calcimycin and piriprost

Because leukotrienes may mediate hypoxic pulmonary vasoconstriction (HPV), we examined the influence of two lipoxygenase inhibitors on HPV in anesthetized pigs. HPV was induced by ventilation with a hypoxic gas mixture (FIO2 at 0.095), resulting in a fall in PaO2 to 23 +/- 2 mm Hg and a rise in pulmonary vascular resistance from 285 +/- 15 to 595 +/- 30 dyne/s/cm-5. After infusion of either U-60,257B (50 mg/kg, n = 13) or BW 755c (20 mg/kg, n = 8), the responses to repeated hypoxic challenges were recorded. After U-60,257B infusion the hypoxic pressor response was eliminated at 10 and 30 min and remained significantly (p less than 0.01) attenuated at 50 min. The pulmonary pressor response to angiotensin II infusion (0.2 micrograms/kg/min) was also ablated, whereas the systemic response was unchanged. In contrast, after BW 755c infusion there was a modest but sustained augmentation of HPV, maximum at 30 min (pulmonary vascular resistance, 158 +/- 23% control, p less than 0.01), and no alteration of the responses to angiotensin II. BW 755c inhibited the A23187-induced release of leukotrienes, but not histamine, from isolated porcine lung cells (IC50, 6.3 x 10(-5) M), whereas U-60,257B inhibited the release of both leukotriene (IC50, 1.1 x 10(-4) M) and histamine. These findings indicate that reduction of HPV by lipoxygenase inhibitors is not necessarily a consequence of inhibition of leukotriene synthesis.

Topics: 4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyrazol-3-amine; Angiotensin II; Animals; Calcimycin; Epoprostenol; Female; Histamine Release; Hypoxia; Lipoxygenase Inhibitors; Lung; Male; Pulmonary Circulation; Pyrazoles; SRS-A; Swine; Vascular Resistance; Vasoconstriction

Arachidonic acid metabolites, notably leukotrienes (LTs), have been postulated to play a role in hypoxic pulmonary vasoconstriction. In the present study, we examined the contribution of arachidonic acid metabolites, via the cyclooxygenase, 5-lipoxygenase and cytochrome P-450 monooxygenase pathways, to the hypoxia (25 +/- 3 torr)- and anoxia (0 +/- 2 torr)-induced contractions of isolated pulmonary venules. Neither the cyclooxygenase inhibitors indomethacin (5 microM) or ibuprofen (10 microM) nor the 5-lipoxygenase inhibitors nordihydroguaiaretic acid (5 microM) or U 60257B (10 microM) affected the contractile responses. Similarly, the LT receptor antagonists FPL 57231 (3 microM) or LY 163443 (1 microM), at concentrations that inhibited LT-induced venular contractions, did not significantly affect the responses to hypoxia or anoxia. In fact, anoxia suppressed spontaneous LT release from the venules. The cytochrome P-450 inhibitor SKF-525A (500 microM) nonselectively depressed venular contractions induced by decreased PO2 and pharmacological agents. Induction of the cytochrome P-450 monooxygenase system with beta-naphthoflavone did not alter venular contractions induced by hypoxia or anoxia. Contractions of isolated guinea pig pulmonary venules elicited by decreased PO2 are not mediated by 5-lipoxygenase or cyclooxygenase metabolites. Furthermore, the data do not support a role for cytochrome P-450 metabolites of endogenous substrates in these contractions.

Topics: Acetophenones; Animals; Benzoflavones; beta-Naphthoflavone; Calcimycin; Chromones; Cytochrome P-450 Enzyme System; Epoprostenol; Guinea Pigs; Hypoxia; Leukotrienes; Lipoxygenase; Lipoxygenase Inhibitors; Lung; Masoprocol; Muscle Contraction; Muscles; Prostaglandin-Endoperoxide Synthases; Pyridines; Veins; Venules

We studied the effect of piriprost, an inhibitor of sulfidopeptide leukotriene (LT) formation, on the generation of the known products of the 5-lipoxygenase pathway of arachidonate metabolism in calcium ionophore A23187-challenged rat basophil leukemia cells and cloned, growth factor-dependent, mouse mast cells. Piriprost inhibited the formation of 5-hydroxyeicosatetraenoic acid (5-HETE), and LTB4, and the sulfidopeptide leukotrienes (LTC4 in the mouse mast cells and both LTC4 and a mixture of LTD4 and LTE4 in the rat basophil leukemia cells) in parallel (IC50 values ranged between 9 and 14 microM for the mouse mast cells and between 15 and 50 microM for the basophil leukemia cells). Our previous observation that piriprost is only a very weak inhibitor of the solubilized LTC synthase of rat basophil leukemia cells was extended to similar enzyme preparations derived from the mouse mast cells (IC50 1.5 mM). The results are consistent with the conclusion that piriprost acts as an inhibitor of the 5-lipoxygenase reaction and that its activity in intact cells is not likely to involve the inhibition of the LTC synthase.

