Medical Student John Hopkins School of Medicine Baltimore, Maryland
Background/Objectives: Idiopathic subglottic stenosis (iSGS) is a progressive disease of the upper airway characterized by dysregulated inflammation and activated fibroblasts. Targeting inflammation has been the center of recent investigation, with most studies focusing on adaptive immunity. The nature and role of innate immunity, specifically macrophages, remains poorly understood despite increased populations in iSGS. The goals of this study are (1) characterize the iSGS macrophage population, (2) assess differentially expressed genes, and (3) characterize the effect of identified biomarkers in-vitro.
Methods: Biopsies from iSGS scar (n=5) and normal (n=3) tissue in iSGS patients were assessed by single-cell RNA sequencing (scRNA-seq). Immunohistochemistry (IHC) of human iSGS (n=3) and control tissue was performed to localize S100A8/A9 protein. Finally, iSGS-derived fibroblasts (n=2) were treated with S100A8/A9 complex (1000ng/ml).
Results: scRNA-seq identified 10 unique macrophage populations in iSGS tissue. Cluster analysis of macrophages in iSGS and control tissue revealed S100A8 and S100A9 in the top 10 differentially expressed gene list (Log2FoldChange=1.55, p=0.017 and Log¬2¬FoldChange = 2.42, p<0.001). IHC exhibited increased expression of S100A8/A9 complex in iSGS compared to normal controls. Furthermore, S100A8/A9 co-localized with the macrophage marker CD11b. iSGS-derived fibroblasts demonstrated increased Collagen-1 and Collagen-3 gene expression (fold change= 2.38±0.66 and 3.28±0.7) when treated with S100A8/A9 protein.
Conclusions: S100A8/A9 proteins are damage associated molecular patterns and promote sustained inflammation. S100A8/A9 stimulated collagen expression in iSGS fibroblasts. A unique and dominant population of macrophages in iSGS demonstrate increased expression of S100A8/A9. This cell population and biomarker may play a critical role in promoting fibrosis in iSGS.