Airway inflammation and bronchial microbial patterns in patients with stable chronic obstructive pulmonary disease.

The effect of bacterial colonization of the bronchi on the progress of airflow limitation is not well known. Therefore, the pattern of airway inflammation in smokers and patients with stable chronic obstructive pulmonary disease (COPD) and its relation to bronchial microbial colonization was assessed. Eight nonsmoking and 18 smoking controls as well as 52 patients with COPD (28 mild, 11 moderate and 13 severe) were studied. All subjects were investigated by means of flexible bronchoscopy including protected specimen brush and bronchoalveolar lavage (BAL) sampling. Differential cell counts, cytokine (interleukin (IL)-1beta, IL-6, IL-8, IL-10 and tumour necrosis factor-alpha(TNF-alpha) concentrations and microbial patterns were determined in BAL fluid. Forced expiratory volume in one second (FEV1) % of the predicted value was inversely correlated with pack-yrs of cigarette smoking (r=-0.47, p<0.0001), the percentage of neutrophil (p=-0.56, p<0.0001) and IL-6 (p=-0.37, p=0.01) and IL-8 concentration (p=-0.43, p=0.004) in BAL fluid. Accordingly, pk-yrs of cigarette smoking (p=0.39, p=0.01) and IL-8 (p=0.69, p<0.0001) and TNFalpha (p=0.4, p<0.005) were positively correlated with the percentage of neutrophils in BAL fluid. Smoking controls and COPD patients were mainly colonized in the bronchial tree (33%) by community endogenous potentially pathogenic micro-organisms (PPMs). Colonization rates and patterns of PPMs were not affected by severity of airflow obstruction. The presence of PPMs was significantly associated with higher percentages of neutrophils (33.2+/-10.4% versus 10.1+/-3.5%, p=0.02) and TNF-alpha concentration (29.9+/-10.8 versus 6.3+/-2.1 pg x mL(-1), p=0.01) in BAL fluid. In conclusion, bronchial neutrophilia is a key inflammatory pattern in chronic obstructive pulmonary disease patients. Bronchial colonization with potentially pathogenic micro-organisms may represent an independent stimulus for additional airway inflammation.     

Small airways dysfunction and neutrophilic inflammation in bronchial biopsies and BAL in COPD

BACKGROUND: The single-breath N(2) test (sbN(2)-test) is closely related to small airways pathology in resected lung specimens of smokers. We investigated whether uneven ventilation and airway closure are associated with specific markers of airway inflammation as obtained by bronchial biopsies, BAL, and induced sputum in patients with manifest COPD. METHODS: Fifty-one patients with stable COPD not receiving corticosteroids were examined in a cross-sectional study (43 men; mean [SD] age, 63 +/- 8 years; exsmokers and smokers; median pack-years, 41 [interquartile range, 31 to 51 pack-years]). Postbronchodilator spirometry (FEV(1), 63 +/- 8% of predicted) and sbN(2)-test (slope of phase III [DeltaN(2)], closing capacity [CC]/total lung capacity [TLC] percentage of predicted) were performed. Inflammatory cell counts were assessed in bronchial biopsies, BAL (only in the first half of patients), and induced sputum. Neutrophil elastase (NE), secretory leukocyte proteinase inhibitor (SLPI), and interleukin-8 levels were determined in BAL fluid. RESULTS: DeltaN(2) increased with subepithelial neutrophil numbers in bronchial biopsies (rs = 0.337, p = 0.017) and with NE levels (rs = 0.443, p = 0.039), NE/neutrophil ratio (rs = 0.575, p = 0.005) and SLPI levels (rs = 0.484, p = 0.022) in BAL. CC/TLC was associated with BAL neutrophil numbers (rs = 0.448, p = 0.048). The sbN(2)-test was not associated with any other inflammatory cell type in BAL or biopsies, nor with inflammatory cell counts in sputum. Of importance, the correlations between DeltaN(2) and BAL NE/neutrophil ratio, and between CC/TLC and BAL neutrophil numbers remained significant when adjusting for FEV(1) percentage of predicted. CONCLUSIONS: The results of the sbN(2)-test are associated with neutrophilic inflammation in bronchial biopsies and BAL in patients with COPD. Our findings support a role of neutrophilic inflammation in the pathogenesis of small airways dysfunction in COPD.     

