Development of a functional point-of-need diagnostic for myeloperoxidase detection to identify neutrophilic bronchitis.

With over 4.8 million Canadians suffering from chronic airway diseases, respiratory exacerbations are currently the leading cause of hospitalization in Canada. In cases of bacterial infection, neutrophil cell density increases from ∼10 million cells per gram to over 15 million cells per gram. As sputum is a direct discharge from the primarily affected areas of respiratory diseases, quantification of granulocytes (including neutrophils) can be used to effectively determine a course of patient treatment. Unfortunately this quantification is currently limited to labour-intensive and time-consuming cell counts. In the present study, we describe a simple one-step lateral flow test (LFT) that can semi-quantitatively determine myeloperoxidase (MPO), a biomarker found in neutrophils, in minimally-processed sputum samples. This point-of-need (PON) diagnostic device provides positive results observable to the naked eye after 15 minutes. 37 human sputum samples were quantified for MPO using the developed LFT and compared to neutrophil levels quantified through traditional cell counting. A trend between sputum MPO concentration and total neutrophils was observed, suggesting that the LFT has the potential to replace cell counting for neutrophil approximation to aid in directing therapies quickly at the point of need.

Series "matrix metalloproteinases in lung health and disease": Matrix metalloproteinases in COPD.

There is considerable evidence that matrix metalloproteinases (MMPs) are up- and/or downregulated in chronic obstructive pulmonary disease (COPD), particularly in emphysema, in which they probably participate in proteolytic attack on the alveolar wall matrix. Recent data suggest that MMPs also have major roles in driving inflammation or shutting it down, as well as modifying the release of fibrogenic growth factors, processes that are important in the genesis of the various lesions of COPD. In cigarette smoke-induced animal models of emphysema, MMP-12 appears to play a consistent and important role, whereas the data for other MMPs are difficult to interpret. In human lungs, evidence for a role for MMPs is more tenuous and there are numerous contradictions in the literature. Little is known about the effects of MMPs in small airway remodelling, smoke-induced pulmonary hypertension and chronic bronchitis, but MMP-12 participates in experimental small airway modelling. To date, the accumulated data suggest that selective inhibition of MMP-12 might be a viable therapy for emphysema and small airway remodelling, but subtle differences in the functions of MMP-12 in animals and humans mandate caution with this approach. Whether inhibition of other MMPs might be useful is unclear.

Role of neprilysin in airway inflammation induced by diesel exhaust emissions.

In this study, we examined the role of neprilysin (NEP), a key membrane-bound endopeptidase, in the inflammatory response induced by diesel exhaust emissions (DEE) in the airways through a number of approaches: in vitro, animal, and controlled human exposure. Our specific aims were (1) to examine the role of NEP in inflammatory injury induced by diesel exhaust particles (DEP) using Nep-intact (wild-type) and Nep-null mice; (2) to examine which components of DEP are associated with NEP downregulation in vitro; (3) to determine the molecular impact of DEP exposure and decreased NEP expression on airway epithelial cells' gene expression in vitro, using a combination of RNA interference (RNAi) and microarray approaches; and (4) to evaluate the effects on NEP activity of human exposure to DEE. We report four main results: First, we found that exposure of normal mice to DEP consisting of standard reference material (SRM) 2975 via intratracheal installation can downregulate NEP expression in a concentration-dependent manner. The changes were accompanied by increases in the number of macrophages and epithelial cells, as well as proinflammatory cytokines, examined in bronchoalveolar lavage (BAL) fluid and cells. Nep-null mice displayed increased and/or additional inflammatory responses when compared with wild-type mice, especially in response to exposure to the higher dose of DEP that we used. These in vivo findings suggest that loss of NEP in mice could cause increased susceptibility to injury or exacerbate inflammatory responses after DEP exposure via release of specific cytokines from the lungs. Second, we found evidence, using in vitro studies, that downregulation of NEP by DEP in cultured human epithelial BEAS-2B cells was mostly attributable to DEP-adsorbed organic compounds, whereas the carbonaceous core and transition metal components of DEP had little or no effect on NEP messenger RNA (mRNA) expression. This NEP downregulation was not a specific response to DEP or its contents because the change also occurred after exposure to urban dust (SRM 1649a), which differs in physical and chemical composition from DEP. Third, we also collected the transcriptome profiles of the concentration-effects of SRM 2975 in cultured BEAS-2B cells through a 2 X 3 factorial design. DEP exposure upregulated 151 genes and downregulated 59 genes. Cells with decreased NEP expression (accomplished by transfecting an NEP-specific small interfering RNA [siRNA]) substantially altered the expression of genes (upregulating 17 and downregulating 14) associated with DNA/protein binding, calcium channel activities, and the cascade of intracellular signaling by cytokines. Data generated from the combined RNAi and microarray approaches revealed that there is a complex molecular cascade mediated by NEP in different subcellular compartments, possibly influencing the inflammatory response. Fourth, in a controlled human exposure study, we observed significant increases in soluble NEP in sputum after acute exposure to DEE, with an average net increase of 31%. We speculate that the change in NEP activity in sputum, if confirmed in larger epidemiologic investigations at ambient exposure levels to DEE, may provide a useful endpoint and promote insight into the mechanism of DEE-induced airway alterations.

