A composite of exhaled LTB4 , LXA4 , FeNO, and FEV1 as an "asthma classification ratio" characterizes childhood asthma.

BACKGROUND: Aberrant generation of eicosanoids is associated with asthma, but the evidence remains incomplete and its potential utility as biomarkers is unclear. Major eicosanoids in exhaled breath condensates (EBCs) were assessed as candidate markers for childhood asthma. METHODS: Ten exhaled eicosanoid species was evaluated using ELISA in the discovery phase, followed by prediction model-building and validation phases. RESULTS: Exhaled LTB4 , LTE4 , PGE2, and LXA4 showed significant difference between asthmatics (N = 60) and controls (N = 20). For validation, an expanded study population consisting of 626 subjects with asthma and 161 healthy controls was partitioned into a training subset to establish a prediction model and a test sample subset for validation. Receiver operating characteristic (ROC) analyses of the training subset revealed the level of exhaled LTB4 to be the most discriminative among all parameters, including FeNO, and a composite of exhaled LTB4 , LXA4 , together with FeNO and FEV1 , distinguishing asthma with high sensitivity and specificity. Further, the Youden index (J) indicated the cut point value of 0.598 for this composite of markers as having the strongest discriminatory ability (sensitivity = 85.2% and specificity = 83.6%). The predictive algorithm as "asthma classification ratio" was further validated in an independent test sample with sensitivity and specificity being 84.4% and 84.8%, respectively. CONCLUSIONS: In a pediatric study population in Taiwan, the levels of exhaled LTB4 , LTE4 , LXA4, and PGE2 in asthmatic children were significantly different from those of healthy controls, and the combination of exhaled LTB4 and LXA4 , together with FeNO and FEV1 , best characterized childhood asthma.

Parameters of lung inflammation in asthmatic as compared to healthy children in a contaminated city.

BACKGROUND: The impact of air pollution on the respiratory system has been estimated on the basis of respiratory symptoms and lung function. However; few studies have compared lung inflammation in healthy and asthmatics children exposed to high levels of air pollution. The aim of the study was to elucidate the modulatory effect of air pollution on Cysteinyl-leukotrienes (Cys-LTs) levels in exhaled breath condensate (EBC) among healthy and asthmatic children. METHODS: We performed a cross-sectional comparative study. Children between 7-12 years of age, asthmatics and non-asthmatics, residents of a city with high levels of PM10 were included. In all cases, forced spirometry, Cys-LTs levels in EBC, and the International Study of Asthma and Allergies in Childhood questionnaire were evaluated. We also obtained average of PM10, CO, SO2 and O3 levels during the period of the study by the State Institute of Ecology. RESULTS: We studied 103 children (51 asthmatics and 52 non-asthmatics). Cys-LTs levels were higher in asthmatics than in non-asthmatics (77.3 ± 21.6 versus 60.3 ± 26.8 pg/ml; p = 0.0005). Also, Cys-LTs levels in children with intermittent asthma were lower than in children with persistent asthma (60.4 ± 20.4 versus 84.7 ± 19.2 pg/ml; p = 0.0001). In the multiple regression model, factors associated with levels of Cys-LTs were passive smoking (ß = 13.1, p 0.04) and to be asthmatic (ß = 11.5, p 0.03). CONCLUSIONS: Cys-LTs levels are higher in asthmatic children than in healthy children in a contaminated city and its levels are also associated with passive smoking.

Menopausal asthma: a new biological phenotype?

BACKGROUND: Female hormones play an important role in women's lung health, especially in asthma pathophysiology. Although a growing interest has recently been aroused in asthma related to short-term reproductive states, menopausal asthma has been little studied in the past. The aim of the present study was to explore airway inflammation in menopausal asthmatic women in a noninvasive manner. METHODS: Forty consecutive women with menopausal asthma, 35 consecutive women with premenopausal asthma and 30 age-matched healthy controls were enrolled in the study. Urinary LTE-4, induced sputum inflammatory cells, and exhaled LTE-4, IL-6, pH, and NO levels were measured in all the subjects enrolled. RESULTS: Women with menopausal asthma showed decreased estradiol concentrations, high sputum neutrophils, and exhaled IL-6. Women with premenopausal asthma presented instead an essentially eosinophilic inflammatory pattern. Higher urine and breath condensate LTE-4 concentrations were found in premenopausal and menopausal asthma compared to controls. CONCLUSION: Our results substantiate the existence of a new biological phenotype of menopausal asthma that is mainly characterized by neutrophilic airways inflammation and shares several characteristics of the severe asthma phenotype.