The effect of processing on inflammatory markers in induced sputum.

The effects of the mucolytic agent, dithioerythritol (DTE), and the temperature at which sputum processing is conducted on cellular and biochemical markers in induced sputum was assessed. Samples from healthy and atopic asthmatic subjects were treated with either DTE or phosphate-buffered saline (PBS) at 22 or 37 degrees C and compared for cell counts and concentrations of histamine, tryptase, eosinophil cationic protein (ECP), free interleukin (IL)-8, immunoglobulin (Ig)A, IL-8/IgA complexes and secretory component (SC). In addition, the influence of DTE on in vitro mediator release from blood eosinophils, basophils and bronchoalveolar lavage (BAL) mast cells was studied. Processing with DTE improved cytospin quality and increased the cell yield and measurable ECP, tryptase, IgA and SC, but reduced levels of histamine in PBS-treated samples and had no effect on IL-8. Cell counts or mediator levels were similar when sputum was processed at 22 or 37 degrees C, even though DTE induced blood basophils and BAL mast cells to release histamine at 37 degrees C. In spiking experiments, recovery of added ECP, tryptase, total IL-8 and histamine from sputum was similar in DTE- and PBS-processed sputum, but reduced for free IL-8 in PBS-treated samples. In conclusion, dithioerythritol improves cell and mediator recovery without causing cell activation when sputum processing is conducted at room temperature. The extent of recovery depends on the mediator studied.     

慢性阻塞性肺疾病患者p38蛋白激酶表达的变化及其临床意义

目的探讨慢性阻塞性肺疾病(COPD)患者诱导痰中p38蛋白激酶(p38MAPK)表达的变化及其临床意义。方法选择COPD急性加重期患者29例、稳定期患者18例及健康对照者15例。3%～5%氯化钠注射液进行痰液诱导,计数诱导痰中细胞总数、细胞分类。分别采用蛋白质免疫印迹、ELISA方法检测诱导痰中细胞核蛋白p38MAPK含量和上清液中IL-8的含量。结果与健康对照者比较,COPD患者诱导痰中细胞总数、中性粒细胞比例、p38MAPK的表达和上清液IL-8浓度明显升高,FEV1%显著降低(P<0.05);各组诱导痰中细胞核蛋白p38 MAPK的表达与上清液IL-8浓度和中性粒细胞比例均呈正相关(r=0.531,0.664,P<0.01),与FEV1呈负相关(r=-0.468,P<0.05)。结论 COPD患者诱导痰中p38 MAPK表达增高,可能与COPD气道炎症发生发展密切相关。     

Serum and bronchial lavage fluid concentrations of IL-8, SLPI, sCD14 and sICAM-1 in patients with COPD and asthma

BACKGROUND: Airway inflammation is associated with an increased expression and release of inflammatory reactants that regulate processes of cell migration, activation and degranulation. The purpose of this study was to quantify bronchial lavage (BAL) fluid and serum levels of chemokine (IL-8), secretory leukocyte protease inhibitor (SLPI), soluble intracellular adhesion molecules-1 (sICAM-1) and sCD14, as surrogate markers of inflammatory and immune response in asthma and chronic obstructive pulmonary disease (COPD) patients with similar disease duration time. METHODS: Biomarkers in serum and BAL fluid from asthma (n=13) and COPD (n=25) patients were measured using commercially available ELISA kits. RESULTS: We found that in asthma and COPD groups the concentrations of IL-8 and SLPI are significantly higher in BAL fluid than in serum, while levels of sICAM-1 and sCD14 in BAL fluid are significantly lower than in serum. Of these 4 measured biomarkers, only the BAL IL-8 was higher in COPD patients when compared to asthma (P<0.05). In both groups, BAL IL-8 correlated with SLPI (r=0.577, P<0.01 and r=0.589, P<0.05, respectively). In patients with COPD the BAL sICAM-1 correlated with sCD14 (r=0.576, P<0.01), while in asthma patients BAL sICAM-1 correlated with FEV(1)/FVC (r=0.418, P<0.01). Moreover, in asthma patients the serum SLPI correlated with sCD14 (r=0.688, P<0.01) and serum sICAM-1 negatively correlated with FEV(1)/FVC (r=-0.582, P<0.05). CONCLUSION: Our findings point to the importance of selecting a correct biological fluid when analyzing specific biomarkers, and also show that of 4 measured biomarkers, only the BAL IL-8 was higher in COPD patients when compared to asthma.     