[Evaluation of matrix metalloproteinases (pro-MMP-1, MMP-2,8) and their inhibitor (TIMP-1) contents in patients with occupational lung diseases].

Studies of matrix metalloproteinases in patients with occupational bronchopulmonary diseases and in individuals exposed to asbestos dust revealed hyperactivated protease system--lower level of MMP-1 proenzyme and increased production of TIMP-1 (metalloproteinases inhibitor)--in all the examinees groups. Patients with pneumoconiosis and occupational dust bronchitis demonstrated increased neutrophilic elastase that is activator of metalloproteinases inducing sclerotic changes and pulmonary fibrosis.

Characterisation of the acute and reversible airway inflammation induced by cadmium chloride inhalation in healthy dogs and evaluation of the effects of salbutamol and prednisolone.

The aims of this study were firstly to characterise a model of subclinical and reversible bronchial inflammation induced by cadmium chloride inhalation in healthy dogs and then to examine the effect of prednisolone or salbutamol treatment on the resulting bronchitis. The model characterisation and the effects of treatment were studied using clinical symptoms, haematology, thoracic radiography, bronchoscopy and bronchoalveolar lavage, barometric whole-body plethysmography and histamine broncho-provocation tests. In addition, the activity of matrix metalloproteinases (MMP)-2 and -9 were determined in serum and bronchoalveolar lavage fluid (BALF). Cadmium inhalation induced: (1) a transient bronchial inflammation, dominated by neutrophils; (2) a neutrophilia of the blood that persisted for up to 4 weeks; (3) a transient increased bronchial reactivity, and (4) a significant increase in MMP-9 activity in the BALF. Prednisolone treatment reduced the influx of inflammatory cells into the BALF, but not significantly, had no effect on pulmonary function, and did not reduce of airway hypersensitivity. Salbutamol had almost no effect on any of the parameters investigated.

Granzyme K: a novel mediator in acute airway inflammation.

BACKGROUND: Granzymes are a subfamily of serine proteases involved in the pathogenesis of many inflammatory disorders. In contrast with granzyme A and B, the role of granzyme K (GrK) in human lung diseases is unknown. Therefore, the release and expression of GrK in allergic asthma, chronic obstructive pulmonary disease (COPD) and bronchopneumonia were investigated. METHODS: Soluble GrK was quantified using an enzyme linked immunosorbent assay in the bronchoalveolar lavage fluid of patients with allergic asthma (before and after segmental allergen challenge), and in patients with mild COPD, pneumonia and in healthy controls. The molecular form of GrK was analysed by western blot. Flow cytometry was performed to determine the cellular expression of GrK. RESULTS: Compared with healthy controls, there were normal levels of soluble GrK in the bronchoalveolar lavage fluid of patients with COPD, and patients with allergic asthma before allergen challenge. In contrast, soluble GrK was strongly increased in the bronchoalveolar lavage fluid of patients with acute bronchopneumonia. In patients with allergic asthma, there was a significant increase in soluble GrK as well as in GrK expressing CD8(+) T cells in the bronchoalveolar lavage fluid 24 h and 72 h after allergen challenge. After allergen challenge, soluble GrK correlated with the percentage of GrK expressing CD8(+) T cells. Finally, it was shown that the endobronchial release of the CCR5 ligand CCL3 might be a mechanism for the recruitment of GrK(+)CD8(+) T cells after allergen challenge. CONCLUSION: These data provide the first evidence that expression of GrK is upregulated in acute airway inflammation, both in infectious and non-infectious diseases.

