The effect of induced sputum and bronchoalveolar lavage fluid from patients with chronic obstructive pulmonary disease on neutrophil migration in vitro.

OBJECTIVE: The aim of study was to investigate a chemotactic effect of induced sputum and bronchoalveolar lavage fluid on blood neutrophils in patients with chronic obstructive pulmonary disease (COPD) and healthy individuals. MATERIAL AND METHODS: Forty-three smokers with COPD, 19 ex-smokers with COPD, 13 healthy smokers, and 17 healthy nonsmokers were recruited to the study. Neutrophils were isolated from peripheral blood of study individuals. For the same experimental conditions, pooled induced sputum and bronchoalveolar lavage fluid of 20 COPD patients were used. Neutrophil chemotaxis in vitro was performed in cell-transmigration chamber. Substances tested for chemoattraction (interleukin-8, induced sputum, bronchoalveolar lavage fluid directly or in addition to interleukin-8) were added to lower wells. Upper wells were filled with 2.5 x 10(6)/mL of neutrophil culture and incubated for 2 hours. Migration was analyzed by flow cytometry. RESULTS: Interleukin-8 (10-100 ng/mL) induced a dose-dependant neutrophil migration in all the groups. Only 100 ng/L of interleukin-8 induced more intensive chemotaxis of neutrophils from COPD smokers as compared to ex-smokers (P<0.05). Such difference between healthy individuals was obtained using 30 ng/mL of interleukin-8 (P<0.05). Induced sputum/interleukin-8 (10-100 ng/mL), as well as induced sputum directly, induced neutrophil migration (P<0.05). Chemotaxis of neutrophils isolated from COPD patients and healthy nonsmokers did not depend on additional interleukin-8 concentration. Bronchoalveolar lavage fluid/interleukin-8 (30-100 ng/mL) induced more intensive migration of neutrophils from COPD patients than bronchoalveolar lavage fluid (P<0.05) alone. CONCLUSIONS: Migration of neutrophils isolated from patients with COPD was more intensive compared to healthy individuals. Induced sputum and bronchoalveolar lavage fluid directly and with addition of interleukin-8 stimulated chemotaxis, and it was higher in neutrophils from COPD patients. Migration of neutrophils did not depend on smoking status.

Smoking affects eotaxin levels in asthma patients.

BACKGROUND: Chronic airway inflammation is most important pathological finding in asthma. Cigarette smoking may modify type of inflammation as well as may influence disease severity and response to the treatment. OBJECTIVE: Thus the aim of this study was to investigate whether cigarette smoking may have an influence on the levels of eotaxin-1, eotaxin-2, eotaxin-3 and IL-5 in patients with stable mild/moderate asthma. METHODS: 45 steroid naive asthmatics (mean age: 55.2 +/- 2.2 yrs) and 23 "healthy" smokers and non-smokers control subjects (mean age: 54.4 +/- 9.7 yrs) were investigated. Asthmatics were divided into two subgroups according to their smoking histories: asthmatic smokers (n = 19) who currently smoke and have a history of > 10 pack-years and asthmatic never-smokers (n = 26). BAL and induced sputum were performed. Cytospins of induced sputum and BAL were stained with May-Grunwald-Giemsa for differential cell counts. Eotaxin-1, eotaxin-2, eotaxin-3 and IL-5 concentrations in serum, sputum and BAL supernatant was measured using a commercial ELISA kit. RESULTS: In sputum supernatant from asthma smokers was significantly higher concentration of eotaxin-1 than in non-smokers asthmatics (203.4 +/- 10.0 vs. 140.2 +/- 9.5 respectively, p < 0.05). In non-smokers asthma patients levels of BAL eotaxin-1 strongly related to percent and absolute numbers of BAL eosinophils and neutrophils (Rs = 0.737 and Rs = 0.514 respectively, p < 0.05). The number and percent of sputum neutrophils and eosinophils, obtained from smokers asthmatics, significantly correlated with eotaxin-2 concentration in sputum supernatant (Rs = 0.58 and Rs = 0.75 respectively, p < 0.05). IL-5 levels in the serum and sputum from asthmatic never-smokers were significantly higher than they were from asthmatic smokers and "healthy" smokers. Asthmatic never-smokers showed a significantly higher amount of IL-5 in serum and sputum than the asthmatic smokers showed. CONCLUSIONS: This study showed the elevated levels of sputum eotaxin-1 as well as serum, sputum and BAL eotaxin-2 in asthmatic smokers without a significant increase of eosinophils compared to asthmatic never-smokers. The eotaxin concentrations were related not only with number of eosinophils but also with the number of neutrophils in all the studied tissue compartments. The data herein permits a suggestion that smoking may influence change in asthmatic airway inflammation by stimulating the production of eotaxins.