[The clinical significance of noninvasive inflammatory markers in exhaled breath condensate and induced sputum in persistent asthmatic patients].

OBJECTIVE: To assess the clinical significance of three different noninvasive airway inflammatory indices in induced sputum and exhaled breath condensate (EBC) from persistent asthmatic patients. METHODS: Moderate and severe asthmatic patients were prescribed inhaled corticosteroids combined with long-acting beta(2) agonists for a month. The symptom scores and percentage of predicted value of forced expiratory volume in one second (FEV(1)) (FEV(1)%pred) were measured while the concentrations of H(2)O(2), NO(3)(-)/NO(2)(-), and cysteinyl-leukotriene E(4) (LTE(4)) in induced sputum and EBC were detected before and after therapy. RESULTS: A total of twenty-five subjects with moderate and severe asthma were enrolled. By combined therapy for one month the asthma symptoms relieved and FEV(1)%pred improved significantly (P < 0.01). The concentrations of H(2)O(2), NO(3)(-)/NO(2)(-) and LTE(4) in induced sputum and EBC declined significantly (P < 0.01) although the concentrations were still higher than those at normal baseline. More marked reduction of H(2)O(2) and NO(3)(-)/NO(2)(-) compared to LTE(4) was observed. It was revealed that the concentrations of H(2)O(2)and NO(3)(-)/NO(2)(-) but not of LTE(4) in EBC were negatively correlated with FEV(1)%pred (P < 0.01) and positively with symptom scores. Such correlations were also found in H(2)O(2) in induced sputum with FEV(1)%pred and symptom scores as well as NO(3)(-)/NO(2)(-) in induced sputum with FEV(1)%pred. The improvement of FEV(1)%pred after treatment was positively correlated with the reduction of H(2)O(2) and NO(3)(-)/NO(2)(-) both in induced sputum and EBC. Correlation analysis also demonstrated three inflammatory indices were equivalent in induced sputum and EBC (correlation coefficient of H(2)O(2), NO(3)(-)/NO(2)(-) and LTE(4), 0.759, 0.826 and 0.653, respectively. P < 0.01). CONCLUSIONS: (1) Combined therapy with inhaled corticosteroid plus long-acting beta(2) agonist significantly improves the clinical symptoms and lung function of patients with moderate and severe asthma companies with marked suppression of airway inflammation. (2) Both of EBC and induced sputum sampling are valuable noninvasive procedures for detecting asthma airway inflammation, however, EBC technique is superior in safety and reproducibility. (3) H(2)O(2) and NO(3)(-)/NO(2)(-) seem to be more sensitive indices in diagnosis and monitoring asthma compared to LTE(4).

Fatty Acid Composition and Eicosanoid Levels (LTE4 and PGE2) of Human Milk from Normal Weight and Overweight Mothers.

