15-hydroxy-5-8-11-13-eicosatetraenoic-acid and Asthma

Topics: Anti-Inflammatory Agents, Non-Steroidal; Aspirin; Asthma; Cyclooxygenase Inhibitors; Dinoprostone; Eosinophils; Humans; Hydroxyeicosatetraenoic Acids; Leukotrienes; Lipoxins; Mast Cells; Prostaglandin-Endoperoxide Synthases

Topics: Arachidonate 15-Lipoxygenase; Arachidonic Acid; Asthma; Bronchi; Eosinophils; Humans; Hydroxyeicosatetraenoic Acids; Leukotrienes; Muscle, Smooth; Neutrophils

Eosinophilic accumulation in the airway is a prominent feature in patients with bronchial asthma (chronic desquamative eosinophilic bronchitis). Recently, arachidonic acid metabolite such as leukotriene, 15-HETE and PAF are proved to play an essential role as chemical mediators in the pathogenesis of bronchial asthma. A close relation is found between eosinophils activation and those mediators. The bronchoconstrictive leukotriene (LT C4, D4, E4) are generated mainly from eosinophils and mast cells, while the chemoattractive LTB4 is produced from neutrophils. Platelet activating factor (PAF) is postulated as a causative mediator of the eosinophilic airway inflammation and bronchial hyperresponsiveness because PAF attracts and activates preferentially eosinophils. Evidence that eosinophils have ability to produce PAF like other inflammatory cells suggests an autocrine mechanisms of the eosinophil activation by PAF.

Topics: Asthma; Bronchial Hyperreactivity; Eosinophils; Humans; Hydroxyeicosatetraenoic Acids; Leukotrienes; Platelet Activating Factor

Although the mechanism of aspirin-induced asthma and rhinitis is unknown, it has been suggested that adverse nasal and bronchial reactions are caused by an increased production of lipoxygenase products. In examining this hypothesis we have measured the release of peptide leukotrienes (PeptLTs), 15-HETE, and prostaglandins in nasal fluids obtained by nasal lavages after instillation of acetylsalycilic acid (ASA) and placebo (saline). Ten ASA-sensitive asthmatics, 10 ASA-insensitive asthmatics, and seven healthy subjects were challenged in a double-blind study with normal saline and 12 mg of ASA. Twelve mg were administered based on the results of a previous study that showed that this dose caused minor to moderate symptoms in ASA-sensitive patients. PeptLTs, LTB4, 15-HETE, PGE2, PGF2 alpha, and PGD2 were measured by radioimmunoassay methods. Significant levels of PeptLTs were detected in sensitive asthmatic patients 60 min after nasal challenge. This change was associated with a significant increase in symptoms. No increase in PeptLTs levels were found, however, in either insensitive patients or healthy subjects. Inhibition of PGE2 and PGF2 alpha release was detected in the three groups after ASA administration. ASA also inhibited PGD2 release in insensitive asthmatic patients but not in both sensitive patients and healthy subjects. These results suggest that an abnormal release of PeptLTs in ASA-sensitive asthmatic patients contributes to nasal and bronchial adverse reactions. The lack of effects on PGD2 release suggests that mast cells from ASA-insensitive patients are more sensitive to ASA than those from sensitive asthmatic patients and healthy subjects.

Topics: Administration, Intranasal; Adult; Albumins; Aspirin; Asthma; Dinoprost; Dinoprostone; Drug Hypersensitivity; Female; Humans; Hydroxyeicosatetraenoic Acids; Leukotriene B4; Leukotrienes; Male; Middle Aged; Nasal Mucosa; Prostaglandin D2

15-(s)-hydroxyeicosatetraenoic acid (15-HETE), a major oxidative metabolite of arachidonic acid in human lungs, has complex actions on the 5-lipoxygenase pathway in different cell systems. We have examined the effect of inhaled 15-HETE on the early and late asthmatic responses (EAR and LAR) and the associated change in nonspecific bronchial responsiveness to inhaled allergen in 10 subjects with atopic asthma. On 2 separate days 3 wk apart, subjects inhaled either 70 nmol 15-HETE or the diluent (sodium phosphate buffer) in a randomized and double-blind fashion, followed by a dose of allergen that had previously been shown to produce a 25% fall in baseline FEV1. Analysis of the area under the FEV1 response time course curves (AUC) in the first hour revealed that preinhalation with 15-HETE increased the EAR by 39% from that achieved after the diluent (p less than 0.05). In the seven subjects who were classified as dual responders by developing a LAR previously (greater than 15% fall in baseline FEV1 3 to 8 h after allergen challenge), 15-HETE did not cause any significant change in the magnitude of the LAR when compared with that observed after placebo. The values of the provocation concentration of histamine causing a 20% fall in FEV1 (Pc20) at 8 h postchallenge were significantly reduced on both study days when compared with the corresponding preallergen challenge values, whether analyzed for the whole group or for the seven dual responders (p less than 0.05). 15-HETE had no effect on this allergen-acquired airway hyperresponsiveness to histamine when compared with the diluent.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adult; Asthma; Bronchial Provocation Tests; Double-Blind Method; Female; Forced Expiratory Volume; Humans; Hydroxyeicosatetraenoic Acids; Male; Mast Cells; Middle Aged; Randomized Controlled Trials as Topic

Topics: Animals; Asthma; Bronchitis, Chronic; Cystic Fibrosis; Goblet Cells; Humans; Hydroxyeicosatetraenoic Acids; Idiopathic Pulmonary Fibrosis; Mucins

Epithelial 15-lipoxygenase 1 (15LO1) and activated ERK are increased in asthma despite modest elevations in IL-13. MAPK kinase (MEK)/ERK activation is regulated by interactions of Raf-1 with phosphatidylethanolamine-binding protein 1 (PEBP1). Epithelial 15LO1 generates intracellular 15-hydroxyeicosatetraenoic acid (15HETE) conjugated to phosphatidylethanolamine (PE) (15HETE-PE). We hypothesized that (i) 15LO1 and its product 15HETE-PE serve as signaling molecules interacting with PEBP1 to activate Raf-1/MEK/ERK and that (ii) this 15LO1-15HETE-PE-regulated ERK activation amplifies IL-4Rα downstream pathways. Our results demonstrate that high epithelial 15LO1 levels correlate with ERK phosphorylation ex vivo. In vitro, IL-13 induces 15LO1, which preferentially binds to PEBP1, causing PEBP1 to dissociate from Raf-1 and activate ERK. Exogenous 15HETE-PE similarly induces dissociation of PEBP1 from Raf-1 independently of IL-13/15LO1. siRNA knockdown of 15LO1 decreases the dissociation of Raf-1 from PEBP1, and the resulting lower ERK activation leads to lower downstream IL-4Rα-related gene expression. Identical protein-protein interactions are observed in endobronchial biopsies and fresh epithelial cells from asthmatics ex vivo. Colocalization of Raf-1 to PEBP1 is low in asthmatic tissue and cells compared with normals, whereas there is striking colocalization of 15LO1 with PEBP1 in asthma. Low 15LO1 levels in normals limit its colocalization with PEBP1. The results confirm a previously unknown signaling role for 15LO1 and its PE-conjugated eicosanoid product in human airway epithelial cells. This pathway enhances critical inflammatory pathways integral to asthma pathogenesis.

