Mucins 3A and 3B Are Expressed in the Epithelium of Human Large Airway

Abstract

Aberrant mucus secretion is a hallmark of chronic obstructive pulmonary disease (COPD). Expression of the membrane-tethered mucins 3A and 3B (MUC3A, MUC3B) in human lung is largely unknown. In this observational cross-sectional study, we recruited subjects 45-65 years old from the general population of Stockholm, Sweden, during the years 2007-2011. Bronchial mucosal biopsies, bronchial brushings, and bronchoalveolar lavage fluid (BALF) were retrieved from COPD patients (n = 38), healthy never-smokers (n = 40), and smokers with normal lung function (n = 40). Protein expression of MUC3A and MUC3B in bronchial mucosal biopsies was assessed by immunohistochemical staining. In a subgroup of subjects (n = 28), MUC3A and MUC3B mRNAs were quantified in bronchial brushings using microarray. Non-parametric tests were used to perform correlation and group comparison analyses. A value of p < 0.05 was considered statistically significant. MUC3A and MUC3B immunohistochemical expression was localized to ciliated cells. MUC3B was also expressed in basal cells. MUC3A and MUC3B immunohistochemical expression was equal in all study groups but subjects with emphysema had higher MUC3A expression, compared to those without emphysema. Smokers had higher mRNA levels of MUC3A and MUC3B than non-smokers. MUC3A and MUC3B mRNA were higher in male subjects and correlated negatively with expiratory air flows. MUC3B mRNA correlated positively with total cell concentration and macrophage percentage, and negatively with CD4/CD8 T cell ratio in BALF. We concluded that MUC3A and MUC3B in large airways may be a marker of disease or may play a role in the pathophysiology of airway obstruction.

Keywords: COPD; bronchoalveolar lavage; epithelium; immunohistochemistry; large airways; lung function; mRNA; microarray; mucin; smoking.

Figure 1

Representative images of the immunohistochemical stainings for MUC3A and MUC3B proteins in bronchial biopsy samples. Scores of the expression were assessed as negative, faint, moderate, strong, or very strong. Faint expression of MUC3A in the large airways of a smoker with COPD (A) and a strong MUC3A expression in the epithelium of a smoker with normal lung function (B). Faint MUC3B expression in ciliated and basal cells in a never-smoker (C) and a strong expression of MUC3B in epithelial cells in a never-smoker (D). Black arrows show the positive ciliated cells in the epithelium, black/red arrows point to the positive cilia of the cells, red arrows show the positive basal cells, and an asterisk is showing negative goblet cells. Scale bar is 50 µm.

Figure 2

The immunohistochemical expression of MUC3A in ciliated cells (A), MUC3B in basal cells (B), and MUC3B in ciliated cells (C) in the four study groups: never-smokers, current smokers with normal lung function, currently smoking COPD patients, and COPD patients who were ex-smokers. The immunohistochemical expression of MUC3A in ciliated cells in subjects with and without emphysema (D). Correlation between the immunohistochemical expression of MUC3A in ciliated cells and macrophage percentage in BALF (E). Correlation between the immunohistochemical expression of MUC3B in basal cells and the immunohistochemical EGFR expression in basal cells (F). Bars represent mean and standard deviation. Kruskal–Wallis test in (A–C). Mann–Whitney U Test in (D). Spearman correlation analysis in (E,F). R: Spearman’s rank correlation coefficient.

Figure 3

MUC3A mRNA (A) and MUC3B mRNA (B) in samples acquired by bronchial brushing across the study groups. Due to lack of biospecimens, mRNA samples from study subjects belonging to the study group of ex-smokers with COPD were missing. MUC3A mRNA (C) and MUC3B mRNA (D) in samples acquired by bronchial brushing in male and female study subjects. Correlation between MUC3A mRNA and MUC3B mRNA (E). Correlation between exhaled carbon monoxide and MUC3A mRNA (F) and MUC3B mRNA (G). Kruskal–Wallis test in (A,B). Bars represent mean and standard deviation. Mann–Whitney U Test in (C,D). Spearman correlation analysis in (E–G). R: Spearman’s rank correlation coefficient.

Figure 4

Correlation between MUC3B mRNA in samples acquired by bronchial brushing and total cell concentration in BALF (A), macrophage percentage (B), lymphocyte percentage (C), neutrophil percentage (D), and CD4/CD8 ratio in BALF (E). Spearman correlation analysis in all. R: Spearman’s rank correlation coefficient.

Figure 5

Correlation between MUC3A mRNA in samples acquired by bronchial brushing and postbronchodilator forced vital capacity in percent predicted (A). Correlation between MUC3B mRNA in samples acquired by bronchial brushing and postbronchodilator forced expiratory volume in 1 s in percent predicted (B) and residual volume in percent of predicted (C), counted according to ECCS reference equations. Spearman correlation analysis in all. R: Spearman’s rank correlation coefficient.