Sputum proteomics identifies elevated PIGR levels in smokers and mild-to-moderate COPD.

Chronic obstructive pulmonary disease (COPD) is one of the leading causes of morbidity and mortality around the world. However, the exact mechanisms leading to COPD and its progression are still poorly understood. In this study, induced sputum was analyzed by cysteine-specific two-dimensional difference gel electrophoresis (2D-DIGE) coupled with mass spectrometry to identify proteins involved in COPD pathogenesis. The comparison of nonsmokers, smokers, and smokers with moderate COPD revealed 15 changed proteins with the majority, including polymeric immunoglobulin receptor (PIGR), being elevated in smokers and subjects with COPD. PIGR, which is involved in specific immune defense and inflammation, was further studied in sputum, lung tissue, and plasma by Western blot, immunohistochemistry/image analysis, and/or ELISA. Sputum PIGR was characterized as glycosylated secretory component (SC). Lung PIGR was significantly elevated in the bronchial and alveolar epithelium of smokers and further increased in the alveolar area in mild to moderate COPD. Plasma PIGR was elevated in smokers and smokers with COPD compared to nonsmokers with significant correlation to obstruction. In conclusion, new proteins in smoking-related chronic inflammation and COPD could be identified, with SC/PIGR being one of the most prominent not only in the lung but also in circulating blood.

Ageing and smoking contribute to plasma surfactant proteins and protease imbalance with correlations to airway obstruction.

BACKGROUND: A significant number of young people start smoking at an age of 13-15, which means that serious smoking-evoked changes may have been occurred by their twenties. Surfactant proteins (SP) and matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) have been linked to cigarette smoke induced lung remodelling and chronic obstructive pulmonary disease (COPD). However, the level of these proteins has not been examined during ageing or in young individuals with short smoking histories. METHODS: Plasma levels of SP-A, SP-D, MMP-9, and TIMP-1 were measured by EIA/ELISA from young (18-23 years) non-smoking controls (YNS) (n = 36), smokers (YS) (n = 51), middle aged/elderly (37-77 years) non-smoking controls (ONS) (n = 40), smokers (OS) (n = 64) (FEV1/FVC >0.7 in all subjects) and patients with COPD (n = 44, 35-79 years). RESULTS: Plasma levels of SP-A increased with age and in the older group in relation to smoking and COPD. Plasma SP-D and MMP-9 levels did not change with age but were elevated in OS and COPD as compared to ONS. The TIMP-1 level declined with age but increased in chronic smokers when compared to ONS. The clearest correlations could be detected between plasma SP-A vs. age, pack years and FEV1/FVC. The receiver operating characteristic (ROC) curve analysis revealed SP-A to be the best marker for discriminating between patients with COPD and the controls (area under ROC curve of 0.842; 95% confidence interval, 0.785-0.899; p < 0.001). CONCLUSIONS: Age has a significant contribution to potential markers related to smoking and COPD; SP-A seems to be the best factor in differentiating COPD from the controls.

Elevation of sputum matrix metalloproteinase-9 persists up to 6 months after smoking cessation: a research study.

BACKGROUND: Smoking cessation is the best possible way to prevent the progression of smoking related airway diseases. However, the effect and time scale of smoking cessation on airway inflammation/remodelling are largely unknown. This prospective study evaluated the effects of smoking cessation on induced sputum (IS) neutrophils, matrix metalloproteinases (MMP-7, -8, -9) and tissue inhibitor of metalloproteinase-1 (TIMP-1). METHODS: A total of 61 subjects participated in the study; 17 stopped smoking for 3 months and 9 for 6 months. The proportion of IS neutrophils and the levels of MMPs and TIMP-1 by ELISA were determined at baseline and at 3 and 6 months after cessation. RESULTS: In the smokers, baseline IS neutrophils, MMPs and TIMP-1 were significantly higher compared to non-smokers. Levels of MMP-7, -8 and TIMP-1 decreased nearly to those of non-smokers but the levels of MMP-9 increased significantly from the baseline of the same subjects at 3 months after cessation (p = 0.009) with no significant decline at 6 months after cessation. CONCLUSIONS: Sputum MMP-9 remained elevated after 6 months of smoking cessation, which may contribute to ongoing lung damage typical of COPD.

Persistence of oxidant and protease burden in the airways after smoking cessation.

