Smoking-associated increase in mucins 1 and 4 in human airways

Abstract

Rationale: Smoking-related chronic obstructive pulmonary disease (COPD) is associated with dysregulated production of mucus. Mucins (MUC) are important both for mucus secretion and epithelial defense. We have examined the distribution of MUC1 and MUC4 in the airway epithelial cells of never-smokers and smokers with and without COPD.

Methods: Mucosal biopsies and bronchial wash samples were obtained by bronchoscopy from age- and sex-matched COPD-patients (n = 38; GOLD I-II/A-B), healthy never-smokers (n = 40) and current smokers with normal lung function (n = 40) from the Karolinska COSMIC cohort (NCT02627872). Cell-specific expressions of MUC1, MUC4 and regulating factors, i.e., epithelial growth factor receptor (EGFR) 1 and 2, were analyzed by immunohistochemistry. Soluble MUC1 was measured by quantitative immunodetection on slot blot.

Results: The levels of cell-bound MUC1 expression in basal cells and in soluble MUC1 in bronchial wash were increased in smokers, regardless of airway obstruction. Patients with chronic bronchitis had higher MUC1 expression. The expression of MUC4 in cells with goblet cell phenotype was increased in smokers. The expression of EGFR2, but not that of EGFR1, was higher in never-smokers than in smokers.

Conclusions: Smoking history and the presence of chronic bronchitis, regardless of airway obstruction, affect both cellular and soluble MUC1 in human airways. Therefore, MUC1 may be a novel marker for smoking- associated airway disease.

Keywords: Bronchus; COPD; Chronic Bronchitis; Epithelium; Mucin; Smoking.

Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Fig. 1

Representative images of the immunohistochemical stainings for cell-bound MUC1 in bronchial biopsy samples. Intensity of the expression was designated as negative, faint, moderate, strong or very strong, and the extent of the positive staining was estimated from 0 to 100% in each cell type present in the airways. Expression of MUC1 in cells suggesting basal cell phenotype in the large airways of a Never-smoker with normal lung functions (a), a Smoker with normal lung function (b), MUC1 expression in basal cells of an ex-smoker with COPD (c) and a Smoker with COPD (d)

Fig. 2

Immunohistochemical scores for cell-bound MUC1 were calculated based on the intensity and extent of the staining. Scores for immunohistochemical expression of MUC1 in basal cells of large airways from smokers (including COPD) and Never-smokers and from ex-smokers with COPD. MUC1 scores in all subjects (a), and scores separately in males (b) and females (c). Results are shown as mean bars with standard error of mean

Fig. 3

Representative images of the immunohistochemical stainings for MUC4 in bronchial biopsy samples. MUC4 was positive mainly in cells suggesting basal cell and ciliated cell phenotypes, in some cases also in cells with goblet cell phenotype. MUC4 expression in ciliated and basal cells in a Never-smoker (a), a positive expression for MUC4 in the main epithelial cell types in a Smoker (b), an ex-smoker with COPD (c) and a smoker with COPD (d)

Fig. 4

Significant results of the scores on the immunohistochemical expression of MUC4 in goblet cells of large airways. MUC4 scores in the goblet cells of all subjects (a) and in males (b). Results are shown as median bars with standard error of median

Fig. 5

Representative images of the immunohistochemical stainings for EGFR1. Expression of EGFR in cells suggesting basal cell phenotype in the large airways of a Never-smoker with normal lung functions (a), a Smoker with normal lung function (b), MUC1 expression in basal cells of an ex-smoker with COPD (c) and a Smoker with COPD (d). Expression scores of EGFR1 in large airways (e)

Fig. 6

Representative images of the immunohistochemical stainings for EGFR 2. Expression of EGFR2 in cells suggesting basal cell and ciliated cell phenotype in the large airways of a Never-smoker with normal lung functions (a), a Smoker with normal lung function (b), MUC1 expression in basal cells of an ex-smoker with COPD (c) and a Smoker with COPD (d). EGFR2 scores of all subjects in basal cell phenotype (e) and in ciliated cell phenotype (f). Results are shown as median bars and standard error of median

Fig. 7

Soluble MUC1 from basal cells or sub-epithelial glands were measured with quantitative immunoblotting. MUC1 concentration in bronchial wash samples. MUC1 concentration in the whole group (a) and in the females (b). 1 unit equals 0.4 μg/mL. Results are shown as mean and standard error of mean

Fig. 8

Cell-bound and soluble MUC1 as well as MUC4 was studied in large airways of non-smokers and smokers. The non-smoker group includes both never-smokers and COPD ex-smokers whereas the smoker group includes smokers with normal lung function and smokers with COPD. a MUC1 scores in cells suggesting basal cell phenotype from immunohistochemical stainings; b MUC4 scores in cells suggesting goblet cell phenotype from immunohistochemical stainings and c. Amount of MUC1 in bronchial wash samples; 1 unit equals 0.4 μg/mL. MUC1 results are shown as mean bars and standard error of mean, MUC4 results are shown as median bars and standard error of median

Fig. 9

Differences in MUC1 expression levels in patients with or without chronic bronchitis. a Scores of cell-bound MUC1 expression in large airways and b. Amount of MUC1 in bronchial wash samples from the patients; 1 unit equals 0.4 μg/mL. MUC1 results are shown as mean bars and standard error, MUC1 results are shown as median bars and standard error of median