Airway inflammation and bronchial bacterial colonization in chronic obstructive pulmonary disease

RATIONALE: Inflammation is now recognized as an integral part of the pathogenesis of chronic obstructive pulmonary disease (COPD). In contrast to the sterile airways of normal lungs, bacterial pathogens are often isolated from the airways in stable COPD. This "colonization" of the tracheobronchial tree, currently believed to be innocuous, could serve as an inflammatory stimulus, independent of current tobacco smoke exposure. Objective: To test the hypothesis that bacterial colonization is associated with airway inflammation in stable COPD. METHODS: Bronchoscopy with bronchoalveolar lavage (BAL) was performed in three groups of subjects: 26 ex-smokers with stable COPD (COPD), 20 ex-smokers without COPD (ex-smokers), and 15 healthy nonsmokers (nonsmokers). Quantitative bacterial cultures, cell counts, chemokine, cytokine, proteinase/antiproteinase, and endotoxin levels in the BAL fluid were compared. RESULTS: Potentially pathogenic bacteria were recovered at > or = 100 cfu/ml in 34.6% of COPD, 0% of ex-smokers, and in 6.7% of nonsmokers (p = 0.003). All values are expressed as median (interquartile range). Subjects with colonized COPD had significantly greater relative (12.0 [28.4] vs. 3.0 [7.8]%, p = 0.03) and absolute (4.98 [5.26] x 10(4)/ml vs. 3.04 [2.82] x 10(4)/ml, p = 0.02) neutrophil counts, interleukin 8 (33.8 [189.8] vs. 16.9 [20.1] pg/ml, p = 0.005), active matrix metalloproteinase-9 (2.16 [4.30] vs. 0.84 [0.99] U/ml, p = 0.03), and endotoxin (36.0 [72.6] vs. 3.55 [7.17] mEU/ml, p = 0.004) levels in the BAL than the subjects with noncolonized COPD. These inflammatory constituents of BAL were also significantly elevated in subjects with colonized COPD when compared with ex-smokers and nonsmokers. CONCLUSIONS: Bacterial colonization is associated with neutrophilic airway lumen inflammation in ex-smokers with COPD and could contribute to progression of airway disease in COPD.     

The eosinophil component of the alveolitis in idiopathic pulmonary fibrosis. Signs of eosinophil activation in the lung are related to impaired lung function

The number of eosinophils and the concentrations of eosinophil cationic protein (ECP), a specific granule constituent of eosinophil granulocytes, were measured in bronchoalveolar lavage (BAL) fluid from patients (n = 22) with idiopathic pulmonary fibrosis (IPF). The median recovery of eosinophils during lavage performance was 4% (range, zero to 49) of the nonepithelial cells and significantly increased compared with the recovery in healthy control subjects (less than 1%) and in control patients with sarcoidosis (1%; range, zero to 7). The median BAL fluid concentration of ECP was in IPF 13.3 micrograms/L (range, 2 to 118) and significantly increased compared with the concentrations in healthy control subjects (2.7 micrograms/L; range, less than 2 to 8) and in control patients (6.6 micrograms/L; range, less than 2 to 64). The BAL fluid concentrations of myeloperoxidase (MPO) were also significantly increased in IPF, indicating a local neutrophil activation. A significant correlation was found between BAL fluid ECP and MPO, suggesting a common activator of eosinophils and neutrophils. BAL fluid eosinophils and ECP correlated with the reduced diffusion capacity of the lung but not with vital capacity or forced expiratory volume. It is concluded that eosinophil activation is part of the inflammatory process in IPF. ECP and other cytotoxic eosinophil products may play a pathophysiologic role for the lung damage in this disease.     

Induced sputum in patients with newly diagnosed sarcoidosis: comparison with bronchial wash and BAL.

