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

Although prostanoids are known to be involved in regulation of the spontaneous beating rate of cultured neonatal rat cardiomyocytes, the various subtypes of prostanoid receptors have not been investigated in detail. In our experiments, prostaglandin (PG)F(2α) and prostanoid FP receptor agonists (fluprostenol, latanoprost and cloprostenol) produced a decrease in the beating rate. Two prostanoid IP receptor agonists (iloprost and beraprost) induced first a marked drop in the beating rate and then definitive abrogation of beating. In contrast, the prostanoid DP receptor agonists (PGD(2) and BW245C) and TP receptor agonists (U-46619) produced increases in the beating rate. Sulprostone (a prostanoid EP(1) and EP(3) receptor agonist) induced marked increases in the beating rate, which were suppressed by SC-19220 (a selective prostanoid EP(1) antagonist). Butaprost (a selective prostanoid EP(2) receptor agonist), misoprostol (a prostanoid EP(2) and EP(3) receptor agonist), 11-deoxy-PGE(1) (a prostanoid EP(2), EP(3) and EP(4) receptor agonist) did not alter the beating rate. Our results strongly suggest that prostanoid EP(1) receptors are involved in positive regulation of the beating rate. Prostanoid EP(1) receptor expression was confirmed by western blotting with a selective antibody. Hence, neonatal rat cardiomyocytes express both prostanoid IP and FP receptors (which negatively regulate the spontaneous beating rate) and prostanoid TP, DP(1) and EP(1) receptors (which positively regulate the spontaneous beating rate).

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Animals, Newborn; Blotting, Western; Cells, Cultured; Cloprostenol; Dibenz(b,f)(1,4)oxazepine-10(11H)-carboxylic acid, 8-chloro-, 2-acetylhydrazide; Dinoprostone; Dose-Response Relationship, Drug; Epoprostenol; Hydantoins; Iloprost; Latanoprost; Myocytes, Cardiac; Prostaglandin D2; Prostaglandins F, Synthetic; Rats; Rats, Sprague-Dawley; Receptors, Prostaglandin; Receptors, Prostaglandin E, EP1 Subtype; Receptors, Thromboxane

TRA-418, a compound with both thromboxane A2 receptor (TP receptor) antagonistic and prostacyclin receptor (IP receptor) agonistic activities, was synthesised in our laboratory as a new antithrombotic agent. In this study, we examined the effects of TRA-418 on platelet-leukocyte interactions in human whole blood. Platelet-leukocyte interactions were induced by U-46619 in the presence of epinephrine (U-46619 + epinephrine) or with thrombin receptor agonist peptide 1-6 (TRAP). Platelet-leukocyte interactions were assessed by flow cytometry, with examination of both platelet-neutrophil and platelet-monocyte complexes. In a control experiment, the TP receptor antagonist SQ-29548 significantly inhibited the induction of platelet-leukocyte complexes by the combination of U-46619 and epinephrine, but not TRAP-induced formation of platelet-leukocyte complexes. Conversely, the IP receptor agonist beraprost sodium inhibited platelet-leukocyte complex formation induced by both methods, although the IC50 values of beraprost sodium for U-46619 + epinephrine were at least 10-fold greater than for TRAP. Under such conditions, TRA-418 inhibited both U-46619 + epinephrine-induced and TRAP-induced platelet-leukocyte complex formation in a concentration-dependent manner, in a similar range. These results suggest that TRA-418 exerts its inhibitory effects on platelet-leukocyte interactions by acting as a TP receptor antagonist as well as an IP receptor agonist in an additive or synergistic manner. These inhibitory effects of TRA-418 on formation of platelet-leukocyte complexes suggest the compound is beneficial effects as an antithrombotic agent.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Blood Platelets; Bridged Bicyclo Compounds, Heterocyclic; Cell Communication; Cell Separation; Cells, Cultured; Epinephrine; Epoprostenol; Fatty Acids, Unsaturated; Flow Cytometry; Humans; Hydrazines; Leukocytes; Oxazines; Peptide Fragments; Platelet Activation; Receptors, Epoprostenol; Receptors, Thromboxane A2, Prostaglandin H2; Thrombosis

