Epithelial and Immune Phenotyping of Distal Airway Cells in Lung Allograft Dysfunction

Abstract

Airway epithelial injury is thought to be the final common pathway leading to airway fibrosis in chronic lung allograft dysfunction (CLAD). While cumulative post-transplant inflammatory insults represent CLAD risk factors, we lack adequate methods to measure epithelial changes during such events. We hypothesize that early detection of allograft epithelial dysregulation and injury will reveal relevant pathways in CLAD pathogenesis and predict subsequent CLAD. We aim to use transbronchial brushings (TBBr) to sample distal airway epithelial cells and assess their characteristics in the context of allograft dysfunction. In this study, TBBr are being collected from patients undergoing routine surveillance bronchoscopies at 3, 6, 9, and 12 months post-transplant, as well as patients with acute lung allograft dysfunction (ALAD), defined as a >=10% drop in FEV1 compared to previous baseline (Figure 1A). Multi-colour flow cytometry panels were developed to identify populations of epithelial and immune cells. Cell cytospins were assessed with immunofluorescence (Figure 1B-C). In this preliminary interim analysis of TBBr from 14 patients (6 stable and 8 with ALAD), we were able to identify important epithelial and mucosal immune cell subsets. In patients with ALAD, there was a trend towards decreased club cells, which act as epithelial cell progenitors and have anti-inflammatory properties (Figure 1D-E). Additionally, there was a trend towards increased epithelial cell expression of CD54 (ICAM), an activation marker (Figure 1E). In contrast, intraepithelial T and B lymphocytes were not found to be different in these patient groups. This preliminary study shows that TBBr can successfully isolate airway epithelial and mucosal immune cells for identification of specific cell populations. Follow-up studies from this project will include recruitment of additional subjects, longitudinal sample analyses, and association of cellular changes with later clinical outcomes.