Patterns of airway inflammation and MMP-12 expression in smokers and ex-smokers with COPD.

BACKGROUND: Smoking activates and recruits inflammatory cells and proteases to the airways. Matrix metalloproteinase (MMP)-12 may be a key mediator in smoke induced emphysema. However, the influence of smoking and its cessation on airway inflammation and MMP-12 expression during COPD is still unknown. We aimed to analyse airway inflammatory cell patterns in induced sputum (IS) and bronchoalveolar lavage (BAL) from COPD patients who are active smokers and who have ceased smoking >2 years ago. METHODS: 39 COPD outpatients - smokers (n = 22) and ex-smokers (n = 17) were studied. 8 'healthy' smokers and 11 healthy never-smokers were tested as the control groups. IS and BAL samples were obtained for differential and MMP-12+-macrophages count analysis. RESULTS: The number of IS neutrophils was higher in both COPD groups compared to both controls. The amount of BAL neutrophils was higher in COPD smokers compared to healthy never-smokers. The number of BAL MMP-12+-macrophages was higher in COPD smokers (1.6 +/- 0.3 x 106/ml) compared to COPD ex-smokers, 'healthy' smokers and healthy never-smokers (0.9 +/- 0.4, 0.4 +/- 0.2, 0.2 +/- 0.1 x 106/ml respectively, p < 0.05). CONCLUSION: The lower amount of BAL neutrophils in COPD ex-smokers, compared to COPD smokers, suggests positive alterations in alveolar compartment after smoking cessation. Smoking and disease itself may stimulate MMP-12 expression in airway compartments (IS and BAL) from COPD patients.

Perception of dyspnea in asthmatics with normal lung function.

UNLABELLED: The perception of dyspnea varies widely among asthmatics and it is influenced by many factors. The aims of our study were to investigate the perception of dyspnea during methacholine-induced bronchoconstriction in asthmatics with normal lung function and to evaluate the influence of bronchial responsiveness, age and gender to dyspnea perception in these patients. A total of 192 outpatients (aged 16-77 years) with stable asthma and normal lung function were examined. Methacholine challenge test was performed to each patient. The provocative dose of methacholine that reduces forced expiratory volume in 1 sec. (FEV1) by 20% (PD20) was estimated. Dyspnea perception of bronchoconstriction was evaluated using a Borg Scale and calculating the perception score corresponding to a fall in FEV1 of 20% (PS20). According to PS20+/-1 standard deviation subjects were divided into three groups: hypoperceivers, normoperceivers and hyperperceivers. From the hypoperceivers group we set up asthmatics with PS20=0 and defined them as nonperceivers. We found out that 43 (22.4%) patients were hypoperceivers, 116 (60.4%)--normoperceivers and 33 (17.2%)--hyperperceivers. The nonperceivers presented 6.3% (n=12) of all subjects. PD20 positively correlated with PS20 (r=0.252, p<0.001). Hypoperceivers showed significantly higher bronchial hyperesponsiveness (PD20=174+/-28 microg) comparing with hyperperceivers (PD20=323+/-50 microg, p=0.013). Bronchial responsiveness to methacholine of nonperceivers (PD20= 106+/-31 microg, range 15-253 microg) was the highest and PD20 was significantly lower comparing with normoperceivers (p=0.005) and hyperperceivers (p=0.001). Age and gender had no significant effect on the perception of bronchoconstriction. CONCLUSION: The part of asthmatics with normal lung function has impaired perception of dyspnea. Dyspnea perception depends on bronchial responsiveness, but not on age and gender of these patients.

Influence of smoking status on cough reflex sensitivity in subjects with COPD.