Topics: Animals; Basophils; Calcimycin; Cells, Cultured; Clone Cells; Epoprostenol; Leukemia, Experimental; Lipoxygenase Inhibitors; Mast Cells; Mice; Rats; SRS-A

Platelets and neutrophils are important in determining the extent of myocardial injury following coronary occlusion. The detrimental effects of these blood elements are mediated in part via release of arachidonate metabolites and oxidative species. A new selective inhibitor of leukotriene formation, piriprost (U-60,257), has been observed to decrease both neutrophil accumulation in the myocardium and infarct size following coronary ligation in experimental animals. Since piriprost may have clinical use, we examined its effects on human platelet and neutrophil functions. This compound was found to exert potent inhibitory effects on epinephrine-induced human platelet aggregation and TXA2 biosynthesis (IC50 0.04 microM). Piriprost also inhibited human neutrophil chemotaxis, oxidative species release, aggregation, and LTB4 synthesis with IC50 of 0.1, 0.04, 10 and 14 microM, respectively. Thus, piriprost inhibits a variety of human platelet and neutrophil functions. Because of its suppressive effects on human platelet and neutrophil functions and protective effects in experimental myocardial infarction, this agent may have clinical applications.

Topics: Blood Platelets; Calcimycin; Cell Aggregation; Epinephrine; Epoprostenol; Humans; In Vitro Techniques; Luminescent Measurements; Neutrophils; Oxidation-Reduction; Platelet Aggregation; SRS-A

Arachidonic acid and the calcium ionophore A23187 are known to provoke a pulmonary artery pressor response, edema formation and release of thromboxane B2 (TxB2) and 6-keto prostaglandin-F1 alpha (6-keto PGFl alpha) into the recirculating perfusion fluid of isolated blood-free perfused rabbit lungs. Here we investigated the release of leukotrienes (LTs) by repetitive 0.1 microM A23187 challenge in the presence or absence of cyclooxygenase and 5-lipoxygenase inhibitors. RP-HPLC analysis of perfusion fluid extracts persistently showed peaks with retention times of authentic LTC4, -D4, -E4 and -B4. Fractionated RP-HPLC eluate subjected to radioimmunoassay (RIA) with LTC4 and LTB4 antibodies showed two major peaks of immunoreactivity corresponding to those compounds and minor immunoreactivity with LTD4 and LTE4 in accordance with the stated cross-reactivities of the LTC4 antibody. Good correlation for both LTB4 and LTC4 levels measured by RP-HPLC versus RIA of collected HPLC peaks was found. Five to ten min after A23187 challenge, LTC4, -D4 and -B4 levels ranged from 800 to 1600 pg/ml perfusate. LTC4 reached a maximum level at 20 min whereas LTB4 slightly increased over a 35 min period. Upon repeated A23187 challenge, interrupted by rinsing phases with fresh perfusion fluid, the LT release was reproducible several times with increasing reaction strength. This performed in presence of increasing concentration of the 5-lipoxygenase inhibitors AA-861 or U-60,257 caused a dose-dependent inhibition of the release of all LTs with an IC50 of approximately 10(-8) to 10(-7) M and 10(-6) M, respectively. Cyclooxygenase inhibition with acetylsalicylic acid at doses completely suppressing the A23187 induced pressor response did not inhibit the peptidoleukotriene release and only slightly depressed LTB4 release.. using a rapid and sensitive extraction and RP-HPLC method isolated lungs are found to release nanomolar amounts of LTs into the perfusate upon repetitive A23187 challenge, suppressed by 5-lipoxygenase inhibition.

Topics: 6-Ketoprostaglandin F1 alpha; Animals; Arachidonate Lipoxygenases; Aspirin; Benzoquinones; Blood Pressure; Calcimycin; Dose-Response Relationship, Drug; Epoprostenol; Leukotriene B4; Lipoxygenase Inhibitors; Lung; Perfusion; Pulmonary Artery; Quinones; Rabbits; Secretory Rate; SRS-A