OBJECTIVES: Sarcoidosis is characterized by a diffuse alveolar inflammatory process, although bronchial airways are often involved. This study compares the cellular profiles of induced sputum (IS), bronchial washing (BW), and BAL in newly diagnosed sarcoidosis patients to those in control subjects, and examines whether inflammatory cell counts from IS are correlated with inflammatory cell counts from BW and BAL in sarcoidosis patients. PATIENTS AND MEASUREMENTS: We recruited 15 untreated patients with stage I and II pulmonary sarcoidosis and 12 healthy volunteers. Sputum was induced with hypertonic saline solution in all individuals. Bronchoscopy was performed on a different occasion in all patients and in five control subjects. RESULTS: Mean lymphocyte counts in IS, BW, and BAL fluid from sarcoidosis patients were significantly higher than in control subjects (9.4% vs 3.8%, p < 0.05; 12.6% vs 3.9%, p < 0.05; 24.1% vs 2.6%, p < 0.05, respectively). Moreover, total cell count and percentage of epithelial cells in IS were significantly higher in sarcoidosis patients than in control subjects (p < 0.01 and p < 0.05, respectively). In sarcoidosis patients, comparison between different samples showed significantly higher percentages of macrophages in BW and BAL than in IS (p < 0.05 and p < 0.01, respectively), whereas the percentage of neutrophils was higher in IS compared with BW and BAL (p < 0.01 and p < 0.001, respectively). Finally, the percentage of lymphocytes in IS was significantly lower than that in BAL (p < 0.05) but not that in BW. CONCLUSIONS: We demonstrated that, compared with IS in healthy control subjects, IS in untreated pulmonary sarcoidosis patients contains more total cells, lymphocytes, and epithelial cells. Although the relative proportion of inflammatory cells in the three samples differed, lymphocyte counts in IS were high. This finding suggests that IS could be used as a valuable alternative to more conventional invasive techniques in clinical assessment of pulmonary sarcoidosis patients.     

慢性阻塞性肺疾病患者诱导痰中白介素8、白三烯B4、髓过氧化物酶的变化以及噻托溴铵对其的影响

目的 探讨慢性阻塞性肺疾病(COPD)患者中自三烯B4(LTB4)、白介素8(IL-8)、髓过氧化物酶(MPO)的变化及噻托溴铵对其的影响,进一步验证噻托溴铵是否有抗炎作用.方法 选中度至极重度COPD稳定期患者30例为噻托溴铵治疗前组,予吸入噻托溴铵30 d,剂量18μg·d-1,为噻托溴铵治疗后组.并选10例健康查体者为对照组,本组无任何干预措施.三组均进行血气分析、肺功能检查和诱导痰沉渣细胞计数和痰上清液ELISA法测定IL-8、LTB4、MPO.采用独立样本t检验及配对t检验,Pearson及Spearman相关等分析.结果 噻托溴铵组与健康对照组痰细胞分类、IL-8[(777.05±392.43)pg·ml-1vs(311.27±135.93)pg·ml-1]、LTB4[(96.70±29.13)pg·ml-1vs(20.60±9.89)pg·ml-1]、MPO[(23.50±16.77)U·ml-1 vs(10.52±3.55)U·ml-1]指标比较,差异均具有统计学意义.噻托澳铵治疗前后痰细胞分类比较,差异无统计学意义,IL-8[(777.05±392.43)pg·ml-1vs(408.93±339.14)pg·ml-1]、LTB4[(96.70±29.13)pg·ml-1 vs(43.69±16.49)pg·ml-1]差异有统计学意义.相关分析示IL-8与PaO2(r=-0.411)、肺功能指标FEV1%pred(r=-0.440)、FVC%pred(r=-0.440)、FEV1/FVC%(r=-0.417)有相关性.结论 COPD患者诱导痰中炎性细胞及炎性因子与健康对照组比较,差异有统计学意义.噻托溴铵对COPD患者诱导痰中IL-8、LTB4有降低作用.     

