Sputum proteomic signature of gastro-oesophageal reflux in patients with severe asthma.

Gastro-oesophageal reflux disease (GORD) has long been associated with poor asthma control without an established cause-effect relationship. 610 asthmatics (421 severe/88 mild-moderate) and 101 healthy controls were assessed clinically and a subset of 154 severe asthmatics underwent proteomic analysis of induced sputum using untargeted mass spectrometry, LC-IMS-MSE. Univariate and multiple logistic regression analyses (MLR) were conducted to identify proteins associated with GORD in this cohort. When compared to mild/moderate asthmatics and healthy individuals, respectively, GORD was three- and ten-fold more prevalent in severe asthmatics and was associated with increased asthma symptoms and oral corticosteroid use, poorer quality of life, depression/anxiety, obesity and symptoms of sino-nasal disease. Comparison of sputum proteomes in severe asthmatics with and without active GORD showed five differentially abundant proteins with described roles in anti-microbial defences, systemic inflammation and epithelial integrity. Three of these were associated with active GORD by multiple linear regression analysis: Ig lambda variable 1-47 (p = 0·017) and plasma protease C1 inhibitor (p = 0·043), both in lower concentrations, and lipocalin-1 (p = 0·034) in higher concentrations in active GORD. This study provides evidence which suggests that reflux can cause subtle perturbation of proteins detectable in the airways lining fluid and that severe asthmatics with GORD may represent a distinct phenotype of asthma.

8-isoprostane in exhaled breath condensate after experimental exposure to wood smoke in humans.

Wood smoke, a well-known indoor and outdoor air pollutant, may cause adverse health effects through oxidative stress. In this study 8-isoprostane, a biomarker of oxidative stress, was measured in exhaled breath condensate (EBC) and urine before and after experimental exposure to wood smoke. The results were compared with measurements of other biomarkers of oxidative stress and inflammation. Thirteen subjects were exposed first to clean air and then, after 1 week, to wood smoke in an exposure chamber during 4-hour sessions. Exhaled breath condensate, exhaled nitric oxide, blood and urine were sampled before and at various intervals after exposure to wood smoke and clean air. Exhaled breath condensate was examined for 8-isoprostane and malondialdehyde (MDA), while exhaled air was examined for nitric oxide, serum for Clara cell protein (CC16) and urine for 8-isoprostane. 8-isoprostane in EBC did not increase after wood smoke exposure and its net change immediately after exposure was inversely correlated with net changes in MDA (r(s)= -0.57, p= 0.041) and serum CC16 (S-CC16) (r(p)= -0.64, p= 0.020) immediately after the exposure. No correlation was found between 8-isoprostane in urine and 8-isoprostane in EBC. In this study controlled wood smoke exposure in healthy subjects did not increase 8-isoprostane in EBC.

Long-acting beta-agonists and their association with inhaled corticosteroids in COPD.

Inhaled bronchodilators, including beta(2)-agonists and antimuscaric receptor antagonists, are the mainstay of pharmacotherapy in chronic obstructive pulmonary disease (COPD). The short-acting beta(2)-agonists, including salbutamol, and fenoterol, have a rapid onset of action, a bronchodilating effect for 3-6 h and are used on demand. The long-acting beta(2)-agonists (LABAs), including salmeterol and formoterol, have 12-hour duration of action and are used with a twice-daily dosing regimen for long-term COPD treatment. Unlike salmeterol, formoterol has a rapid onset of action. Pharmacological characteristics required by novel inhaled LABAs include 24 h bronchodilator effect in vivo which would make them suitable for once daily administration (ultra-LABA), high potency and selectivity for beta(2)-adrenoceptors, rapid onset of action, low oral bioavailability (< 5%) after inhalation, and high systemic clearance. Indacaterol, which has been approved for long-term treatment of COPD in Europe and in the USA, has a 24-h duration of action and a once-daily dosing regimen. Newer ultra-LABAs, including olodaterol, vilanterol, milveterol, carmoterol, and abediterol, are in development. Combination with ICS (fluticasone/salmeterol, budesonide/formoterol, beclomethasone/formoterol) appears to provide an additional benefit over the monocomponent therapy, although the extent of this benefit is variable and often not clinically significant in all the endpoints assessed. In patients with COPD, treatment with ICS is associated with increased risk of pneumonia which should be carefully considered when assessing the risk/benefit ratio of ICS/LABA combinations. Subphenotyping of patients with COPD (e.g., frequent exacerbations, sputum eosinophilia, mixed asthma/COPD phenotype) might help identify those patients who are most likely to benefit from addition of ICS to bronchodilating treatment. Ultra-LABA/ long-acting muscarinic receptor antagonist (LAMA) combination treatment is under development and is likely to become a standard pharmacological strategy for COPD. Dual-pharmacology inhaled muscarinic antagonist-beta(2) agonist (MABA) molecules provide a new approach to the treatment of COPD.

