Involvement of superoxide and nitric oxide on airway inflammation and hyperresponsiveness induced by diesel exhaust particles in mice.

We previously demonstrated that chronic intratracheal instillation of diesel exhaust particles (DEP) induces airway inflammation and hyperresponsiveness in the mouse, and that these effects were partially reversed by the administration of superoxide dismutase (SOD). In the present study, we have investigated the involvement of superoxide in DEP-induced airway response by analyzing the localization and activity of two enzymes: (1) a superoxide producer, NADPH cytochrome P-450 reductase (P-450 reductase), and (2) a superoxide scavenger, SOD, in the lungs of the exposed mice and controls. P-450 reductase was detected mainly in ciliated cells and clara cells: its activity was increased by the repeated intratracheal instillation of DEP. While CuZn-SOD and Mn-SOD were also present in the airway epithelium, their activity was significantly decreased following DEP instillation. Exposure to DEP doubled the level of nitric oxide (NO) in the exhaled air. DEP exposure also increased the level of constitutive NO synthase (cNOS) in the airway epithelium and inducible NO synthase (iNOS) in the macrophages. Pretreatment with N-G-monomethyl L-arginine, a nonspecific inhibitor of NO synthase, significantly reduced the airway hyperresponsiveness induced by DEP. These results indicate that superoxide and NO may each contribute to the airway inflammation and hyperresponsiveness induced by the repeated intratracheal instillation of DEP in mice.

Dissociation of cell death from covalent binding of paracetamol by flavones in a hepatocyte system.

Paracetamol metabolism and toxicity were studied in isolated rat hepatocytes. Cell damage, due to paracetamol, was shown to be dose dependent and was worse in cells from animals pre-treated with phenobarbitone. Exposure to 10 mM paracetamol for 1 hr caused a loss of intracellular reduced glutathione (GSH) and a later progressive leakage of isocitrate dehydrogenase (ICD). Treatment with (+)catechin, 3-O-methyl(+)catechin and promethazine reduced or prevented the paracetamol-induced ICD leakage. Similarly, studies on covalent binding of paracetamol showed that 3-O-methyl(+)catechin, which "protected" the cells, did so without affecting the amount of material bound covalently to cellular protein. Incubation in tissue culture for 24 hr, after prior treatment with paracetamol +/- the protective agent, showed that the protected cells remained viable and attached to tissue culture plates much better than did the "unprotected" cells. These results suggest that the protective effect is much more than just a temporarily delayed cell death. GSH loss and covalent binding of paracetamol metabolites to cell protein are not sufficient causes of cell death, although they may act as starting points in the chain of events leading to cell death.

New insights into the understanding of asthma.

The prevalence of asthma is increasing, despite better understanding of its pathogenesis and improved treatments. During the past 10 years, the perception of asthma has shifted from a disease primarily characterized by altered smooth muscle function to one mainly characterized by chronic inflammation. This article reviews the evidence supporting the relationship of inflammation in both the upper and lower airways, focusing on intermittent seasonal disease as well as on the more chronic and severe forms of asthma, including that associated with aspirin intolerance. It also presents evidence to support a pivotal role for the epithelial cell, together with the mast cell and the eosinophil, in initiating and maintaining inflammation in the upper and lower airways.

Histamine synthesis by respiratory tract micro-organisms: possible role in pathogenicity.

Five bacterial species considered to be potential pathogens in acute exacerbations of chronic bronchitis, cystic fibrosis, and pneumonia--Branhamella catarrhalis, Haemophilus parainfluenzae, Pseudomonas aeruginosa, Staphylococcus aureus and Streptococcus pneumoniae--were evaluated for their potential to synthesise histamine in vitro. Bacterial species commonly isolated from infected sputum but generally not considered to be pathogenic--Enterobacteriacae, Neisseria pharyngis, coagulase negative staphylococci, alpha-haemolytic streptococci, and Candida albicans--were similarly studied. Of the "pathogens", the Gram negative species B catarrhalis, H parainfluenzae and Ps aeruginosa synthesised clinically important amounts of histamine; this was not the case for the Gram positive species S aureus and S pneumoniae. Of the "non-pathogenic" species, only the Enterobacteriacae, as a group, were found to synthesise clinically important amounts of histamine. These results show that some Gram negative bacteria, associated with acute exacerbations in respiratory infections, produce histamine and possibly other inflammatory mediators, which may contribute to their pathogenecity in the lower respiratory tract in vivo.