Background: Maternal obesity is known to affect human milk composition. Long-chain polyunsaturated fatty acids (LCPUFA) are vital nutrients to the nervous system development and precursors of eicosanoids related to obesity (prostaglandin E2-PGE2-and leukotriene E4-LTE4). The aim of the present research was to study the lipid profiles, with particular emphasis to LCPUFAs, and the concentrations of eicosanoids PGE2 and LTE4, involved in adipose tissue development, in human milk from overweight mothers compared with normal weight mothers. Materials and Methods: Study including 46 overweight and 86 normal weight breastfeeding volunteers was carried out. Fatty acids and eicosanoids (PGE2 and LTE4) were analyzed in mature human milk. Fatty acids quantification was determined by gas chromatography and mass spectrometry. PGE2 and LTE4 were measured by immununoassay. Results: Human milk of overweight mothers had lower contents of n-3 LCPUFA, including eicosapentaenoic acid (20:5n-3, EPA) and docosahexaenoic acid (22:6n-3, DHA) and higher levels of total n-6 LCPUFA, compared with normal weight mothers (0.45 ± 0.23 versus 0.58 ± 0.38, p = 0.016; 0.05 ± 0.04 versus 0.08 ± 0.08, p = 0.005; 0.26 ± 0.15 versus 0.34 ± 0.22, p = 0.015; 0.84 ± 0.25 versus 0.74 ± 0.20, p = 0.029; respectively). Multiple regression analyses showed that maternal overweight was associated with human milk fatty acid profile. The levels of PGE2 and LTE4 in human milk did not show significant differences between groups. Conclusions: Our findings support the hypothesis that mother weight status influences human milk n-3 LCPUFA lipid composition, but not its relationship with PGE2 and LTE4 levels.

Increased leukotriene E4 in the exhaled breath condensate of children with mild asthma.

BACKGROUND: Chronic airway inflammation is a feature of asthma. Increased levels of cysteinyl leukotrienes (cys-LTs; leukotriene [LT]C(4), LTD(4), LTE(4)) have been shown in the exhaled breath condensate (EBC) of children with moderate-to-severe asthma. The aim of this study was to examine the relationship between EBC cys-LTs (LTE(4)) levels and bronchial hyperreactivity in children with mild asthma in order to evaluate the clinical utility of measuring EBC cys-LTs levels. METHODS: We measured LTE(4) levels in the EBC of children aged 8 to 18 years, including healthy nonasthmatic children (n = 6) and children with mild asthma (n = 37). Patients with mild asthma were classified into the following three groups: group 1, participants who had been asymptomatic (no wheezing/symptoms of asthma) for > 6 months prior to examination (n = 12); group 2, participants who were asymptomatic but had had wheezing/symptoms of asthma within 6 months before examination (n = 18); and group 3, patients with current wheeze and/or mild symptoms of asthma exacerbation at the time of examination. RESULTS: Exhaled LTE(4) levels were increased in all children with mild asthma compared with nonasthmatic control subjects (5.69 +/- 9.62 pg/20 min vs 0.74 +/- 0.79 pg/20 min, p < 0.05) [mean +/- SD]. In particular, the EBC LTE(4) levels in group 2 (4.99 +/- 6.70 pg/20 min) and group 3 (14.66 +/- 17.11 pg/20 min) were increased compared with control subjects and group 1 (1.50 +/- 1.69 pg/20 min). The EBC LTE(4) levels negatively correlated with the provocative concentration of methacholine causing a 15% fall in FEV(1) (r = - 0.454, p = 0.012). CONCLUSION: EBC cys-LTs may be useful as a noninvasive marker assessing airway inflammation and hyperreactivity in children with asthma.

Evaluation and interference of serum and skin lesion levels of leukotrienes in patients with eczema.

To evaluate the levels of leukotrienes (LTs), interleukin-2 (IL-2), interleukin-4 (IL-4), interferon-gamma (IFN-gamma) in patients with eczema and observe the effects inversed by mizolastine. Serum LTB4, LTC4, IL-2, IL-4, IFN-gamma and urinary LTE4 levels were detected by enzyme-linked immunosorbent assay (ELISA) and LTB4, LTC4, LTE4 concentrations of cutis tissue were measured by reverse-phase high-pressure liquid chromatography (RP-HPLC) in 10 eczema patients and 10 healthy volunteers. Eczema patients received mizolastine 10 mg once a day for 5 days, respectively, for comparison between before and after treatment. The above markers were assayed again after treatment. Serum LTB4, LTC4, IL-2, IFN-gamma and urinary LTE4 and skin tissue LTB4, LTC4, LTE4 levels in patients are higher than those in healthy volunteers significantly (P < 0.05). But serum IL-4 level did not show significant difference between patients and normal controls (P > 0.05). Mizolastine significantly reduced serum LTB4 and IFN-gamma levels as well as skin lesion LTB4, LTC4, LTE4 concentrations. LTs are involved in the pathogenesis of eczema. Mizolastine clearly reduces LTs levels in skin lesion.