Inhaled muscarinic acetylcholine receptor antagonists for treatment of COPD.

Bronchodilators, generally administered via metered dose or dry powder inhalers, are the mainstays of pharmacological treatment of stable COPD. Inhaled long-acting beta-agonists (LABA) and anticholinergics are the bronchodilators primarily used in the chronic treatment of COPD. Anticholinergics act as muscarinic acetylcholine receptor antagonists and are frequently preferred over beta-agonists for their minimal cardiac stimulatory effects and greater efficacy in most studies. Their therapeutic efficacy is based on the fact that vagally mediated bronchoconstriction is the major reversible component of airflow obstruction in patients with COPD. However, bronchodilators are effective only on the reversible component of airflow obstruction, which by definition is limited, as COPD is characterized by a fixed or poorly reversible airflow obstruction. Inhaled anticholinergic antimuscarinic drugs approved for the treatment of COPD include ipratropium bromide, oxitropium bromide and tiotropium bromide. Ipratropium bromide, the prototype of anticholinergic bronchodilators, is a short-acting agent. Oxitropium bromide is administered twice a day. Tiotropium bromide, the only long-acting antimuscarinic agent (LAMA) currently approved, is administered once a day. Newer LAMAs including aclidinium bromide and glycopyrrolate bromide are currently in phase III development for treatment of COPD. Some new LAMAs, including glycocpyrrolate, are suitable for once daily administration and, unlike tiotropium, have a rapid onset of action. New LAMAs and their combination with ultra-LABA and, possibly, inhaled corticosteroids, seem to open new perspectives in the management of COPD. Dual-pharmacology muscarinic antagonist-beta2 agonist (MABA) molecules present a novel approach to the treatment of COPD by combining muscarinic antagonism and beta2 agonism in a single molecule.

Exhaled breath condensate, nasal eosinophil cationic protein level and nasal cytology during immunotherapy for cypress allergy.

Interest in cypress allergy is widely rising: an increasing number of studies have pointed out the efficacy of immunotherapy to reduce cypress-related symptoms and drug use. Cypress immunotherapy is well tolerated, but there are few studies dealing with its sub-clinical effects on the airways. The aim of this investigation is to assess the effects of immunotherapy on airways by the analysis of exhaled breath condensate (EBC), nasal lavage fluid (NAL) and nasal cytology. Fifteen mono-sensitized to cypress pollen patients have been observed, among them 9 have been treated with sub-cutaneous immunotherapy (SCIT), 3 with sub-lingual immunotherapy (SLIT) and 3 which were not treated underwent EBC, NAL and nasal cytology out of the pollen season. 8-isoprostane in EBC, Eosinophil cationic protein (ECP) and inflammatory cells in nasal cytology were also evaluated. The median value of 8-isoprostane in EBC was 18.58 pg/ml in patients who did not undergo immunotherapy, 49.38 pg/ml in SCIT patients and 13.41 pg/ml in SLIT subjects. The median value of ECP in nasal lavage was higher in non- treated subjects (27.3 mg/l) than in those treated with SCIT (1 mg/l)(p less than 0,05) or SLIT (2.6 mg/l). All nasal cytology specimens did not show any sign of inflammation. In conclusion SLIT seems to be well tolerated and to reduce significantly the levels of ECP in nasal lavage. In addition the levels of 8-isoprostane in EBC among SCIT patients were unexpectedly high and need to be further evaluated.

