thromboxane-b2 and isosorbide-5-mononitrate

Nitric oxide (NO) donors are capable of ripening the human cervix during pregnancy. The purpose of this study was to examine how NO donors may be involved in this process. Cervical biopsies were obtained from pregnant women randomized to receive isosorbide mononitrate (n = 10) or no treatment (n = 10) prior to suction termination. Enzyme-linked immunosorbent assays (ELISA) were performed on culture supernatant for interleukin (IL)-1, IL-6, IL-8, IL-10, IL-15, tumour necrosis factor-alpha, monocyte chemoattractant protein-1 and prostaglandin metabolites. Immunohistochemistry was performed to localize these cytokines, cyclooxygenase (COX)-1, COX-2 and prostaglandin dehydrogenase in cervical tissue and reverse transcription-polymerase chain reaction (RT-PCR) to identify COX-1 and COX-2 expression. Biopsies treated with the NO donor isosorbide mononitrate (IMN) produced significantly greater amounts of prostaglandin F(2alpha) in culture and lower amounts of thromboxane B(2) than controls (572.8 versus 34.9 pg/ml, P < 0.05; 53.3 pg/ml versus 530.9 pg/ml, P < 0.01 respectively). The release of other prostaglandins and of cytokines was not affected by treatment with NO. Inflammatory mediators were localized to cervical tissue and COX-1 and COX-2 expression was confirmed by RT-PCR. In conclusion, the mechanism of NO donor-induced cervical ripening during pregnancy may be mediated in part via increased prostaglandin F(2alpha) synthesis.

Topics: Base Sequence; Cervical Ripening; Cervix Uteri; Cyclooxygenase 1; Cyclooxygenase 2; Cytokines; Dinoprost; DNA Primers; Female; Humans; Immunohistochemistry; Isoenzymes; Isosorbide Dinitrate; Membrane Proteins; Nitric Oxide Donors; Pregnancy; Pregnancy Trimester, First; Prostaglandin-Endoperoxide Synthases; Prostaglandins; Reverse Transcriptase Polymerase Chain Reaction; Thromboxane B2

The purpose of the present work was to investigate the ex vivo platelet-inhibiting properties of the nitric oxide-containing vasodilator, molsidomine, and the organic nitrate, isosorbide-5-mononitrate, in comparison with placebo. Ex vivo platelet aggregation in 11 healthy volunteers was measured before, as well as 30 and 60 min after, the intake of either 4 mg molsidomine, 20 mg isosorbide-5-mononitrate (ISMN) or placebo in a randomized double-blind fashion. The release of thromboxane was also determined. Threshold doses of platelet-activating factor (PAF) to induce irreversible aggregation were significantly increased by 100 and 120% 30 and 60 min after molsidomine. Slopes of aggregation curves (aggregation induced with 50 and 200 nM PAF) were significantly reduced after molsidomine (P less than 0.01). Small platelet-inhibiting effects were also observed after ISMN and after placebo intake. The release of thromboxane was not influenced when platelets were maximally stimulated either during clotting of whole blood or during aggregation of platelet-rich plasma with a high dose of PAF. Thromboxane release with a low dose of PAF was reduced 30 and 60 min after drug intake, independent of whether molsidomine, ISMN or placebo was applied. The data indicate that the usual clinical doses of molsidomine, but not of ISMN inhibit platelet aggregation in healthy man.

Topics: Adult; Bleeding Time; Cyclic GMP; Dose-Response Relationship, Drug; Double-Blind Method; Female; Humans; Isosorbide Dinitrate; Male; Molsidomine; Platelet Activating Factor; Platelet Aggregation Inhibitors; Random Allocation; Reference Values; Thromboxane B2