Active monomers of human beta-tryptase have expanded substrate specificities.

beta-Tryptase, a product of the TPSAB1 and TPSB2 genes, is a trypsin-like serine protease that is a major and selective component of the secretory granules of all human mast cells, accounting for as much as 25% of cell protein. Once mast cells are activated, beta-tryptase is released along with histamine and heparin proteoglycan. beta-Tryptase is a unique enzyme with a homotetrameric structure in which active sites face into the central cavity of the four monomers, stabilized by heparin-proteoglycan. This structure makes beta-tryptase resistant to most biological inhibitors of serine proteases. Without stabilization, at neutral pH beta-tryptase converts to inactive monomers. Tryptase levels are elevated in bronchoalveolar lavage (BAL) fluid obtained from atopic asthmatics and in serum during systemic anaphylactic shock. Several synthetic small molecular weight beta-tryptase inhibitors reduced Ag-induced airway hypersensitivity in animals, suggesting that beta-tryptase is involved in the pathogenesis of airway inflammation. Although the major biologic substrate(s) of beta-tryptase remain ambiguous, the protease can digest several proteins of potential biologic importance, including fibrinogen, fibronectin, pro-urokinase, pro-matrix metalloprotease-3 (proMMP-3), protease activated receptor-2 (PAR2) and complement component C3. Recently, monomers of beta-tryptase with enzymatic activity have been detected in vitro. Here we discuss how beta-tryptase monomers with enzymatic activity were identified as well as their potential role in vivo.

CF-Emerging therapies: Modulation inflammation.

Persistent and dysregulated inflammation, combined with an exaggerated host response is a major contributor to CF lung disease. As lung disease progresses, neutrophil accumulation in the airways ensues. Modulation of CF airway inflammation may result in either beneficial or deleterious side effects, resulting in more harm than good. Antibiotics, in particular, macrolides which act as a long-term anti-inflammatory agent with an excellent safety profile, and dornase alpha, are very interesting agents; steroids are not indicated in CF except in very special situations, and other promising agents such as leukotriene modifiers, high-dose N-acetylcysteine, anti-elastase and anti-cytokines require further research. Research should focus on early treatment, before lung damage has occurred.

[Analysis of tryptase levels in infants and children with wheezy bronchitis]. / Analiza stezenia tryptazy u niemowlat i malych dzieci chorych na obturacyjne zapalenie oskrzeli.

UNLABELLED: Wheezy bronchitis is one of hospitalization causes of infants and young children. Significant problem is recurrent wheezing. Wheezy bronchitis can be the first sign of bronchial asthma. New markers are taken into consideration in aspect of recurrent bronchitis prevention. Such marker seems to be tryptase. Tryptase is released from mastocytes in early phase of allergic reaction. The aim of the study was to assess tryptase and ECP concentrations among infants and young children with wheezy bronchitis. MATERIAL AND METHODS: Ninety-four patients with wheezy bronchitis in age from between 1 month to 36 months old were included into the study (47 with the first episode and 47 with at least third episode). Forty-three patients hospitalized due to other causes, from the same age group (these patients haven't already had any wheezy bronchitis) were included into the control group. Among all patients concentrations of tryptase and ECP were evaluated (among patients from the study group in the acute phase of disease) by fluoroimmunoenzymatic method (FEIA) with the the use of UniCAP 100 set (Pharmacia & Upjohn Diagnostics AB). RESULTS: No statistically significant differences of tryptase concentrations in blood plasma during wheezy bronchitis among infants and young children in correlation to the control group were found. Statistically significant differences were identified in aspect of tryptase concentrations during wheezy bronchitis between patients with the first episode of wheezing and patients with recurrent wheezing and higher concentrations were observed among patients with the first episode. No statistically significant influence of family history of allergy and symptoms of allergy were identified in aspect of tryptase concentrations. CONCLUSIONS: Obtained results lead to conclusion that tryptase concentrations have a little significance in wheezy bronchitis.

Inhibition of p38 mitogen activated protein kinase controls airway inflammation in cystic fibrosis.

