prostaglandin-d2 and ramatroban

COPD (Chronic Obstructive Pulmonary Disease) and bronchial asthma are two severe lung diseases which represent a major problem of world public health. Leukotrienes and prostanoids play an important role in the pathogenesis of pulmonary diseases. Prostanoids: prostaglandins (PGs) and thromboxane A2 (TXA2), the cyclooxygenase metabolites of arachidonic acid are implicated in the inflammatory cascade that occurs in asthmatic airways. Recently, the roles played by isoprostanes or prostaglandin-like compounds nonenzymatically generated via peroxidation of membrane phospholipids by reactive oxygen species, in particular F2-isoprostanes, in pulmonary pathophysiology have been highlighted. This article aims to provide an overview of the role of prostanoids and isoprostanes in the pathogenesis of COPD and asthma and to discuss the pharmacological strategies developed in prevention and/or treatment of these pathologies.

Topics: Animals; Asthma; Benzoquinones; Carbazoles; Enzyme Inhibitors; F2-Isoprostanes; Heptanoic Acids; Humans; Methacrylates; Prostaglandin Antagonists; Prostaglandin D2; Pulmonary Disease, Chronic Obstructive; Randomized Controlled Trials as Topic; Receptors, Immunologic; Receptors, Prostaglandin; Receptors, Thromboxane A2, Prostaglandin H2; Sulfonamides; Thromboxane A2; Thromboxane-A Synthase

Prostaglandin D2 (PGD2) is a potent bronchoconstrictor, and is thought to have a role in the pathogenesis of asthma. PGD2 causes vasodilation acting via the prostaglandin (DP) receptor on vascular smooth muscle, and myocontraction acting via the thromboxane (TP) receptor on bronchial smooth muscle. To determine the relative contribution of these mechanisms we have studied the degree to which a potent TP receptor antagonist inhibits PGD2-induced bronchoconstriction. Twelve atopic asthmatic subjects underwent baseline PGD2 bronchial challenges to determine the cumulative concentration of PGD2 required to reduce forced expiratory volume in one second (FEV1) by 20%. At four subsequent randomized visits, subjects received this concentration of PGD2 90 min after dosing with placebo or 20, 50 or 100 mg of BAY u 3405, a potent competitive TP receptor antagonist. Serum was taken for drug assay at 90 min. After each dose of PGD2, FEV1 was measured for 30 min, and the area under the percentage fall in the FEV1/time curve (AUC) was calculated. The mean +/- SEM AUC for placebo was 414 +/- 68, and for the 20, 50 and 100 mg doses of BAY u 3405 was 169 +/- 33, 173 +/- 59 and 135 +/- 63, respectively. There were no significant differences between the AUCs for any of the drug doses, whilst all three doses were significantly different from placebo. The plateau response achieved with increasing doses of the antagonist suggests complete blockade of the TP receptor. These data demonstrate that thromboxane receptor blockade only partially inhibits the airway narrowing response to PGD2, and support the existence of a vascular component to PGD2-induced acute airway narrowing in asthma.

Topics: Adult; Asthma; Bronchial Provocation Tests; Bronchoconstriction; Carbazoles; Double-Blind Method; Forced Expiratory Volume; Humans; Male; Prostaglandin D2; Receptors, Prostaglandin; Receptors, Thromboxane; Sulfonamides; Thromboxanes

Nasal lavage and challenge studies in allergic rhinitis implicate prostaglandin (PG) D2 in the genesis of nasal blockage. PG D2 is known to act via at least two receptors, the thromboxane prostanoid receptor and the PG D2 prostanoid (DP) receptor; the lower airway effects are mediated chiefly by the TP receptor. The receptor involved in the genesis of PG D2-induced nasal blockage is unknown. BAY u 3405 is a potent selective competitive TP receptor antagonist, which inhibits the lower airway response to PG D2, and shifts the dose-response curve to the right by up to 16-fold.. The efficacy of a single oral dose of 20 mg of BAY u 3405 was examined in comparison with PG D2 nasal insufflation in a randomized, double-blind, placebo-controlled crossover study, with objective measurement of nasal resistance by active posterior rhinomanometry.. BAY u 3405 afforded no protection against PG D2-induced nasal blockage.. This suggests that PG D2-induced nasal blockage may be mediated by the DP receptor rather than the TP receptor and that TP receptor antagonists are unlikely to be of benefit in the treatment of allergic rhinitis. In vivo investigation with specific potent DP receptor antagonists is awaited.

Topics: Adult; Aged; Airway Resistance; Carbazoles; Dose-Response Relationship, Drug; Double-Blind Method; Humans; Male; Middle Aged; Nasal Provocation Tests; Nose; Prostaglandin D2; Receptors, Thromboxane; Rhinitis, Allergic, Perennial; Sulfonamides

1. The potent bronchoconstrictors prostaglandin (PG) D2, PG F2 alpha and thromboxane A2 are thought to have a role in the pathogenesis of asthma, mediated via the thromboxane (TP) receptor. 2. BAY u 3405 is a new potent selective competitive TP receptor antagonist. 3. The effect of single oral doses of 20 mg and 50 mg BAY u 3405 was examined against histamine and PG D2 bronchial provocation at 90 min after drug ingestion and, for the 20 mg dose alone, at 60 min after ingestion, in randomised, double-blind placebo controlled crossover studies. A time course study was performed with the 20 mg dose. 4. BAY u 3405 protected against PG D2 bronchial provocation. The 20 mg dose increased the amount of PG D2 required to produce a fall of 20% in the forced expiratory volume in 1 s by 6-fold and 16-fold at 60 min and 90 min after ingestion respectively, and the 50 mg dose by 14-fold at 90 min after ingestion. 5. The specificity of the drug was confirmed in vivo in that there was no significant protection against histamine bronchial provocation at either dose or at either time point. 6. The time course study showed significant protection against PG D2 bronchial provocation at 1 h and at 3 h after a single 20 mg oral dose. 7. There was no correlation between subjects in plasma BAY u 3405 concentration and drug effect. Within the subjects performing the time course study there was a strong correlation in time between drug effect and plasma BAY u 3405 concentration.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adult; Asthma; Bronchial Provocation Tests; Bronchoconstriction; Carbazoles; Female; Histamine; Histamine Antagonists; Humans; Male; Middle Aged; Prostaglandin D2; Respiratory Function Tests; Sulfonamides; Thromboxanes

