IL-25 drives remodelling in allergic airways disease induced by house dust mite

Abstract

Background: Overexpression of the transforming growth factor β family signalling molecule smad2 in the airway epithelium provokes enhanced allergen-induced airway remodelling in mice, concomitant with elevated levels of interleukin (IL)-25.

Objective: We investigated whether IL-25 plays an active role in driving this airway remodelling.

Methods: Anti-IL-25 antibody was given to mice exposed to either inhaled house dust mite (HDM) alone, or in conjunction with an adenoviral smad2 vector which promotes an enhanced remodelling phenotype.

Results: Blocking IL-25 in allergen-exposed mice resulted in a moderate reduction in pulmonary eosinophilia and levels of T helper type 2 associated cytokines, IL-5 and IL-13. In addition, IL-25 neutralisation abrogated peribronchial collagen deposition, airway smooth muscle hyperplasia and airway hyperreactivity in control mice exposed to HDM and smad2-overexpressing mice. IL-25 was shown to act directly on human fibroblasts to induce collagen secretion. Recruitment of endothelial progenitor cells to the lung and subsequent neovascularisation was also IL-25 dependent, demonstrating a direct role for IL-25 during angiogenesis in vivo. Moreover, the secretion of innate epithelial derived cytokines IL-33 and thymic stromal lymphopoietin (TSLP) was completely ablated.

Conclusions: In addition to modulating acute inflammation, we now demonstrate a role for IL-25 in orchestrating airway remodelling. IL-25 also drives IL-33 and TSLP production in the lung. These data delineate a wider role for IL-25 in mediating structural changes to the lung following allergen exposure and implicate IL-25 as a novel therapeutic target for the treatment of airway remodelling in asthma.

Figure 1

Interleukin (IL)-25 neutralisation abolishes house dust mite (HDM)-induced pulmonary remodelling. (A) IL-25 (brown stained) section. (B) Sirius red stained perivascular and peribronchial collagen. (C) Quantitative analysis of subepithelial peribronchiolar collagen density. (D) α-Smooth muscle actin (brown stained) smooth muscle cells and myofibroblasts. (E) Peribronchial smooth muscle thickness. (F) Proliferating cell nuclear antigen (PCNA)-positive (brown stained) proliferating cells. (G) Percentage of PCNA-positive peribronchial mesenchymal cells. (H) Periodic acid-Schiff (PAS)-positive (pink/purple coloured) mucin-containing cells. (I) Epithelial PAS scoring. Original magnification ×40. Scale bar=50 µm. *p<0.05 compared with phosphate-buffered saline (PBS) control group. §p<0.05 AdS HDM Ig compared with AdC HDM Ig control group. †p<0.05 HDM anti-IL-25 compared with HDM Ig control group. Plots depict the median and IQR and minimum and maximum values. Data are generated from two independent experiments (n=8–12). IHC, immunohistochemistry. This figure is only reproduced in colour in the online version.

Figure 2

Interleukin (IL)-25 acts directly on mesenchymal cells to induce remodelling. (A) Activin A quantification in lung homogenate. (B) Biologically active pulmonary transforming growth factor (TGF)-β levels. (C) IL-25-induced extracellular collagen secretion by normal human lung fibroblasts (NHLFs) in vitro. (D) IL-17RB (brown stained) lung section: (i) negative Ig control; (ii)–(iv) IL-17RB-positive cells. *p<0.05 compared with phosphate-buffered saline (PBS) control group. §p<0.05 AdS house dust mite (HDM) Ig compared with AdC HDM Ig control group. †p<0.05 HDM anti-IL-25 compared with HDM Ig control group. Plots depict the median and IQR and minimum and maximum values. Data are generated from two independent experiments (n=8–12). IHC, immunohistochemistry. This figure is only reproduced in colour in the online version.

Figure 3

Allergen-induced vascular remodelling. (A) von Willebrand factor (vWF)-positive (brown stained) peribronchial vessels. (B) vWF-positive vessels per square millimetre. (C) Vascular endothelial growth factor (VEGF) levels in the lung homogenate. (D) Pulmonary endothelial progenitor cell (EPC) colony assay. (E) Levels of CXCL2 in lung. *p<0.05 compared with phosphate-buffered saline (PBS) control group. †p<0.05 house dust mite (HDM) anti-interleukin (IL)-25 compared with HDM Ig control group. Plots depict the median and IQR and minimum and maximum values. Data are generated from two independent experiments (n=8–12). This figure is only reproduced in colour in the online version.

Figure 4

Airway hyperreactivity is dependent on interleukin (IL)-25. (A) Airway resistance in mice. (B) Airway resistance (R_L) at 100 mg/ml methacholine (MCh). *p<0.05 compared with phosphate-buffered saline (PBS) control group. §p<0.05 AdS house dust mite (HDM) Ig compared with AdC HDM Ig control group. †p<0.05 HDM anti-IL-25 compared with HDM Ig control group. Plots depict the median and IQR and minimum and maximum values. Data are generated from two independent experiments (n=8–12).

Figure 5

Blocking interleukin (IL)-25 reduces allergen-induced pulmonary inflammation. (A) H&E stained lung sections. Scale bar=50 μm. (B) Total lung inflammatory cells. (C) Pulmonary eosinophil numbers. (D) T helper type 2 (Th2) cells in the lung. *p<0.05 compared with phosphate-buffered saline (PBS) control group. †p<0.05 house dust mite (HDM) anti-IL-25 compared with HDM Ig control group. Plots depict the median and IQR and minimum and maximum values. Data are generated from two independent experiments (n=8–12). This figure is only reproduced in colour in the online version.

Figure 6

Allergen-induced inflammatory mediator production is partially decreased by blocking interleukin (IL)-25. (A) Pulmonary IL-4, (B) IL-5, (C) IL-13 and (D) interferon (IFN)-γ levels. (E) Serum total IgE and (F) house dust mite (HDM)-specificIgE determined by optical density (OD). *p<0.05 compared with phosphate-buffered saline (PBS) control group. †p<0.05 HDM anti-IL-25 compared with HDM Ig control group. Plots depict the median and IQR and minimum and maximum values. Data are generated from two independent experiments (n=8–12).

Figure 7

Allergen-induced epithelial cytokine release is completely abrogated by blocking interleukin (IL)-25. Levels of (A) IL-33, (B) thymic stromal lymphopoietin (TSLP), (C) CCL20 and (D) IL-25 in the lung. *p<0.05 compared with phosphate-buffered saline (PBS) control group. †p<0.05 house dust mite (HDM) anti-IL-25 compared with HDM Ig control group. Plots depict the median and IQR and minimum and maximum values. Data are generated from two independent experiments (n=8–12).