bay-u9773 and verlukast

Cysteinyl leukotrienes (cys-LTs) are potent asthma-related mediators that function through their G protein-coupled receptors, cys-LT receptor type 1 (CysLT1R) and cys-LT receptor type 2 (CysLT2R).. Because many G protein-coupled receptors transactivate the epidermal growth factor receptor (EGFR) through metalloprotease-mediated ligand shedding, we investigated the effects of cys-LTs on signal transduction and proliferation of bronchial fibroblasts.. Human bronchial fibroblasts were grown from biopsy specimens of healthy subjects. Mitogenesis was assessed on the basis of tritiated methylthymidine incorporation.. Leukotriene (LT) D(4) alone did not increase mitogenesis but dose-dependently increased thymidine incorporation and cell proliferation in the presence of epidermal growth factor (EGF). The enhancement was not prevented by CysLT1R antagonists (MK-571 and montelukast) or by a dual antagonist (BAY u9773), which is consistent with the lack of detectable mRNA for CysLT1R and CysLT2R in bronchial fibroblasts. LTD(4) did not cause EGFR transphosphorylation nor was the synergism blocked by the metalloprotease inhibitor GM6001. The EGFR-selective kinase inhibitor AG1478 suppressed the synergy between LTD(4) and EGF but had no effect on synergistic interactions of LTD(4) with other receptor tyrosine kinase growth factors. The effect of LTD(4) involved a pertussis toxin-sensitive and protein kinase C-mediated intracellular pathway, leading to sustained growth factor-dependent phosphorylation of extracellular signal-regulated kinase 1/2 and protein kinase B (PKB/Akt).. Cys-LTs do not transactivate EGFR but have a broader capability to synergize with receptor tyrosine kinase pathways.. This study implies a critical role of cys-LTs in airway fibrosis in asthma and other chronic airway diseases, which might not be blocked by therapy with current LT receptor antagonists.

Topics: Acetates; Bronchi; Cell Proliferation; Cells, Cultured; Cyclopropanes; ErbB Receptors; Fibroblasts; Humans; Intercellular Signaling Peptides and Proteins; Leukotriene C4; Leukotriene D4; Membrane Proteins; Propionates; Protein Serine-Threonine Kinases; Quinolines; Receptors, Leukotriene; RNA, Messenger; SRS-A; Sulfides

The cysteinyl leukotrienes (CysLTs) are lipid mediators that have been implicated in the pathogenesis of allergic diseases, and their actions are mediated via specific receptors named CysLT1 receptor (CysLT1R) and CysLT2 receptor (CysLT2R). Little information is known about the role of T(H)2 cytokines in the regulation of both CysLT1R and CysLT2R expression.. To investigate the possible modulation of both CysLT1R and CysLT2R messenger RNA (mRNA) expression, we have developed a real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) assay based on the TaqMan fluorescence method to quantify CysLT1R and CysLT2R mRNA in human monocytes.. Human monocytes were stimulated with leukotriene D4 or interleukin (IL) 4 or IL-13, and the levels of CysLT1R and CysLT2R mRNA were measured by the quantitative RT-PCR.. CysLT1R and CysLT2R mRNA was increased after stimulation with leukotriene D4. CysLT1R mRNA was augmented 150-fold after treatment with IL-4; however, no significant increase was observed in CysLT2R mRNA level. IL-13 could induce a biphasic augmentation of CysLT1R mRNA level. In contrast to IL-4, IL-13 enhanced CysLT2R mRNA level, with a maximal effect at 2 hours of incubation.. CysLT1R and CysLT2R expression can be regulated by CysLT itself and T(H)2 cytokines at the transcriptional level.

Topics: Cells, Cultured; Cytokines; Flow Cytometry; Gene Expression; Humans; Interleukin-13; Interleukin-4; Leukotriene Antagonists; Leukotriene D4; Membrane Proteins; Monocytes; Propionates; Quinolines; Receptors, Leukotriene; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; SRS-A; Up-Regulation