Non-invasive biomarkers of lung inflammation in smoking subjects.

Cigarette smoking is the most important risk factor for the development of chronic obstructive pulmonary disease (COPD) and lung cancer, but only a part of smoking subjects develop these respiratory pathologies. Therefore, it is necessary to find sensible parameters to detect early lung alterations due to chronic tobacco smoke exposure. Long-term cigarette smoking is associated with a persistent inflammatory response in the lung that leads to tissue injury and dysfunction. Bronchoscopy and bronchial biopsies are the gold standard techniques for assessing pulmonary inflammation, but are invasive and not routinely used. Cellular analysis of induced sputum and measurement of fraction of exhaled nitric oxide (F(E)NO) are validated non-invasive techniques for assessing respiratory inflammation. Measurement of biomolecules in sputum supernatants and exhaled breath condensate (EBC) are used as a research tool, but require standardization of procedures and, generally, analytical validation. Electronic nose differentiates healthy smokers from healthy nonsmokers based on breath volatile organic compounds (VOC) patterns. These techniques are potentially useful for identifying biomarkers of pulmonary inflammation and oxidative stress. Induced sputum, F(E)NO, EBC and electronic nose are suitable for longitudinal sampling, thereby facilitating monitoring of lung damage process. This approach could enable an early identification of subgroups of healthy smokers at higher risk for tobacco-induced lung damage and prompt planning of secondary prevention strategies.

Leukotriene modifiers for asthma treatment.

Leukotrienes (LTs), including cysteinyl LTs (CysLTs) and LTB(4) , are potent lipid mediators that have a role in the pathophysiology of asthma. At least two receptor subtypes for CysLTs, CysLT(1) and CysLT(2) , have been identified. The activation of the CysLT(1) receptor is responsible for most of the pathophysiological effects of CysLTs in asthma, including increased airway smooth muscle activity, microvascular permeability, and airway mucus secretion. LTB(4) might have a role in severe asthma, asthma exacerbations, and the development of airway hyperresponsiveness. CysLT(1) receptor antagonists can be given orally as monotherapy in patients with mild persistent asthma, but these drugs are generally less effective than inhaled glucocorticoids. Combination of CysLT(1) receptor antagonists and inhaled glucocorticoids in patients with more severe asthma may improve asthma control and enable the dose of inhaled glucocorticoids to be reduced while maintaining similar efficacy. The identification of subgroups of asthmatic patients who respond to CysLT(1) receptor antagonists is relevant for asthma management as the response to these drugs is variable. CysLT(1) receptor antagonists have a potential anti-remodelling effect that might be important for preventing or reversing airway structural changes in patients with asthma. This review discusses the role of LTs in asthma and the role of LT modifiers in asthma treatment.

Novel perspectives in the detection of oral and nasal oxidative stress and inflammation in pediatric united airway diseases.