Antigen specific quantification of sulfidoleukotrienes in patients allergic to Betalactam antibiotics.

BACKGROUND: After in vitro allergen-specific stimulation, basophils become activated and release sulfidoleukotrienes LTC4, LTD4 and LTE4. This can be detected by means of the CAST assay. We assessed the positivity criteria and the reliability of antigen-specific sulfidoleukotriene production (CAST) in the in vitro diagnosis of betalactam (BL) allergic patients. MATERIAL AND METHODS: We studied a sample of 67 patients (age 48.94 +/- 15.76 years) who had presented with anaphylaxis or urticaria-angioedema within the first 60 minutes after administration of Amoxicillin (54/67), Penicillin G (7/67), Cefuroxime (5/67) or Cefazoline (1/67). All of them had a positive skin test to at least one of the antigenic determinants of Penicillin. As control group 30 adults with negative skin tests who tolerated BL were included. All of them underwent skin tests, oral provocation tests, specific IgE (CAP-FEIA, Pharmacia) and CAST. RESULTS: Positivity criteria were established by means of ROC curves: a sLT release induced by Betalactams of at least 100 pg/ml and greater than or equal to 3 times the basal value. The overall sensitivity of CAST is 47.7% and specificity 83.3%. Sensitivity of specific IgE is 37.8% and specificity 83.3%. CONCLUSIONS: We have established validated positivity criteria for the CAST technique in patients allergic to Betalactams. This technique is a useful in vitro diagnostic method in patients with IgE-mediated allergy to Betalactam antibiotics.

Effects of pranlukast on chemical mediators in induced sputum on provocation tests in atopic and aspirin-intolerant asthmatic patients.

BACKGROUND: Leukotrienes (LTs) are important in asthma, and LT modifiers modulate antigen-induced asthma. Overproduction of LT by suppression of cyclooxygenase activity is involved in patients with aspirin-intolerant asthma (AIA). METHODS: House dust mite (HDM) inhalation provocation tests were performed in HDM-sensitive asthmatic inpatients without AIA (HDM group; n = 6), and aspirin oral provocation tests were performed in AIA patients (ASA group; n = 7). Tests were repeated using the same regimen after 7 days of treatment with pranlukast, an LT receptor antagonist (LTRA). The effects of pranlukast on changes in sputum LTC(4)-LTD(4), eosinophil cationic protein (ECP), eosinophil count, urinary LTE(4)/creatinine, 11-dehydrothromboxane B(2) (11-dhTXB(2))/creatinine, serum LTC(4)-LTD(4), ECP, and peripheral blood eosinophil count, during immediate asthmatic reaction (IAR) and late asthmatic reaction (LAR) in the HDM group and during IAR in the ASA group for each test, were compared in each group. RESULTS: In the HDM group, IAR and LAR were observed. Sputum LTC(4)-LTD(4) and urinary LTE(4)/creatinine increased significantly both during IAR and LAR. Sputum ECP increased during IAR and further increased during LAR. Eosinophil count in the sputum did not increase during IAR but significantly increased during LAR. Pranlukast suppressed the fall in FEV(1) both during IAR and LAR (73.8% and 51.9%, respectively) and inhibited the increase in sputum eosinophil count during LAR and sputum ECP during IAR and LAR. In the ASA group, aspirin-induced IAR was associated with a fall in urinary 11-dhTXB(2)/creatinine, increased the levels of sputum LTC(4)-LTD(4) and ECP and urinary LTE(4)/creatinine. Pranlukast suppressed IAR and inhibited the increase of the level of sputum ECP, but failed to change aspirin-induced LT production in the sputum and urine. The levels of sputum LTC(4)-LTD(4) and urinary LTE(4)/creatinine in the stable phase in the ASA group were significantly greater than those in the HDM group. CONCLUSION: Our results indicated that HDM-provoked asthma is associated with overproduction of LT with an antigen-antibody reaction, while AIA is associated with overproduction of LT with a shift to the 5-lipoxygenase series of the arachidonate cascade. LTRA may be useful against both types of asthma through inhibition of LT activity and eosinophilic inflammation of the airways.