BACKGROUND: Cystic fibrosis (CF) airways are characterised by chronic inflammation, increased interleukin (IL)-8 secretion, and neutrophil activation which are considered the principal factors of morbidity and mortality in CF patients. Optimising management of this chronic inflammatory response is therefore a key issue of basic and clinical CF research. Several reports have addressed ways to manage CF airways inflammation, and an attractive therapeutic strategy may be the inhibition of the p38-mitogen activated protein kinase (p38-MAP-k) pathway. METHODS: A new ex vivo model was used to study the mucosal inflammatory response to environmental airways stimuli. Nasal biopsy tissues from CF patients and controls were cultured ex vivo for 20 minutes, 4 hours, and 24 hours in the presence of lipopolysaccharide (LPS) from Pseudomonas aeruginosa (PA) with and without the p38-MAP-k inhibitor SB203580. Quantitative mRNA assessment, immunohistochemistry, and Western blots were used to detect the expression and modulation of inflammatory markers. RESULTS: PA-LPS challenge induced a time dependent mucosal inflammation indicated by rapid epithelial activation, IL-8 release, COX-2 upregulation, and neutrophil migration to the upper mucosal layers. Some of these LPS induced changes (IL-8 release and neutrophil migration) were specific to CF tissues. SB203580 significantly controlled all LPS induced mucosal changes in CF tissues. CONCLUSION: These findings provide a rationale and proof of principle for the potential use of p38-MAP-k inhibitors to control inflammation in patients with CF.

Increased expression of p38 MAPK in human bronchial epithelium after lipopolysaccharide exposure.

Bacterial endotoxin (lipopolysaccharides (LPS)) is normally present in the wall of Gram-negative bacteria and has potent pro-inflammatory properties. Exposure to LPS has been shown to induce neutrophilic airway inflammation in humans. The aim of this investigation was to study the early inflammatory responses to LPS exposure in human airway mucosa in vivo. In total, 15 healthy nonsmoking volunteers participated. Bronchoscopy was performed on two separate occasions, 3 h after saline inhalation and after inhalation of 50 mug LPS in saline. Endobronchial mucosal biopsy specimens were taken and stained immunohistochemically using a panel of monoclonal antibodies directed against mitogen-activated protein kinases (MAPKs), transcription factors, cytokines, adhesion molecules and inflammatory cells. Expression of p38 MAPK increased as a consequence of LPS exposure, as determined by both total epithelial staining and nuclear location. These two responses were strongly associated. Epithelial expression of interleukin-8 showed a tendency towards a significant increase after LPS compared to saline. Epithelial mast cell numbers were increased after LPS, whereas neutrophil numbers were unchanged. Inhalation of lipopolysaccharide induced activation of the bronchial epithelium, as demonstrated 3 h after exposure by increased expression of p38 mitogen-activated protein kinase and interleukin-8, and may represent early regulatory steps in the subsequent development of a neutrophilic bronchial inflammation.

Specific enzyme activities in bronchoalveolar lavage fluid as an aid to diagnosis of tracheobronchitis and bronchopneumonia in dogs.

Lactate dehydrogenase (LDH), alkaline phosphatase (ALP), alanine aminotransferase and gamma-glutamyl transferase enzyme activities, and total protein (TP), calcium, inorganic phosphate, urea nitrogen (UN) and creatinine concentrations in bronchoalveolar lavage fluid were investigated for their relative importance in the diagnosis of respiratory diseases in dogs. Bronchoalveolar lavage (BAL) fluid was obtained from 26 dogs (20 with respiratory diseases and six controls) following anaesthesia with sodium pentothal. Enzyme activities and biochemical parameters were measured in BAL fluid. LDH and ALP levels were significantly increased in 12 dogs with bronchopneumonia, but not in eight dogs with tracheobronchitis. Insignificant and variable levels of TP and UN concentrations were found in both groups. It was concluded that LDH and ALP enzyme activities could be considered as pointers to pulmonary inflammation and/or damage while TP and UN measurements in BAL fluid may have a place in the identification of changes in respiratory and vascular permeability.

Involvement of haemoxygenase-1 in ozone-induced airway inflammation and hyperresponsiveness.