United airway disease (UAD) concept proposed that asthma and rhinitis are both different clinical manifestation of a single inflammatory process. The aim of this study is to assess in upper and lower airways the level of inflammation and oxidative stress and to investigate the relationship between biomarkers in persistent allergic rhinitis (PER) and in concomitant asthma with PER. By a crosssectional study we measured oral and nasal (FENO) and oral and nasal EBC 8-isoprostane, LTB4 and PGE2 in children with PER (n=14) and with PER and concomitant intermittent asthma (IA; n=25), mild persistent asthma (mA; n=28), moderate persistent asthma (MA; n=13) and in Healthy Controls (HCs; n=13). Oral and nasal FENO concentrations were increased in children with PER, IA, mA and MA when compared with HCs. Nasal 8-isoprostane was higher in EBC of children with PER and asthma than in HCs. Oral and nasal LTB4 were higher in EBC of children with PER and mA than in HCs. Oral and nasal PGE2 concentrations were higher in EBC of children with PER than in HCs. Positive correlations between oral and nasal biomarkers were found in IA for LTB4 and PGE2, in mA for FENO, 8-isoprostane, LTB4 and PGE2, and in MA for PGE2. No correlations were observed in children with PER and HCs. Our results suggest that non-invasive markers of inflammation and oxidative stress might be useful to study the relationships between oral and nasal compartments in allergic children with PER and concomitant asthma with the aim of defining the UAD.

Induced sputum, exhaled breath condensate and nasal lavage fluid in electroplating workers exposed to chromium.

Occupational exposure to chromium may cause airway inflammation and bronchial asthma. In this study we investigated the effect of chromium on the respiratory tract of exposed and non-exposed electroplating workers using spirometry and analysis of induced sputum (IS), exhaled breath condensate (EBC) and nasal lavage fluid (NLF). In both groups spirometry was normal; chromium in induced sputum was higher in exposed workers (7.90 +/- 0.855 microg/L, vs 1.78 +/- 0.075 microg/L; p<0.001); no significant difference was found in induced sputum cellularity. Median nitrite concentration in EBC was significantly higher in exposed subjects (4.35 micromol/L, 5 degrees -95 degrees percentile: 1.88-10.13 vs 0.11 micromol/L, 5-95 percentile: 0-0.72) (p<0.001). IL-6 and TNF-alpha were not detectable in EBC. Median IL-6 concentration in nasal lavage fluid was higher in exposed workers (5.72 pg/ml, 5-95 percentile: 0-65.25 pg/ml vs 0.28 pg/ml, 5-95 percentile: 0-1.7 pg/ml) (p<0.01). No differences in Eosinophil Cationic Protein concentration were found. TNF-alpha was not detectable in NLF. Chromium in induced sputum correlated with nitrites in EBC. For the first time three non-invasive methods were used to assess changes in respiratory tract in workers exposed to chromium. The results suggest chromium exerts an inflammatory/irritative action on airways.

Effects of cyclo-oxygenase inhibition on exhaled eicosanoids in patients with COPD.

BACKGROUND: Leukotriene (LT) B4 concentrations are increased and prostaglandin (PG) E2 concentrations are decreased in exhaled breath condensate (EBC) in patients with chronic obstructive pulmonary disease (COPD). A study was undertaken to investigate the short term effects of cyclo-oxygenase (COX) inhibition on exhaled LTB4 and PGE2 concentrations in patients with COPD and to identify the COX isoform responsible for exhaled PGE2 production. METHODS: Two studies were performed. A double blind, crossover, randomised, placebo controlled study with ibuprofen (400 mg qid for 2 days), a non-selective COX inhibitor, was undertaken in 14 patients with stable COPD, and an open label study with oral rofecoxib (25 mg once a day for 5 days), a selective COX-2 inhibitor, was undertaken in a different group of 16 COPD patients. EBC was collected before and after drug treatment. Exhaled LTB4 and PGE2 concentrations were measured with specific immunoassays. RESULTS: All patients complied with treatment as indicated by a reduction in ex vivo serum thromboxane B2 concentrations (ibuprofen) and a reduction in lipopolysaccharide induced increase in ex vivo plasma PGE2 values (rofecoxib) of more than 80%. Exhaled LTB4 was increased after ibuprofen (median 175.5 (interquartile range 128.8-231.5) pg/ml v 84.0 (70.0-98.5) pg/ml, p < 0.001) and exhaled PGE2 was reduced (93.5 (84.0-105-5) pg/ml v 22.0 (15.0-25.5) pg/ml, p < 0.0001). Rofecoxib had no effect on exhaled LTB4 (p = 0.53) or PGE2 (p = 0.23). CONCLUSIONS: Non-selective COX inhibition decreases PGE2 and increases LTB4 in EBC, whereas selective COX-2 inhibition has no effect on these eicosanoids. PGE2 in EBC is primarily derived from COX-1 activity, and COX inhibition may redirect arachidonic acid metabolism towards the 5-lipoxygenase pathway.

