Correlation of Apical Fluid-Regulating Channel Proteins with Lung Function in Human COPD Lungs

et al. (2014) Correlation of Apical Fluid-Regulating Channel Proteins with Lung Function in Human COPD Lungs. PLoS ONE 9(10): e109725. doi:10.1371/journal.pone.0109725 Correlation of Apical Fluid-Regulating Channel Proteins with Lung Function in Human COPD Lungs Runzhen Zhao 0 1 Xinrong Liang 0 1 Meimi Zhao 0 1 Shan-Lu Liu 0 1 Yao Huang 0 1 Steven Idell 0 1 Xiumin Li 0 1 Hong-Long Ji 0 1 Estelle Cormet-Boyaka, The Ohio State University, United States of America 0 Current address: School of Pharmacy, China Medical University , Liaoning Shenyang , China 1 1 Department of Cellular and Molecular Biology, University of Texas Health Science Center at Tyler, Tyler, Texas, United States of America, 2 Medicine, University of Texas Health Science Center at Tyler, Tyler, Texas, United States of America, 3 Texas Lung Injury Institute, University of Texas Health Science Center at Tyler, Tyler, Texas, United States of America, 4 Department of Molecular Microbiology and Immunology, Bond Life Sciences Center, University of Missouri , Columbia , Missouri, United States of America, 5 Department of Obstetrics and Gynecology, St. Joseph's Hospital and Medical Center, Phoenix, Arizona, United States of America, 6 Xinxiang Medical University , Xinxiang, Henan , China Links between epithelial ion channels and chronic obstructive pulmonary diseases (COPD) are emerging through animal model and in vitro studies. However, clinical correlations between fluid-regulating channel proteins and lung function in COPD remain to be elucidated. To quantitatively measure epithelial sodium channels (ENaC), cystic fibrosis transmembrane conductance regulator (CFTR), and aquaporin 5 (AQP5) proteins in human COPD lungs and to analyze the correlation with declining lung function, quantitative western blots were used. Spearman tests were performed to identify correlations between channel proteins and lung function. The expression of a and b ENaC subunits was augmented and inversely associated with lung function. In contrast, both total and alveolar type I (ATI) and II (ATII)-specific CFTR proteins were reduced. The expression level of CFTR proteins was associated with FEV1 positively. Abundance of AQP5 proteins and extracellular superoxide dismutase (SOD3) was decreased and correlated with spirometry test results and gas exchange positively. Furthermore, these channel proteins were significantly associated with severity of disease. Our study demonstrates that expression of ENaC, AQP5, and CFTR proteins in human COPD lungs is quantitatively associated with lung function and severity of COPD. These apically located fluid-regulating channels may thereby serve as biomarkers and potent druggable targets of COPD. - Funding: This work was supported by National Institutes of Health grants HL87017 and HL095435. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: Hong-Long Ji is currently serving as an academic editor. This does not alter the authors adherence to PLOS ONE Editorial policies and criteria. . These authors contributed equally to this work. Mucus hypersecretive COPD is the fourth and will be the third leading cause of death by 2020 worldwide [13]. As a widely acknowledged heterogeneous disease, COPD encompasses small airway disease, emphysema, and chronic obstructive bronchitis. Notwithstanding the complexity, genetic studies have associated several genes with COPD, including SOD3, GSTM1, TGFB1, TNF, GSTP1, etc [46]. Defective mucin hydration has been confirmed in the airways of cystic fibrosis sufferers. Abnormal bioelectric properties including hyperactive ENaC activity and deficient cystic fibrosis transmembrane conductance regulator (CFTR) are well-known in cystic fibrosis lungs [7,8]. CFTR interactively regulates other proteins activities [911]. These epithelial channel proteins, together with aquaporins and Na+-K+-ATPases finely adjust luminal surface fluid in the airways and air spaces [12,13]. Mall and co-workers successfully established a COPD-like mouse strain by genetically over-expressing b ENaC in airway epithelia [1417]. These transgenic mice exhibit dehydrated airways and severe mucus obstruction in the trachea. In addition, the authors noted goblet cell hyperplasia and neutrophilic inflammation, commonly observed in COPD [14,18,19]. In surviving mice, increased mucus concentration and delayed mucus transport in the conducting airways were observed [14]. Clinical pathologic examination revealed that chronic mucus obstruction was found in human distal airways and lungs, accompanied by goblet cell metaplasia, increased mucin expression, persistent neutrophilic airway inflammation, and transient eosinophilic airway filtration [20]. Furthermore, mice over-expressing multiple ENaC subunits developed emphysema with increased lung volumes, distal airspace enlargement, and decreased lung compliance [14,18]. The correlations between ENaCs and lung function in human COPD, however, are not yet known. Given the interregulation of ENaC, CFTR, and aquaporins, we aimed to quantitatively analyze the expression of fluid-regulating ion channel proteins, including ENaC, CFTR, and AQP5 in human COPD lungs. We then sought to correlate their expression levels with declined lung function. Our novel results demonstrate that the expression of these channel proteins in human COPD lungs is significantly associated with declined spirometry test, gas exchange, and severity of COPD. Materials and Methods Human lung tissues and clinical data The use of lung tissues curated by the Lung Tissue Research Consortium (LTRC), National Institute of Health has been approved by the Institutional Review Board (EXEMPT 07013) of the University of Texas Health Science Center at Tyler. Written informed consents from the donor or the next of kin were obtained for use of these samples in research. Frozen human lung tissue blocks for protein extractions, fixed human lung slices for histology, and anonymous lung function data from COPD patients and healthy individuals were provided by the LTRC. Of the thirty four smokers, twenty-four were diagnosed with moderate to severe COPD/emphysema, and ten healthy subjects were controls according to FEV1 pre-bronchodilator % predicted values (FEV1pd1a): control $80%, moderate 5080%, and severe , 50%. The patients demographics, spirometric results, and gas diffusion capacity are summarized in Table 1. Questionnaires All subjects completed standardized questionnaires to assess respiratory history and symptoms, respiratory medicines, smoking history, family history and other medical histories. The LTRC database curated by four clinical centers (The Mayo Clinic chaired by Dr. Andrew H. Limper, Temple University by Dr. Gerard J. Criner, University of Michigan by Dr. Fernando J. Martinez, and University of Pittsburgh by Dr. Frank C. Sciurba) includes data from CT scans, chest X-rays, tissue types, and b (...truncated)