Isosorbide monitrates (IS-2-MN and IS-5-MN), hepatic metabolites of isosorbide dinitrate, inhibit platelet function in vitro very differently, with IS-2-MN being much more potent than IS-5-MN. To assess their antiplatelet properties in vivo and to compare time and dosage requirements, we infused both IS-2-MN and IS-5-MN for 30 minutes, on 2 separate days, into nine patients with stable coronary artery disease, at rates of 4 mg/hr (n = 4) and 8 mg/hr (n = 5). Two additional patients received IS-5-MN at 16 mg/hr. Platelet aggregation and thromboxane (TX) B2 generation in response to various agonists, drug plasma concentrations, and blood pressure were monitored throughout the study. A significant decrease in platelet aggregation and TXB2 production by adenosine diphosphate and adrenaline occurred in seven of nine patients receiving IS-2-MN and in 7 of 11 patients receiving IS-5-MN. Response was dose related, with more patients responding at 8 mg/hr to IS-2-MN (five of five) than to IS-5-MN (three of five), and was maximum at the end of the infusion time, corresponding to peak plasma levels. Patients responding to drug infusions with an inhibition of platelet function were characterized by a greater vascular responsiveness compared to nonresponders, since the decrease in systolic blood pressure (mean +/- SEM) was significantly greater in the former (15.4 +/- 3.2) than in the latter (2.5 +/- 2.1, p less than 0.05). Therefore both mononitrates, when administered at infusion rates between 8 and 16 mg/hr, are accompanied by a consistent inhibition of adenosine diphosphate- and adrenaline-induced aggregation and TX generation.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Coronary Disease; Dose-Response Relationship, Drug; Female; Hemodynamics; Humans; Infusions, Intravenous; Isosorbide Dinitrate; Male; Middle Aged; Platelet Aggregation; Platelet Aggregation Inhibitors; Thromboxane B2

Nitroglycerin and isosorbide dinitrate (ISDN), the most commonly used nitrate vasodilators, have been shown to possess antiplatelet properties. It has also been shown, interestingly, that their inhibition of aggregation (mostly upon ADP and adrenaline) occurs in vivo at concentrations 1-2 log orders lower than in vitro, and in the full therapeutic range. Many different hypotheses may explain such an in vivo/in vitro difference: 1) that nitrates induce prostacyclin synthesis; 2) that they synergize prostacyclin; 3) that they give rise to more active in vivo metabolites; 4) that some other requirement for their action, such as the availability of reducing thiols, may be a limiting factor in the in vitro setting. The discrimination among such hypotheses should contribute new insights into nitrate action at the platelet level. On the basis of experiments on cultured endothelial cells and vascular fragments, we had previously concluded against prostacyclin induction by nitrates. On the other hand, ISDN may decrease the IC50 for prostacyclin in aggregation by suprathreshold doses of various aggregating agents, and therefore, be synergistic with this endogenous antiplatelet substance. Compared to ISDN, the two longer-lived metabolites isosorbide-2- and isosorbide-5-mononitrates (IS-2-MN, IS-5-MN) appear remarkably different in terms of antiplatelet potency in vitro (minimum effective concentrations 10(-7)-10(-6)M for IS-2-MN, 10(-4)M for IS-5-MN, for ADP-and adrenaline-induced aggregation and thromboxane (TX) B2 production).(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Dose-Response Relationship, Drug; Epoprostenol; Humans; Isosorbide Dinitrate; Platelet Aggregation; Platelet Aggregation Inhibitors; Thromboxane B2; Vasodilator Agents

Recently the possibility that nitrates inhibit platelet function in man has been explored in vitro and in vivo. We have studied the effect of isosorbide-5-mononitrate (ISMN), a stable and long-acting organic nitrate, on platelet function in vivo. Given orally within the current therapeutic range, the drug has practically no effect on platelet aggregation nor thromboxane generation in platelet-rich plasma in response to ADP, collagen, arachidonate, epinephrine and PAF. Synergistic effects of prostacyclin and ISMN on inhibition of ADP-induced platelet aggregation have been observed. Thus, local inhibition of platelet aggregation might not have been detectable, due to the short half-life in vitro of prostacyclin.

Topics: Adult; Angina Pectoris; Blood Platelets; Epoprostenol; Female; Humans; In Vitro Techniques; Isosorbide Dinitrate; Male; Middle Aged; Platelet Aggregation; Thromboxane B2

