Rapid Proteome Analysis of Bronchoalveolar Lavage Samples of Lifelong Smokers and Never-Smokers by Micro-Scale Liquid Chromatography and Mass Spectrometry

Abstract Background: The aim of this study was to determine whether relative qualitative and quantitative differences in protein expression could be related to smoke exposure or smoke-induced airway inflammation. We therefore explored and characterized the protein components found in bronchoalveolar lavage (BAL) fluid sampled from either lifelong smokers or never-smokers. Methods: BAL fluid samples obtained by bronchoscopy from 60-year-old healthy never-smokers (n = 18) and asymptomatic smokers (n = 30) were analyzed in either pooled or individual form. Initial global proteomic analysis used shotgun digestion approaches on unfractionated BAL fluid samples (after minimal sample preparation) and separation of peptides by gradient (90-min) liquid chromatography (LC) coupled with on-line linear ion trap quadropole mass spectrometry (LTQ MS) for identification and analysis. Results: LTQ MS identified 481 high- to low-abundance proteins. Relative differences in patterns of BAL fluid proteins in smokers compared with never-smokers were observed in pooled and individual samples as well as by 2-dimensional gel analysis. Gene ontology categorization of all annotated proteins showed a wide spectrum of molecular functions and biological processes. Conclusions: The described method provides comprehensive qualitative proteomic analysis of BAL fluid protein expression from never-smokers and from smokers at risk of developing chronic obstructive pulmonary disease. Many of the proteins identified had not been detected in previous studies of BAL fluid; thus, the use of LC-tandem MS with LTQ may provide new information regarding potentially important patterns of protein expression associated with lifelong smoking.

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Protein Expression Patterns Associated with Progression of Chronic Obstructive Pulmonary Disease in Bronchoalveolar Lavage of Smokers

Clinical Chemistry ◽

10.1373/clinchem.2006.076075 ◽

2007 ◽

Vol 53 (4) ◽

pp. 636-644 ◽

Cited By ~ 30

Author(s):

Amelie Plymoth ◽

Claes-Göran Löfdahl ◽

Ann Ekberg-Jansson ◽

Magnus Dahlbäck ◽

Per Broberg ◽

...

Keyword(s):

Chronic Obstructive Pulmonary Disease ◽

Protein Expression ◽

Bronchoalveolar Lavage ◽

Pulmonary Disease ◽

Disease Stage ◽

Chronic Obstructive ◽

Obstructive Pulmonary Disease ◽

Mass Spectrometry Identification ◽

Never Smokers

Abstract Background: We modeled the expression of proteins in baseline bronchoalveolar lavage (BAL) samples from asymptomatic 60-year-old lifelong current smokers or healthy never-smokers, who were reevaluated after 6 to 7 years to record clinical outcome. Methods: Applying a technology toolbox consisting of replicate 2-dimensional gel separations, image annotation, and mass spectrometry identification, we catalogued a global set of proteins that were differentially expressed in individuals by presence, absence, and intensity scores. Results: By use of multivariate analysis, we selected a subset of proteins that accurately separated smokers from never-smokers based on composite scoring. Follow-up after 6 to 7 years identified a group of individuals who had progressed to chronic obstructive pulmonary disease (COPD), Global Initiative for Chronic Obstructive Lung Disease stage 2. The baseline BAL samples of these eventual COPD patients shared a distinct protein expression profile that could be identified using partial least-squares discriminant analysis. This pattern was not observed in BAL samples of asymptomatic smokers free of COPD at 6- to 7-year follow-up. Conclusions: Our model suggests that certain patterns of protein expression occurring in the airways of long-term smokers may be detected in smokers susceptible to a progression of COPD disease, before disease is clinically evident.

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