Histidine decarboxylases from bacteria that colonise the human respiratory tract.

We investigated whether production of histamine by bacteria isolated from sputum of patients with infective lung diseases could be attributed to the presence of histidine decarboxylase (HD). Twenty gram-positive and 20 gram-negative organisms were studied for their ability to decarboxylate 14C-histidine in vitro over the pH range 4.5-7.5. Of the bacteria investigated, lysates from the gram-negative species Haemophilus influenzae, H. parainfluenzae, Moraxella (Branhamella) catarrhalis and Pseudomonas aeruginosa liberated 14CO2 and histamine from 14C-histidine in the presence of the cofactor pyridoxal phosphate. In contrast, results obtained in the absence of cofactor were similar to those of negative (lysate-free) controls suggesting that the HD enzymes of these species resembled those previously described in other gram-negative bacteria. No HD activity was detected over this pH range in lysates from gram-positive species. This finding correlated with earlier observations that these gram-positive organisms did not produce histamine in vitro.

Culture and comparison of human bronchial and nasal epithelial cells in vitro.

Human nasal and bronchial epithelial cells were cultured in vitro and compared morphologically and functionally. Morphologic assessment by both light and electron microscope and indirect immunoperoxidase staining techniques confirmed the identity of the two cell types as being epithelial. Light microscopy of confluent cultures revealed tightly packed cell monolayers, whilst electron microscopy showed that cells were linked by tight junctions. Estimation of cell size by planimetry found these cells to have a mean width of 10.6 +/- 1.1 microns for nasal cells and a mean width of 10.2 +/- 1.0 microns for bronchial cells. A high proportion of both the nasal and the bronchial cells exhibited features of the mature ciliated cell types, and constituted between 50 and 76% of the total cells at the earlier stages of culture although this decreased to between 16 and 23% of the total by 4 weeks in culture. The ciliary beat frequencies of the nasal and bronchial cells were found to be similar at 10.8 +/- 0.7 Hz and 11.8 +/- 2.3 Hz, respectively. The cilial beat on adjacent cells was synchronous, suggesting the presence of intercellular communication between the neighbouring cells. These studies demonstrated that there was little difference between the cultured nasal and bronchial epithelial cells with respect to either their morphology or ciliary activity.

The effects of topical fluticasone propionate on allergen-induced immediate nasal airways response and eosinophil activation: preliminary results.

Nasal application of grass pollen allergen in atopic individuals with seasonal rhinitis leads to an early rise in nasal airways resistance. The effects of fluticasone propionate, a powerful, topically active glucocorticosteroid, on nasal airways resistance and cellular infiltration of the nasal mucous membrane were investigated. Fluticasone propionate blunted the rise in nasal airway resistance following allergen challenge (P = 0.089). Although this glucocorticosteroid did not affect the total number of eosinophils in biopsies of nasal mucous membrane, the number of activated eosinophils was significantly reduced (P less than 0.05).

Air pollutants and respiratory hypersensitivity.