Haemoxygenase catalyses the degradation of haem to bilirubin, and the inducible form of haemoxygenase, haemoxygenase-1, is highly induced in response to oxidative stress in vitro. The effect of haemoxygenase-1 in oxidant stress in vivo is not known. We determined the effect of exposure to ozone on haemoxygenase-1 expression, and the modulation of haemoxygenase-1 expression on ozone-induced lung neutrophilia and bronchial hyperresponsiveness in rats. Ozone caused a significant induction of lung haemoxygenase-1. Pretreatment of rats with haemoglobin, a potent inducer of haemoxygenase-1, resulted in a large induction of haemoxygenase-1 expression, and inhibited ozone-induced neutrophilia and bronchial hyperresponsiveness. Tin protoporphyrin, a competitive inhibitor of haemoxygenase, reduced the expression of haemoxygenase-1 induced by haemoglobin. It enhanced ozone-induced neutrophilia, but not the bronchial hyperresponsiveness, and reduced the protective effect of haemoglobin. Overall, there was an association between bronchial hyperresponsiveness and the neutrophilic response. These data indicate that haemoxygenase-1 plays an important role in modulating the effects of an oxidant, such as ozone in the lungs.

[Effect of the preparation Wobe-Mugos E on parameters of the antioxidant defence system and morphofunctional erythrocyte status in patients with chronic obstructive bronchitis]. / Vplyv preparatu "Wobe-Mugos E" na pokaznyky antyoksydantnoï systemy zakhystu i morfofunktsional'nogo stanu erytrotsytiv u khvorykh na khronichnyi obstruktyvnyi bronkhit.

In the examination of 64 patients with chronic obstructive bronchitis, some specificities of functioning of decompensation mechanisms in the glutathione antiradical system were established as were changes in morphofunctional and receptor properties of erythrocytes in chronic obstructive bronchitis. The patients were shown to have derived benefit from a combined treatment involving the use of the enzymic drug preparation Wobe-Mugos E which was found to make for improvement of rheological properties of erythrocytes, their capability of depositing and transporting catecholamines.

[On the pathogenesis of chronic dust-related bronchitis (according to the status of immune and thrombin-plasmin systems]. / K patogenezu khronicheskogo pylevogo bronkhita (po sostoianiiu immunoi i trombin-plazminovoi sistem).

As a result of the performed investigations there was revealed the injury of T-link of the systemic immunity, as well, as the reduction of the immunoglobulin A. And in the sick with chronic dust bronchitis the state of local immunity was characterized by the increase of the per cent content of neutrophilic granulocytes in the bronchoalveolar washout, and also by the reduction of adgesive and absorptive ability of the alveolar macrophages at the background of significant activation of the oxygendependent metabolism. The investigation of coagulant blood system revealed the predomination of thrombinogenesis over the processes of fibrinolysis.

Bronchial cartilage atrophy in chronic bronchitis: observations on chondrolytic processes.

The status of bronchial cartilage degeneration in chronic bronchitis is unclear, and little is known about the chondrolytic mechanisms involved. The potential contributions of various inflammatory cells, chondrocytes and cartilage-degrading enzymes to cartilage atrophy have been examined. Bronchial cartilage specimens were obtained at autopsy from lobar secondary bronchi from chronic bronchitics and age-matched controls; each was examined by light microscopy and immunohistology for the distributions of mast cells, macrophages, eosinophils, collagenase 1, collagenase 3, and degradation products of cartilage collagen. Most bronchitic specimens showed hypertrophic chondrocytes, some of which were immunostained for collagenase 3, and occasionally for collagenase 1. Evidence for collagen degradation products was demonstrated around the lacunae of a proportion of chondrocytes, and both collagenases were also observed in the soft inflammatory tissues in close association with the cartilage surface, together with variable distributions of mast cells and macrophages. Such observations were generally absent or very much reduced in the control, non-bronchitic specimens. Degenerative changes, atrophy and loss of bronchial cartilage were common features of most chronic bronchitic specimens, this usually being related to intrinsic changes in the chondrocyte phenotype, including proliferative and matrix-degrading properties. Mast cells and macrophages were often observed in close association with the bronchial cartilage, suggesting that inflammatory cells may also contribute to the mechanisms of bronchial cartilage degradation and loss. These observations of bronchial cartilage degeneration were generally lacking in age-matched non-bronchitic control specimens.

Sputum metalloproteinase-9/tissue inhibitor of metalloproteinase-1 ratio correlates with airflow obstruction in asthma and chronic bronchitis.