Cysteinyl leukotrienes in the bile of patients with obstructive jaundice.

BACKGROUND: Cysteinyl leukotrienes (LTs) are potent proinflammatory mediators. They are predominantly excreted from blood by hepatobiliary elimination. To explore the clinical significance of biliary cysteinyl LTs, we determined their concentration changes in bile during treatment in patients with obstructive jaundice. METHODS: Bile samples were obtained during endoscopic or transhepatic biliary drainage. Leukotrienes C(4), D(4), and E(4) were quantified by two-step reversed-phase high-performance liquid chromatography and subsequent radioimmunoassay. RESULTS: The increased excretion of cysteinyl LTs (LTC(4) + LTD(4) + LTE(4)) decreased between day 1 and 14 after drainage (means, 171 pmol/h to 79 pmol/h; P < 0.02). During drainage, the excretion was higher when there was additional cholangitis (mean, 225 and 86 pmol/h, with and without cholangitis, respectively; P < 0.001). The concentrations of LTD(4) and LTE(4) were also higher with additional cholangitis than without (LTD(4), mean 6.0 vs 2.0 nM; P < 0.05; LTE(4), 6.8 vs 2.4 nM; P < 0.02, respectively). Biliary LTC(4) was detected only in patients with cholangitis. The biliary excretion of cysteinyl LTs was positively correlated with leukocyte concentration ( r = 0.68; P < 0.005) and C-reactive protein ( r = 0.73; P < 0.005) in blood. Furthermore, only in the absence of cholangitis, the excretion was positively correlated with serum gamma-glutamyl transferase ( r = 0.76; P < 0.02) and alanine aminotransferase ( r = 0.72; P < 0.02). CONCLUSIONS: The excretion of biliary cysteinyl LTs increases with the severity of cholestasis and hepatic inflammation in patients with obstructive jaundice. An additional increase of cysteinyl LTs was observed during bacterial cholangitis. The increased biliary excretion of biologically active cysteinyl LTs may contribute to the aggravation of cholestasis and inflammatory reaction in obstructive jaundice.

On-line coupling of microdialysis sampling with liquid chromatography for the determination of peptide and non-peptide leukotrienes.

An automated on-line sampling method was developed using microdialysis as the simultaneous sampling and sample pre-treatment technique. The extraction fraction values of microdialysis probes sampling different eicosanoids were investigated. The impact of cyclodextrins in the perfusion liquid used for sampling hydrophobic eicosanoids in biological systems was also studied. The total time for one analysis was 7.6 min allowing seven measurements per hour for monitoring kinetic changes in biological systems.

Eosinophil influx into the airways in patients with exercise-induced asthma.

Exercise-induced asthma is a common phenomenon, the mechanism of which is undetermined. Eosinophils have been suggested as playing a role in its occurrence. We studied the effect of exercise-induced asthma on the cellular and mediator composition of spontaneously obtained sputum. Twenty-five patients with bronchial asthma were investigated by studying sputum spontaneously obtained before and following challenge. One group with (n=9) and one without (n=9) exercise-induced asthma performed exercise challenge. A third group (n=7) performed methacholine challenge. The sputum was analysed using Giemsa staining for differential cell count, measuring eosinophil cationic proteins and mixtures of leukotrienes (D4, E4 and C4) in the liquid phase using ELISA. The group with exercise-induced asthma had a mean drop of 23.7+/-7.4% in FEV1, significantly (P=0.001) higher than the group without it. Following challenges, there were significant increases in sputum eosinophils only in the group with exercise-induced asthma (from 8.1+/-13.9% to 18.3+/-20.2%, P=0.0017) and not in control groups (from 0.9+/-0.9% to 1.5+/-15%) or in those who had methacholine challenge (from 23.6+/-27.2% to 22.3+/-23.8%). Eosinophil cationic proteins did not change significantly in any group. In the liquid phase of the sputum, the amount of leukotrienes increased following exercise in six of the seven patients with exercise-induced asthma in whom it was measured. The influx of eosinophils to the airway in patients who develop exercise-induced asthma can be partially explained by the leukotrienes in the airways of those patients.