In the pathogenesis of exercise-induced bronchoconstriction (EIB), prostaglandin D2 (PGD2) may play a role as a newly generated, mast cell-derived mediator. As the bronchoconstrictor effects of PGD2 are predominantly mediated via stimulation of thromboxane receptors in the lung, we studied a novel, orally effective, thromboxane-receptor antagonist, BAY u 3405, on EIB in 12 male subjects with mild asthma. On 4 study days, we determined, in a randomized, double-blind, placebo-controlled, crossover fashion, the effects of 20 mg of BAY u 3405 administered orally 1 hour before PGD2 and exercise challenges, respectively. Increasing dosages of PGD2 were inhaled to establish dose-response curves that allowed determination of the provocative concentration necessary to decrease FEV1 by at least 20% (PC20) and to increase specific airway resistance (SR(aw)) by 100% (PC100). EIB was measured as a maximal fall/increase in postexertional FEV1/SR(aw) after bicycle exercise and cold-air breathing. Prechallenge lung-function values were similar on all four occasions. BAY u 3405 did not elicit any effect on resting bronchial tone. After placebo, the geometric means (SD) of PC20 and PC100 were 0.0380 (2.6) and 0.0266 (2.4) mg/ml, increasing to 0.554 (5.9) and 0.143 (8.1) mg/ml after BAY u 3405 (p = 0.0002). Mean (SD) maximal postexertional decrease in FEV1 and increase in SR(aw) after placebo was 29.4% (16.4%) and 280% (135%), and after BAY u 3405, 31.4% (18.1%) and 379% (281%) (not significant). No clinically relevant BAY u 3405-related side effects were observed. From these results we conclude that BAY u 3405 is highly effective in attenuating PGD2-induced bronchoconstriction.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Air; Airway Resistance; Asthma, Exercise-Induced; Bronchial Provocation Tests; Bronchoconstriction; Carbazoles; Cold Temperature; Dose-Response Relationship, Drug; Double-Blind Method; Drug Evaluation; Forced Expiratory Volume; Humans; Male; Prostaglandin D2; Receptors, Prostaglandin; Receptors, Thromboxane; Sulfonamides; Thromboxanes; Time Factors

Eosinophilic esophagitis is characterized by eosinophil-predominant inflammation in the esophagus. How eosinophils migrate and infiltrate into the esophagus, however, is less clear. Our previous study demonstrated that mast cell activation led to eosinophil infiltration in the esophagus. Prostaglandin D2 (PGD2) is an important mediator released from activated mast cells. The present study aims to determine whether PGD2 induces eosinophil infiltration into the esophagus via a d-type prostanoid receptor 2 (DP2) receptor-dependent mechanism. Using an in vivo guinea pig model, PGD2, d-type prostanoid receptor 1 (DP1) agonist, or DP2 agonist were injected into the esophagus. Esophageal tissues were removed 2 hours after injections and proceeded to either hematoxylin-eosin (HE) staining or immunofluorescent staining of eosinophil major basic protein (MBP) to compare each treatment-induced eosinophil infiltration in the esophagus. In a separate study, ovalbumin (OVA)-sensitized guinea pigs were pretreated with either DP2 or DP1 antagonists, followed by inhalation of OVA to induce mast cell activation. Esophageal tissues were then processed for immunofluorescent staining of MBP. PGD2 injection in the esophagus led to an increase of eosinophil infiltration in esophageal epithelium at the injection site as revealed by HE staining. Increased infiltration of eosinophils was further confirmed by the increased presence of MBP-labeled immunopositive (MBP-LI) cells in esophageal epithelium. Injection with DP2 agonist 15(R)-PGD2, but not DP1 agonist BW 245C, mimicked the PGD2-induced response. In OVA-sensitized animals, antigen inhalation increased MBP-LI cells in esophageal epithelium. Pretreatment with DP2 antagonist BAY-u3405, but not DP1 antagonist BW 868C, inhibited the antigen inhalation-induced increase of MBP-LI cells in esophageal epithelium. These data support the hypothesis that PGD2 induces eosinophil trafficking into the esophageal epithelium via a DP2-mediated pathway, suggesting a role of DP2 antagonist in the prevention of eosinophilic esophagitis.