NCX-4016 is an acetylsalicylic acid (ASA) derivative containing a nitric oxide-releasing moiety. Compared with ASA, NCX-4016 has a broader spectrum of antithrombotic and antiinflammatory activities. We hypothesized that NCX-4016 might inhibit in vivo lipopolysaccharide (LPS)-induced expression of tissue factor (TF).. Rats were administered 90 mg/kg NCX-4016 orally for 5 days. Placebo, 50 mg/kg ASA, and 80 mg/kg isosorbide-5-mononitrate (ISMN) were used in control groups. On day 5, rats were injected intraperitoneally with 100 microg/kg LPS and killed 6 hours later. The expression of TF in monocytes was measured by flow cytometry and Western blot analysis. Reverse transcriptase-polymerase chain reaction was performed to assess expression of TF and cyclooxygenase-2 (COX-2) genes. Plasma concentrations of interleukin-1beta and tumor necrosis factor-alpha were measured. Urine samples were collected to evaluate the excretion of the thromboxane metabolite 11-dehydro-thromboxane (TX)B2. Gastric mucosa was inspected. LPS injection was followed by synthesis TF and COX-2 mRNAs in circulating monocytes, which were blunted by NCX-4016 but not by ASA or ISMN. Both NCX-4016 and ISMN reduced TF expression on surface of circulating monocyte. LPS increased the excretion 11-dehydro-TXB2, and this was prevented by NCX-4016 and ASA. Unlike ASA, NCX-4016 reduced plasma interleukin-1beta and tumor necrosis factor-alpha. In addition, NCX-4016 almost completely prevented mucosal damage, whereas ASA increased the extension of gastric lesions in LPS-injected rats.. NCX-4016 prevents monocyte TF expression; this is accompanied by inhibition of TX and cytokine biosynthesis. These additive effects of nitric oxide release and COX inhibition may help explain efficacy and tolerability of NCX-4016.

Topics: Administration, Oral; Animals; Anti-Inflammatory Agents, Non-Steroidal; Aspirin; Blotting, Western; Cyclooxygenase 2; Cysteine Proteinase Inhibitors; Drug Administration Schedule; Flow Cytometry; Gastric Mucosa; Interleukin-1; Isoenzymes; Isosorbide Dinitrate; Male; Monocytes; Nitric Oxide; Platelet Aggregation Inhibitors; Prostaglandin-Endoperoxide Synthases; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Thromboplastin; Thromboxane B2; Tumor Necrosis Factor-alpha

We investigated two possible reasons for the greater antiplatelet efficacy of isosorbide dinitrate (ISDN) in vivo as compared to in vitro. ISDN and its two main hepatic metabolites, isosorbide-2-mononitrate (IS-2-MN) and isosorbide-5-mononitrate (IS-5-MN) were compared for their ability to inhibit platelet aggregation and thromboxane B2 generation in vitro in response to threshold concentrations of ADP, adrenaline, collagen, thrombin and arachidonic acid. We also determined the concentration of prostacyclin required to inhibit by 50% platelet aggregation (IC50) in response to supra-threshold doses of aggregating agents. Out of the three nitrates tested, IS-2-MN was more effective than ISDN in inhibition of platelet aggregation and thromboxane B2 formation after ADP (minimum effective concentration: 10(-7) M) and adrenaline (minimum effective concentration: 10(-6) M). In addition, ISDN 10(-4)-10(-6) M decreased IC50 for prostacyclin from 2.7 +/- 1.2 to 0.36 +/- 0.2 nM. Generation of a platelet-active species, IS-2-MN, and synergism with prostacyclin are novel properties of ISDN and may account for in vivo-in vitro differences in antiaggregatory properties and in vivo mechanism of action.

Topics: Adult; Epoprostenol; Female; Humans; Iloprost; In Vitro Techniques; Isosorbide Dinitrate; Male; Middle Aged; Platelet Aggregation Inhibitors; Thromboxane B2

Isosorbide dinitrate inhibits platelet function in vivo at concentrations about 10 times lower than in vitro (10(-7)-10(-6) vs. 10(-6)-10(-5) M). We investigated two possible reasons for this difference. Isosorbide dinitrate and its in vivo longer-lived metabolites, isosorbide-2- and isosorbide-5-mononitrate were incubated for 5 min with human platelet-rich plasma or washed platelets; irreversible aggregation was induced with threshold doses of ADP, adrenaline, collagen, arachidonic acid and thrombin, and thromboxane (TX) B2 production was measured by radioimmunoassay. Moreover, the concentration of exogenous prostacyclin required to inhibit platelet aggregation by 50% (IC50) after preincubation with isosorbide dinitrate or vehicle was determined. At 10(-7) M, only isosorbide-2-mononitrate inhibited aggregation (-12%, p less than 0.05) and TX production (-36%, p less than 0.01) by ADP. At 10(-6) M isosorbide-2-mononitrate inhibited aggregation by adrenaline more than the dinitrate (-41% vs. -25%, p less than 0.05). In addition, at supra-threshold doses of all the aggregating agents, isosorbide dinitrate decreased IC50 of prostacyclin from 2.7 +/- 1.2 to 0.36 +/- 0.2 nM. Generation of a platelet-active metabolite and synergism with prostacyclin are new properties of isosorbide dinitrate that may account for antiplatelet effects in vivo.

