Acid; Blood Platelets; Bronchoconstriction; Guinea Pigs; Imidazoles; Lung; N-Formylmethionine Leucyl-Phenylalanine; Pentanoic Acids; Platelet Aggregation; Prostaglandin Endoperoxides, Synthetic; Pyridines; Radioimmunoassay; Serum Albumin; Suprofen; Thrombocytopenia; Thromboxane A2; Thromboxane B2; Thromboxane-A Synthase; Tretoquinol

Ridogrel (6.3 x 10(-6) to 10(-4) M) inhibited contractions of isolated rat caudal arteries and rabbit femoral arteries caused by U-46619. The slope of an Arunlakshana-Schild plot (pA2-value: 3.4 x 10(-6) M) on the caudal artery was slightly higher than one (1.14). This effect was maximal within 20 min of incubation of the blood vessel with the compound and easily reversible. Ridogrel antagonised contractions of isolated rabbit femoral arteries caused by prostaglandin F2 alpha in the same concentration range. Ridogrel also inhibited contractions induced by aggregating rat platelets on isolated rat caudal arteries (in the presence of ketanserin 4 x 10(-7) M) and on isolated rabbit pulmonary and femoral arteries (in the absence of ketanserin). Ridogrel had no effect on Ca2(+)-induced contractions in depolarised isolated rabbit femoral arteries, and at 10(-4) M antagonised serotonin-induced contractions in this blood vessel. Its effect on serotonin-induced contractions was statistically significant but very small on isolated rat caudal arteries. These observations indicate that ridogrel is an antagonist of prostaglandin endoperoxide/thromboxane A2 and prostaglandin F2 alpha receptors on vascular smooth muscle.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Arteries; In Vitro Techniques; Pentanoic Acids; Platelet Activation; Prostaglandin Endoperoxides, Synthetic; Pyridines; Rabbits; Rats; Receptors, Prostaglandin; Receptors, Thromboxane; Serotonin Antagonists; Thromboxane-A Synthase; Thromboxanes; Vasoconstriction

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Biphenyl Compounds; Collagen; Fatty Acids, Monounsaturated; Heptanoic Acids; Humans; In Vitro Techniques; Pentanoic Acids; Platelet Aggregation; Prostaglandin Endoperoxides, Synthetic; Pyridines; Thromboxane-A Synthase; Valerates