Automated quantitation of circulating neutrophil and eosinophil activation in asthmatic patients.

BACKGROUND: Asthma has been associated with eosinophil activation, measured in serum, sputum, bronchoalveolar lavage (BAL) fluid, and urine. A whole blood automated method was developed to assess eosinophil and neutrophil activity in terms of peroxidase content and cell morphology using the Bayer haematology analyser. The method was applied to an in vitro stimulation model when fMLP was added to whole blood and the samples were then analysed for changes in granularity and shape. In addition, cells stimulated with interleukin (IL)-8 were examined by electron microscopy. METHODS: A cross sectional analysis was performed on venous blood from non-atopic, non-asthmatic normal subjects (n = 37), mild (n = 46) and symptomatic (n = 22) asthmatic patients on inhaled beta(2) agonist only, and more severe asthmatic patients (n = 17) on inhaled and oral corticosteroid therapy. Samples were analysed by the haematology analyser and peroxidase leucograms gated using the WinMDI software program. RESULTS: There were significant differences in the amount of light scatter by the neutrophil populations in the symptomatic (p = 0.007) and severe asthmatic (p = 0.0001) groups compared with the control group. However, abnormalities in eosinophil populations were not observed. In vitro activation of whole blood with fMLP caused similar changes in neutrophil light scatter, suggesting that neutrophil activation is present in peripheral blood of symptomatic asthmatic patients. IL-8 caused a change in shape of the neutrophils seen using transmission electron microscopy. CONCLUSIONS: Evidence of neutrophil activation can be seen in whole blood from patients with asthma using a novel automated method. This may potentially be applied to other inflammatory diseases.

Comparison of salmeterol and albuterol-induced bronchoprotection against adenosine monophosphate and histamine in mild asthma.

Short-acting beta(2)-agonists provide greater protection to bronchoconstriction induced by adenosine-5'-monophosphate (AMP) than does methacholine. Because AMP produces bronchoconstriction through release of mediators from mast cells, and methacholine directly constricts airway smooth muscle, this suggests that beta(2)-agonists stabilize mast cells in vivo. This in vivo property has not been demonstrated with long-acting beta(2)-agonists. We undertook two double-blind, randomized, crossover, placebo-controlled studies to investigate the effects of salmeterol and albuterol on airway responsiveness (AR) to AMP and histamine in patients with mild asthma. In the first study, 19 patients attended on four occasions to inhale salmeterol 50 micrograms or placebo 2 h before challenge with AMP or histamine. In the second study 16 patients (13 of whom had participated in the first study) were studied in a similar fashion but inhaled albuterol 400 micrograms or placebo 30 min prior to challenge. Salmeterol reduced AR to AMP and histamine by 3.4 +/- 0.3 and 3.9 +/- 0.3 doubling doses, respectively (NS). In contrast, albuterol demonstrated a greater protective effect on AMP than on histamine, reducing AR by 5.1 +/- 0.3 and 3.8 +/- 0.2 doubling doses, respectively (p < 0.005). Thus, in contrast to albuterol, salmeterol did not demonstrate mast-cell stabilizing properties in vivo at a time corresponding to maximal bronchodilatation. These findings might be explained by the unique pharmacologic profile of salmeterol in combination with the differential beta(2)-adrenoceptor pharmacology of bronchial mast cells and bronchial smooth muscle.

Tolerance to the nonbronchodilator effects of inhaled beta 2-agonists in asthma.

BACKGROUND: Tolerance to the direct bronchodilator effects of beta 2-agonists does not appear to occur in asthma. However, it is not known whether this is true for the nonbronchodilator effects of these agents, which protect the airways against bronchoconstrictive stimuli. METHODS: We investigated whether tolerance develops to the protective effect of inhaled terbutaline on airway responsiveness to the bronchoconstrictors methacholine (which acts directly on airway smooth muscle) and AMP (which acts indirectly by stimulating the release of mediators from mast cells) during sustained treatment with terbutaline. In a randomized, double-blind, crossover study, 12 patients with mild asthma each inhaled a single dose of terbutaline (500 micrograms) or placebo before a challenge with a series of doubling doses of inhaled methacholine or AMP, before and after treatment for seven days with 500 micrograms of terbutaline four times daily or placebo. RESULTS: Before the seven days of treatment with terbutaline, a single dose of terbutaline reduced airway responsiveness to methacholine by 2.7 doubling doses (95 percent confidence interval, 1.9 to 3.5), but it had an even greater protective effect against AMP, reducing airway responsiveness by 3.8 doubling doses (95 percent confidence interval, 2.7 to 4.9; P less than 0.001). After seven days of treatment with terbutaline, the protective effect of terbutaline against methacholine decreased to 2.2 doubling doses (95 percent confidence interval, 1.3 to 3.0; P = 0.04), and that against AMP decreased even more, to 1.7 doubling doses (95 percent confidence interval, 1.1 to 2.4; P less than 0.001). By contrast, the bronchodilator response to terbutaline was unchanged during seven days of treatment with this agent. CONCLUSIONS: We observed tolerance to the nonbronchodilator actions of the inhaled beta 2-agonist terbutaline in patients with mild asthma, an effect that may be more pronounced in mast cells than in bronchial smooth muscle. This property of beta-agonists may constitute a drawback to their regular use in patients with asthma.