Topics: Animals; Carbazoles; Cell Movement; Eosinophil Major Basic Protein; Eosinophilic Esophagitis; Eosinophils; Epithelium; Esophagus; Guinea Pigs; Hydantoins; Male; Mast Cells; Ovalbumin; Prostaglandin Antagonists; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides

Chemoattractant receptor homologous molecule expressed on T helper type 2 cells (CRTH2) is a PGD2 receptor found on eosinophils, basophils, and Th2 type T cells which exhibits chemotaxis and functions in activation cascades. However, while a number of CRTH2 antagonists, including ramatroban, are known to exert activity in certain animal models, activity in a guinea pig model of EA-induced airway hyperresponsiveness has not been demonstrated. The newly developed CRTH2 antagonist ASP5642 has shown antagonistic activity against human and guinea pig CRTH2 in previous studies and has also been found effective in treating guinea pig models of airway inflammation and airway hyperresponsiveness. While previous studies have used animals such as rats and mice to evaluate CRTH2 antagonist effects, ours is the first attempt to evaluate CRTH2 function in a guinea pig asthma model, which may prove useful in evaluating the compound's effects in humans, given the comparable airway function between the two species taken together, these data from the present study strongly suggest the utility of ASP5642 in investigating the role of CRTH2 in inflammatory responses and as a drug treatment for human asthma.

Topics: Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Antigens; Benzhydryl Compounds; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Carbazoles; Cell Count; Eosinophilia; Guinea Pigs; HEK293 Cells; Humans; K562 Cells; Male; Ovalbumin; Pneumonia; Prostaglandin D2; Pyridazines; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides

Prostaglandin D2 (PGD2) is the most abundant prostaglandin in brain but its effect on neuronal cell death is complex and not completely understood. PGD2 may modulate neuronal cell death via activation of DP receptors or its metabolism to the cyclopentenone prostaglandins (CyPGs) PGJ2, Δ(12)-PGJ2 and 15-deoxy-Δ(12,14)-PGJ2, inducing cell death independently of prostaglandin receptors. This study aims to elucidate the effect of PGD2 on neuronal cell death and its underlying mechanisms. PGD2 dose-dependently induced cell death in rat primary neuron-enriched cultures in concentrations of ≥10μM, and this effect was not reversed by treatment with either DP1 or DP2 receptor antagonists. Antioxidants N-acetylcysteine (NAC) and glutathione which contain sulfhydryl groups that can bind to CyPGs, but not ascorbate or tocopherol, attenuated PGD2-induced cell death. Conversion of PGD2 to CyPGs was detected in neuronal culture medium; treatment with these CyPG metabolites alone exhibited effects similar to those of PGD2, including apoptotic neuronal cell death and accumulation of ubiquitinated proteins. Disruption of lipocalin-type prostaglandin D synthase (L-PGDS) protected neurons against hypoxia. These results support the hypothesis that PGD2 elicits its cytotoxic effects through its bioactive CyPG metabolites rather than DP receptor activation in primary neuronal culture.

Topics: Animals; Apoptosis; Carbazoles; Cells, Cultured; Cerebral Cortex; Cyclopentanes; Dose-Response Relationship, Drug; Embryo, Mammalian; Hypoxia; Intramolecular Oxidoreductases; Lipocalins; Mice; Mice, Knockout; Neurons; Prostaglandin D2; Rats; Rats, Sprague-Dawley; Receptors, Prostaglandin; Sulfonamides

PGD2 (prostaglandin D2) is a mediator in various pathophysiological processes, including inflammation and tumorigenesis. PGD2 can be converted into active metabolites and is known to activate two distinct receptors, DP (PGD2 receptor) and CRTH2/DP2 (chemoattractant receptor-homologous molecule expressed on Th2 cells). In the past, PGD2 was thought to be involved principally in the process of inflammation. However, in recent years, several studies have shown that PGD2 has anti-proliferative ability against tumorigenesis and can induce cellular apoptosis via activation of the caspase-dependent pathway in human colorectal cancer cells, leukaemia cells and eosinophils. In the lung, where PGD2 is highly released when sensitized mast cells are challenged with allergen, the mechanism of PGD2-induced apoptosis is unclear. In the present study, A549 cells, a type of NSCLC (non-small cell lung carcinoma), were treated with PGD2 under various conditions, including while blocking DP and CRTH2/DP2 with the selective antagonists BWA868C and ramatroban respectively. We report here that PGD2 induces A549 cell death through the intrinsic apoptotic pathway, although the process does not appear to involve either DP or CRTH2/DP2. Similar results were also found with H2199 cells, another type of NSCLC. We found that PGD2 metabolites induce apoptosis effectively and that 15d-PGJ2 (15-deoxy-Δ12,14-prostaglandin J2) is a likely candidate for the principal apoptotic inducer in PGD2-induced apoptosis in NSCLC A549 cells.

Topics: Apoptosis; Carbazoles; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Cell Survival; Chromatography, High Pressure Liquid; Cytochromes c; Cytosol; DNA Fragmentation; Humans; Hydantoins; Mitochondria; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides

Mast cells release mediators upon stimulation that contribute to the pathogenesis of chronic airway disease, including the recruitment and activation of Th2 lymphocytes. The objective was to determine the involvement of prostaglandin D(2) (PGD(2)) and its receptors in the chemotaxis of Th2 cells, using nasal polyp tissue.. Tissue explants from ten patients with nasal polyposis were incubated with RPMI alone or RPMI containing IgE/anti-IgE for 30 min. Some samples were treated with diclofenac to inhibit the production of PGD(2). Supernatants were assayed for PGD(2) content and for their ability to promote human Th2 cell chemotaxis in the presence and absence of a CRTH2 antagonist. Transcript levels of D protanoid receptor type 1 (DP(1)), chemoattractant receptor-homologous receptor expressed on Th2 cells (CRTH2) and PGD(2) synthase were analysed by real time PCR.. Increased release of PGD(2) by nasal polyp tissue treated with IgE/anti-IgE was significantly inhibited by preincubation of the tissue with diclofenac. Transcript levels of PGD(2) synthase, DP(1) and CRTH2 receptors increased after stimulation with IgE/anti-IgE. Supernatants from IgE/anti-IgE-stimulated nasal polyp tissue caused significantly increased chemotaxis of Th2 cells. The levels of PGD(2) produced and the degree of Th2 cell chemotaxis were highly correlated. Diclofenac inhibited the production of Th2 cell chemotactic activity, and the chemotactic effect of the supernatant on Th2 cells was inhibited by the CRTH2 antagonist ramatroban.. These data suggest that in immunologically activated nasal polyp tissue, PGD(2) produced by mast cells promotes the migration of Th2 cells through a CRTH2 dependent mechanism.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Antibodies, Anti-Idiotypic; Carbazoles; Chemotaxis, Leukocyte; Diclofenac; Humans; Immunoglobulin E; Lymphocyte Activation; Mast Cells; Nasal Polyps; Platelet Aggregation Inhibitors; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides; Th2 Cells