Topics: Arachidonate 15-Lipoxygenase; Asthma; Binding, Competitive; Bronchi; Cell Membrane; Chemokine CCL26; Chemokines, CC; Demography; Enzyme Activation; Epithelial Cells; Extracellular Signal-Regulated MAP Kinases; Humans; Hydroxyeicosatetraenoic Acids; Interleukin-13; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase Kinases; Mucin 5AC; Nitric Oxide Synthase Type II; Phosphatidylethanolamine Binding Protein; Phosphorylation; Protein Binding; Proto-Oncogene Proteins c-raf

Increased levels of leukotrienes (LTs) in exhaled breath condensate (EBC) are associated with asthma and bronchial hyperresponsiveness (BHR), whereas eicosanoids generated through the 15-lipoxygenase (LO) pathway (15-hydroxyeicosatetraenoic acid [HETE] and eoxins) have been less studied.. We investigated whether metabolites of the 5- and 15-LO pathways in EBC are associated with childhood asthma, asthma severity, and clinical parameters.. The present study included 131 school-aged children (27 children with problematic severe asthma, 80 children with mild-to-moderate asthma, and 24 healthy children) from the Severe Asthma Recognized in Childhood study and 19 children with other nonasthmatic chronic lung diseases. Clinical work-up included spirometry, fractional exhaled nitric oxide measurements, skin prick testing, and methacholine challenge. Eicosanoids were analyzed in EBC by using mass spectrometry and are reported as concentrations (in picograms per milliliter) and eicosanoid/palmitic acid (PA) ratios.. Eoxin C₄/PA, eoxin D₄/PA, eoxin E₄/PA, 15-HETE/PA, and LTC₄/PA ratios were significantly increased in asthmatic versus healthy children. Eoxin D₄/PA and LTE₄/PA ratios were also significantly higher in children with BHR. A nonsignificant trend was observed toward higher eoxin/PA ratios with increasing asthma severity. In contrast to asthma, children with chronic lung disease had the highest 15-HETE/PA, LTC₄/PA, LTE₄/PA, and LTB₄/PA ratios.. The results point to increased activity of the 15-LO inflammatory pathway in childhood asthma. Mass spectrometric analyses of EBC demonstrate that increased eoxin levels not only accompany the increased 5-LO product LTC₄ but are also associated with BHR. These markers might represent a new therapeutic target for asthma treatment.

Topics: Adolescent; Arachidonate 15-Lipoxygenase; Asthma; Breath Tests; Bronchial Hyperreactivity; Child; Exhalation; Female; Humans; Hydroxyeicosatetraenoic Acids; Inflammation; Leukotriene C4; Leukotriene E4; Leukotrienes; Male; Mass Spectrometry; Severity of Illness Index

Airway inflammation is common in severe asthma despite antiinflammatory therapy with corticosteroids. Lipoxin A(4) (LXA(4)) is an arachidonic acid-derived mediator that serves as an agonist for resolution of inflammation.. Airway levels of LXA(4), as well as the expression of lipoxin biosynthetic genes and receptors, in severe asthma.. Samples of bronchoalveolar lavage fluid were obtained from subjects with asthma and levels of LXA(4) and related eicosanoids were measured. Expression of lipoxin biosynthetic genes was determined in whole blood, bronchoalveolar lavage cells, and endobronchial biopsies by quantitative polymerase chain reaction, and leukocyte LXA(4) receptors were monitored by flow cytometry.. Individuals with severe asthma had significantly less LXA(4) in bronchoalveolar lavage fluids (11.2 +/- 2.1 pg/ml) than did subjects with nonsevere asthma (150.1 +/- 38.5 pg/ml; P < 0.05). In contrast, levels of cysteinyl leukotrienes were increased in both asthma cohorts compared with healthy individuals. In severe asthma, 15-lipoxygenase-1 mean expression was decreased fivefold in bronchoalveolar lavage cells. In contrast, 15-lipoxgenase-1 was increased threefold in endobronchial biopsies, but expression of both 5-lipoxygenase and 15-lipoxygenase-2 in these samples was decreased. Cyclooxygenase-2 expression was decreased in all anatomic compartments sampled in severe asthma. Moreover, LXA(4) receptor gene and protein expression were significantly decreased in severe asthma peripheral blood granulocytes.. Mechanisms underlying pathological airway responses in severe asthma include lipoxin underproduction with decreased expression of lipoxin biosynthetic enzymes and receptors. Together, these results indicate that severe asthma is characterized, in part, by defective lipoxin counterregulatory signaling circuits.