Concentrations of cysteinyl leukotrienes in urine and bronchoalveolar lavage fluid of cats with experimentally induced asthma.

OBJECTIVE: To evaluate changes in cysteinyl leukotriene (LT) concentrations in urine and bronchoalveolar lavage fluid (BALF) in cats with experimentally induced asthma. ANIMALS: 19 cats with experimentally induced asthma and 5 control cats. PROCEDURE: Cats were sensitized to Bermuda grass or house dust mite allergen, and phenotypic features of asthma were confirmed with intradermal skin testing, evaluation of BALF eosinophil percentages, and pulmonary function testing. A competitive ELISA kit for LTC4, LTD4, and LTE4 was used for quantitative analysis of LTs. Urinary creatinine concentrations and BALF total protein (TP) concentrations were measured, and urinary LT-to-creatinine ratios and BALF LT-to-TP ratios were calculated. RESULTS: Mean urinary LT-to-creatinine ratios did not differ significantly between control cats and allergen-sensitized cats before or after sensitization and challenge exposure with saline (0.9% NaCl) solution or allergen, respectively. In BALF the mean LT-to-TP ratio of control cats did not differ significantly before or after sensitization and challenge exposure with saline. Asthmatic cats had BALF LT-to-TP ratios that were significantly lower than control cats at all time points, whereas ratios for asthmatic cats did not differ significantly among the various time points. CONCLUSIONS AND CLINICAL RELEVANCE: Although LTs were readily detectable in urine, no significant increases in urinary LT concentrations were detected after challenge in allergen-sensitized cats. Spot testing of urinary LT concentrations appears to have no clinical benefit for use in monitoring the inflammatory asthmatic state in cats. The possibility that cysteinyl LTs bind effectively to their target receptors in BALF and, thus, decrease free LT concentrations deserves further study.

Prostaglandins, leukotrienes and perennial rhinitis.

Prostaglandins and leukotrienes are implicated in conditions of both the upper and lower airways. In the former they are deranged in nasal polyposis, intrinsic rhinitis and allergic rhinitis while in the latter they are involved in the pathogenesis of asthma. The aim of the present study was to measure mucosal eicosanoid levels in the three types of rhinitis and compare with controls. In addition, the effect of topical steroids on eicosanoid levels in rhinitis was examined. The levels of prostaglandins E(2) (PGE(2)) and D(2) (PGD(2)) and of leukotrienes E(4) (LTE(4)) and B(4) (LTB(4)) were measured in nasal biopsies from the inferior turbinates of patients suffering from perennial rhinitis and a control group. Rhinitis patients were classified into three categories: perennial allergic rhinitis (PAR), non-allergic rhinitis with eosinophilia (NARES) and noneosinophilic non-allergic rhinitis (NENAR) on the basis of symptoms, secretion eosinophilia, nasal resistance and allergy testing. Patients with rhinitis were randomized into two groups. One received fluticasone propionate nasal spray (FPANS) and the other a placebo (PNS) over a period of six weeks prior to the biopsies. One hundred and one patients with PAR, NARES or NENAR were recruited sequentially and the control group consisted of 21 patients with no evidence of rhinitis but with nasal obstruction due to septal deviation. Untreated rhinitics had significantly lower levels of PGE(2), PGD(2) and LTE(4) than non-rhinitic controls. Six-weeks' treatment with FPANS significantly increased the levels of those eicosanoids in patients with PAR and NARES but they were still significantly below normal. Levels of LTB(4) in all three rhinitis groups were not significantly different from controls and treatment with topical steroids had no effect. Their findings are contrary to current thinking that increased levels of eicosanoids, in particular cysteinyl-leukotrienes, play an important role in the pathogenesis of chronic, non-infective upper airway inflammation.