Exhaled breath condensate: methodological recommendations and unresolved questions.

Collection of exhaled breath condensate (EBC) is a noninvasive method for obtaining samples from the lungs. EBC contains large number of mediators including adenosine, ammonia, hydrogen peroxide, isoprostanes, leukotrienes, nitrogen oxides, peptides and cytokines. Concentrations of these mediators are influenced by lung diseases and modulated by therapeutic interventions. Similarly EBC pH also changes in respiratory diseases. The aim of the American Thoracic Society/European Respiratory Society Task Force on EBC was to identify the important methodological issues surrounding EBC collection and assay, to provide recommendations for the measurements and to highlight areas where further research is required. Based on the currently available evidence and the consensus of the expert panel for EBC collection, the following general recommendations were put together for oral sample collection: collect during tidal breathing using a noseclip and a saliva trap; define cooling temperature and collection time (10 min is generally sufficient to obtain 1-2 mL of sample and well tolerated by patients); use inert material for condenser; do not use resistor and do not use filter between the subject and the condenser. These are only general recommendations and certain circumstances may dictate variation from them. Important areas for future research involve: ascertaining mechanisms and site of exhaled breath condensate particle formation; determination of dilution markers; improving reproducibility; employment of EBC in longitudinal studies; and determining the utility of exhaled breath condensate measures for the management of individual patients. These studies are required before recommending this technique for use in clinical practice.

Gas chromatography/mass spectrometry analysis of exhaled leukotrienes in asthmatic patients.

BACKGROUND: Leukotriene-like immunoreactivity has been detected in exhaled breath condensate (EBC), but definitive evidence for the presence of leukotrienes (LTs) in this biological fluid is not available. A study was undertaken to determine whether LTC(4), LTD(4), LTE(4), and LTB(4) are measurable in EBC by gas chromatography/mass spectrometry and to quantify exhaled LTs in adults and children with asthma and in control subjects. METHODS: Twenty eight adults and 33 children with mild to moderate persistent asthma treated with inhaled corticosteroids and age matched healthy controls (50 adults and 50 children) were studied. LTB(4), LTC(4), LTD(4), and LTE(4) in EBC were measured by gas chromatography/mass spectrometry. RESULTS: LTD(4), LTE(4), and LTB(4) were detectable in all samples. Concentrations of LTC(4) in EBC were either close to or below the detection limit of 1 pg/ml. Median exhaled LTD(4), LTE(4), and LTB(4) concentrations in asthmatic adults were increased 4.1-fold (p<0.001), 1.8-fold (p<0.01), and 2.6-fold (p<0.001), respectively, compared with values in healthy adults. Median exhaled LTD(4), LTE(4), and LTB(4) concentrations in asthmatic children were increased 2.8-fold (p<0.001), 1.3-fold (p<0.001), and 1.6-fold (p<0.001), respectively, compared with those in healthy children. In patients with asthma there was a correlation between exhaled LTD(4) and LTE(4) in both adults (r = 0.87, p<0.0001) and children (r = 0.78, p<0.0001). CONCLUSIONS: Gas chromatography/mass spectrometry can be used to accurately quantify exhaled LTs which are increased in asthmatic adults and children compared with controls.

Exhaled leukotrienes and prostaglandins in COPD.