To investigate the effect of prostaglandin D2 receptor antagonists on the airway inflammation in rats with asthma.. Forty male SD rats were randomly divided into four groups: Group A (normal control), Group B (asthma group), Group C (CRTH2 antagonist BAYu3405 treatment group), Group D (DP1 antagonist BWA868C treatment group). Asthma was induced by ovalbumin (OVA) challenge. The rats in each group were sacrificed 24 h after the last challenge of OVA.DP1/CRTH2 receptors on eosinophils (EOS) were measured by radiological binding assay (RBA). The left lungs were used for histological examinations and bronchoalveolar lavage fluid (BALF) was collected from the right lungs. The total cell numbers, EOS absolute count and differential cell counts in BALF were performed. Serum concentrations of IL-4, 5 and IFN-gamma were measured by ELISA.. Rats in BAYu3405 treatment group showed profoundly decreased infiltrates of EOS and lymphocytes in the wall of bronchus when compared with those of asthma group and BWA868C treatment group. Serum concentrations of IFN-gamma in rats of BAYu3405 treatment group increased, but IL-4 and IL-5 decreased significantly when compared with those in rats of asthma group and BWA868C treatment group (P<0.01), and BALF EOS count was decreased significantly (P<0.01). Peripheral blood EOS count was higher than that in rats of normal control group, but was not significantly different from that in rats of asthma group and BWA868C treatment group. The combining capacity of CRTH2 and DP total combining capacity on EOS in asthma group, BAYu3405 treatment group and BWA868C treatment group were significantly higher than those in Group A (P<0.01). There was no significant difference in DP1 among all the groups (P>0.05).. CRTH2, but not DP1 antagonist can effectively ameliorate airway inflammation in rats with asthma.

Topics: Animals; Asthma; Bronchi; Carbazoles; Inflammation; Male; Ovalbumin; Prostaglandin D2; Random Allocation; Rats; Rats, Sprague-Dawley; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides

Prostaglandin D(2) (PGD(2)) has been shown to cause eosinophil, basophil and Th2 cell chemotaxis in vitro and in vivo through an action on the prostaglandin CRTH2 receptor. In the present study, the dependence of PGD(2)-induced eosinophil accumulation in vivo on interleukin-5 (IL-5) blood eosinophilia was investigated.. Guinea-pigs were exposed to aerosols of 13,14di-hydro 15-keto PGD(2) (DK-PGD(2)) or platelet activating factor (PAF) and the eosinophil content of the bronchoalveolar lavage fluid or blood determined. In some experiments, DK-PGD(2) was administered systemically and eosinophilia measured.. Aerosols of DK-PGD(2) caused eosinophil accumulation in the lungs 24h after exposure. DK-PGD(2) (10 microg x ml(-1)) -induced airway eosinophilia was inhibited when animals were treated with the CRTH2 receptor antagonist ramatroban. 1-4h after exposure to DK-PGD(2) a significant decrease in blood eosinophil count was measured. The anti-IL-5 antibody TRFK-5 had no inhibitory effect of DK-PGD(2)-induced airway eosinophilia, but abolished airway eosinophilia induced by exposure of guinea-pigs to aerosols of PAF. Intracardiac injection of DK-PGD(2) induced a dose-related increase in blood eosinophil numbers.. It is concluded that, in the guinea-pig, DK-PGD(2)-induced airway eosinophilia is mediated by an action on prostaglandin CRTH2 receptors and that this response appears to be independent of IL-5.

Topics: Animals; Carbazoles; Eosinophilia; Female; Guinea Pigs; Humans; Interleukin-5; Lung; Platelet Aggregation Inhibitors; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides

Chemoattractant receptor homologous molecule of Th2 cells (CRTH2) has been shown to mediate the chemotaxis of eosinophils, basophils and Th2-type T lymphocytes. The major mast cell product prostaglandin (PG) D(2) is considered to be the principal ligand of CRTH2.. We developed a novel CRTH2 antagonist, AZ11665362 [2,5-dimethyl-3-(8-methylquinolin-4-yl)-1H-indole-1-yl]acetic acid, and characterized its efficacy in binding assay in HEK293 cells, eosinophil and basophil shape change assay and migration assay, platelet aggregation and eosinophil release from guinea pig bone marrow. The effects were compared with ramatroban, the sole CRTH2 antagonist clinically available to date.. AZ11665362 bound with high affinity to human and guinea pig CRTH2 expressed in HEK293 cells and antagonized eosinophil and basophil shape change responses to PGD(2). AZ11665362 was without effect on the PGD(2)-induced inhibition of platelet aggregation. In contrast, AZ11665362 effectively inhibited the in vitro migration of human eosinophils and basophils towards PGD(2). The release of eosinophils from the isolated perfused hind limb of the guinea pig was potently stimulated by PGD(2), and this effect was prevented by AZ11665362. In all assays tested, AZ11665362 was at least 10 times more potent than ramatroban.. AZ11665362 is a potent CRTH2 antagonist that is capable of blocking the migration of eosinophils and basophils, and the rapid mobilization of eosinophils from bone marrow. AZ11665362 might hence be useful for the treatment of allergic diseases.