Topics: Adult; Asthma; Bronchi; Bronchoalveolar Lavage Fluid; Female; Flow Cytometry; Humans; Hydroxyeicosatetraenoic Acids; Lipoxins; Male; Middle Aged; Polymerase Chain Reaction; Receptors, Formyl Peptide; Receptors, Lipoxin

Cyclooxygenase-1 (COX-1) is a key enzyme involved in generation of prostanoids, important mediators and modulators of asthmatic inflammation. In a subpopulation of aspirin-sensitive asthmatics (ASA) inhibition of COX-1 by nonsteroidal anti-inflammatory drugs results in activation of inflammatory cells and development of symptoms. Alternatively spliced variants of COX-1 lacking 111 bp from exon 9 were described previously but have never been identified in human leucocytes peripheral blood leucocytes (PBL) or upper airway epithelial cells. We aimed to assess the expression of spliced variants of COX-1 mRNA in PBLs from patients with asthma and in healthy subjects (HS) referring the expression to patients characteristics (including ASA-sensitivity) and to aspirin-triggered 15-hydroxyeicosatetraenoic acid (15-HETE) generation.. The study included 30 patients with ASA, 30 patients with aspirin-tolerant asthma (ATA) and 30 HS serving as controls. Nasal polyps for epithelial cell cultures were obtained from 10 patients with chronic rhinosinusitis. Expression of full length and spliced variants of COX-1 enzyme was detected by RT-PCR and presented as the ratio of full-length COX-1 to alternatively spliced COX-1 mRNA [COX-1 alternative splicing index (COX-1 AS index)]. Release of eicosanoids (PGE(2) and 15-HETE) by PBLs was measured with enzyme immunoassay.. In both PBLs and airway epithelial cells the expression of full-length product prevailed over spliced variants of COX-1 enzyme. Cyclooxygenase-1 AS index was significantly lower in asthmatics as compared to HS (1.96 +/- 0.71 vs 2.41 +/- 0.99, P < 0.05) indicating the relatively higher expression of the alternative transcript in asthmatic patients. Cyclooxygenase-1 AS index was not different between ASA and ATA groups (mean 1.90 +/- 0.66 vs 2.02 +/- 0.76, respectively, P = 0.39). There was no significant association between COX-1 AS index and mean daily dose of inhaled glucocorticosteroids or pulmonary function tests (FEV(1), FVC) but in ASA group a weak correlation with daily dose of oral glucocorticosteroids was found (r = 0.39; P = 0.03). In ASA patients there was a significant positive correlation between the COX-1 AS index and the percentage of aspirin-triggered increase in 15-HETE generation (r = 0.51; P < 0.03).. Alternatively spliced variants of COX-1 mRNA are differently expressed in patients with bronchial asthma and may be associated with aspirin-triggered 15-HETE generation.

Topics: Adult; Aged; Alternative Splicing; Anti-Inflammatory Agents, Non-Steroidal; Aspirin; Asthma; Cyclooxygenase 1; Drug Hypersensitivity; Female; Gene Expression; Humans; Hydroxyeicosatetraenoic Acids; Hypersensitivity; Isoenzymes; Leukocytes; Male; Middle Aged; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Skin Tests

Mast cells play a key role in the pathophysiology of asthma. These cells exert their effector functions by releasing a variety of proinflammatory and immunoregulatory compounds. Mast cells infiltrate the bronchial epithelium and smooth muscle to a higher degree in patients with asthma compared to control subjects. 15-Lipoxygenase type-1 (15-LO-1) is a prooxidant enzyme which is expressed in asthmatic lungs leading to formation of pro- and anti-inflammatory mediators. Here we report that interleukin-4 (IL-4) induced the expression of 15-LO-1 in human cord blood derived mast cells (CBMC) as demonstrated by RT-PCR, western blot and immunocytochemistry. The major metabolite of arachidonic acid formed via the 15-LO pathway in IL-4 treated CBMC was identified as 15-ketoeicosatetraenoic acid (15-KETE, also named 15-oxo-ETE) with smaller amounts of 15-hydroxyeicosatetraenoic acid (15-HETE) as identified by HPLC and mass spectrometry (MS/MS). Furthermore, immunohistochemical stainings demonstrated the expression of 15-LO-1 in mast cells in lung and skin in vivo. Osmotic activation of CBMC with mannitol resulted in activation of the 15-LO-1 pathway. In conclusion, the expression of 15-LO-1 and release of 15-LO-1 derived products by mast cells may contribute to the role of these cells in asthma and other inflammatory diseases.

Topics: Arachidonate 15-Lipoxygenase; Arachidonic Acids; Asthma; Fetal Blood; Humans; Hydroxyeicosatetraenoic Acids; Interleukin-4; Isoenzymes; Leukotrienes; Lipid Peroxides; Lung; Mannitol; Mast Cells; Skin; Tryptases

We have previously demonstrated that aspirin triggers specific generation of 15-hydroxyeicosateraenoic acid (15-HETE) from nasal polyp epithelial cells and peripheral blood leukocytes (PBL) from aspirin-sensitive (AS) but not aspirin-tolerant (AT) patients with asthma/rhinosinusitis. The goal of this study was to assess the diagnostic value of ASA-induced 15-HETE generation measurement to identify AS patients.. PBL were obtained from 43 AS patients with asthma and rhinosinusitis, 35 AT asthmatics and 17 healthy control (HC) subjects. PBL were incubated with 2-200 muM aspirin (ASA) and 15-HETE release was measured in cell supernatants with competitive ELISA.. Unstimulated PBL from all three groups of patients generated similar amount of 15-HETE. Incubation with 200 microM ASA resulted in an increase in an 15-HETE generation (mean increase +421%) in AS-asthmatics but small and nonsignificant response in AT-asthmatics or control subjects. Receiver operating curve (ROC) analysis revealed that the sensitivity of the test for confirmation of ASA-sensitivity was 83% and the specificity 82%. Positive predictive value was 0.79 and negative predictive value was 0.86. Naproxen induced a significant increase in 15-HETE only in some AS-asthmatics, but not in AT-asthmatics.. Our data demonstrate that ASA-induced 15-HETE generation by PBL is a specific and sensitive aspirin-sensitive patients identification test (ASPITest).

Topics: Adult; Aged; Aspirin; Asthma; Drug Hypersensitivity; Female; Humans; Hydroxyeicosatetraenoic Acids; Immunologic Tests; Leukocytes; Male; Middle Aged; Predictive Value of Tests; Rhinitis; Sinusitis