BACKGROUND: The role of eicosanoids, including leukotrienes (LTs) and prostaglandins (PGs), in chronic obstructive pulmonary disease (COPD) is uncertain. The aim of this study was to investigate whether eicosanoids are measurable in exhaled breath condensate (EBC), a non-invasive method of collecting airway secretions, in patients with stable mild to moderate COPD, and to show possible differences in their concentrations compared with control subjects. METHODS: LTB(4), LTE(4), PGE(2), PGD(2)-methoxime, PGF(2alpha), and thromboxane B(2) (TxB(2)) were measured in EBC in 15 healthy ex-smokers, 20 steroid naïve patients with COPD who were ex-smokers, and in 25 patients with COPD who were ex-smokers and who were treated with inhaled corticosteroids. The study was of cross sectional design and all subjects were matched for age and smoking habit. RESULTS: LTB(4) and PGE(2) concentrations were increased in steroid naïve (LTB(4): median 100.6 (range 73.5-145.0) pg/ml, p<0.001; PGE(2): 98.0 (range 57.0-128.4) pg/ml, p<0.001) and steroid treated patients with COPD (LTB(4): 99.0 (range 57.9-170.5) pg/ml, p<0.001; PGE(2): 93.6 (range 52.8-157.0) pg/ml, p<0.001) compared with control subjects (LTB(4): 38.1 (range 31.2-53.6) pg/ml; PGE(2): 44.3 (range 30.2-52.1) pg/ml). Both groups of patients had similar concentrations of exhaled LTB(4) (p=0.43) and PGE(2) (p=0.59). When measurable, LTE(4) and PGD(2)-methoxime concentrations were similar in COPD patients and controls, whereas PGF(2alpha) concentrations were increased in the former. TxB(2)-LI was undetectable in any of the subjects. CONCLUSIONS: There is a selective increase in exhaled LTB(4) and PGE(2) in patients with COPD which may be relatively resistant to inhaled corticosteroid therapy.

Validation of leukotriene B4 measurements in exhaled breath condensate.

OBJECTIVE: To qualitatively validate an enzyme immunoassay to measure leukotriene B4 in exhaled breath condensate. Exhaled breath condensate is a new non-invasive method to monitor airway inflammation. SUBJECTS: Twenty-two subjects with different lung diseases attended the outpatient clinic on one occasion for exhaled breath condensate collection. METHODS: Samples were pooled together and purified by reverse-phase high-performance liquid chromatography. The fractions eluted were assayed for leukotriene B4 by enzyme immunoassay. RESULTS: A single peak of leukotriene B4-like immunoreactivity co-eluting with leukotriene B4 standard (retention time: 24 min) was identified by enzyme immunoassay. Reverse phase-high performance liquid chromatography peak of leukotriene B4 was clearly separated from those of 6-trans-leukotriene B4 (retention time: 14 min) and leukotriene B5 (retention time: 18 min) for which the antiserum used in the enzyme immunoassay had the highest cross-reactivity. Leukotriene B4 recovery was 64%. CONCLUSIONS: This study provides evidence for the presence of leukotriene B4 in the exhaled breath condensate and the specificity of the enzyme immunoassay used.

Immediate allergic reactions to beta-lactams: diagnosis and therapy.

Beta-lactams are the antibiotics which most frequently provoke adverse reactions mediated by specific immunological mechanisms. These reactions, classifiable as immediate or non-immediate, can be produced by the four classes of beta-lactams (penicillins, cephalosporins, carbapenems and monobactams) currently available, which share a common beta-lactam ring structure. Immediate reactions occur within the first hour after drug administration and are characterized by urticaria, angioedema, rhinitis, bronchospasm, and anaphylactic shock. Immediate reading skin tests are the quickest and most reliable method for demonstrating the presence of beta-lactam specific IgE antibodies. It is crucial to use in diagnosis the suspected beta-lactams themselves, particularly cephalosporins, in addition to penicillin determinants. Serum specific IgE assays can be used as complementary tests. Negative test results should be interpreted in light of the time elapsed from the last exposure to the responsible beta-lactam. In fact, both in vivo and in vitro test sensitivity is known to decrease over time. In some diagnostic work-ups, patients with a positive history and negative skin and in vitro tests with classic reagents undergo a controlled administration of the suspected beta-lactam. The management of immediate allergic reactions should take into consideration their severity and type. Adrenaline is the drug of choice in the treatment of anaphylactic shock. In addition to adrenaline, corticosteroids and antihistamines should be administered. Histamine H(1) receptor antagonists are the mainstay of the treatment of immediate allergic reactions such as urticaria, rhinitis and conjunctivitis.