Topics: Animals; Basophils; Bone Marrow; Carbazoles; Cell Movement; Chemotaxis; Guinea Pigs; Humans; Platelet Aggregation Inhibitors; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides; Th2 Cells

PGD(2) is the major prostanoid produced during the acute phase of allergic reactions. Two PGD(2) receptors have been isolated, DP and CRTH2 (chemoattractant receptor-homologous molecule expressed on Th2 cells), but whether they participate in the pathophysiology of allergic diseases remains unclear. We investigated the role of CRTH2 in the initiation of allergic rhinitis in mice. First, we developed a novel murine model of pollinosis, a type of seasonal allergic rhinitis. Additionally, pathophysiological differences in the pollinosis were compared between wild-type and CRTH2 gene-deficient mice. An effect of treatment with ramatroban, a CRTH2/T-prostanoid receptor dual antagonist, was also determined. Repeated intranasal sensitization with Cry j 1, the major allergen of Cryptomeria japonica pollen, in the absence of adjuvants significantly exacerbated nasal hyperresponsive symptoms, Cry j 1-specific IgE and IgG1 production, nasal eosinophilia, and Cry j 1-induced in vitro production of IL-4 and IL-5 by submandibular lymph node cells. Additionally, CRTH2 mRNA in nasal mucosa was significantly elevated in Cry j 1-sensitized mice. Following repeated intranasal sensitization with Cry j 1, CRTH2 gene-deficient mice had significantly weaker Cry j 1-specific IgE/IgG1 production, nasal eosinophilia, and IL-4 production by submandibular lymph node cells than did wild-type mice. Similar results were found in mice treated with ramatroban. These results suggest that the PGD(2)-CRTH2 interaction is elevated following sensitization and plays a proinflammatory role in the pathophysiology of allergic rhinitis, especially pollinosis in mice.

Topics: Allergens; Animals; Antigens, Plant; Carbazoles; Cryptomeria; Cytokines; Disease Models, Animal; Eosinophils; Immunoglobulins; Mice; Mice, Inbred BALB C; Mice, Mutant Strains; Nasal Mucosa; Nasal Septum; Plant Proteins; Pollen; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Rhinitis, Allergic, Seasonal; Sulfonamides; Th2 Cells

This study was undertaken to investigate the interactive effect of histamine and prostaglandin D(2) in nasal allergic symptoms in rats. The intranasal application of histamine at doses lower than 10 mumol/site caused no sneezing or nasal rubbing. In addition, prostaglandin D(2) also showed no significant increase in these responses, even at a dose of 10 nmol/site. On the other hand, the simultaneous instillation of histamine and prostaglandin D(2) resulted in a 1000 times more potent effect in inducing nasal symptoms than the administration of histamine alone. Thus, prostaglandin D(2) enhanced the actions of histamine in inducing sneezing and nasal rubbing in a dose-dependent manner, and significant effects were observed at doses higher than 1 nmol/site. The responses induced by the simultaneous application of histamine and prostaglandin D(2) were inhibited by chlorpheniramine, cyproheptadine, BW A868C and ramatroban. Chlorpheniramine and cyproheptadine showed the dose-related inhibition of nasal symptoms induced by the combined administration of histamine (10 nmol) and prostaglandin D(2) (10 nmol), but the effect of cyproheptadine was relatively weak compared with chlorpheniramine. Moreover, BW A868C and ramatroban also showed the inhibition of nasal symptoms induced by the simultaneous administration of histamine and prostaglandin D(2) in a dose-dependent manner. BW A868C was more potent in inhibiting the nasal symptoms than ramatroban. These results clearly indicate that prostaglandin D(2) showed a synergistic effect on sneezing and nasal rubbing induced by histamine in rats, and its effect occurred through both prostaglandin D(2) and CRTH2 (chemoattractant receptor-homologous molecule expressed on TH2 cells) receptors.

Topics: Administration, Intranasal; Administration, Oral; Animals; Carbazoles; Chlorpheniramine; Cyproheptadine; Dose-Response Relationship, Drug; Drug Synergism; Histamine; Histamine Agents; Histamine H1 Antagonists; Hydantoins; Injections, Intravenous; Nasal Mucosa; Prostaglandin D2; Rats; Rats, Wistar; Receptors, Immunologic; Receptors, Prostaglandin; Rhinitis; Sneezing; Sulfonamides

Activation of human CRTH2(+) CD4(+) T helper type 2 (Th2) cells with anti-CD3/anti-CD28 led to time-dependent production of prostaglandin D(2) (PGD(2)) which peaked at 8 hr. The production of PGD(2) was completely inhibited by cotreatment with the cyclo-oxygenase inhibitor diclofenac (10 microm) but was not affected by cotreatment with ramatroban, a dual antagonist of both the thromboxane-like prostanoid (TP) receptor and the chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2). Supernatants from activated CRTH2(+) CD4(+) Th2 cells caused a concentration-dependent increase in the migration of naive CRTH2(+) CD4(+) Th2 cells compared to supernatants from unstimulated CRTH2(+) CD4(+) Th2 cells. The level of chemotactic activity peaked at 8 hr after activation, corresponding to the peak levels of PGD(2), but production of chemotactic activity was only partially inhibited by the cyclo-oxygenase inhibitor diclofenac. In contrast, ramatroban completely inhibited the chemotactic responses of naive Th2 cells to supernatants from activated CRTH2(+) CD4(+) Th2 cells collected up to 8 hr after activation, although supernatants collected 24 hr after activation were less sensitive to inhibition by ramatroban. The selective TP antagonist SQ29548 did not inhibit migration of Th2 cells, implicating CRTH2 in this response. These data suggest that CRTH2 plays an important paracrine role in mediating chemotactic activation of Th2 cells. Interestingly, although PGD(2) is produced from Th2 cells and contributes to this paracrine activation, it appears that additional CRTH2 agonist factors are also produced by activated Th2 cells and the production of these factors occurs independently of the cyclo-oxygenase pathway of the arachidonic acid metabolism.