Cough reflex sensitivity has not been studied extensively in patients with chronic obstructive pulmonary disease (COPD). The aim of the study was to evaluate cough reflex sensitivity to capsaicin in current and former smokers with COPD and examine its association with potentially protussive mediators. Fifteen active smokers and 18 ex-smokers with moderate to severe COPD, 14 healthy active smokers, and 13 healthy never smokers were enrolled. Capsaicin aerosol was administered in order of ascending concentration until the concentrations inducing two or more coughs (C(2)) and five or more coughs (C(5)) were attained. The concentrations of leukotriene E(4) (LTE(4)), leukotriene B(4) (LTB(4)), and interleukin-8 (IL-8) in bronchoalveolar lavage (BAL) fluid were analyzed by ELISA. Cough reflex sensitivity in COPD smokers [mean log C(2) = 1.20 +/- 0.23 (SEM) microM; log C(5) = 1.85 +/- 0.21 microM] did not differ from that in COPD ex-smokers (log C(2) = 1.15 +/- 0.14 microM; log C(5) = 2.10 +/- 0.19 microM; p > 0.05). Mean C(2) and C(5) in both COPD groups were significantly lower than in healthy active smokers, but higher compared with the healthy never-smokers. BAL fluid concentrations of LTE(4) and LTB(4) were similar in all groups. IL-8 concentrations did not differ between COPD smokers, COPD ex-smokers, and healthy active smokers, but were significantly higher in all three groups compared with healthy never smokers. Cough reflex sensitivity to capsaicin does not differ between smokers and ex-smokers with COPD.

Changes in cough reflex sensitivity after cessation and resumption of cigarette smoking.

Previous studies have shown that healthy cigarette smokers have diminished cough reflex sensitivity compared to healthy nonsmokers. We have recently demonstrated that cough reflex sensitivity is enhanced soon after smoking cessation, suggesting that diminished cough sensitivity in smokers results from chronic cigarette smoke-induced desensitization of airway cough receptors. In this study, we evaluated cough reflex sensitivity to capsaicin (C(5)) in 11 chronic smokers who had discontinued smoking for at least 2 weeks, and then resumed smoking. Two weeks after smoking cessation there was a significant enhancement of cough reflex sensitivity; mean (+/-SEM) log C(5) decreased from 1.77+/-0.18 to 1.47+/-0.14 (p=0.01). All subjects resumed smoking after 2-12 weeks of abstinence. Repeat capsaicin cough challenge was performed 14-23 days after resumption of smoking. Mean log C(5) increased compared to the last value obtained during the smoking cessation period: 1.42+/-0.15 vs. 1.77+/-0.16 (p=0.0004). Mean log C(5) after resumption of smoking returned to almost exactly the baseline value. Our findings suggest that the sensitivity of airway cough receptors is a dynamic phenomenon, promptly affected and modulated by changes in environmental conditions, such as the presence or absence of cigarette smoke.

Dyspnea perception and reversibility of methacholine-induced unlimited airway narrowing in asthmatics.

The hypothesis was that asthmatics might experience impaired perception of dyspnea and salbutamol-induced reversibility during unlimited airway narrowing. A total of 38 asthmatics (18 to 59 years of age) were examined. All patients underwent the methacholine challenge test. According to the dose-response curve to methacholine, they were categorized as having either unlimited airway narrowing (UAN group) (n = 20) or response plateau (RP group) (n = 18). Reversibility of methacholine-induced bronchoconstriction was measured 20 minutes after the inhalation of 400 microg of salbutamol to compare postbronchodilator FEV1 with baseline FEV1. Dyspnea perception was evaluated using the Borg Scale to calculate a perception score at a 20% decrease in FEV1 (PS20) and the slope alpha of the regression line between the changes in Borg scores and the reduction in FEV1 as percentage of the baseline value. Subjects in the UAN group exhibited significantly lower PS20 compared with the RP group (1.45+/- 0.23 vs. 2.84 +/- 0.35, p = 0.002); the mean of the slope values was higher in the RP group than it was in the UAN group (0.150 +/- 0.015 vs. 0.095 +/- 0.006, p = 0.003). Salbutamol-induced reversibility was significantly lower in the UAN group (81 +/- 1.4 % of baseline FEV1) compared with patients from the RP group (91 +/- 1.1% of baseline FEV1; p < 0.001). In conclusion, asthmatics during methacholine-induced unlimited airway narrowing exhibit diminished perception of dyspnea and lower bronchial reversibility to the baseline 20 minutes after inhalation of salbutamol. This suggests that more careful monitoring of the lung function for timely recognition of asthma deteriorations and adequate bronchodilatory therapy during severe acute attacks should be recommended for such patients.