Conventional intracellular microelectrodes, neuronal tracer injection techniques and immunohistochemistry were used to study the actions of cysteinyl leukotrienes (CysLTs) on electrical and synaptic behavior of enteric neurons in guinea-pig stomach and small intestine. Bath application of leukotriene C(4), leukotriene D(4) or leukotriene E(4) evoked a slowly activating depolarizing response in most of the myenteric and submucous plexus neurons in the small intestine while no effect was observed in gastric neurons. The depolarization evoked by cysteinyl leukotrienes in intestinal neurons was associated with increased input resistance and enhanced excitability. Suppression of hyperpolarizing after-potentials occurred in AH type neurons. The depolarizing action of cysteinyl leukotrienes was resistant to tetrodotoxin and cyclooxygenase inhibitors. Neither the CysLT(1) receptor antagonists (E)-3-[[[3-[2-(7-chloro-2-quinolinyl)ethenyl]phenyl][[3-dimethylamino)-3-oxopropyl]thio]methyl]thio]-propanoic acid (MK 571), 1-[2-hydroxy-3-propyl-4-[4-(1H-tetrazol-5-yl)butoxy]phenyl]-ethanone (LY 171883) and alpha-pentyl-3-(2-quinolinylmethoxy)-benzenemethanol (REV 5901), nor the dual CysLT(1)/CysLT(2) receptor antagonist 6(R)-(4'-carboxyphenylthio)-5(S)-hydroxy-7(E),9(E),11(Z),14(Z)-eicosatetraenoic acid (BAY u9773) significantly altered the depolarizing action of the cysteinyl leukotrienes. Neurotransmission was unaffected by the cysteinyl leukotrienes. The results suggested involvement of cysteinyl leukotrienes in enteric immuno-neural communication through excitatory actions on enteric neurons. The receptor mediating these effects was distinct from currently recognized cysteinyl leukotriene receptor subtypes (CysLT(1) and CysLT(2) receptors) and may represent a new receptor subtype.

Topics: Acetophenones; Animals; Cysteine; Enteric Nervous System; Guinea Pigs; In Vitro Techniques; Intestine, Small; Leukotriene Antagonists; Leukotrienes; Male; Membrane Potentials; Neurons; Propionates; Quinolines; SRS-A; Stomach; Synaptic Transmission; Tetrazoles

Cysteinyl-leukotrienes (Cys-LTs) increase intrahepatic vascular resistance in normal rat livers. CCl4 cirrhotic rat livers have increased Cys-LT production and 5-lipoxygenase messenger RNA (mRNA) expression. The aim of this study was to investigate the role of 5-lipoxygenase-derived eicosanoids regulating intrahepatic vascular tone in control and CCl4-induced cirrhotic rat livers.. In different groups of portally perfused control and cirrhotic rat livers, the following were analyzed: a portal perfusion pressure (PP) dose-response curve to LTD4; the effects on PP caused by either vehicle, the selective 5-lipoxygenase inhibitor AA-861, the selective Cys-LT1 receptor antagonist MK-571, or the dual Cys-LT1 and Cys-LT2 receptor antagonist BAY u9773; and immunohistochemistry for 5-lipoxygenase in liver sections of cirrhotic and control livers.. Cirrhotic livers have a hyperesponse to LTD4. In control livers, AA-861 and MK-571 produced a moderate and similar reduction in PP. In cirrhotic livers, 5-lipoxygenase inhibition produced a marked and significantly greater reduction in PP than in controls. However, no effect on PP was observed after MK-571 or BAY u9773. 5-Lipoxygenase-positive cells were markedly increased in cirrhotic livers.. Our results suggest that 5-lipoxygenase-derived eicosanoids may contribute to the increased intrahepatic vascular resistance of cirrhotic rat livers and therefore the pathogenesis of portal hypertension.

Topics: Animals; Arachidonate 5-Lipoxygenase; Benzoquinones; Carbon Tetrachloride; Cysteine; Leukotriene Antagonists; Leukotrienes; Lipoxygenase Inhibitors; Liver Circulation; Liver Cirrhosis; Male; Propionates; Quinolines; Rats; Rats, Wistar; SRS-A; Vascular Resistance

Contractions of guinea-pig tracheal preparations to cysteinyl-leukotrienes (LTC(4), LTD(4) and LTE(4)) were characterized in organ baths, and cysteinyl-leukotriene metabolism was studied using radiolabelled agonists and RP-HPLC separation. In the presence of S-hexyl GSH (100 microM) the metabolism of [(3)H]-LTC(4) into [(3)H]-LTD(4) was inhibited and the LTC(4)-induced contractions were resistant to CysLT(1) receptor antagonism but inhibited by the dual CysLT(1)/CysLT(2) receptor antagonist BAY u9773 (0.3 - 3 microM) with a pA(2)-value of 6.8+/-0.2. In the presence of L-cysteine (5 mM), the metabolism of [(3)H]-LTD(4) into [(3)H]-LTE(4) was inhibited and the LTD(4)-induced contractions were inhibited by the CysLT(1) receptor antagonist ICI 198,615 (1 - 10 nM) with a pA(2)-value of 9.3+/-0.2. However, at higher concentrations of ICI 198,615 (30 - 300 nM) a residual contraction to LTD(4) was unmasked, and this response was inhibited by BAY u9773 (1 - 3 microM). In the presence of the combination of S-hexyl GSH with L-cysteine, the LTD(4)-induced contractions displayed the characteristics of the LTC(4) contractile responses, i.e. resistant to CysLT(1) receptor antagonism, increased maximal contractions and slower time-course. This qualitative change of the LTD(4)-induced contraction was also observed in the presence of S-decyl GSH (100 microM), GSH (10 mM) and GSSG (10 mM). S-hexyl GSH, S-decyl GSH, GSH and GSSG all stimulated a formation of [(3)H]-LTC(4) from [(3)H]-LTD(4). In conclusion, GSH and GSH-related compounds changed the pharmacology of the LTD(4)-induced contractions by stimulating the conversion of LTD(4) into LTC(4). Moreover, the results indicate that, in addition to the metabolism of LTC(4) into LTD(4) and LTE(4), also the formation of LTC(4) from LTD(4) may regulate cysteinyl-leukotriene function.