Topics: Carbazoles; CD28 Antigens; CD3 Complex; Cells, Cultured; Chemotaxis, Leukocyte; Culture Media, Conditioned; Cyclooxygenase Inhibitors; Cytokines; Diclofenac; Humans; Lymphocyte Activation; Paracrine Communication; Prostaglandin D2; Receptors, Formyl Peptide; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides; Th2 Cells

Several chemoattractants can regulate the recruitment of eosinophils to sites of inflammation, but the hierarchy among them is unknown. We observed here that eosinophil chemotaxis towards eotaxin or 5-oxo-6,8,11,14-eicosatetraenoic acid (5-oxo-ETE) was amplified up to sixfold in the presence of prostaglandin (PG) D2. This effect was only seen in eosinophils, and not in neutrophils or basophils. Pretreatment with the chemoattractant receptor-homologous molecule expressed on TH2 cells (CRTH2) antagonist ramatroban prevented the PGD2 enhancement of eosinophil migrations. In contrast, eotaxin or 5-oxo-ETE inhibited the migration of eosinophils towards PGD2. 5-oxo-ETE enhanced the chemotaxis to eotaxin, while eotaxin had no effect on 5-oxo-ETE-induced migration. 5-oxo-ETE induced the phosphorylation of p38 mitogen-activated protein kinase, and inhibition of p38 mitogen-activated protein kinase by SB-202190 converted the effect of 5-oxo-ETE on the chemotaxis to PGD2 from inhibition to enhancement. The presence of blood or plasma markedly decreased the sensitivity of eosinophils to eotaxin or 5-oxo-ETE, while responses to PGD2 were unaltered. In conclusion, PGD2 might be an initial chemoattractant, since it maintains its potency in the circulation and augments the responsiveness of eosinophils to other chemoattractants. In contrast, eotaxin seems to be an end-point chemoattractant, since it has reduced efficacy in blood and is capable of down-modulating eosinophil responsiveness to other chemoattractants.

Topics: Arachidonic Acids; Carbazoles; Chemokine CCL11; Chemokines, CC; Chemotactic Factors, Eosinophil; Chemotaxis, Leukocyte; Eosinophils; Humans; p38 Mitogen-Activated Protein Kinases; Platelet Aggregation Inhibitors; Prostaglandin D2; Sulfonamides

Human cultured mast cells, immunologically activated with immunoglobuin E (IgE)/anti-IgE, released a factor(s) that promoted chemotaxis of human CRTH2+ CD4+ T helper type 2 (Th2) lymphocytes. Mast cell supernatants collected at 20 min, 1 hr, 2 hr and 4 hr after activation caused a concentration-dependent increase in the migration of Th2 cells. The effect of submaximal dilutions of mast-cell-conditioned media was inhibited in a dose-dependent manner by ramatroban (IC50 = 96 nm), a dual antagonist of both the thromboxane-like prostanoid (TP) receptor and the chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2), but not by the selective TP antagonist SQ29548, implicating CRTH2 in mediating the chemotactic response of these Th2 cells. The effect of mast-cell-conditioned media was mimicked by prostaglandin D2 (PGD2) and this eicosanoid was detected in the conditioned media from activated mast cells in concentrations sufficient to account for the activity of the mast cell supernatants. Treatment of the mast cells with the cyclo-oxygenase inhibitor diclofenac (10 microm) inhibited both the production of PGD2 and the CRTH2+ CD4+ Th2-stimulatory activity, while addition of exogenous PGD2 to conditioned media from diclofenac-treated mast cells restored the ability of the supernatants to promote chemotaxis of these Th2 cells. The degree of inhibition caused by diclofenac treatment of the mast cells was concordant with the degree of inhibition of chemotactic responses afforded by CRTH2 blockade. These data suggest that PGD2, or closely related metabolites of arachidonic acid, produced from mast cells may play a central role in the activation of CRTH2+ CD4+ Th2 lymphocytes through a CRTH2-dependent mechanism.

Topics: Carbazoles; Cells, Cultured; Chemotaxis, Leukocyte; Culture Media, Conditioned; Cyclooxygenase Inhibitors; Diclofenac; Dose-Response Relationship, Immunologic; Humans; Immunoglobulin E; Lymphocyte Activation; Mast Cells; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Receptors, Thromboxane; Sulfonamides; Th2 Cells

