Involvement of IL-26 in bronchiolitis obliterans syndrome but not in acute rejection after lung transplantation

Abstract

Background: The main long-term complication after lung transplantation is bronchiolitis obliterans syndrome (BOS), a deadly condition in which neutrophils may play a critical pathophysiological role. Recent studies show that the cytokine interleukin IL-26 can facilitate neutrophil recruitment in response to pro-inflammatory stimuli in the airways. In this pilot study, we characterized the local involvement of IL-26 during BOS and acute rejection (AR) in human patients.

Method: From a biobank containing bronchoalveolar lavage (BAL) samples from 148 lung transplant recipients (LTR), clinically-matched patient pairs were identified to minimize the influence of clinical confounders. We identified ten pairs (BOS/non-BOS) with BAL samples harvested on three occasions for our longitudinal investigation and 12 pairs of patients with and without AR. The pairs were matched for age, gender, preoperative diagnosis, type of and time after surgery. Extracellular IL-26 protein was quantified in cell-free BAL samples using an enzyme-linked immunosorbent assay. Intracellular IL-26 protein in BAL cells was determined using immunocytochemistry (ICC) and flow cytometry.

Results: The median extracellular concentration of IL-26 protein was markedly increased in BAL samples from patients with BOS (p < 0.0001) but not in samples from patients with AR. Intracellular IL-26 protein was confirmed in alveolar macrophages and lymphocytes (through ICC and flow cytometry) among BAL cells obtained from BOS patients.

Conclusions: Local IL-26 seems to be involved in BOS but not AR, and macrophages as well as lymphocytes constitute cellular sources in this clinical setting. The enhancement of extracellular IL-26 protein in LTRs with BOS warrants further investigation of its potential as a target for diagnosing, monitoring, and treating BOS.

Keywords: Acute rejection; Bronchiolitis obliterans syndrome; Cytokine; Interleukin-26; Lung transplantation; Neutrophil.

Fig. 1

Interleukin (IL)-26 protein concentration (logarithmic scale) in cell-free bronchoalveolar lavage (BAL) fluid from lung transplant recipients (LTRs) with (n = 20) and without (n = 20) bronchiolitis obliterans syndrome (BOS) quantified by enzyme-linked immunosorbent assay (ELISA). Bars represent the median

Fig. 2

Interleukin (IL)-26 protein concentration (logarithmic scale) in cell-free bronchoalveolar lavage fluid (BAL) from lung transplant recipients (LTRs) with (n = 12) and without (n = 12) acute rejection (AR) quantified by enzyme-linked immunosorbent assay (ELISA). Bars represent the median

Fig. 3

Interleukin (IL)-26 protein concentration (logarithmic scale) in cell-free bronchoalveolar (BAL) fluid from lung transplant recipients (LTRs) with (n = 10) or without (n = 10) bronchiolitis obliterans syndrome (BOS) quantified by enzyme-linked immunosorbent assay (ELISA). Time one and two before and time three at BOS diagnosis (with corresponding time points in the BOS free group

Fig. 4

Representative images showing flow cytometry of stained bronchoalveolar lavage (BAL) cells from patients with bronchiolitis obliterans syndrome (BOS), CD4 + cells A and CD8 + cells B as well as from patients without BOS, CD4 + cells C and CD8 + cells (D)”

Fig. 5

Immunostaining of IL-26 in bronchoalveolar lavage (BAL) cells from lung transplant recipients with (n = 5; a–c and e) and without (n = 4; d) bronchiolitis obliterans (BOS). Immunostaining was performed using a an IgG2b isotype control antibody, a polyclonal CD68 antibody (b; brown), a monoclonal specific IL-26 antibody (c; purple) or a monoclonal specific IL-26 antibody in combination with a polyclonal CD68 antibody (d and e)