Although the mechanisms of aspirin-induced rhinosinusitis-asthma appear to be related to arachidonic acid abnormalities, only recently has a specific aspirin-triggered enhancement of 15-hydroxyeicosatetraenoic acid (15-HETE) generation in nasal polyp epithelial cells from aspirin-sensitive patients been demonstrated.. The aim of this study was to assess generation of 15-HETE and other eicosanoids by peripheral blood leukocytes (PBLs) from aspirin-sensitive and aspirin-tolerant asthmatic patients and modulation of 15-HETE generation by a prostaglandin (PG) E(1) analogue (misoprostol).. Twenty-four aspirin-sensitive patients with asthma-rhinosinusitis and 18 aspirin-tolerant asthmatic patients were studied, and eicosanoids released from PBLs were assessed by means of enzyme immunoassays.. Unstimulated PBLs from aspirin-sensitive and aspirin-tolerant patients generated similar amounts of PGE(2), leukotriene C(4), and 15-HETE, but lipoxin A(4) release was significantly less in aspirin-sensitive patients (300 +/- 70 pg/mL) in comparison with that seen in aspirin-tolerant patients (690 +/- 100 pg/mL, P <.05). Cell incubation with 2, 20, or 200 micromol/L aspirin resulted in a dose-dependent increase in 15-HETE generation (mean change of +85%, +189%, and +284% at each aspirin concentration, respectively) only in aspirin-sensitive asthmatic patients. Naproxen stimulated 15-HETE generation in aspirin-sensitive asthmatic patients, but indomethacin or specific COX-2 inhibitors (NS-398 and celecoxib) did not affect 15-HETE release. A synthetic PGE(1) analogue (misoprostol) inhibited aspirin-induced 15-HETE release but enhanced 15-HETE generation by aspirin in leukocytes from aspirin-tolerant patients. After preincubation with misoprostol, aspirin induced a dose-dependent production of lipoxin A(4) in both groups.. PBLs from patients with aspirin-sensitive rhinosinusitis-asthma might be specifically triggered by aspirin to generate 15-HETE. Metabolism of 15-HETE is differentially regulated by misoprostol in aspirin-tolerant and aspirin-sensitive asthmatic patients.

Topics: Adult; Aged; Anti-Inflammatory Agents, Non-Steroidal; Aspirin; Asthma; Dinoprostone; Drug Tolerance; Eicosanoids; Female; Humans; Hydroxyeicosatetraenoic Acids; In Vitro Techniques; Ionophores; Leukocytes; Leukotriene C4; Lipoxins; Male; Middle Aged; Misoprostol

Although 15(S)-hydroxyeicosatetraenoic acid (15(S)-HETE), a product of 15-lipoxygenase (15-LO), may be involved in mild to moderate asthma, little is known about its potential roles in severe asthma.. This study was performed to evaluate 15(S)-HETE levels in bronchoalveolar lavage fluid (BALF) from severe asthmatics with and without airway eosinophils and from the control groups. In addition, 15-LO protein expression was examined in endobronchial biopsy, while its expression and activation were evaluated in BAL cells.. While 15(S)-HETE levels in BALF were significantly higher in all severe asthmatics than normal subjects, severe asthmatics with airway eosinophils had the highest levels compared with mild, moderate asthmatics and normal subjects. 15(S)-HETE levels were associated with tissue eosinophil numbers, sub-basement membrane thickness and BALF tissue inhibitor of metalloproteinase-1 levels, and were accompanied by increased 15-LO expression in bronchial epithelium. In addition, activation of 15-LO was suggested by the increased proportion of 15-LO in the cytoplasmic membrane of alveolar macrophages from severe asthmatics.. The data suggest that severe asthmatics with persistent airway eosinophils manifest high levels of 15(S)-HETE in BALF, which may be associated with airway fibrosis. It is likely that 15-LO expression and activation by airway cells explain the increased 15(S)-HETE levels.

Topics: Adult; Arachidonate 15-Lipoxygenase; Asthma; Bronchi; Bronchoalveolar Lavage Fluid; Collagen; Eosinophils; Female; Fibrosis; Humans; Hydroxyeicosatetraenoic Acids; Male; Middle Aged; Signal Transduction

The mechanism of aspirin (acetylsalicylic acid [ASA]) sensitivity associated with severe asthma and chronic rhinosinusitis with nasal polyps ("aspirin triad") has been attributed to arachidonic metabolism alternations, although the putative biochemical defects have not been elucidated. The aim of this study was assessment of the hypothesis that local production of eicosanoids in the respiratory epithelium in patients with ASA-sensitive asthma/rhinosinusitis (ASARS) differs from that of ASA-tolerant patients with rhinosinusitis (ATRS). Nasal polyps were obtained from 10 patients with ASARS and 15 with ATRS during routine polypectomies, and epithelial cells (ECs) were cultured on bovine collagen type I matrix (Vitrogen 100), in medium supplemented with growth factors. The generation of eicosanoids in supernatants of confluent ECs (6 to 8 d of culture; purity > 98%) was quantified by immunoassays. Unstimulated ECs from ASARS patients generated significantly less prostaglandin E(2)(PGE(2)) compared with ATRS (0.8 +/- 0.3 versus 2. 4 +/- 0.5 ng/microg double-stranded deoxyribonucleic acid [dsDNA], respectively), although a similar relative increase in response to calcium ionophore and inhibition with ASA was observed in both groups. Basal levels of 15-hydroxyeicosatetraenoic acid (15-HETE) were not different between groups, and calcium ionophore enhanced its production to a similar extent. However, cells incubation with 200 microM ASA for 60 min resulted in a significant increase (mean +359%) in 15-HETE generation only in ASARS patients, whereas no effect of ASA on 15-HETE generation in ATRS patients was observed. PGF(2alpha) generation was similar in both groups, and no significant generation of PGD(2) or leukotriene C(4) (LTC(4)) was observed in epithelial cell cultures from either group. Our results indicate that nasal polyps ECs from ASA-sensitive patients have significant abnormality in basal and ASA-induced generation of eicosanoids which may be causally related to the mechanism of ASA sensitivity.

Topics: Adult; Aged; Animals; Anti-Inflammatory Agents, Non-Steroidal; Arachidonic Acid; Aspirin; Asthma; Cattle; Cells, Cultured; Dinoprostone; Drug Hypersensitivity; Female; Humans; Hydroxyeicosatetraenoic Acids; Male; Middle Aged; Nasal Polyps; Prostaglandin D2; Respiratory Mucosa

Recent evidence shows that 15(S)-hydroxy-eicoisatetraenoic acid (15[S]-HETE) can be released and rapidly reincorporated into cellular lipids. These mechanisms exert several immunoregulatory functions that may be relevant in airway inflammation.. Our purpose was to evaluate the levels of both soluble and cell-associated 15(S)-HETE and to examine 15-lipoxygenase (15-LO) messenger RNA (mRNA) expression in sputum samples obtained from 10 control and 18 asthmatic subjects.. Levels of 15(S)-HETE were measured by reverse-phase HPLC separation followed by RIA in supernatants and in cell membrane-extracted phospholipids after acid hydrolysis. 15-LO mRNA was evaluated by primed in situ hybridization (PRINS). Combined immunocytochemistry and PRINS was used to identify the phenotype of cells bearing 15-LO transcripts.. Levels of both soluble and cell-associated 15(S)-HETE were higher in asthmatic than in control subjects (P <.0001). The percentage of cells expressing 15-LO mRNA was higher in asthmatic than in control subjects (P <.01). On double staining for specific cell-type markers and 15-LO mRNA, macrophages were the major source for 15-LO.. This study shows that the induced sputum technique allows the evaluation of 15-LO activity and that soluble, cell-associated 15(S)-HETE and 15-LO levels are higher in asthmatic than in control subjects. In addition, this study indicates that, in induced sputum, airway macrophages are the major source of 15(S)-HETE in asthma.

