Delay of airway epithelial wound repair in COPD is associated with airflow obstruction severity

Abstract

Background: Airway epithelium integrity is essential to maintain its role of mechanical and functional barrier. Recurrent epithelial injuries require a complex mechanism of repair to restore its integrity. In chronic obstructive pulmonary disease (COPD), an abnormal airway epithelial repair may participate in airway remodeling. The objective was to determine if airway epithelial wound repair of airway epithelium is abnormal in COPD.

Methods: Patients scheduled for lung resection were prospectively recruited. Demographic, clinical data and pulmonary function tests results were recorded. Emphysema was visually scored and histological remodeling features were noted. Primary bronchial epithelial cells (BEC) were extracted and cultured for wound closure assay. We determined the mean speed of wound closure (MSWC) and cell proliferation index, matrix metalloprotease (MMP)-2, MMP-9 and cytokines levels in supernatants of BEC 18 hours after cell wounding. In a subset of patients, bronchiolar epithelial cells were also cultured for wound closure assay for MSWC analyze.

Results: 13 COPD and 7 non COPD patients were included. The severity of airflow obstruction and the severity of emphysema were associated with a lower MSWC in BEC (p = 0.01, 95% CI [0.15-0.80]; p = 0.04, 95% CI [-0.77;-0.03] respectively). Cell proliferation index was decreased in COPD patients (19 ± 6% in COPD vs 27 ± 3% in non COPD, p = 0.04). The severity of COPD was associated with a lower level of MMP-2 (7.8 ± 2 10(5) AU in COPD GOLD D vs 12.8 ± 0.13 10(5) AU in COPD GOLD A, p = 0.04) and a lower level of IL-4 (p = 0.03, 95% CI [0.09;0.87]). Moreover, higher levels of IL-4 and IL-2 were associated with a higher MSWC (p = 0.01, 95% CI [0.17;0.89] and p = 0.02, 95% CI [0.09;0.87] respectively). Clinical characteristics and smoking history were not associated with MSWC, cell proliferation index or MMP and cytokines levels. Finally, we showed an association of the MSWC of bronchial and corresponding bronchiolar epithelial cells obtained from the same patients (p = 0.02, 95% CI [0.12;0.89]).

Conclusion: Our results showed an abnormal bronchial epithelial wound closure process in severe COPD. Further studies are needed to elucidate the contribution and the regulation of this mechanism in the complex pathophysiology of COPD.

Figures

Figure 1

Wound closure assay in bronchial epithelial cells from COPD and non COPD patients. A mechanical injury was performed on confluent bronchial epithelial cell monolayer. Representative photographs at 0, 6, 12 and 18 h in a non COPD and a COPD GOLD D spirometric GOLD 3 patient (A). The mean percentage of remaining wound area in non COPD patients (n = 7), spirometric COPD GOLD 1 (n = 5), GOLD 2 (n = 5) and GOLD 3–4 patients (n = 3) are presented (B). *p < 0.05 vs non COPD.

Figure 2

Relationships between mean speed of bronchial epithelial wound closure and FEV1. Bronchial epithelial wound closure in non COPD (triangle, n = 7) and COPD patients (square, n = 13) was monitored for 18 h, and the mean speed of wound closure (MSWC) was calculated. p = 0.01.

Figure 3

Cell proliferation during epithelial wound closure in non COPD and COPD patients. Representative photographs (x10 enlargement) of Ki-67 and DAPI staining, as well as merge images in non COPD and COPD bronchial epithelial cells after 18 h of repair (A). Quantification of the number of Ki-67 positive cells expressed as a percentage of total cell number at 0 h, 6 h, 12 h and 18 h of repair (B).

Figure 4

Association between the mean speed of bronchiolar and bronchial epithelial wound closure. Paired bronchial and bronchiolar epithelial cells were obtained in 12 patients and analysed in wound closure assay. Representative photographs at 0 and 18 h in a non COPD and a COPD GOLD D spirometric GOLD 3 patient (A) are presented. The association between the mean speed of wound closure of bronchiolar epithelial cells and corresponding bronchial epithelial cells is shown (B). p = 0.02.