BACKGROUND: Oxidative stress is associated with the pathogenesis of cigarette smoke related lung diseases, but longitudinal effects of smoking cessation on oxidant markers in the airways are unknown. METHODS: This study included 61 smokers; 21 with chronic bronchitis or COPD, 15 asthmatics and 25 asymptomatic smokers followed up for 3 months after smoking cessation. Fractional exhaled nitric oxide (FeNO), sputum neutrophil counts, sputum 8-isoprostane, nitrotyrosine and matrix metalloproteinase-8 (MMP-8) were investigated at baseline and 1 and 3 months after smoking cessation. RESULTS: After 3 months 15 subjects had succeeded in quitting of smoking and in these subjects symptoms improved significantly. Unexpectedly, however, sputum neutrophils increased (p = 0.046) after smoking cessation in patients with chronic bronchitis/COPD. At baseline, the other markers did not differ between the three groups so these results were combined for further analysis. Sputum 8-isoprostane declined significantly during the follow-up at 3 months (p = 0.035), but levels still remained significantly higher than in non-smokers. The levels of FeNO, nitrotyrosine and MMP-8 did not change significantly during the 3 months after smoking cessation. CONCLUSION: Whilst symptoms improve after smoking cessation, the oxidant and protease burden in the airways continues for months.

The value of sputum 8-isoprostane in detecting oxidative stress in mild asthma.

BACKGROUND: Exhaled nitric oxide and induced sputum eosinophils are well established as direct markers of inflammation/oxidative stress in asthma. Recently, it has been proposed that sputum 8-isoprostane concentrations may present a reliable index for measuring oxidative stress in asthmatic patients. We assessed the value of sputum 8-isoprostane in mild asthma in children and adolescents. METHODS: Patients with newly diagnosed asthma (children, n = 23; adults, n = 14) and age-matched healthy controls (children, n = 13; adults, n = 15) were studied. Lung function was measured by spirometry, sputum was induced by hypertonic saline, and fractional exhaled nitric oxide (FeNO) was measured with standard methods. Cell differential counts were obtained from sputum slides and the concentration of 8-isoprostane was measured with an enzyme immunoassay from sputum supernatants. RESULTS: High-quality sputum specimens could be obtained from 10 children and 10 adults, and the sputum analyses were conducted only for the representative specimens. Asthmatics had increased FeNO (children 35.5 vs. 11.9 ppb; adults 81.1 vs. 16.6 ppb; p < 0.001) and sputum eosinophils (children 2.4% vs. 1.4%; adults 10.4% vs. 0.2%; p = 0.005) compared to healthy controls. There was a significant correlation between FeNO and eosinophils (R = 0.65; p < 0.0001). Sputum 8-isoprostane was not elevated in asthmatics compared to healthy subjects (children 81.1 vs. 89.9 and adults 76.9 vs. 73.4 pg/mL) and did not correlate with lung function or other measurements of airway inflammation. However, increased 8-isoprostane levels were detected in patients with chronic obstructive pulmonary disease (n = 11, 184.7 pg/mL, used as controls for assays). CONCLUSIONS: In agreement with earlier studies, FeNo is sensitive in detecting oxidative/nitrosative stress in asthmatic airways. However, our results suggest that 8-isoprostane may not be sensitive in reflecting oxidant burden in mild asthma.

Airway biomarkers of the oxidant burden in asthma and chronic obstructive pulmonary disease: current and future perspectives.

The pathogenesis of asthma and chronic obstructive pulmonary disease (COPD) has been claimed to be attributable to increased systemic and local oxidative stress. Detection of the oxidant burden and evaluation of their progression and phenotypes by oxidant biomarkers have proved challenging and difficult. A large number of asthmatics are cigarette smokers and smoke itself contains oxidants complicating further the use of oxidant biomarkers. One of the most widely used oxidant markers in asthma is exhaled nitric oxide (NO), which plays an important role in the pathogenesis of asthma and disease monitoring. Another oxidant marker that has been widely investigated in COPD is 8-isoprostane, but it is probably not capable of differentiating asthma from COPD, or even sensitive in the early assessment of these diseases. None of the current biomarkers have been shown to be better than exhaled NO in asthma. There is a need to identify new biomarkers for obstructive airway diseases, especially their differential diagnosis. A comprehensive evaluation of oxidant markers and their combinations will be presented in this review. In brief, it seems that additional analyses utilizing powerful tools such as genomics, metabolomics, lipidomics, and proteomics will be required to improve the specificity and sensitivity of the next generation of biomarkers.