Spatial phenotyping of human bronchial airways in obstructive lung disease.

Abstract

Chronic respiratory diseases such as asthma and COPD involve interactions between multiple resident and immune cell types within bronchial airways, resulting in structural and functional changes. Thus cellular heterogeneity, arrangements and associated neighborhoods as well as interactions between cells and matrices represent intriguing yet challenging areas of study. Spatial phenotypic profiling facilitates exploration of these issues of the cellular microenvironment and identification of context-dependent cell-cell interactions. Utilizing spatial phenotyping, we interrogated the features and cellular landscape of lungs from non-asthmatics, asthmatics, and COPD in FFPE samples by developing a 10-plex antibody panel for the Akoya PhenoCycler^®-Fusion system, focused on immune cells (CD45, CD3, CD4, CD8), proliferative cells (Ki67, PCNA), angiogenesis (CD34), epithelium (E-cadherin), smooth muscle (SMA) and extracellular matrix (collagen). We performed cell segmentation on multiplex immunofluorescence images and quantified marker intensity in each cell. Phenotypes were manually identified after normalization, integration, and clustering cells across samples. The composition, cell profiling, and distribution varied significantly between asthmatics and COPD compared to non-asthmatics emphasizing disease heterogeneity. Spatially agnostic analysis revealed that the matrix cluster was more abundant in COPD compared to non-asthmatics and asthmatics, consistent with a greater role for fibrosis. However, asthmatic patients had a higher proportion of unclassified and CD8 + clusters highlighting immune responses. Co-localization analysis showed near random distribution in non-asthmatics. But strong spatial interaction between T cells and other immune or matrix cells in asthma, and a higher avoidance of smooth muscle and immune cells, and of proliferative markers in both asthmatic and COPD. Niche analysis demonstrated different recurrent cell-cell interactions in asthmatic and COPD cohorts. In COPD, the matrix cell-enriched niche was more abundant, while in asthmatics, the unclassified cell-enriched niche was more prevalent compared to non-asthmatics. These findings provide insights into differential spatial organization of cells and tissues in asthma and COPD, with immune and epithelial mechanisms suggesting active inflammation and remodeling in asthma, but fibrotic processes in COPD, and potential role for vascular processes in both conditions.