Topics: Adult; Aged; Arachidonate 15-Lipoxygenase; Asthma; Cell Count; Forced Expiratory Volume; Humans; Hydroxyeicosatetraenoic Acids; Immunohistochemistry; In Situ Hybridization; Middle Aged; RNA, Messenger; Saliva; Solubility; Sputum

Asthma is characterized by chronic airway inflammation resulting from overproduction of pro-inflammatory mediators, such as leukotrienes (LT). The authors questioned the biosynthetic capacity of asthmatic patients for lipoxins (LX) and 15-epimer lipoxins (15-epi-LX), endogenous regulators of inflammatory responses that inhibit pro-inflammatory events. Levels of LXA4, 15-epi-LXA4 and LTC4 were determined in 14 clinically characterized aspirin-intolerant asthmatics (AIA), 11 aspirin-tolerant asthmatics (ATA) and eight healthy volunteers using a stimulated whole blood protocol. Both LXA4 and 15-epi-LXA4 were generated in whole blood activated by the divalent cation ionophore, A23187. Higher levels of LXA4 were produced in ATA than either AIA or healthy volunteers. Exposure of AIA whole blood to interleukin-3 prior to A23187 did not elevate their reduced capacity to generate LXA4. Generation of a bronchoconstrictor, LTC4, was similar in both AIA and ATA. Consequently, the ratio of LXA4:LTC4 quantitatively favoured the bronchoconstrictor for AIA and differed from both ATA and healthy subjects. In addition, the capacity for 15-epi-LXA4 generation was also diminished in AIA, since whole blood stimulated in the presence of aspirin gave increased levels only in samples from ATA. The present results indicate that asthmatics possess the capacity to generate both lipoxins and 15-epimer-lipoxins, but aspirin-intolerant asthmatics display a lower biosynthetic capacity than aspirin-tolerant asthmatics for these potentially protective lipid mediators. This previously unappreciated, diminished capacity for lipoxin formation by aspirin-intolerant asthmatic patients may contribute to their more severe clinical phenotype, and represents a novel paradigm for the development of chronic inflammatory disorders.

Topics: Adult; Aspirin; Asthma; Chromatography, High Pressure Liquid; Enzyme-Linked Immunosorbent Assay; Female; Humans; Hydroxyeicosatetraenoic Acids; Leukotriene C4; Lipoxins; Male; Middle Aged; Stereoisomerism

Topics: Arachidonic Acid; Asthma; Calcimycin; Cell Culture Techniques; Eicosanoids; Glucocorticoids; Humans; Hydroxyeicosatetraenoic Acids; Ionophores; Macrophages, Alveolar; Monocytes; Neutrophils

IL-4 modulates the synthesis of IgE, the expression of CD23, and the release of 15(S)-hydroxyeicosatetraenoic (15[S]-HETE).. We evaluated the release of 15(S)-HETE by IL-4-stimulated monocytes and verified whether the observed increase in 15(S)-HETE release after passive sensitization and anti-IgE challenge of IL-4-treated monocytes was secondary to an increased CD23 expression.. Human monocytes were incubated for 24, 48, and 72 hours with IL-4 (10 ng/mL) with or without an IgE-anti-IgE stimulation. We evaluated CD23 expression by immunocytochemistry and 15(S)-HETE release by HPLC and RIA. To prove that the increase in 15(S)-HETE release was due to the effect of IL-4 on CD23, we performed experiments with an anti-CD23 blocking mAb.. CD23 expression and 15(S)-HETE release were significantly increased by IL-4, reaching a peak after 72 hours (P <.02). After passive sensitization with human IgE and anti-IgE challenge, IL-4-stimulated monocytes released higher amounts of 15(S)-HETE than IL-4-unstimulated monocytes (P <.02). Pretreatment with the anti-human B-cell CD23 MHM6 mAb caused a dose-dependent inhibition of 15(S)-HETE release.. This study shows that immunologic challenge of IL-4-treated, passively sensitized monocytes results in a CD23-dependent additional increase of 15(S)-HETE release, indicating the presence of a synergistic effect of IL-4 on CD23 expression and 15(S)-HETE production.

Topics: Antibodies, Anti-Idiotypic; Antigen-Antibody Complex; Asthma; Humans; Hydroxyeicosatetraenoic Acids; Immunoglobulin E; Interleukin-4; Monocytes; Phagocytes; Receptors, IgE

Bronchial epithelial cells are the first cells to come into contact with inhaled pneumoallergens. It has been suggested that these cells may play an important role in the allergic response, and indeed bronchial epithelial cells of some atopic asthmatic subjects have been shown to express the low-affinity receptor for IgE on their surface. In this report we demonstrate, using bronchial biopsies, that bronchial epithelial cells of some asthmatic subjects express both the alpha and gamma chains of the high-affinity receptor for IgE (Fcepsilon RI) on their surface and that they are capable of fixing IgE. Second, using reverse transcription-polymerase chain reaction, we show that both control and asthmatic subjects have messenger RNA for Fcepsilon RI. Finally, we demonstrate that this receptor may be functional since stimulation of the cells with the antibody to the alpha chain of Fcepsilon RI results in the liberation of 15-hydroxyeicosatetraenoic acid from epithelial cells of asthmatic, but not control, subjects or subjects suffering from chronic bronchitis. These data suggest that bronchial epithelial cells from at least some asthmatic subjects express a functional high-affinity receptor for IgE and it is therefore possible that these cells may be able to interact directly with inhaled allergens.