The effects of five inhibitors of the lipoxygenase pathway were evaluated on oxygen radical production, degranulation, chemotaxis, leukotriene B4 (LTB4) production by neutrophils. The lipoxygenase inhibitors tested were nordihydroguaiaretic acid (NDGA), esculetin, eicosatetraynoic acid (ETYA), 2-(12-hydroxydodeca-5,10-diynyl)-3,5,6-trimethyl-1,4-benzoqu inone (AA-861), and 6,9-deepoxy-6, 9-(phenylimino)-delta 6.8-prostaglandin I1 (U-60,257). Neutrophils were activated by n-formyl-methionyl-leucyl-phenylalanine (fMLP), phorbol myristate acetate (PMA), A23187, or platelet activating factor (PAF). The effects of these inhibitors on NADPH oxidase activity and phospholipase A2 activity of isolated particulate fraction of neutrophils were also evaluated. ETYA inhibited neutrophil function induced by all the stimulators except PMA. AA-681 was unique in that it did not inhibit PAF-induced neutrophil activation. U-60,257 had virtually no effect on oxygen radical production and degranulation, but chemotaxis was moderately suppressed. NDGA effectively inhibited neutrophil function, except for chemotaxis. Esculetin inhibited only oxygen radical production, but this was due to inhibition on NADPH oxidase activity of neutrophil membrane. The inhibitory effect on neutrophil function and that of LTB4 production were not closely correlated. It is suggested that lipoxygenase inhibitors may modify neutrophil function by the mechanism not involving the lipoxygenase pathway. It is also suggested that LTB4 may not be a mediator in neutrophil oxygen radical production and degranulation induced by the stimulators used in the present study.

Topics: 5,8,11,14-Eicosatetraynoic Acid; Benzoquinones; Calcimycin; Chemotaxis, Leukocyte; Epoprostenol; Humans; Hydroxides; Hydroxyl Radical; Leukotriene B4; Lipoxygenase Inhibitors; Masoprocol; N-Formylmethionine Leucyl-Phenylalanine; NADH, NADPH Oxidoreductases; NADPH Oxidases; Neutrophils; Phospholipases A; Phospholipases A2; Platelet Activating Factor; Quinones; Tetradecanoylphorbol Acetate; Umbelliferones

6,9-Deepoxy-6-9-(phenylimino)-delta 6,8-prostaglandin I1, a prostacyclin analogue reported to inhibit sulphidopeptide leukotriene formation in animals, was evaluated for its pharmacological activity against eicosanoid and histamine release from human dispersed lung cells (HDLC). In the absence of drug, challenge of HDLC with A23187 (2.5 microM) increased immunoreactive eicosanoid generation by factors of 7.6 for prostaglandin (PG) D2, 9.1 for TXB2, 3.2 for PGF2 alpha, 2.0 for 5-HETE, 6.3 for LTC4, in association with a twofold increase in histamine release. When exogenous [14C]-arachidonic acid was added to HDLC simultaneously with A23187 challenge, radiolabelled eicosanoids were recovered in the supernatant, but on separating the products by radio-thin layer chromatography the proportions of individual eicosanoids were not significantly different from unchallenged cells. With endogenous arachidonate, U-60,257 was a potent inhibitor of i-LTC4 generation at 1 microM, but between 3 and 300 microM there was a concentration-related reversal of this inhibition. The effects of U-60,257 on the metabolism of exogenous [14C]-arachidonic acid were also studied. Under these circumstances the drug was a potent inhibitor of both 5-HETE and 5,12-diHETE formation, without significantly affecting the formation of other mono-HETES. In agreement with previous endogenous substrate experiments there was a concentration-dependent inhibition of TxB2 formation from exogenous arachidonic acid. These findings highlight the complex pharmacological actions of U-60,257 which appear dependent on the source of arachidonic acid substrate.

Topics: Arachidonic Acid; Arachidonic Acids; Calcimycin; Cells, Cultured; Chromatography, High Pressure Liquid; Eicosanoic Acids; Epoprostenol; Humans; Kinetics; Lipoxygenase; Lung; SRS-A

Ovalbumin (OA) and calcium ionophore A23187 were used to induce contractions of sensitized guinea-pig tracheal and lung parenchymal preparations in the presence and absence of indomethacin. This model was used to examine the properties of a series of compounds reported to inhibit 5-lipoxygenase or to antagonize lipoxygenase products at the receptor level. FPL55712 and piriprost appeared to act as pharmacological antagonists because they rapidly reduced tracheal tone established by OA. The prolonged phase (i.e. greater than 10 min post-challenge) of airways contractions induced by OA is assumed to be lipoxygenase-dependent and was inhibited by nordihydroguaiaretic acid (NDGA), FPL55712, nafazatrom and benoxaprofen, in order of potency. Piriprost had similar inhibitory effects on the trachea, but not on lung parenchyma. The inhibitory effects of NDGA and FPL55712 were reduced, and those of nafazatrom increased by indomethacin. Indomethacin decreased the inhibitory effect of piriprost on the trachea, but facilitated inhibition by this agent on the parenchyma. A roughly similar pattern was seen on A23187-induced contractions, but FPL55712 did not modify these contractions and benoxaprofen enhanced the response of the trachea. BW755C enhanced the overall contractile response of OA- and A23187-induced tracheal contractions (but not parenchymal contractions) in a bell-shaped manner, an effect not seen in the presence of indomethacin. This indicated that BW755C could be acting as a cyclo-oxygenase inhibitor. The results suggested that, although inhibitors of the lipoxygenase pathway were partially effective in inhibiting the contractions of the airways induced by OA or A23187, other mediators in addition to those of the lipoxygenase pathway contribute to these contractions.

