Human neutrophil elastase (HNE) dose-dependently increases the load of nontypeable <i>Haemophilus influenzae</i> (NTHi) in cultured human airway epithelial cells.

<p>Primary normal human tracheobronchial epithelial cells (N = 3) were differentiated into the mucociliary phenotype under air-liquid interface culture for 10 days, and then infected with NTHi for 48 hrs. (A) – NTHi in apical supernatants; (B) – NTHi associated with epithelial cells. Data are expressed as means ± SEM.</p

SPLUNC1 protein is decreased in the lungs of patients with chronic obstructive pulmonary disease (COPD).

<p>SPLUNC1 protein was measured in bronchoalveolar lavage (BAL) fluid of healthy non-smokers (n = 5), healthy smokers (n = 9) and COPD smokers (n = 4). The horizontal solid red lines indicate medians.</p

Human neutrophil elastase (HNE) decreases SPLUNC1 protein in cultured human airway epithelial cells.

<p>Primary normal human tracheobronchial epithelial cells (N = 3) were differentiated into the mucociliary phenotype under air-liquid interface culture for 10 days. After 4 (<b>A</b>) and 48 (<b>B</b>) hrs of HNE treatment, SPLUNC1 protein in apical supernatants was measured by an ELISA. Data are expressed as means ± SEM.</p

SPLUNC1 knockout (SPLUNC1^−/−) mice increase the load of nontypeable <i>Haemophilus influenzae</i> (NTHi) in the lung.

<p>SPLUNC1^−/− and wild-type mice (n = 7 or 8 mice/group) were intranasally inoculated with NTHi (10⁵ CFUs/mouse). After 24 hrs of infection, left lungs were homogenized to quantify NTHi load by culture. The horizontal lines represent means of CFUs.</p

Degradation of SPLUNC1 protein by human neutrophil elastase (HNE).

<p>(<b>A</b>) Western blot of recombinant human SPLUNC1 protein (10 µg/ml) that was incubated with HNE for 30 minutes. HNE treatment dose-dependently reduces the native SPLUNC1 protein (25 kD). A fragment of 22-kD SPLUNC1 was seen in HNE-treated samples. (<b>B</b>) Mass spectrometry analysis of NHE-treated recombinant human SPLUNC1 protein revealed a 25 amino acid peptide F21-T45 (FGGLPVPLDQTLPLNVNPALPLSPT).</p

Recombinant human SPLUNC1 protein significantly reduces the load of nontypeable <i>Haemophilus influenzae</i> (NTHi) in apical supernatants of human neutrophil elastase (HNE)-treated human airway epithelial cells.

<p>Primary normal human tracheobronchial epithelial cells (N = 5) were differentiated into the mucociliary phenotype under air-liquid interface culture for 10 days, and then infected with NTHi (10³ CFU/transwell) in the presence or absence of HNE (10 µg/ml). After 2 hrs of NTHi and HNE treatment, recombinant human SPLUNC1 protein (10 µg/ml) was added to the apical surface of epithelial cells. After 46 hrs of SPLUNC1 treatment or after 48 hrs of NTHi infection, apical supernatants and cells were harvested for NTHi quantification. (A) – NTHi in apical supernatants; (B) – NTHi associated with epithelial cells. Each connected color line represents the data collected from cells of an individual donor. The horizontal dotted red lines indicate medians of NTHi fold changes (HNE treatment versus vehicle solution control).</p

Time course study of the rescuing effects of recombinant human SPLUNC1 protein on SPLUNC1 protein levels in human neutrophil elastase (HNE)-treated human airway epithelial cells.

<p>Primary normal human tracheobronchial epithelial cells (N = 3) were differentiated into the mucociliary phenotype under air-liquid interface culture for 10 days, and then treated with HNE (10 µg/ml) in the presence or absence of recombinant human SPLUNC1 protein (10 µg/ml). At indicated time points, SPLUNC1 protein in the apical supernatants was measured by using an ELISA. Data are expressed as means ± SEM.</p

Characteristics of human subjects.

<p><b>HNS</b> = Healthy non-smokers; <b>HS</b> = Healthy smokers; <b>COPD</b> = Chronic obstructive pulmonary diseases; <b>FEV₁</b> =  forced expiratory volume in the 1^st second; <b>FVC</b> = forced vital capacity.</p

Expression of asthma susceptibility genes in bronchial epithelial cells and bronchial alveolar lavage in the Severe Asthma Research Program (SARP) cohort

<p><i>Objective</i>: Genome-wide association studies (GWASs) have identified genes associated with asthma, however expression of these genes in asthma-relevant tissues has not been studied. This study tested expression and correlation between GWAS-identified asthma genes and asthma or asthma severity. <i>Methods</i>: Correlation analyses of expression levels of GWAS-identified asthma genes and asthma-related biomarkers were performed in cells from human bronchial epithelial biopsy (BEC, <i>n</i> = 107) and bronchial alveolar lavage (BAL, <i>n</i> = 94). <i>Results</i>: Expression levels of asthma genes between BEC and BAL and with asthma or asthma severity were weakly correlated. The expression levels of <i>IL18R1</i> were consistently higher in asthma than controls or in severe asthma than mild/moderate asthma in BEC and BAL (<i>p</i> < 0.05). In <i>RAD50-IL13</i> region, the expression levels of <i>RAD50</i>, not <i>IL4, IL5</i>, or <i>IL13</i>, were positively correlated between BEC and BAL (ρ = 0.53, <i>P</i> = 4.5 × 10⁻⁶). The expression levels of <i>IL13</i> were positively correlated with <i>IL5</i> in BEC (ρ = 0.35, <i>P</i> = 1.9 × 10⁻⁴) and <i>IL4</i> in BAL (ρ = 0.42, <i>P</i> = 2.5 × 10⁻⁵), respectively. rs3798134 in <i>RAD50</i>, a GWAS-identified SNP, was correlated with <i>IL13</i> expression and the expression levels of <i>IL13</i> were correlated with asthma (<i>P</i> = 0.03). rs17772583 in <i>RAD50</i> was significantly correlated with <i>RAD50</i> expression in BAL and BEC (<i>P</i> = 7.4 × 10⁻⁷ and 0.04) but was not associated with asthma. <i>Conclusions</i>: This is the first report studying the expression of GWAS-identified asthma genes in BEC and BAL. <i>IL13</i>, rather than <i>RAD50, IL4</i>, or <i>IL5</i>, is more likely to be the asthma susceptibility gene. Our study illustrates tissue-specific expression of asthma-related genes. Therefore, whenever possible, disease-relevant tissues should be used for transcription analysis.</p

Density plots of the distribution of asthma hospitalizations stratified by gender and race.

<p>Panel A shows that asthma hospitalization is more frequent among young boys and middle age women. Panel B shows a bi-modal distribution of asthma severity across different races. Panel A and B are abstracted from NIS 2012.</p