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

PGD2, produced by mast cells, has been detected in high concentrations at sites of allergic inflammation. It can stimulate vascular and other inflammatory responses by interaction with D prostanoid receptor (DP) and chemoattractant receptor-like molecule expressed on Th2 cells (CRTH2) receptors. A significant role for PGD2 in mediating allergic responses has been suggested based on the observation that enhanced eosinophilic lung inflammation and cytokine production is apparent in the allergen-challenged airways of transgenic mice overexpressing human PGD2 synthase, and PGD2 can enhance Th2 cytokine production in vitro from CD3/CD28-costimulated Th2 cells. In the present study, we investigated whether PGD2 has the ability to stimulate Th2 cytokine production in the absence of costimulation. At concentrations found at sites of allergic inflammation, PGD2 preferentially elicited the production of IL-4, IL-5, and IL-13 by human Th2 cells in a dose-dependent manner without affecting the level of the anti-inflammatory cytokine IL-10. Gene transcription peaked within 2 h, and protein release peaked approximately 8 h after stimulation. The effect of PGD2 was mimicked by the selective CRTH2 agonist 13,14-dihydro-15-keto-PGD2 but not by the selective DP agonist BW245C, suggesting that the stimulation is mediated by CRTH2 and not DP. Ramatroban, a dual CRTH2/thromboxane-like prostanoid receptor antagonist, markedly inhibited Th2 cytokine production induced by PGD2, while the selective thromboxane-like prostanoid receptor antagonist SQ29548 was without effect. These data suggest that PGD2 preferentially up-regulates proinflammatory cytokine production in human Th2 cells through a CRTH2-dependent mechanism in the absence of any other costimulation and highlight the potential utility of CRTH2 antagonists in the treatment of allergic diseases.

Topics: Base Sequence; Bridged Bicyclo Compounds, Heterocyclic; Carbazoles; Cells, Cultured; Cytokines; DNA; Fatty Acids, Unsaturated; Humans; Hydantoins; Hydrazines; Inflammation; Inflammation Mediators; Interleukin-13; Interleukin-4; Interleukin-5; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides; Th2 Cells; Up-Regulation

Thromboxane (TX) A(2), a cyclooxygenase-derived mediator involved in allergic responses, is rapidly converted in vivo to a stable metabolite, 11-dehydro-TXB(2), which is considered to be biologically inactive. In this study, we found that 11-dehydro-TXB(2), but not the TXA(2) analogue U46,619 or TXB(2), activated eosinophils and basophils, as assayed by flow cytometric shape change. 11-Dehydro-TXB(2) was also chemotactic for eosinophils but did not induce, nor inhibit, platelet aggregation. Chemoattractant receptor-homologous molecule expressed on TH2 cells (CRTH2) is an important chemoattractant receptor expressed by eosinophils, basophils, and TH2 lymphocytes, and prostaglandin (PG)D(2) has been shown to be its principal ligand. 11-Dehydro-TXB(2) induced calcium flux mainly from intracellular stores in eosinophils, and this response was desensitized after stimulation with PGD(2) but not other eosinophil chemoattractants. Shape change responses of eosinophils and basophils to 11-dehydro-TXB(2) were inhibited by the thromboxane (TP)/CRTH2 receptor antagonist ramatroban, but not the selective TP antagonist SQ29,548, and were insensitive to pertussis toxin. The phospholipase C inhibitor U73,122 attenuated both 11-dehydro-TXB(2)- and PGD(2)-induced shape change. 11-Dehydro-TXB(2) also induced the chemotaxis of BaF/3 cells transfected with hCRTH2 but not naive BaF/3 cells. At a threshold concentration, 11-dehydro-TXB(2) had no antagonistic effect on CRTH2-mediated responses as induced by PGD2. These data show that 11-dehydro-TXB(2) is a full agonist of the CRTH2 receptor and hence might cause CRTH2 activation in cellular contexts where PGD-synthase is not present. Given its production in the allergic lung, antagonism of the 11-dehydro-TXB(2)/CRTH2axis may be of therapeutic relevance.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Basophils; Bridged Bicyclo Compounds, Heterocyclic; Calcium; Carbazoles; Cell Size; Chemotaxis, Leukocyte; Enzyme Inhibitors; Eosinophils; Fatty Acids, Unsaturated; Flow Cytometry; Humans; Hydrazines; Pertussis Toxin; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides; Thromboxane B2; Transfection; Type C Phospholipases

Chemoattractant receptor-homologous molecule expressed on Th2 lymphocytes, CRTH2, is a cognate receptor for prostaglandin (PG) D(2) and, in humans, is suggested to play a functional role in Th2-dependent allergic inflammation. While peripheral blood leukocytes expressing high levels of surface CRTH2 have been detected in disease, little is known of the functional significance of CRTH2 in disease etiology. We have utilized a Th2-dependent murine model of FITC-induced contact hypersensitivity to assess the role, if any, CRTH2-PGD(2) may play in the elicitation or maintenance of such pathobiology. Expression of both PGD(2) and CRTH2 in lesional skin was paralleled by the release of the chemoattractants LTB(4) and the chemokine KC, as well as a profuse dermal neutrophilic and eosinophilic infiltrate, closely paralleling the acute inflammatory pathology observed in human atopic dermatitis. A small molecule CRTH2 antagonist, but not a selective PGD(2)R (DP) receptor antagonist, was able to completely abrogate these responses. Inflammatory cascades mediated by CRTH2 ligation may therefore represent an important early step in the elicitation and maintenance of Th2-dependent skin inflammation.