Levels of IL-32 in Serum,Induced Sputum Supernatant,and Bronchial Lavage Fluid of Patients with Chronic Obstructive Pulmonary Disease

Interleukin-32 (IL-32) is a newly described cytokine which is expected to have an important role in autoimmune disorders. It was shown that chronic obstructive pulmonary disease (COPD) has a component of autoimmunity, though the role of IL-32 in its pathogenesis is not known. The aim of this study was to estimate IL-32 concentrations in serum, induced sputum (IS) supernatant and bronchoalveolar lavage (BAL) fluid from patients with COPD, and to compare asthma patients with and healthy subjects. Outpatients with COPD (63.7 ± 8.4 years, n = 51), asthma (58.3 ± 12.4 years, n = 31), and healthy subjects (59.8 ± 8.2 years, n = 9) were studied. The levels of IL-32 in serum, BAL fluid, and IS supernatant samples were analyzed by ELISA. Concentrations of IL-32 were higher in all the studied materials from patients with COPD (BAL 22.46 ± 2.48 pg/ml, IS 19.66 ± 1.69 pg/ml, serum 26.77 ± 2.56 pg/ml) in comparison with patients with asthma (BAL 6.25 ± 1.08 pg/ml, IS 5.82 ± 1.15 pg/ml, serum 6.09 ± 1.16 pg/ml, p < 0.05 respectively) as well as healthy subjects (BAL 4.21 ± 1.13 pg/ml, IS 3.59 ± 0.66 pg/ml, serum 4.63 ± 1.03 pg/ml, p < 0.05 respectively). Moreover, the level of IL-32 was higher in COPD smokers than in COPD ex-smokers in investigated respiratory tissue compartments and serum, and correlated with smoking history. Increased level of IL-32 in serum, IS supernatant, and BAL fluid from patients with COPD in comparison with asthma patients and healthy subjects suggest that IL-32 may play an important role in the pathogenesis of COPD, which depends on the smoking history.     

Impaired neutrophil chemotaxis in chronic obstructive pulmonary disease

RATIONALE: Neutrophilic airway inflammation is considered to be a major factor in the pathogenesis of chronic obstructive pulmonary disease (COPD), with sputum and bronchoalveolar lavage neutrophil counts broadly correlating with disease severity. The mechanisms responsible for neutrophil accumulation are poorly understood, but they could involve increased influx and/or survival of these cells. OBJECTIVES: To investigate whether neutrophil chemotactic responsiveness and/or chemotactic activity in airway secretions are increased in subjects with COPD. METHODS: Chemotaxis experiments were performed using induced sputum supernatants from subjects with and without COPD as a source of chemotactic activity, and neutrophils from healthy donors as responder cells. In addition, chemotactic responses to N-formyl-Met-Leu-Phe (fMLP) and interleukin-8 (IL-8/CXCL8) were studied using neutrophils from healthy subjects and subjects with COPD. MEASUREMENTS AND MAIN RESULTS: As reported in the literature, sputum neutrophil counts were significantly increased in subjects with COPD compared with healthy subjects. However, this was associated with reduced chemotactic activity in sputum in COPD, as judged by reduced chemotaxis to the fluid phase of sputum from subjects with COPD compared with healthy subjects. Furthermore, whereas neutrophils from subjects with stage I COPD had normal responses to fMLP and IL-8, subjects with more severe stage II-IV COPD showed reduced levels of spontaneous migration and chemotaxis to fMLP and IL-8. CONCLUSIONS: Neither increased chemotactic activity in the airways nor increased chemotactic responsiveness of neutrophils explains the increased number of these cells in subjects with stable COPD. The implications of the observed reduction in neutrophil chemotactic activity remain to be established.     

急性发作期老年晚发哮喘患者气道炎症特征分析

目的检测急性发作期老年晚发哮喘(LOA)诱导痰细胞学、嗜酸细胞阳离子蛋白(ECP)和白介素-5(IL-5)、白介素-8(IL-8)水平,观察LOA气道炎症特征。方法检测86例急性发作期LOA患者诱导痰中细胞学分类计数、ECP和IL-5、IL-8水平;选择30例健康老年人作为对照。结果以诱导痰中EOS数量≥3%作为临界值,86例急性发作期LOA患者中,79例(81%)诱导痰中嗜酸细胞(EOS)数量增高,为痰EOS增高组;17例(19%)痰中性细胞数量增高,为痰非EOS增高组。痰非EOS增高组诱导痰中性细胞和IL-8水平显著高于痰EOS增高组和健康老年组(P0.01);而痰中EOS数量、ECP和IL-5水平显著低于痰EOS增高组(P0.01),但与健康老年组比较差异无统计学意义(P0.05)。结论急性发作期LOA患者存在气道嗜酸细胞和中性细胞两种炎症类型,测定患者气道炎症类型有助于指导治疗。     