Topics: Adolescent; Adult; Aged; Asthma; Bronchi; Bronchitis; Chronic Disease; Epithelial Cells; Gene Expression; Histamine Release; Humans; Hydroxyeicosatetraenoic Acids; Immunoenzyme Techniques; Immunoglobulin E; Middle Aged; Receptors, IgE; RNA, Messenger

The aim of this study was to investigate eicosanoid metabolism by human peripheral blood monocytes (PBM) from steroid-dependent asthmatic patients as compared to control subjects and untreated asthmatic patients. Eicosanoid biosynthesis by PBM isolated from venous blood using Percoll gradient centrifugation was evaluated following stimulation of 5 x 10(6) cells with calcium ionophore A23187, with or without exogenous 15(S)-hydroxyeicosatetraenoic acid (15(S)-HETE), and analyzed by reverse phase high performance liquid chromatography (RP-HPLC). Without 15(S)-HETE, PBM synthesized leukotriene B4 (LTB4) only (40 +/- 12 ng and 59 +/- 11 ng for untreated and steroid-dependent asthmatics, respectively). In the presence of 15(S)-HETE, PBM produced six-fold smaller amounts of leukotriene B4 (P < 0.0001). They also released 5(S),15(S)-dihydroxyeicosatetraenoic acid (5(S),15(S)-diHETE) in similar amounts for all the populations, whereas low amounts of lipoxins (LXs) were produced by PBM from asthmatics only (2.7 +/- 0.7 ng and 4.6 +/- 2.8 ng for untreated and steroid-dependent asthmatics, respectively). Moreover, PBM were also able to release an unknown compound containing conjugated triene chromophore. Cells from steroid-dependent asthmatic patients synthesized this unknown metabolite in higher amounts than controls and untreated asthmatics (133 +/- 18 ng vs 52 +/- 19 ng and 68 +/- 15 ng, respectively, P < 0.02). This work shows for the first time that human PBM are able to metabolize 15(S)-HETE and lead to lipoxins and to an unknown metabolite, with the amounts of the latter being enhanced by long-term corticosteroid treatment.

Topics: Adrenal Cortex Hormones; Adult; Aged; Asthma; Eicosanoids; Humans; Hydroxyeicosatetraenoic Acids; Leukotriene B4; Middle Aged; Monocytes

5-Lipoxygenase activation of human blood polymorphonuclear cells (PMN) from asthmatic patients (asthmatics) was studied to investigate whether differences may exist with healthy subjects (controls). The respective cell capacities to produce lipoxins (LXs), leukotrienes, and 5(S), 15(S)-dihydroxyeicosatetraenoic acid [5(S),15(S)-diHETE] were compared under in vitro stimulation by ionophore A23187, with or without exogenous 15(S)-hydroxyeicosatetraenoic acid [15(S)-diHETE]. Eicosanoids were analyzed by elution with an isocratic reverse-phase high performance liquid chromatography system, and their profiles, detected by simultaneous monitoring at 302, 280, and 246 nm, were evaluated on the basis of chromatographic behavior: UV spectral characteristics and coelution with synthetic standards. In the presence of exogenous 15(S)-HETE, human PMN were able to produce LXs and 5(S),15(S)-diHETE, PMN from asthmatics were able to produce 5(S), 5(S),15(S)-diHETE, and LXs from endogenous sources, whereas in the same experimental conditions, no detectable amounts of these compounds were released by PMN from controls. The levels of 5(S),15(S)-diHETE, and LXs biosynthesized from endogenous arachidonic acid were highly correlated. Two different LX patterns were observed involving two possible metabolic pathways: (a) via the intermediate 5,6-epoxytetraene alone for LXs generation from exogenous 15(S)-HETE; and (b) via 5,6- and/or 14,15-epoxytetraenes leading to the formation of an enzyme-bound delocalized carbocation for LXs generation from endogenous arachidonate, respectively. The enhanced 5-lipoxygenase activation of blood PMN from asthmatics and the metabolism of exogenous 15(S)-HETE may reflect a priming induced by various mediators released from environmental cells, and could be considered as a model of transcellular signalization between PMN and endothelial cells.

Topics: Arachidonate 5-Lipoxygenase; Arachidonic Acids; Asthma; Calcimycin; Chromatography, High Pressure Liquid; Female; Humans; Hydroxyeicosatetraenoic Acids; Male; Models, Biological; Neutrophil Activation; Substrate Specificity

Products of the 15-lipoxygenase (15-LO) pathway of arachidonic acid metabolism such as the mono- and di-hydroxyeicosatetraenoic acids (HETEs) may contribute to the pathophysiology of allergic airway inflammation through the recruitment and activation of inflammatory cells and stimulation of glandular secretion. In this study we have examined the expression of 15-LO and its cellular localization in the asthmatic and normal bronchial mucosa. Bronchial mucosal biopsies were obtained by fiberoptic bronchoscopy from 10 patients with symptomatic allergic asthma and six normal control subjects and processed into glycolmethacrylate resin. Sections 2 microns thick were immunostained using a specific rabbit polyclonal antihuman 15-LO antibody. Strong immunoreactivity for 15-LO was present throughout the epithelium in both the asthmatic and the normal subjects, with no difference between the two groups. Cells expressing 15-LO immunoreactivity were also present in the submucosa of both groups, with a significantly greater number present in the asthmatic group (median, 15.3 cells/mm2) than in the normal group (median, 6.9 cells/mm2) (p = 0.01). The majority (85%) of the submucosal 15-LO+ cells were eosinophils. Patchy 15-LO immunoreactivity was also seen in the vascular endothelium in both groups. These findings demonstrated increased 15-LO expression in the bronchial submucosa of asthmatic subjects, and they suggest that 15-LO products in asthma originate from both bronchial epithelium and infiltrating eosinophils.

Topics: Adult; Arachidonate 15-Lipoxygenase; Asthma; Bronchi; Eosinophils; Epithelium; Female; Humans; Hydroxyeicosatetraenoic Acids; Male; Mucous Membrane

Bronchial epithelial cells may be involved in the pathogenesis of asthma releasing several inflammatory mediators such as interleukins and lipoxygenase products. In this study we evaluated the spontaneous and A23128-induced release of 15-HETE, PGE2 and fibronectin as well as the spontaneous expression of HLA-DR and ICAM-1 molecules by bronchial epithelial cells obtained by bronchial brushing from 35 asthmatic and 27 normal subjects. The release of fibronectin and 15-HETE was studied using the EIA and RIA techniques. The expression of HLA-DR and ICAM-1 molecules was studied using the APAAP and the immunofluorescence methods. Bronchial epithelial cells from asthmatics released higher amounts of 15-HETE and fibronectin both spontaneously (p < 0.002, p < 0.05, respectively) or after stimulation with calcium ionophore compared with normals. On the other hand, PGE2 release was significantly higher only after stimulation with calcium ionophore (p < 0.002). The spontaneous expression of HLA-DR and ICAM-1 (p < 0.001) was significantly higher on epithelial cells from asthmatics than in normal subjects. Finally, the severity of asthma significantly correlated with the release of 15-HETE (p < 0.02) and the expression of HLA-DR and ICAM-1 respectively (p < 0.001 and p < 0.002, respectively). This study indicates that bronchial epithelial cells are in an activated state in asthma and that the degree of activation is correlated to the severity of the disease.