Validation of 8-isoprostane and prostaglandin E(2) measurements in exhaled breath condensate.

OBJECTIVE: To qualitatively validate radioimmunoassays for 8-isoprostane and prostaglandin (PG) E(2) in exhaled breath condensate. SUBJECTS: Twenty-two subjects with different lung diseases attended the outpatient clinic on one occasion for exhaled breath condensate collection. METHODS: Samples were pooled together and purified by reverse phase high performance liquid chromatography (RP-HPLC). The eluted fractions were assayed for 8-isoprostane-like immunoreactivity and PGE(2)-like immunoreactivity by radioimmunoassays. In addition, simultaneous measurements of exhaled breath condensate unextracted samples with two anti-8-isoprostane and anti-PGE(2) sera with different cross-reactivity were performed. RESULTS: A single peak of 8-isoprostane-like immunoreactivity and PGE(2)-like immunoreactivity co-eluting with 8-isoprostane (retention time: 13 min) and PGE(2) (retention time: 21 min) standards, respectively, was identified by radioimmunoassays. Testing with two different antisera showed similar results for both 8-isoprostane-like immunoreactivity (limits of agreement = 4.5 pg/ml and - 4.1 pg/ml, n = 12) and PGE(2)-like immunoreactivity (limits of agreement = 6.1 pg/ ml and - 6.1 pg/ml, n = 12). CONCLUSION: This study provides evidence for the specificity of the radioimmunoassays for 8-isoprostane and PGE(2) in exhaled breath condensate. This is critical for proposing these markers as a non-invasive way for monitoring airway inflammation.

Dose-dependent onset and cessation of action of inhaled budesonide on exhaled nitric oxide and symptoms in mild asthma.

BACKGROUND: Dose dependent anti-inflammatory effects of inhaled corticosteroids in asthma are difficult to demonstrate in clinical practice. The anti-inflammatory effect of low dose inhaled budesonide on non-invasive exhaled markers of inflammation and oxidative stress were assessed in patients with mild asthma. METHODS: 28 patients entered a double blind, placebo controlled, parallel group study and were randomly given either 100 or 400 micro g budesonide or placebo once daily, inhaled from a dry powder inhaler (Turbohaler), for 3 weeks followed by 1 week without treatment. Exhaled nitric oxide (NO), exhaled carbon monoxide (CO), nitrite/nitrate, S-nitrosothiols, and 8-isoprostanes in exhaled breath condensate were measured four times during weeks 1 and 4, and once a week during weeks 2 and 3. RESULTS: A dose-dependent speed of onset and cessation of action of budesonide was seen on exhaled NO and asthma symptoms. Treatment with 400 micro g/day reduced exhaled NO faster (-2.06 (0.37) ppb/day) than 100 micro g/day (-0.51 (0.35) ppb/day; p<0.01). The mean difference between the effect of 100 and 400 micro g budesonide was -1.55 ppb/day (95% CI -2.50 to -0.60). Pretreatment NO levels were positively related to the subsequent speed of reduction during the first 3-5 days of treatment. Faster recovery of exhaled NO was seen after stopping treatment with budesonide 400 micro g/day (1.89 (1.43) ppb/day) than 100 micro g/day (0.49 (0.34) ppb/day, p<0.01). The mean difference between the effect of 100 and 400 micro g budesonide was 1.40 ppb/day (95% CI -0.49 to 2.31). Symptom improvement was dose-dependent, although symptoms returned faster in patients treated with 400 micro g/day. A significant reduction in exhaled nitrite/nitrate and S-nitrosothiols after budesonide treatment was not dose-dependent. There were no significant changes in exhaled CO or 8-isoprostanes in breath condensate. CONCLUSION: Measurement of exhaled NO levels can indicate a dose-dependent onset and cessation of anti-inflammatory action of inhaled corticosteroids in patients with mild asthma.