Asthma and chronic bronchitis are inflammatory diseases with extracellular matrix (ECM) remodeling and collagen deposition. Collagen homeostasis is controlled by metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). We evaluated MMP and TIMP balance in induced sputum of 10 control, 31 untreated asthmatic, and 16 chronic bronchitic subjects. We first performed zymographic analysis to identify the profile of MMPs. Zymography revealed a similar MMPs profile in all populations studied and that MMP-9 was the major enzyme released. We then measured, using enzyme immunoassay, the concentrations of MMP-9 and of its inhibitor TIMP-1 and evaluated whether airflow limitation may be associated with an imbalance between these enzymes. MMP-9 and TIMP-1 concentrations were greater in sputum of patients with asthma and chronic bronchitis than in control subjects. The molar ratio between MMP-9 and TIMP-1 was lower in asthmatics and chronic bronchitics than in control subjects, and positively correlated with FEV1 values. In asthma, MMP-9 levels were significantly correlated with the number of macrophages and neutrophils. This study shows that airway inflammation in asthma and chronic bronchitis is associated with an imbalance between MMP-9 and TIMP-1 which may have a role in the pathogenesis of ECM remodeling and airflow obstruction.

Bronchial subepithelial fibrosis and expression of matrix metalloproteinase-9 in asthmatic airway inflammation.

BACKGROUND: Bronchial asthma is characterized by airway structural changes, including mucosal inflammation and subepithelial collagen deposition. An imbalance between the matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) is thought to play a critical role in the synthesis or degradation of the extracellular matrix of the airway architecture. However, the relationship between subepithelial basement membrane thickness and these enzymes in asthma has not been determined. OBJECTIVE: We compared the thickness of collagen and tenascin deposition and the expression of MMP-9 and TIMP-1 in bronchial biopsy specimens from subjects with asthma and control subjects. METHODS: Bronchial biopsy specimens were obtained from 25 subjects with asthma and 10 healthy control subjects to estimate the extent of collagen and tenascin deposition in subepithelial reticular basement membrane by immunohistochemical staining. Using a computer-assisted image analysis system, we quantitated expression of both epithelial and submucosal MMP-9 and TIMP-1. The numbers of inflammatory cells were also determined. RESULTS: Subjects with asthma exhibited greater thickness of collagen III (P <.01), collagen V (P <.01), and tenascin (P <.01) deposition in reticular basement membrane than did control subjects. The proportions of epithelium and submucosa immunoreactive to MMP-9 and TIMP-1 were significantly higher in the subjects with asthma than in the control subjects (each P <.001). Submucosal expression of MMP-9 was significantly higher than that of TIMP-1 in subjects with asthma (P <.01). Significant correlations were found between the number of myofibroblasts and thicknesses of collagen III (rs = 0. 70, P <.001), collagen V (rs = 0.67, P <.001), and tenascin (rs = 0. 58, P <.01) in subjects with asthma. On the other hand, the number of eosinophils was correlated with degree of mucosal expression of MMP-9 (rs = 0.43, P <.05) and TIMP-1 (rs = 0.69, P <.001). In subjects with asthma, a significant inverse correlation was found between subepithelial fibrosis and FEV1 (type III collagen, rs = -0. 89, P <.001; type V collagen, rs = -0.90, P <.001; tenascin, rs = -0. 88, P <.001), and airway responsiveness (type III collagen, rs = -0. 59, P <.01; type V collagen, rs = -0.47, P <.05; tenascin, rs = -0. 48, P <.05). CONCLUSION: These findings suggest that collagen III, collagen V, and tenascin deposition in basement membrane in subjects with bronchial asthma are associated with increased expression of MMP-9, which may be produced by eosinophils, and that airway remodeling in subjects with asthma may be related to air-flow obstruction and airway hyperresponsiveness.

[The NADPH-diaphorase activity of the bronchial epithelium in chronic lung diseases]. / Aktivnost' NADPH-diaforazy épiteliia bronkhov pri khronicheskikh zabolevaniiakh legkikh.

Topochemistry and activity of NADP-H diaphorase co-localized with NO synthase was examined in operative material of lungs from patients with bronchial asthma (BA), chronic nonobstructive bronchitis (CNO) and chronic obstructive bronchitis. The enzyme activity was found to be dependent upon the types of obstruction and inflammation. In CNO the state of NO synthase was not changed. In conditions of progressive irreversible airway obstruction the enzyme activity was augmented in small bronchi epithelium and alveolar macrophages (AM). In reversible obstruction the activity of NO synthase was not changed in the epithelium but appeared high in resident cells of inflammation--AM and mast cells.