Topics: Adult; Alkaline Phosphatase; Asthma; Bronchi; Calcimycin; Cell Adhesion Molecules; Dinoprostone; Epithelium; Fibronectins; Fluorescent Antibody Technique; HLA-DR Antigens; Humans; Hydroxyeicosatetraenoic Acids; Immunoenzyme Techniques; Intercellular Adhesion Molecule-1

Airways epithelial cells may be involved in the pathogenesis of asthma, but their role remains to be determined. Epithelial cells can release large amounts of 15-hydroxy-eicosatetranoic acid (15-HETE) and smaller amounts of prostaglandin E2 (PGE2) as well as fibronectin, a mediator involved in epithelial repair after injury. Epithelial cells obtained after bronchial brushing of 16 asthmatic (age 38 +/- 5 yr) and 11 normal subjects (age 36 +/- 5 yr) were studied. The percentage of epithelial cells was assessed by immunocytochemistry using an anti-cytokeratin antibody. The viability of the cells was assessed by trypan blue exclusion. The release of 15-HETE PGE2 and fibronectin was studied in resting cells and after A23187 calcium ionophore stimulation. Epithelial cells always comprised more than 86% of cells recovered, and the viability of epithelial cells was significantly (p < 0.001, Mann-Whitney U test) greater in normal subjects (54 +/- 5%) compared with asthmatic subjects (13 +/- 1%). The release of 15-HETE and fibronectin by resting epithelial cells was significantly greater in asthmatics (p < 0.05, Mann-Whitney U test) than in normal subjects. A23187 significantly (p < 0.05, Wilcoxon W test) increased the release of 15-HETE and fibronectin. There was no significant difference in the release of PGE2 by resting cells from either asthmatics or normal subjects, but challenge with A23187 induced a significant (p < 0.03, Wilcoxon W test) increase in PGE2 from cells of asthmatics but not from cells of normal subjects. This study shows that epithelial cells are activated and less viable in asthma and suggests a role for these cells in asthma.

Topics: Adult; Albuterol; Asthma; Bronchi; Bronchoalveolar Lavage Fluid; Bronchoscopy; Calcimycin; Cell Survival; Dinoprostone; Epithelial Cells; Epithelium; Fiber Optic Technology; Fibronectins; Humans; Hydroxyeicosatetraenoic Acids

Human alveolar macrophages (AM) from bronchoalveolar lavage of asthmatic patients (AP) and healthy volunteers (HS) were compared for their respective capacities to produce lipoxins and leukotrienes when stimulated by calcium ionophore A23187 with or without 15(S)-HETE. The metabolites were analyzed using an isocratic RP-HPLC system and their formation profiles evaluated on the basis of chromatographic behaviour, UV spectral characteristics and co-elution with synthetic standards. Without 15-HETE, AM from AP produced more LTB4 and 5-HETE than those from HS. In the presence of 15-HETE, human AM were able to produce 5,15-diHETE and lipoxins. Moreover, the total amount of lipoxins synthesized by AM from AP was 2 fold higher than that synthesized by AM from HS, thus showing an enhanced cell activation via the 5-lipoxygenase (5-LO) pathway. These results presented AM as in vitro 15-HETE metabolizing cells and suggested some hypothesis about human AM 5-LO regulation mechanism. The enhanced 5-LO activity in AM from AP suggested that in vivo they could participate in cell to cell interaction mechanisms involved in inflammatory lung diseases and might also take up and transform 15-HETE predominantly released by airway epithelial cells.

Topics: Adult; Asthma; Bronchoalveolar Lavage Fluid; Calcimycin; Cells, Cultured; Chromatography, High Pressure Liquid; Epithelial Cells; Epithelium; Humans; Hydroxyeicosatetraenoic Acids; Leukotriene B4; Lipoxins; Lung; Macrophage Activation; Macrophages, Alveolar; Middle Aged

Topics: Asthma; Bronchi; Calcimycin; Cell Survival; Dinoprostone; Epithelium; Fibronectins; HLA-DR Antigens; Humans; Hydroxyeicosatetraenoic Acids; Middle Aged

Airway obstruction and hyperreactivity are characteristics of human asthma and of "heaves," a naturally occurring respiratory disorder of horses and ponies. We measured pulmonary function and plasma immunoreactive 15-hydroxyeicosatetraenoic acid (i15-HETE) concentrations in simultaneously collected carotid artery and right ventricle blood samples in five affected ponies and their age- and gender-matched control ponies. Measurements and sampling were performed before (Period A), during (Period B), and following recovery from (Period C) acute airway obstruction precipitated by housing ponies in a barn and exposing them to hay dust. Pulmonary resistance increased significantly, and dynamic compliance and PaO2 decreased significantly in affected ponies at Period B. Plasma i15-HETE concentrations were greater in carotid artery samples compared with right ventricle samples in affected ponies at each measurement period, suggesting that the lung was a source of i15-HETE. Carotid artery i15-HETE concentrations were significantly greater in affected ponies than in control ponies and increased at Period B. There was a significant negative correlation between changes in plasma i15-HETE and changes in dynamic compliance between measurement Periods A and B. We conclude that the lung is a source of i15-HETE in ponies with heaves, that these ponies produce greater quantities of i15-HETE than control ponies, and that exposing affected ponies to a barn environment produces acute airway obstruction and increased plasma concentrations of i15-HETE.