Clinical Implication of Protein Levels of IL-5 in Induced Sputum in Asthmatic Patients

To determine whether protein levels of interleukin-5 (IL-5) in induced sputum reflect the degree of eosinophilic inflammation, we evaluated the role of IL-5 on clinical characteristics in stable asthmatic patients. IL-5 level, differential eosinophil count, and level of eosinophil cationic protein (ECP) in induced sputum were all significantly higher for asthmatics than for normal controls. Both eosinophil counts and ECP levels in induced sputum were inversely correlated with the degree of airflow limitation (FEV₁/FVC). In addition, patients with measurable IL-5 in sputum had significantly more eosinophils, higher levels of ECP in sputum, and lower FEV₁ (percent predicted) than did patients with levels of IL-5 beneath the limit of detection. However, we found no significant difference in IL-5 levels between atopic and nonatopic asthmatics. IL-5 level in induced sputum is a good indicator of eosinophilic inflammation in atopic and nonatopic asthmatic patients.     

Inflammatory cytokine levels in induced sputum and bronchoalveolar lavage fluid in pulmonary sarcoidosis

BACKGROUND: The immune inflammatory process in patients with sarcoidosis is not only compartmentalized within the alveolar walls, but also involves the bronchial airways. Analysis of induced sputum has been used as a non-invasive tool for investigating the airways and may reflect the endobronchial and parenchymal inflammation in patients with sarcoidosis. This present study was designed to measure the soluble pro-inflammatory cytokine levels interleukin-1 (IL-1), interleukin-6 (IL-6), tumuor necrosis factor-alpha (TNF-alpha) and percentage of macrophages expressing these cytokines in induced sputum and bronchoalveolar lavage (BAL) fluid in patients with pulmonary sarcoidosis. METHODS: Sputum induction and BAL was carried out in 27 patients with newly diagnosed sarcoidosis. Control group consisted of six patients with a normal chest radiograph (three patients with carcinoma esophagus and three patients with doubtful history of hemoptysis). Induced sputum was also obtained from 10 non-smoking, non-atopic healthy controls. RESULTS: Percentage of macrophages expressing pro-inflammatory cytokines and soluble cytokine levels in induced sputum were higher in patients with sarcoidosis compared to both groups of controls. There was good correlation between IL-6 and TNF-alpha levels (r = 0.49, 0.58 p < 0.05) and percentage of macrophages expressing all three cytokines (r = 0.56-0.71, p < 0.01) between induced sputum and BAL fluid. Mild positive correlation between cytokine levels in sputum and age was also noted (r = 0.33-0.38, p < 0.05). CONCLUSIONS: Induced sputum may reflect changes in cytokine milieu in BAL in sarcoidosis.     

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-Grünwald-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.     

Association between sputum smear status and local immune responses at the site of disease in HIV-infected patients with pulmonary tuberculosis

Infection with human immunodeficiency virus (HIV) may affect the clinical presentation of pulmonary tuberculosis (TB). To investigate the association between sputum smear status at presentation and local pulmonary immune responses in HIV-infected patients with pulmonary TB, we compared the cellular and cytokine profiles in bronchoalveolar lavage (BAL) fluid obtained from the site of lung disease in 22 sputum smear- and culture-positive, and 17 sputum smear-negative but culture-positive pulmonary TB patients. Smear-positive patients had significantly higher BAL fluid concentrations of IL-6 (p=0.007), IL-8 (p=0.02), IL-10 (p=0.03) and IFN-gamma (p=0.008) than smear-negative patients. No significant differences in the proportions of examined BAL cells were found. We concluded that sputum smear-positive TB was associated with greater pro-inflammatory and immunomodulatory cytokine responses at the site of lung disease than sputum smear-negative disease. The local immune responses may affect the clinical presentation of active pulmonary TB in HIV-infected patients.     

Effect of saliva contamination on induced sputum cell counts, IL-8 and eosinophil cationic protein levels.