Acute hemodynamic effects of beraprost sodium were tested in a canine vasoconstrictive pulmonary hypertension model induced by the continuous infusion of U-46619, a thromboxane A(2)mimetic. The effects of beraprost were compared with those of prostaglandin E(1), nitroglycerin and nifedipine. Beraprost and nitroglycerin decreased pulmonary arterial pressure. On the other hand, prostaglandin E(1)and nifedipine increased pulmonary arterial pressure. All drugs except nitroglycerin increased cardiac output and decreased pulmonary vascular resistance. Beraprost was selective to pulmonary circulation, while nitroglycerin, prostaglandin E(1), and nifedipine showed poor selectivity for the pulmonary vasculature. These results suggest that the vasodilative effect of beraprost is the most selective for the pulmonary circulation among these four vasodilators.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Alprostadil; Animals; Antihypertensive Agents; Disease Models, Animal; Dogs; Dose-Response Relationship, Drug; Epoprostenol; Humans; Hypertension, Pulmonary; Male; Nifedipine; Nitroglycerin; Oxygen; Thromboxane A2; Vasoconstrictor Agents; Vasodilator Agents

We evaluated the inhibitory effects of beraprost, a stable prostacyclin analogue, on platelet aggregation, assessed by two methods of platelet aggregometry. The conventional aggregometry detects changes in light transmission (LT) of a platelet suspension, and a recently developed aggregometry based upon a particle counting principle detects light scattering (LS) generated by platelet aggregates. Since LS is more sensitive than LT in detecting platelet aggregates of small size, the minimal concentrations of agonists (ADP, epinephrine, collagen, and U46619) to induce detectable aggregate formation were consistently lower with LS (1/2 to 1/6) than with LT. The effects of beraprost were evaluated on platelet aggregation induced by the optimal concentrations of agonists thus determined for each sample. The IC50 values of beraprost on platelet aggregation, as assessed by LS, were 1/2 to 1/10 of those assessed by LT. In suppressing platelet aggregation assessed by LS, beraprost was especially potent with IC50 of 0.2-0.5 nM when platelets were activated by U46619, a thromboxane A2 analogue, or low concentrations of collagen which activates platelets through thromboxane A2 production. The IC50 values were 2-5 nM with ADP and epinephrine, which induce the formation of small aggregates independently of thromboxane A2 production. These findings suggest that LS can detect inhibitory effects of lower concentrations of antiplatelet agents, since it detects the formation of small aggregates induced by agonists in the lower concentration range than LT. It is also suggested that beraprost potently inhibits thromboxane A2-elicited initial signal transduction pathway, reflected by the formation of small aggregates.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Adenosine Diphosphate; Binding Sites; Blood Platelets; Cholesterol; Collagen; Dose-Response Relationship, Drug; Down-Regulation; Epinephrine; Epoprostenol; Humans; Iodine Radioisotopes; Lipoproteins, HDL; Methods; Platelet Aggregation; Platelet Aggregation Inhibitors; Platelet Count; Protein Kinase C; Scattering, Radiation; Vasoconstrictor Agents

The vascular contractile mechanism of prostacyclin (PGI2) was investigated using beraprost sodium (BPS), a stable PGI2 analog. Ring strips without endothelium isolated from canine femoral veins and arteries were used. BPS induced a dose-dependent contraction without precontraction and after precontraction with norepinephrine (NE) or 60 mM K+ in the veins. In contrast, BPS induced a dose-dependent relaxation after precontraction with U46619, a thromboxane A2 (TXA2) analog, or prostaglandin F2 alpha (PGF2 alpha) in the veins. In the arteries, BPS induced contraction at higher concentrations after precontraction with NE. However, BPS relaxed arteries dose-dependently after precontraction with PGF2 alpha. By pretreatment with 13-azaprostanoic acid (13-APA), a TXA2/endoperoxide receptor antagonist, the dose-response curve of BPS in the veins was shifted to the right. Schild plot analysis resulted in a linear regression with a slope of 0.86 +/- 0.13, which was not significantly different from unity, and the pA2 value for 13-APA against BPS was 7.10 +/- 0.06. By pretreatment with BPS, the dose-response curve of U46619 in the veins was shifted to the right. Kaumann plot analysis resulted in a linear regression with a slope of 0.89 +/- 0.09, which was not significantly different from unity, and the pA2 value for BPS against U46619 was 5.68 +/- 0.04. These findings indicate that BPS is a partial agonist for the TXA2/endoperoxide receptors.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Acetylcholine; Animals; Dinoprost; Dogs; Dose-Response Relationship, Drug; Epoprostenol; Female; Femoral Artery; Femoral Vein; In Vitro Techniques; Male; Norepinephrine; Potassium; Prostaglandin Endoperoxides, Synthetic; Prostanoic Acids; Thromboxane A2; Vasoconstriction; Vasoconstrictor Agents