Topics: Animals; Asthma; Chromatography, High Pressure Liquid; Disease Models, Animal; Horse Diseases; Horses; Hydroxyeicosatetraenoic Acids; Lung; Lung Diseases, Obstructive; Radioimmunoassay; Respiratory Function Tests

Topics: Arachidonate 15-Lipoxygenase; Arachidonate 5-Lipoxygenase; Arachidonic Acid; Arachidonic Acids; Asthma; Bronchi; Calcimycin; Epithelium; Humans; Hydroxyeicosatetraenoic Acids; Lung

15(S)-Hydroxy-5,8,11,13-eicosatetraenoic acid (15-HETE) was by far the most abundant metabolite of arachidonic acid in chopped human bronchi, as identified by reverse phase HPLC, uv spectrometry, and GC/MS. The quantitation of monohydroxyeicosatetraenoic acids (mono-HETEs) was performed by the use of 16(S)-hydroxy-9(Z),12(Z),14(E)-heneicosatrienoic acid as internal standard. Thus, significant amounts of 15-HETE were obtained in incubations of bronchi in buffer alone, but the addition of exogenous arachidonic acid (3-100 microM), dose-dependently increased the formation, with maximal levels reached at around 10 min. In contrast, challenge with ionophore A23187 or anti-human IgE did not stimulate the production of 15-HETE in the bronchi. Nordihydroguaiaretic acid inhibited the production of 15-HETE, whereas indomethacin did not. Small amounts of 8,15-diHETEs were detected in incubations with exogenous 15H(P)ETE. Lipoxins were however not detected under any of the incubation conditions used. Furthermore, removal of the airway epithelium substantially diminished the production of 15-HETE in the bronchi. Finally, bronchi were obtained from three patients with asthma, and the amounts of 15-HETE in these specimens were significantly higher than those found in tissues from nonasthmatics. Also, in peripheral lung parenchyma and pulmonary blood vessels 15-HETE was the major mono-HETE after stimulation with arachidonic acid but the levels were about 10 times lower than in the bronchi. As another difference, challenge of the parenchyma with the ionophore A23187 made 5-HETE the predominant mono-HETE. Taken together, airway epithelium appears to be the major source of 15-HETE in the human lung and the findings in specimens of asthmatics raise the possibility that 15-HETE somehow is involved in airway inflammation.

Topics: Arachidonic Acid; Arachidonic Acids; Asthma; Bronchi; Chromatography, High Pressure Liquid; Epithelium; Humans; Hydroxyeicosatetraenoic Acids

15-hydroxyeicosatetraenoic acid (15-HETE) is the predominant oxidative metabolite of arachidonic acid in human lung. We have studied its effects on airway calibre and non-specific bronchial responsiveness (NSBR) in eight normal and eight asthmatic subjects. 15-HETE, at doses up to 70 nmol, had no effect on airway calibre in either group of subjects. However, 3 h after its administration, 15-HETE reduced NSBR in the normal subjects (geometric mean methacholine PD40 Vp30 increased by 2.29-fold from baseline compared with a corresponding 1.14-fold increase after diluent, p less than 0.05). Similarly, 4 h after inhaled 15-HETE, the spontaneous increase in NSBR in the asthmatics was completely inhibited (geometric mean histamine PD40 Vp30 decreased significantly to 0.41-fold of baseline after diluent (p less than 0.01) compared with a 1.1-fold increase after 15-HETE, p less than 0.01). These data suggest 15-HETE may play an autacoid role in airway function.

Topics: Administration, Inhalation; Adult; Asthma; Bronchi; Bronchial Provocation Tests; Female; Forced Expiratory Volume; Histamine; Humans; Hydroxyeicosatetraenoic Acids; Male; Methacholine Compounds; Research Design; Time Factors

Among the many possible mediators of the early asthmatic response, prostaglandin D2, a bronchoconstrictor, is the principal cyclooxygenase metabolite of arachidonic acid that is released upon the activation of mast cells and is also synthesized by human alveolar macrophages. We performed bronchoalveolar lavage in five patients with chronic stable asthma, before and up to nine minutes after local provocative challenge with Dermatophagoides pteronyssinus. The lavage fluid was analyzed for products of arachidonic acid metabolism. Prostaglandin D2 levels in all five patients rose an average of 150-fold, from less than 8 to 332 +/- 114 pg per milliliter (mean +/- SEM; P less than 0.050), after local instillation of the antigen. Levels of 15-hydroxyeicosatetraenoic acid, which may also have a role in the pulmonary allergic response, were detectable in lavage fluid before challenge and increased after provocation with the antigen in four of the five patients. The activity of beta-glucuronidase, an enzyme released by macrophages and mast cells upon stimulation, tended to increase in the lavage fluid after provocation in all patients. These studies provide evidence that the release of prostaglandin D2 into the airways is an early event after the instillation of D. pteronyssinus in patients who are sensitive to this antigen.

Topics: Antigens; Asthma; Bronchi; Bronchial Provocation Tests; Gas Chromatography-Mass Spectrometry; Glucuronidase; Humans; Hydroxyeicosatetraenoic Acids; Male; Mites; Prostaglandin D2; Prostaglandins D; Pulmonary Alveoli; Therapeutic Irrigation

Extreme sensitivity of airways to multiple stimuli characterizes asthma. Airway hyperresponsiveness can be produced experimentally in otherwise healthy subjects or animals by inflammatory damage (e.g., induced by respiratory viruses or by inhaled oxidants). Evidence is presented that cell-to-cell interactions play an important role in experimental hyperreactivity and that similar inflammatory cascades may play a similar role in clinical asthma. Although the importance of epithelial cells and neutrophils has been identified in the present studies, other inflammatory mechanisms (e.g., sensory nerve release of substance P, epithelial mast cells, eosinophils) may also play key roles. In exercise-induced bronchospasm, the stimulus (e.g., cooling or drying) must affect a cell (e.g., one near the epithelial surface) by decreasing temperature or by increasing osmolality. This signal may cause mediator release and a subsequent cascade, leading to contraction of smooth muscle. Environmental irritants (e.g., ozone) inhaled during exercise may potentiate these effects by producing further inflammation.

Topics: Animals; Arachidonic Acids; Asthma; Cell Communication; Chemotactic Factors; Dinoprostone; Dogs; Epithelial Cells; Epithelium; Humans; Hydroxyeicosatetraenoic Acids; Inflammation; Interleukin-8; Leukotriene B4; Neutrophils; Ozone; Prostaglandins E; Sputum

Topics: Arachidonic Acids; Asthma; Biotransformation; Culture Techniques; Female; Humans; Hydroxyeicosatetraenoic Acids; Lung; Male