Increased gastric juice leukotriene B4, C4 and E4 concentrations in children with Helicobacter pylori colonization.

During recent years, the role of inflammatory lipid mediators in the pathophysiology of Helicobacter pylori (H. pylori) infections has been investigated in several studies. The concentrations of leukotrienes (LTs) in gastric juice from H. pylori positive (n = 13) and negative (n = 18) children with recurrent abdominal pain were studies in order to determine whether these lipid inflammatory mediators are involved in local and systemic biological actions. Gastric juice samples and biopsy specimens of mucosa were obtained endoscopically from 31 patients with recurrent abdominal pain for assessment of LTs and histopathological examination. In this study, all children with recurrent abdominal pain were investigated by rapid urease test and histological assessment for H. pylori colonization. Leukotriene levels were measured by high performance liquid chromatography (HPLC) and radioimmunoassay (RIA) in gastric juice samples. Gastric juice LTB4, LTC4, and LT4 levels were significantly higher in patients with H. pylori colonization than in children without H. pylori colonization. These results indicate that increased gastric content of proinflammatory mediators (LTB4, LTC4, and LT4) may be related to the pathogenesis of H. pylori-associated gastritis.

Immunomagnetic molecular probe with UHPLC-MS/MS: a promising way for reliable bronchial asthma diagnostics based on quantification of cysteinyl leukotrienes.

A sensitive and precise method for simultaneous quantification of cysteinyl leukotrienes (=cys LTs) - leukotriene C4 (=LTC4), leukotriene D4 (=LTD4) and leukotriene E4 (=LTE4) - essential biomarkers of bronchial asthma present in exhaled breath condensate (=EBC) was developed. An immunomagnetic molecular probe was prepared by anchoring cysteinyl leukotrienes antibody on the surface of functionalized monodispersed magnetic particles and used to selectively isolate cys LTs from biological matrices - EBC, plasma and urine. Immobilization and the immunoaffinity capture procedures were optimized to maximize the amount of separated cys LTs, which were detected "off-beads" after acidic elution by UHPLC-ESI-MS/MS operated in a multiple reaction monitoring mode. The developed method was characterized with high precision ≤13.6% (intra-day precision determined as RSD) and ≤14.5% (inter-day precision determined as RSD), acceptable accuracy ≤18.5% (determined as RE), and high recovery of immunoseparation (≥93.1%) in aforementioned biological matrices. The applicability of the method was demonstrated on EBC, plasma and urine clinical samples of patients with various subtypes of bronchial asthma (occupational, steroid-resistant, moderate with and without corticosteroids therapy) and healthy subjects where reasonable differences in cys LTs concentration levels were found. Combining extremely selective immunomagnetic separation with highly sensitive and precise detection step, the developed method was used to aid diagnosis, predict the most effective therapy, and monitor the response to treatment. The detection of elevated inflammatory mediators (cys LTs) in EBC of subjects with relatively asymptomatic asthma and normal pulmonary function tests could offer a novel way for monitoring the lung inflammation and perhaps initiating treatment in an earlier stage.

Aspirin challenge in patients with chronic rhinosinusitis with polyps correlates with local and systemic inflammatory markers.