Epidemiological evidence suggests that an increase in liquid petroleum derived pollutants is associated with exacerbation of allergic airway disease, and that the effects of pollution may occur 1-2 days later. Laboratory based studies have demonstrated that the pollutants responsible for the adverse effects on respiratory health include nitrogen dioxide (NO2), sulphur dioxide (SO2), ozone (O3) and respirable particulates (PM10). More recently, studies of asthmatic individuals exposed to O3, NO2 and a combination of NO2 and SO2 have indicated that these agents increase the airway responsiveness of these individuals to inhaled allergen, and that this effect may be maximal 24 h after exposure to the pollutants. Studies investigating the putative mechanisms underlying the effects of these pollutants suggest that exposure to these agents may lead to perturbation of the airway epithelium and release of pro-inflammatory mediators from the epithelial cells, which then influence the activity of inflammatory cells, such as eosinophils.

Effect of Variable Inspiratory Flow Rate on the Performance of the Budesonide Rhinocort Turbuhaler™.

OBJECTIVE: To investigate the effect of variable nasal inspiratory flow rates in vitro on total drug delivery and deposition patterns of budesonide delivered from the Rhinocort Turbuhaler™. METHODS: The total dose of budesonide delivered at flow rates of 15, 30 and 60 L/min and the particle size distribution of the delivered drug at flow rates of 28 and 60 L/min were determined at regular intervals throughout the life of six Rhinocort Turbuhalers™, each containing 200 × 100µ,g doses of budesonide. The delivered dose was determined by drawing individual doses of budesonide through separate G0120 Filtrete electrostatic filters. Acascade impactor was used to determine the particle size distribution of the delivered drug. RESULTS: The amount of budesonide delivered from each Turbuhaler™ device was variable throughout the life of the devices and was dependent on the inspiratory flow rate. Variability was greatest at the lower flow rates and decreased slightly with increasing inspiratory flow rate. Similarly, the particle size distribution of budesonide throughout the cascade impactor was variable and dependent on the flow rate. CONCLUSION: This study suggested that the efficiency of the Rhinocort Turbuhaler™ device in the management of allergic rhinitis may be influenced by the severity of nasal symptoms, particularly rhinorrhoea and nasal blockage, which determine the peak nasal inspiratory flow rates in symptomatic individuals.

Human nasal and bronchial epithelial cells in culture: an overview of their characteristics and function.

Our studies have demonstrated it is possible to culture both human nasal and bronchial epithelial cells to confluency under similar conditions in vitro. These cells are similar morphologically, histologically, and functionally and resemble the cells in vivo. Studies of ciliary activity demonstrate that these cells react toward various agonist and antagonist agents in a similar manner as the cells reported in vivo and consequently will prove to be a valuable model for the evaluation of various therapeutic agents used in the management of infective lung diseases. Similarly studies on the effect of histamine have demonstrated that histamine probably does lead to an increase in bronchial epithelial permeability in vivo and that this effect is likely to be mediated via stimulation of H-1 receptors. Again this could be of clinical relevance when formulating appropriate therapy. The finding that human tracheal and nasal epithelial cells in culture are capable of synthesizing PGE2 could have important clinical implications because this compound is thought to be a putative epithelium-derived relaxing factor. Although this finding awaits confirmation, similar studies with bronchial epithelial cells in vitro would provide a useful tool for study of the function of the bronchial epithelium and its role in the pathogenesis of diseases such as asthma and chronic bronchitis.

Asthma. Epithelial cells.

The airway epithelium is the first line of defence against air-borne dusts, vapour, gases and fumes and plays a vital role in the maintenance of physico-chemical homoeostasis. Perturbation of the epithelial cells which predominate within this barrier, may bring about adverse changes in and around the surrounding tissues playing a part in the pathogenesis of asthma. Although the specific mechanism(s) which partake in pathogenesis are not clear, several have been proposed. These include: (1) increased synthesis and release of inflammatory mediators and decreased synthesis of protective mediators; (2) synthesis of inflammatory cytokines; (3) modulation of cell adhesion molecules, important in determination of epithelial architecture; and (4) immunoregulation of the inflammatory cell types. It is likely that these operate together and interact to produce, initially functional changes in the epithelium, which lead eventually to the structural changes that are characteristic of the airway epithelium of asthmatics.