Topics: Animals; Borates; Cysteine; Dicarboxylic Acids; Dose-Response Relationship, Drug; Glutathione; Guinea Pigs; In Vitro Techniques; Indazoles; Leukotriene Antagonists; Leukotriene C4; Leukotriene D4; Leukotriene E4; Leukotrienes; Male; Membrane Proteins; Muscle Contraction; Propionates; Quinolines; Receptors, Leukotriene; Serine; SRS-A; Trachea

Angiotensin II (Ang II) is a vasopressor peptide involved in the pathogenesis of cardiovascular diseases associated with diabetes mellitus. We have previously reported that the 5-lipoxygenase-derived products, particularly the cysteinyl leukotrienes (CysLTs), are involved in Ang II-induced contraction. In this study, we demonstrated that CysLTs contribute to the contraction elicited by Ang II in isolated aortas from streptozotocin-induced diabetic (SS) rats but not from insulin-treated diabetic rats, fructose-fed rats, or control rats. In an organ bath, pretreatment with the 5-lipoxygenase inhibitor (AA861, 10 micromol/L) reduced by 37.6+/-8.2% and 30.1+/-10.9% the Ang II-induced contractions in intact and endothelium-denuded aortic rings, respectively, from SS rats. In contrast, the CysLT(1) receptor antagonist (MK571, 1 micromol/L) or the dual CysLT(1)/CysLT(2) receptor antagonist (BAY-u9773, 0.1 micromol/L) did not affect Ang II-induced contraction. In addition, Ang II induced a 6.2+/-1.5-fold increase in CysLT release through the stimulation of the Ang II type 1 receptor. Furthermore, the urinary excretion of leukotriene E(4) was increased in SS rats (leukotriene E(4), 13.7+/-2.9 ng/24 h [SS rats, n=10] versus 1.5+/-0.5 ng/24 h [control rats, n=6]; P<0.0004). These data suggest the activation of the 5-lipoxygenase pathway in SS rats and the involvement of 5-lipoxygenase-derived products, particularly the CysLTs, in Ang II-induced contraction in aortas from SS rats through stimulation of CysLT receptors different from the well-characterized CysLT(1) or CysLT(2) receptor.

Topics: Angiotensin II; Animals; Aorta; Benzoquinones; Diabetes Mellitus, Experimental; Insulin; Leukotriene Antagonists; Leukotriene E4; Lipoxygenase Inhibitors; Male; Organ Culture Techniques; Potassium Chloride; Propionates; Quinolines; Rats; Rats, Wistar; SRS-A; Vasoconstriction; Vasoconstrictor Agents

The contractile response to cysteinyl-leukotrienes was studied in isolated human pulmonary arterial rings. Concentration-response curves for leukotriene C(4) were significantly potentiated by the cyclooxygenase inhibitor indomethacin (1.7 microM) and after endothelial denudation. Measurements of 6-keto prostaglandin F(1alpha) showed that cysteinyl-leukotrienes stimulated the release of prostacyclin. A single concentration (1 microM) of either leukotriene C(4) or leukotriene D(4) resulted in both contraction and relaxation. Indomethacin abolished the relaxant phase and enhanced the amplitude of the contraction, supporting that cysteinyl-leukotriene-induced contractions of the human pulmonary artery may be functionally antagonised by the release of prostacyclin. The contractions induced by leukotriene C(4) were resistant to the two cysteinyl-leukotriene receptor antagonists MK 571 ((3-(-2(7-chloro-2-quinolinyl)ethenyl)phenyl)((3-(dimethylamino-3-oxo propyl)thio)methyl)thio propanoic acid, 1 microM) and BAY u9773 (6(R)-(4'-carboxyphenylthio)-5(S)-hydroxy-7(E),9(E), 11(Z)14(Z)-eicosatetrenoic acid, 3 microM), both in the absence and presence of indomethacin. These findings suggest a functional cysteinyl-leukotriene receptor in the human pulmonary artery with antagonist properties not previously described in human tissue.