Exhaled carbon monoxide and nitric oxide in COPD.

STUDY OBJECTIVES: To investigate whether exhaled carbon monoxide (CO) and nitric oxide (NO) could be used as noninvasive in vivo biomarkers of oxidative stress in the lungs of patients with COPD. DESIGN: Single-center cross-sectional study. PATIENTS: Ten healthy nonsmokers, 12 smokers, 15 stable ex-smokers with COPD, and 15 stable current smokers with COPD. INTERVENTIONS: Subjects attended the outpatient clinic on one occasion for pulmonary function tests and exhaled CO and NO measurements. MEASUREMENTS AND RESULTS: Mean (+/- SEM) CO levels in ex-smokers with COPD were higher (7.4 +/- 1.9 ppm; p < 0.05) than in nonsmoking control subjects (3.0 +/- 0.3 ppm) but were lower than in current smokers with COPD (20.0 +/- 2.6 ppm; p < 0.001). There was no correlation between exhaled CO and NO. There was no correlation between CO and lung function tests in any group of patients. Exhaled NO was higher in ex-smokers with COPD (12.0 +/- 1.0 parts per billion [ppb]; p < 0.001) than in healthy nonsmokers (6.5 +/- 0.6 ppb) and in current smokers with COPD (7.6 +/- 1.1 ppb; p < 0.01) compared to healthy smokers (3.3 +/- 0.4 ppb). Ex-smokers with COPD had higher exhaled NO levels than did current smokers with COPD (p < 0.001) There was a negative correlation between exhaled NO and FEV(1) in both ex-smokers with COPD (r = -0.60; p < 0.02) and current smokers with COPD (r = -0.59; p < 0.02). CONCLUSION: The measurement of exhaled CO and NO may represent a new method for the noninvasive monitoring of airway inflammation and oxidant stress in COPD ex-smokers. Exhaled CO and NO are strongly affected by cigarette smoking, which limits their usefulness as biomarkers in current smokers.

Increased nitrosothiols in exhaled breath condensate in inflammatory airway diseases.

Nitrosothiols (RS-NOs) are formed by interaction of nitric oxide (NO) with glutathione and may limit the detrimental effect of NO. Because NO generation is increased in airway inflammation, we have measured RS-NOs in exhaled breath condensate in patients with asthma, cystic fibrosis, or chronic obstructive pulmonary disease (COPD). We also measured exhaled NO and nitrite (NO(2-)) in the same subjects. RS-NOs were detectable in exhaled breath condensate of all subjects. RS-NOs were higher in subjects with severe asthma (0.81 +/- 0.06 microM) when compared with normal control subjects (0.11 +/- 0.02 microM, p < 0.01) and with subjects with mild asthma (0.08 +/- 0.01 microM, p < 0.01). Elevated RS-NOs values were also found in patients with cystic fibrosis (0.35 +/- 0.07 microM, p < 0.01), in those with COPD (0.24 +/- 0.04 microM, p < 0.01) and in smokers (0.46 +/- 0.09 microM, p < 0.01). In current smokers there was a correlation (r = 0.8, p < 0.05) between RS-NOs values and smoking history (pack/year). We also found elevated concentrations of NO(2-) in patients with severe asthma, cystic fibrosis, or COPD, but not in smokers or patients with mild asthma. This suggests that exhaled NO(2-) is less sensitive than exhaled RS-NOs. This study has shown that RS-NOs are detectable in exhaled breath condensate of healthy subjects and are increased in patients with inflammatory airway diseases. As RS-NOs concentrations in exhaled breath condensate vary in the different airway diseases and increase with the severity of asthma, their measurement may have clinical relevance as a noninvasive biomarker of nitrosative stress.