Air pollution and airway epithelial cells.

Recent epidemiological evidence suggests that air pollution, resulting from vehicle exhaust emissions and burning of liquid petroleum gas or kerosene, may play an important role in the development of allergic airways disease. Further, agents such as O3 and NO2 are thought to be major proponents of disease and act by damaging the airway epithelium. It is hypothesised that this pollution-induced damage results in the generation of mediators which lead to inflammation and possibly airway hyperreactivity.

Effect of antihistamines on epithelial cells.

Antihistamines have mostly been used in the management of allergic rhinitis, primarily for their symptomatic relief. Recent studies, however, have suggested that the non-sedating second-generation antihistamines also possess anti-inflammatory activity, and consequently may be useful in the management of inflammation in allergic airways disease. Several in vivo studies have demonstrated that antihistamines decrease inflammatory cell infiltration in allergic disease, mediator release from mast/basophil cells, and the expression of adhesion molecules on epithelial cells. Similarly, in vitro studies have demonstrated that antihistamines decrease the migration and activation of eosinophils and the release of pro-inflammatory mediators from mast/basophil cells, induced by immunological and non-immunological stimuli. More recent evidence suggests that the antihistamines may modulate airway inflammation by influencing the activity of airway epithelial cells, which due to their spatial arrangement and predominance in the airways, are thought to play a pivotal role in the aetiology of airway disease. We and others have demonstrated that antihistamines attenuate allergen- or chemical-induced expression and/or release of mediators which influence the activity of inflammatory cells, such as eosinophils, mast cells, basophils and lymphocytes, known to be involved in the pathogenesis of allergic airway diseases. Collectively, these studies suggest that second-generation H1-histamine receptor antagonists may have potential use either as safe anti-inflammatory alternatives to corticosteroids, or as rescue medication in combination with corticosteroids, for the management of severe airway disease.

Airway response of asthmatic subjects to inhaled allergen after exposure to pollutants.

BACKGROUND: Recent studies have suggested that air pollutants resulting from vehicle exhaust emissions and burning of fossil fuels, either in combination or individually, may enhance the airway response of asthmatic subjects to inhaled allergen. It was hypothesised that the airway response to inhaled allergen after exposure to a combination of 400 ppb nitrogen dioxide (NO2) and 200 ppb sulphur dioxide (SO2) is increased 24-48 hours after exposure. METHODS: Thirteen mild atopic asthmatic volunteers were exposed for six hours to a single exposure of air and three exposures of the combination of 400 ppb NO2 + 200 ppb SO2 in randomised order, and then challenged with increasing concentrations of Dermatophagoides pteronyssinus allergen either immediately after exposure to air, or immediately, 24 hours or 48 hours after exposure to the combination of the two pollutants, until a 20% fall in forced expiratory volume in one second (FEV1) was recorded. RESULTS: Exposure to 400 ppb NO2 + 200 ppb SO2 significantly decreased the dose of D pteronyssinus allergen required to produce a 20% fall in FEV1 (PD20FEV1) at all times after exposure when compared with air. The mean percentage changes in allergen PD20FEV1 immediately, 24 hours, and 48 hours after exposure to 400 ppb NO2 + 200 ppb SO2 were -37% (95% confidence intervals (CI) -50 to -23), -63% (CI -75 to -51), and -49% (CI -75 to -28.8), respectively, when compared with the PD20FEV1 after air exposure and were significant at all time points studied. The allergen PD20FEV1 at 24 hours after exposure to the combination of the two pollutants was also found to be significantly lower when compared with that immediately after exposure to the two pollutants. CONCLUSION: These results demonstrate that exposure to a combination of NO2 and SO2, at concentrations which can be encountered during episodes of increased outdoor and indoor air pollution, enhances the airway response to inhaled allergen in asthmatic subjects. This effect persists over a period of 24-48 hours and is maximal 24 hours after exposure to these air pollutants.