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Large-scale, ion-current-based proteomics investigation of bronchoalveolar lavage fluid in chronic obstructive pulmonary disease patients.

ABSTRACT: Proteomic analysis of bronchoalveolar lavage fluid (BALF) in chronic obstructive pulmonary disease (COPD) patients may provide new biomarkers and deeper understanding of the disease mechanisms but remains challenging. Here we describe an ion-current-based strategy for comparative analysis of BALF proteomes from patients with moderate and stable COPD versus healthy controls. The strategy includes an efficient preparation procedure providing quantitative recovery and a nano-LC/MS analysis with a long, heated column. Under optimized conditions, high efficiency and reproducibility were achieved for each step, enabling a "20-plex" comparison of clinical subjects (n = 10/group). Without depletion/fractionation, a total of 423 unique protein groups were quantified under stringent criteria with at least two quantifiable peptides. Seventy-six proteins were determined as significantly altered in COPD, which represent a diversity of biological processes such as alcohol metabolic process, gluconeogenesis/glycolysis, inflammatory response, proteolysis, and oxidation reduction. Interestingly, altered alcohol metabolism responding to oxidant stress is a novel observation in COPD. The prominently elevated key enzymes involved in alcohol metabolism (e.g., ADH1B, ALDH2, and ALDH3A1) may provide a reasonable explanation for a bewildering observation in COPD patients known for decades: the underestimation of the blood alcohol concentrations through breath tests. These discoveries could provide new insights for identifying novel biomarkers and pathological mediators in clinical studies.

SUBMITTER: Tu C

PROVIDER: S-EPMC4073647 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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Large-scale, ion-current-based proteomics investigation of bronchoalveolar lavage fluid in chronic obstructive pulmonary disease patients.

Tu Chengjian C Mammen Manoj Jacob MJ Li Jun J Shen Xiaomeng X Jiang Xiaosheng X Hu Qiang Q Wang Jianmin J Sethi Sanjay S Qu Jun J

Journal of proteome research 20131202 2

Proteomic analysis of bronchoalveolar lavage fluid (BALF) in chronic obstructive pulmonary disease (COPD) patients may provide new biomarkers and deeper understanding of the disease mechanisms but remains challenging. Here we describe an ion-current-based strategy for comparative analysis of BALF proteomes from patients with moderate and stable COPD versus healthy controls. The strategy includes an efficient preparation procedure providing quantitative recovery and a nano-LC/MS analysis with a l ...[more]

PMID: 24188068

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Project description:Micro RNAs (MiRNAs) act as a key regulator participating in various biological process, and the roles of that play in chronic obstructive pulmonary disease (COPD) are discovered. However, recent pharmacological treatment for COPD focus on alleviating symptoms and reducing the risk events. The heterogeneous COPD causes variable responses to pharmacological interventions. COPD treatment has gradually developed into precision medicine, integrating clinical and biomarker information to optimize personalized therapy. Thus, targeting miRNAs represents a promising strategy for COPD individual therapy. Twelve COPD patients, 7 community-acquired pneumonia and 4 normal people were recruited. Total RNAs were collected from the bronch alveolar lavage cells and peripheral blood plasma of each participant. miRNAs were profiled by microarray and systematically compared between patients with different groups. Bioinformatic analysis identified pathways relevant to the pathogenesis of COPD. Next, the target pathway networks were mapped. Compared different groups, we obtain differential expression of miRNAs (Q value (Adjusted P value) < .05 and |log2FC| >2). Gene ontology enrichment analyses showed that differentially expressed miRNAs function as regulators in different modules of cellular component, molecular function and biological process. Kyoto Encyclopedia of Genes and Genomes enrichment analyses suggested that signals, such as MAPK signaling pathway, Ras signaling pathway, FoxO signaling pathway and oxidative stress may participate in the pathogenesis of COPD. In the miRNAs target pathway networks, novel-hsa-miR26-3p or hsa-miR-3529-3p/CDC42/MAPK signaling pathway may play a role in regulating COPD. Our findings demonstrate critical roles of the miRNAs in COPD molecular pathology. The data support a plausible mechanism that miRNAs may be involved in the development of COPD by affecting the inflammatory and oxidative stress. Moreover, hsa-miR-4748/CDC42/MAPK signaling pathway may contribute to the pathogenesis of COPD, providing a potential novel therapeutic strategy in COPD.

Project description:BackgroundIt is now possible to comprehensively characterize the microbiota of the lungs using culture-independent, sequencing-based assays. Several sample types have been used to investigate the lung microbiota, each presenting specific challenges for preparation and analysis of microbial communities. Bronchoalveolar lavage fluid (BALF) enables the identification of microbiota specific to the lower lung but commonly has low bacterial density, increasing the risk of false-positive signal from contaminating DNA. The objectives of this study were to investigate the extent of contamination across a range of sample densities representative of BALF and identify features of contaminants that facilitate their removal from sequence data and aid in the interpretation of BALF sample 16S sequencing data.ResultsUsing three mock communities across a range of densities ranging from 8E+?02 to 8E+?09 16S copies/ml, we assessed taxonomic accuracy and precision by 16S rRNA gene sequencing and the proportion of reads arising from contaminants. Sequencing accuracy, precision, and the relative abundance of mock community members decreased with sample input density, with a significant drop-off below 8E+?05 16S copies/ml. Contaminant OTUs were commonly inversely correlated with sample input density or not reproduced between technical replicates. Removal of taxa with these features or physical concentration of samples prior to sequencing improved both sequencing accuracy and precision for samples between 8E+?04 and 8E+?06 16S copies/ml. For the lowest densities, below 8E+?03 16S copies/ml BALF, accuracy and precision could not be significantly improved using these approaches. Using clinical BALF samples across a large density range, we observed that OTUs with features of contaminants identified in mock communities were also evident in low-density BALF samples.ConclusionRelative abundance data and community composition generated by 16S sequencing of BALF samples across the range of density commonly observed in this sample type should be interpreted in the context of input sample density and may be improved by simple pre- and post-sequencing steps for densities above 8E+?04 16S copies/ml.

Dataset Information

Large-scale, ion-current-based proteomics investigation of bronchoalveolar lavage fluid in chronic obstructive pulmonary disease patients.

Publications

Large-scale, ion-current-based proteomics investigation of bronchoalveolar lavage fluid in chronic obstructive pulmonary disease patients.

Similar Datasets

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