Topics: Animals; Carbazoles; Chemokine CXCL1; Chemokines; Chemokines, CXC; Cytokines; Dermatitis, Allergic Contact; Dermatitis, Atopic; Eosinophils; Female; Inflammation; Leukotriene B4; Mice; Mice, Inbred BALB C; Neutrophil Activation; Neutrophils; Platelet Aggregation Inhibitors; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Signal Transduction; Sulfonamides; Th2 Cells

We cloned, expressed, and characterized in vitro and in vivo the gene encoding the rat ortholog of chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2), a G protein-coupled receptor for prostaglandin D2 (PGD2). Quantitative reverse transcription-polymerase chain reaction analysis demonstrated highest CRTH2 expression in the lung, brain, ovary, and spleen. Pharmacologically, rat CRTH2 stably transfected in mouse preB lymphoma L1.2 cells behaved very similar compared with the mouse and human orthologs, showing a binding affinity for PGD2 of 11 nM, a functional calcium mobilization when exposed to agonist, and similar sensitivity to agonists and antagonists. In vivo, selective activation of CRTH2 by 13,14-dihydro-15-keto (DK)-PGD2 injection into rats led to a dose- and time-dependent increase of the number of leukocytes in the peripheral blood. Specifically, eosinophils, lymphocytes, and neutrophils were recruited with maximum effects seen 60 min after the injection of 300 microg of DK-PGD2 per rat. Pretreatment of the animals with the CRTH2/thromboxane A2 receptor antagonist, ramatroban, completely abrogated DK-PGD2-induced eosinophilia, suggesting that CRTH2 might have a physiological and/or pathophysiological role in controlling leukocyte migration.

Topics: Amino Acid Sequence; Animals; Base Sequence; Bone Marrow; Carbazoles; Cloning, Molecular; Eosinophilia; Humans; Leukocyte Count; Leukocytes; Mice; Molecular Sequence Data; Prostaglandin D2; Rats; Rats, Wistar; Receptors, Immunologic; Receptors, Prostaglandin; Sequence Homology, Amino Acid; Sulfonamides; Th2 Cells

Ramatroban (Baynas, BAY u3405), a thromboxane A(2) (TxA(2)) antagonist marketed for allergic rhinitis, has been shown to partially attenuate prostaglandin (PG)D(2)-induced bronchial hyperresponsiveness in humans, as well as reduce antigen-induced early- and late-phase inflammatory responses in mice, guinea pigs, and rats. PGD(2) is known to induce eosinophilia following intranasal administration, and to induce eosinophil activation in vitro. In addition to the TxA(2) receptor, PGD(2) is known as a ligand for the PGD(2) receptor, and the newly identified G-protein-coupled chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2). To fully characterize PGD(2)-mediated inflammatory responses relevant to eosinophil activation, further analysis of the mechanism of action of ramatroban has now been performed. PGD(2)-stimulated human eosinophil migration was shown to be mediated exclusively through activation of CRTH2, and surprisingly, these effects were completely inhibited by ramatroban. This is also the first report detailing an orally bioavailable small molecule CRTH2 antagonist. Our findings suggest that clinical efficacy of ramatroban may be in part mediated through its action on this Th2-, eosinophil-, and basophil-specific chemoattractant receptor.

Topics: Administration, Oral; Biological Availability; Calcium; Carbazoles; Cell Movement; Drug Interactions; Eosinophils; Humans; In Vitro Techniques; Platelet Aggregation Inhibitors; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides; Transfection

1. The novel thromboxane (TX) antagonist, BAY u3405, has been evaluated against bronchoconstriction induced by the TXA2 mimetic U-46619, prostaglandin D2 (PGD2), 5-hydroxytryptamine (5-HT), leukotriene D4 (LTD4) and histamine in the guinea-pig in vivo by use of a modification of the model described by Konzett & Rössler. 2. When given intravenously (i.v.) at 30 or 100 micrograms kg-1, U-46619 caused 80% maximal bronchoconstriction in most animals. In contrast, PGD2 caused a smaller 40%-50% maximal bronchoconstriction at the highest dose tested (300 micrograms kg-1, i.v.). 3. BAY u3405, given intravenously, orally (p.o.) or by aerosol antagonized U-46619-induced bronchoconstriction in a dose-related manner. The approximate ID50 values were 600 micrograms kg-1, i.v., 1.7 mg kg-1 p.o. and 0.1% w/v 20 breaths by aerosol. 4. BAY u3405 had similar inhibitory activities against U-46619-induced bronchoconstriction and hypertension suggesting that it had no preferential activity on the airways. 5. When given intravenously BAY u3405 antagonized the bronchoconstrictor effect of intravenous PGD2 with ID50 values between 30-100 micrograms kg-1. 6. The action of BAY u3405 (10 mg kg-1, p.o.) was long lasting, causing significant inhibition of U-46619-induced bronchoconstriction 7 h after dosing. 7. At 1 mg kg-1, i.v., a dose that abolished the response to U-46619 and PGD2, BAY u3405 had no effect on histamine-, 5-HT- or LTD4-induced bronchoconstriction. 8. BAY u3405 potently and selectively antagonized U-46619- or PGD2-induced bronchoconstriction in the Konzett-Rössler model of guinea-pig lung function. It should therefore prove to be a useful tool for defining the role of TXA2- and PGD2 in airway diseases such as asthma.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Administration, Oral; Aerosols; Animals; Blood Pressure; Bronchoconstriction; Carbazoles; Guinea Pigs; Histamine; In Vitro Techniques; Injections, Intravenous; Male; Prostaglandin D2; Prostaglandin Endoperoxides, Synthetic; Serotonin; SRS-A; Sulfonamides; Thromboxane A2; Thromboxanes

Topics: Animals; Bronchoconstriction; Carbazoles; Dose-Response Relationship, Drug; Guinea Pigs; Indomethacin; Lung Volume Measurements; Male; Prostaglandin D2; Receptors, Prostaglandin; Receptors, Thromboxane; Sotalol; Sulfonamides; Thromboxanes

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Carbazoles; Dinoprost; Dose-Response Relationship, Drug; Guinea Pigs; In Vitro Techniques; Muscle Contraction; Muscle, Smooth; Prostaglandin D2; Prostaglandin Endoperoxides, Synthetic; Sulfonamides; Thromboxanes; Trachea