IL-25 and type 2 innate lymphoid cells induce pulmonary fibrosis

Abstract

Disease conditions associated with pulmonary fibrosis are progressive and have a poor long-term prognosis with irreversible changes in airway architecture leading to marked morbidity and mortalities. Using murine models we demonstrate a role for interleukin (IL)-25 in the generation of pulmonary fibrosis. Mechanistically, we identify IL-13 release from type 2 innate lymphoid cells (ILC2) as sufficient to drive collagen deposition in the lungs of challenged mice and suggest this as a potential mechanism through which IL-25 is acting. Additionally, we demonstrate that in human idiopathic pulmonary fibrosis there is increased pulmonary expression of IL-25 and also observe a population ILC2 in the lungs of idiopathic pulmonary fibrosis patients. Collectively, we present an innate mechanism for the generation of pulmonary fibrosis, via IL-25 and ILC2, that occurs independently of T-cell-mediated antigen-specific immune responses. These results suggest the potential of therapeutically targeting IL-25 and ILC2 for the treatment of human fibrotic diseases.

Keywords: cytokine; inflammation; innate response; therapy.

Fig. 1.

Decreased pulmonary collagen deposition in IL-25–deficient mice following S. mansoni egg challenge. Pulmonary granulomas were induced in WT, Il25, and Il17br deficient mice. (A) Egg granuloma volume on hematoxylin and eosin-stained slides. (B) Total lung collagen expressed per mg of lung protein. (C) Representative histology images depicting collagen deposition within the granuloma using Masson’s Trichrome stain (scale, 20 μM). (D) Levels of IL-4, IL-13, TGFβ, and IL-17A in lung digests expressed per mg of lung protein. (E) Expression of ILC2 [lineage (CD3, CD4, CD8, CD11b, CD11c, CD19, Gr-1, F4/80, FcεR1)⁻IL-7Rα⁺Sca-1⁺T1/ST2⁺ICOS⁺] in lungs and MLN. Data are representative of mean ± SEM (n = 3–10) from three individual experimental replicates (*P < 0.05, **P < 0.01).

Fig. 2.

IL-25 induces pulmonary collagen deposition in WT but not Rora^sg/sg mice. WT and Rora^sg/sg were treated with recombinant mouse IL-25 i.n. on days 0, 3, 7, 10, and 13. (A) Representative histology images depicting collagen deposition within the lung using Masson’s Trichrome stain (scale, 100 μM). (B) Total collagen in lung digests expressed per mg of lung protein. (C) Levels of IL-13, IL-17A, and TGFβ in lung digests expressed per mg of lung protein. Data are representative of mean ± SEM (n = 3–5) from two individual experimental replicates (ns, not significant; *P < 0.05, **P < 0.01).

Fig. 3.

ILC2 regulate pulmonary collagen deposition. Disparate CD90 chimeras were generated by transferring CD90.1 T (CD3⁺) and B (CD19⁺) cells to Rag-1^−/− mice expressing the CD90.2 allele, which were subsequently treated with anti-CD90.2 or an isotype control mAb. (A) Lung frequency of CD4 and ILC2 in Rag-1^−/−, disparate CD90 chimeras treated with anti-IgG and disparate CD90 chimeras treated with anti-CD90.2. (B) Granuloma volume and representative histology images stained with Masson’s Trichrome (scale, 100 μM). Lung collagen (C) and cytokines (D) expressed per mg of lung protein. Data are representative of mean ± SEM (n = 3–5) from two individual experimental replicates (ns, not significant; *P < 0.05, **P < 0.01).

Fig. 4.

Transferring IL-13–expressing ILC2 induces pulmonary collagen deposition in IL-13–deficient mice. ILC2 isolated from WT and Il13^−/− were transferred into unchallenged and egg-challenged IL-13–deficient mice. (A) Representative histology images depicting Masson’s Trichrome staining from PBS and egg-challenged animals (scale, 100 μM) and (B) egg-associated granuloma volume. (C) Lung collagen expressed per mg of lung protein. Data are representative of mean ± SEM (n = 2–4) from two individual experimental replicates (*P < 0.05).

Fig. 5.

IPF patients have increased expression of IL-25 and ILC2 in the BAL. BAL fluid was collected from patients with IPF at diagnosis and at a 1-y follow-up (IPF + 1 y) and patients diagnosed with erythema nodosum (EN; nonprogressive patients). (A) IL-25, IL-13, TGFβ, and IL-17A in BAL fluid expressed as fold-increase relative to EN patients. (B) Correlation between BAL fluid levels of IL-25, IL-13, TGFβ, or IL-17A and periostin was plotted and Pearson’s correlation calculated and displayed on the graph. (C) Representative plots of ILC2 [lineage (CD3, CD4, CD8, CD11b, CD11c, CD19, CD14, CD56, CD123, FcεR1) negative, expressing CRTH2, T1/ST2, CD45, ICOS, IL-7Rα, and IL-17BR] in the BAL of nonfibrotic patients and IPF patients, with expression of these cells displayed as a percentage of live cells per sample. Data are representative of mean ± SEM (n = 3, EN; n = 14, IPF and IPF + 1 y) (ns, not significant; *P < 0.05, **P < 0.01).