Topics: 6-Ketoprostaglandin F1 alpha; Cysteine; Dose-Response Relationship, Drug; Endothelium, Vascular; Epoprostenol; Female; Humans; In Vitro Techniques; Indomethacin; Leukotriene Antagonists; Leukotriene C4; Leukotriene D4; Leukotrienes; Male; Membrane Proteins; Middle Aged; Propionates; Pulmonary Artery; Quinolines; Receptors, Leukotriene; SRS-A; Vasoconstriction

The effects of BAY u9773 (6(R)-(4'-carboxyphenylthio)-5(S)-hydroxy-7(E),9(E), 11(Z),14(Z)-eicosatetraenoic acid), a cysteinyl-leukotriene analogue, were investigated on a variety of smooth muscle preparations in order to determine its profile as a cysteinyl-leukotriene receptor antagonist. The tissues were contracted with leukotriene C4 or leukotriene D4 and their receptor characteristics defined as either 'typical' or 'atypical' according to the activity or inactivity, respectively, of the selective antagonists ICI 198615, MK 571 and SKF 104353. BAY u9773 antagonised 'typical' cysteinyl-leukotriene receptors with pA2 (or pKB) values in the range 6.8-7.4 and also antagonised 'atypical' receptors with pA2 values in the range 6.8-7.7. However, BAY u9773 had no effect at 10(-6) M against a selection of non-leukotriene stimuli in the same preparations. BAY u9773 competitively displaced [3H]leukotriene D4 binding to guinea-pig lung homogenate, with a pKi of 7.0 +/- 0.1. In the guinea-pig lung strip, BAY u9773 was found to be inactive at 10(-6)M against leukotriene C4- and leukotriene D4-induced contractions, which may suggest the existence of a third type of cysteinyl-leukotriene receptor. These data demonstrate that BAY u9773 is a selective cysteinyl-leukotriene receptor antagonist with comparable activity at both 'typical' and 'atypical' receptors and as such represents a valuable tool for the study of cysteinyl-leukotriene receptors.

Topics: Animals; Binding, Competitive; Bronchi; Dicarboxylic Acids; Ferrets; Guinea Pigs; In Vitro Techniques; Indazoles; Leukotriene C4; Leukotriene D4; Lung; Male; Muscle Contraction; Muscle Relaxation; Muscle, Smooth; Propionates; Quinolines; Rats; Rats, Wistar; Receptors, Immunologic; Sheep; Spleen; SRS-A; Trachea

Leukotrienes (LT) are potent spasmogenic agents in human isolated bronchial and pulmonary venous muscle preparations. Treatment of human isolated pulmonary veins with the L-serine borate complex (45 mM; 30 min) did not alter the LTC4 pD2 values in these preparations. The cysteinyl LT antagonists, ICI 198615, MK 571 and SKF 104353, significantly shifted to the right the LT concentration-effect curves in airways with pKB values against LTC4 of 8.4 for ICI 198615, 8.6 for MK 571 and 8.0 for SKF 104353. Similar results were found against LTD4. In contrast, these antagonists did not inhibit the LTC4 and LTD4 contractions in human pulmonary veins. LTE4 was a partial agonist on the human pulmonary veins and blocked the contractions with a pKp value of 6.3 against LTD4 and 6.6 against LTC4. An LT analog, BAY u9773, also blocked the LT contractions in bronchial and venous muscle preparations with pKp values against LTD4 and LTC4 of 6.5 and 6.7, respectively. These data provide pharmacological evidence for a second cysteinyl LT receptor in the human lung. One LT receptor (LT-1) is stimulated by all cysteinyl LT, found on airways and inhibited by the LT-1 antagonists, and a second receptor (LT-2) can also be stimulated by all cysteinyl LT and is found on pulmonary veins, resistant to LT-1 antagonists but blocked by LTE4 and the dual LT-1/LT-2 antagonist BAY u9773.

Topics: Adult; Aged; Cysteine; Dicarboxylic Acids; Female; Humans; Leukotriene E4; Lung; Male; Middle Aged; Models, Biological; Muscle Contraction; Propionates; Pulmonary Veins; Quinolines; Receptors, Immunologic; Receptors, Leukotriene; Receptors, Leukotriene B4; SRS-A