Excessive salivary contamination of induced sputum samples prevents the satisfactory examination of lower airway inflammation. The effects of salivary contamination on different sputum fluid phase measures and the levels of salivary contamination preventing analysis are not defined. The present study sought to examine this by investigating the effect of increasing salivary contamination on induced sputum samples. Sputum and saliva samples from subjects with asthma and healthy controls were collected, and treated with dithiothreitol (DTT). Saliva was then added to aliquots of dispersed sputum in increasing proportions (0% to 100%). The effect of increasing saliva contamination was assessed on sputum total cell count, viability, differential cell count and fluid phase levels of interleukin (IL)-8, eosinophil cationic protein (ECP) and total protein. The addition of saliva to induced sputum reduced total cell counts and absolute cell counts but did not change the differential cell count. Levels of fluid phase ECP and IL-8 were significantly reduced with increased salivary contamination. There was a progressive reduction in ECP and IL-8, which reached significance at 70% and 80% saliva contamination, respectively. IL-8 levels corrected for total protein showed no change with increasing saliva concentrations. Induced sputum differential cell counts expressed as the proportion of nonsquamous cells are robust measures that are not influenced by salivary contamination. Studies reporting total and absolute cell counts and fluid phase mediator levels require control for squamous contamination.     

Cellular profiles of induced sputum in children with stable cystic fibrosis: comparison with BAL.

Neutrophil-dominated endobronchial inflammation is a major characteristic of cystic fibrosis (CF) and there is increasing demand for easy-to-perform noninvasive monitoring for prediction and intervention. Fourteen stable paediatric CF patients (8-17 yrs; mean forced expiratory volume in one second 86.7% of the predicted value) were investigated once by fractional bronchoalveolar lavage (BAL) and by sputum induction on three occasions, 2-6 weeks apart. Sputum was induced by consecutive 10-min inhalations of 3, 4 and 5% saline. CF sputum cellular profiles were compared with BAL fluid cell counts and samples from age-matched healthy children, and between different time points to assess reproducibility. Adequate sputum was recovered on >95% of occasions. In all sputum fractions, CF patients showed higher neutrophil counts than healthy children. Neutrophil percentages were highest in the first BAL fraction (median 92%), followed by sputum, in which the percentages decreased in consecutive fractions (72, 66 and 64%), whereas counts were lowest in the pooled BAL fraction (53%). Increasing percentages of macrophages mirrored the decreases in neutrophil percentage. Results of sputum induction at different time points in the CF patients showed good reproducibility and nonoverlap with counts from healthy children. In conclusion, the results of sputum induction in children with mild stable cystic fibrosis adequately describe airway inflammation by providing cellular profiles with lower relative neutrophil counts than in the first ("bronchial") bronchoalveolar lavage fraction and higher relative neutrophil counts than in subsequent pooled ("more peripheral") bronchoalveolar lavage fractions.     

Evidence of mast-cell activation in a subset of patients with eosinophilic chronic obstructive pulmonary disease.

Although asthma has been viewed mainly as an eosinophilic disease, and chronic obstructive pulmonary disease (COPD) as a neutrophilic disease, recent studies have shown increased neutrophil counts in severe asthma and sputum eosinophilia in some COPD patients. In an attempt to further characterise these two syndromes according to pathology, the current authors have conducted a study of induced sputum in 15 subjects with COPD, 17 asthmatics, and 17 nonatopic healthy individuals. Sputum was analysed for cytology and levels of eosinophil cationic protein (ECP), albumin, tryptase and soluble intercellular adhesion molecule-1. The COPD subjects differed from the asthmatics as they had higher sputum neutrophil and lower columnar epithelial cell counts, but there were no differences in any soluble marker studied. When compared to control subjects, both the asthmatic and COPD subjects had raised eosinophil counts and ECP levels. In a subset of COPD subjects with sputum eosinophilia (>3% of total cells), significantly increased levels of tryptase were detected. In conclusion, although chronic obstructive pulmonary disease is a more neutrophilic disease than asthma, the two diseases are difficult to distinguish on the basis of sputum levels of the soluble markers traditionally associated with asthma. However, a subset of patients with chronic obstructive pulmonary disease with airway eosinophilia and mast-cell activation might represent a distinct pathological phenotype.