BACKGROUND: Acetylsalicylic acid (ASA; aspirin) is a well-known inducer of pseudoallergic response in patients with chronic rhinosinusitis with polyps (CRSwPs). The mechanism that leads to this response remains unclear. This study was designed to measure and compare the local and systemic inflammatory response to aspirin challenge in patients with CRSwPs who develop either a nasobronchial response (NBR) or a nasal response (NR), and compare it with nonresponders (non-Rs). METHODS: The three groups underwent nasal wash before ASA challenge, and inflammatory mediators were measured in the nasal wash as well as in serum. RESULTS: A total of 25 CRSwP patients were enrolled. The NBR patients (n = 13) had a significantly longer mean disease duration and a higher mean serum leukotriene E4 (LTE4) level than the NR (n = 6) and non-R (n = 6) patients (39.2 ± 9.7 months, 21 ± 8.8 months, and 22.8 ± 11.2 months, respectively, and 4221 ± 1205 pg/mL, 1430 ± 605 pg/mL, and 857 ± 461 pg/mL, respectively). The NBR and NR patients had a larger mean number of nasal eosinophils than the non-R group (52.8 ± 28.8 cells/µL, 47 ± 21.3 cells/µL, and 19.3 ± 13.4 cells/µL, respectively). The tryptase, albumin, nasal LTE4, and prostaglandin E2 levels were not significantly different between the three groups in any examined combination. CONCLUSION: The nasal eosinophil and serum LTE4 levels correlate with aspirin sensitivity.

Leukotrienes B4, C4, D4 and E4 in the exhaled breath condensate (EBC), blood and urine in patients with pneumoconiosis.

Leukotrienes (LTs) are involved in the pathogenesis of lung fibrosis and were increased in exhaled breath condensate (EBC) of the patients with pneumoconiosis. However the possible influence of extra-pulmonary disorders on the EBC markers is not known. Therefore in parallel with EBC, LTs' levels in the plasma and urine were measured in patients with pneumoconiosis (45 × asbestos exposure, 37 × silica exposure) and in 27 controls. Individual LTs B4, C4, D4 and E4 were measured by liquid chromatography - electrospray ionization - tandem mass spectrometry (LC-ESI-MS/MS). In EBC, LT D4 and LT E4 were increased in both groups of patients (p<0.001 and p<0.05), comparing with the controls. Both LT B4 and cysteinyl LTs were elevated in asbestos-exposed subjects (p<0.05). Asbestosis with more severe radiological signs (s1/s2-t3/u2) and lung functions impairment has shown higher cysteinyl LTs and LT C4 in the EBC (p<0.05) than mild asbestosis (s1/s0-s1/s1). In addition, in the subjects with asbestosis, cysteinyl LTs in EBC correlated with TLC (-0.313, p<0.05) and TLCO/Hb (-0.307, p<0.05), and LT C4 with TLC (-0.358, p<0.05). In pneumoconioses, EBC appears the most useful from the 3 fluids studied.

Exhaled leukotrienes and bronchial responsiveness to methacholine in patients with seasonal allergic rhinitis.

BACKGROUND: Allergic rhinitis and bronchial asthma can coexist and affect each other. OBJECTIVE: To investigate the relationship between the postseasonal increase in the concentration of leukotriene (LT) B4 and LTE4 in exhaled breath condensate (EBC) and bronchial responsiveness to methacholine (BRM) in patients with seasonal allergic rhinitis (SAR). METHODS: In 28 patients with SAR and 50 healthy study patients, the leukotrienes were measured in EBC during and after the pollen season by gas chromatography/mass spectrometry. The BRM was determined after the pollen season. RESULTS: In 7 patients with SAR, significantly increased concentrations of both the leukotrienes were found in EBC during and 5 months after the pollen season. The following seasonal and postseasonal median values were measured in patients with SAR in comparison with control patients: LTB4: 131 and 90 pg/mL vs 80 and 79 pg/mL, P < .001 and P = .03, respectively; LTE4: 122 and 86 pg/mL vs 76 and 74 pg/mL, P < .001 and P = .02, respectively. Five months after the pollen season, the concentrations of LTB4 and LTE4 decreased with respect to their seasonal values (90 and 86 pg/mL, respectively, P < .001, for both leukotrienes). In 7 patients with SAR and leukotriene levels exceeding the reference limits, significantly increased BRM was also found (LTB4: P = .02; LTE4: P = .002). CONCLUSIONS: The seasonal and postseasonal increases in LTB4 and LTE4 concentrations in EBC of the patients with SAR correlated significantly with the later increase in BMR. This relationship could provide a useful predictive parameter for early inflammatory processes in the lower airways of patients with allergic rhinitis.