Topics: 4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyrazol-3-amine; Animals; Arachidonate Lipoxygenases; Calcimycin; Catechols; Chromones; Drug Interactions; Epoprostenol; Guinea Pigs; Indomethacin; Lipoxygenase Inhibitors; Lung; Male; Masoprocol; Muscle Contraction; Ovalbumin; Pyrazoles; Pyrazolones; Trachea

When chopped porcine pulmonary arteries were incubated with calcium ionophore A23187 (1) in the presence of indomethacin there was a time dependent generation of a substance which produced contractions of superfused strips of guinea-pig ileum smooth muscle (GPISM) which were indistinguishable from those induced by LTD4. This material however had a different retention time from LTD4 when subjected to HPLC and co-chromatographed with synthetic LTE4. In addition to LTE4 a substance which had properties indistinguishable from those of LTB4 when assayed on a combination of guinea-pig lung parenchymal strips (GPP) and GPISM (2) was generated from the pulmonary artery. This substance co-chromatographed with synthetic LTB4. The adventitia and intima were the richest source of LTE4, the adventitia releasing slightly more than the intima. The output of LTB4 and LTE4 was inhibited by 6,9-deepoxy-6,9-(phenylimino)-delta 6,8 prostaglandin I (U-60,257). Nordihydroguaiaretic acid (NDGA) inhibited the generation of LTE4.

Topics: Animals; Calcimycin; Epoprostenol; Indomethacin; Kinetics; Leukotriene B4; Leukotriene E4; Pulmonary Artery; SRS-A; Swine

6,9-deepoxy- 6,9-(phenylimino)-delta 6,8-Prostaglandin I1 (U-60,257), a prostaglandin analogue known to inhibit leukotriene formation in a number of cell systems, potentiates mast cell release of prostaglandin D2 from human dispersed lung cells activated with ionophore A23187. Over the same concentration range of 30-300 microM there was a related inhibition of ionophore-induced generation of thromboxane B2 (r = 0.93, P less than 0.01). As both prostaglandin D2 and thromboxane A2 are potent bronchoconstrictors, these observations may be relevant to the potential of this drug in the treatment of asthma.

Topics: Calcimycin; Dinoprost; Epoprostenol; Humans; In Vitro Techniques; Lung; Mast Cells; Prostaglandin D2; Prostaglandins D; Prostaglandins F; Thromboxane B2; Thromboxanes

6,9-Deepoxy-6,9-(phenylimino)-delta 6,8-prostaglandin I1 (U-60257) and its methyl ester (U-56467) are selective inhibitors of leukotriene C and D biosynthesis both in vitro and in vivo. In this study, we demonstrated that the principal site of inhibition may be arachidonate 5-lipoxygenase, the initial enzyme of leukotriene biosynthesis. U-60257 and its methyl ester block LTB4 synthesis in human peripheral neutrophils with an ID50 of 1.8 and 0.42 microM respectively. This inhibitory action of U-60257 on neutrophil 5-lipoxygenase can be reduced or reversed by a high concentration of exogenous arachidonic acid. U-60257 at 100 microM has no apparent effect on the following enzymes. 1) cyclooxygenase of sheep vesicular gland or human platelets; 2) 12-lipoxygenase of human platelets and 3) soybean 15-lipoxygenase. Thus, we conclude that U-60257 and its methyl ester are potent and selective inhibitors of arachidonate 5-lipoxygenase.

Topics: Arachidonate Lipoxygenases; Calcimycin; Chromatography, High Pressure Liquid; Chromatography, Thin Layer; Epoprostenol; Humans; Lipoxygenase Inhibitors; Neutrophils; SRS-A

Topics: Calcimycin; Chemotaxis, Leukocyte; Cytochalasin B; Epoprostenol; Glucuronidase; Humans; Kinetics; Leukotriene B4; Muramidase; N-Formylmethionine; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Oligopeptides; Prostaglandins; Superoxides