Topics: Adult; Blood Platelets; Drug Synergism; Epoprostenol; Female; Humans; In Vitro Techniques; Isosorbide Dinitrate; Male; Nitrates; Platelet Aggregation; Thromboxane B2

In an experimental model in which cultured endothelial cells (EC) and platelets were incubated with autologous plasma, we investigated the pharmacological modulations by isosorbide nitrates (ISN) [isosorbide dinitrate (ISDN) + 2-isosorbide mononitrate (2-ISMN) + 5-ISMN] of the EC-induced inhibition of platelet aggregation; and the associated changes in prostanoid profile of these mixed EC-platelet suspensions. ISDN antiplatelet activities were found to be magnified profoundly by EC, being dependent upon both ISDN concentration and EC number, e.g., 5.10(-5) M ISDN in the presence of 2.10(4) cells, fully arrested ADP-induced aggregation, whereas the same ISDN concentration induced 30% inhibition in control platelet activities. In contrast, there were no significant changes in 2- and 5-ISMN antiaggregating properties, whether incubated in the presence or absence of EC. Thromboxane B2 accumulated noticeably after aggregation, whereas 6-keto-prostaglandin (PG) F1 alpha and PGE2 accumulated poorly in the medium. In the presence of EC, thromboxane B2 accumulation fell in parallel to the extent of aggregation, whereas 6-keto-PGF2 alpha and PGE2 accumulated in the medium. Aspirin-treated, washed ECs still inhibited platelet aggregation. ISDN was the only ISN capable of inducing PG-accumulation profile changes. These results demonstrate the existence of an endothelium-dependent ISDN antiplatelet activity. Furthermore, this effect is specific to ISDN not being shown by its mononitrate metabolites. These results suggest that PG accumulation changes may be a consequence rather than a cause of the inhibition of platelet activity by (ISDN-stimulated) EC.

Topics: 6-Ketoprostaglandin F1 alpha; Adenosine Diphosphate; Adult; Arachidonic Acid; Arachidonic Acids; Cell Extracts; Cells, Cultured; Dinoprostone; Endothelium; Female; Humans; Isosorbide Dinitrate; Male; Platelet Aggregation; Prostaglandins E; Thromboxane B2; Tissue Extracts

The hypothesis that nitrates evoke prostacyclin production by vascular endothelium has been reevaluated on cultured umbilical vein endothelial cells and in vascular fragments, both obtained from humans. Endothelial cell monolayers (passages 1 and 2) were washed free of culture medium and exposed for 3 to 5 min to buffer or nitroglycerin (NTG), isosorbide dinitrate (ISDN), or isosorbide-5-mononitrate (ISMN) over a range of concentrations (10(-9)M to 10(-6)M) encompassing those usually attained in vivo, with or without 25 microM sodium arachidonate. Basal prostacyclin production, measured by radioimmunoassay of the stable metabolite 6-keto-PGF1 alpha, depended on cell density in the endothelial monolayer (being higher in preconfluent cultures) and on incubation time. Basal prostacyclin, however, was not altered by incubation with NTG (3.3 +/- 2.0 pg/1000 cells without drug vs 3.9 +/- 3.8 pg/1000 cells with drug, mean +/- SD), ISDN (3.1 +/- 1.9 vs 3.1 +/- 2.2), or ISMN (2.0 +/- 0.9 vs 2.3 +/- 1.5) at 10(-7)M (all differences NS). Also, long-term incubation (2, 6, and 24 hr) with ISDN and ISMN did not alter prostacyclin production over control. Over a 30-fold increase (p less than .001) in prostacyclin production was obtained with arachidonate stimulation, but incubation with nitrates did not significantly modify the stimulated production. Saphenous vein, mesenteric artery, and atrial appendage fragments incubated at 37 degrees C for 20 min in a shaking water bath with a control buffer produced 27.8 +/- 13.9, 189.7 +/- 75.2, and 662.3 +/- 390.6 pg 6-keto-PGF1 alpha/mg tissue, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 6-Ketoprostaglandin F1 alpha; Blood Vessels; Cells, Cultured; Endothelium; Epoprostenol; Humans; In Vitro Techniques; Isosorbide Dinitrate; Nitrates; Nitroglycerin; Thromboxane B2; Vasodilator Agents