Determining drivers of mast cell expansion and function during human airway disease

Project Summary/Abstract: Mast cells (MCs) expand within the airway epithelium and sub-epithelium during prevalent and burdensome human respiratory disease, including asthma and nasal polyposis, where they are thought to play a central role in disease pathobiology. MCs take on discrete protease expression profiles in each location, with sup-epithelial MCs co-expressing tryptase and chymase (MCTC) while epithelial MCs express tryptase alone (MCT). Neither the mechanisms underlying this expansion, the signals directing the epithelial and sub-epithelial MC phenotypes, nor the differential contribution of MC phenotypes to tissue inflammation are well understood. This application focuses on understanding the mechanisms through which airway tissue structural cells differentially direct the expansion and differentiation of MC progenitors (MCps), based on preliminary studies identifying a central role for fibroblasts and epithelial cells in both processes. This proposal tests the central hypothesis that MC hyperplasia in inflamed human airway mucosa is driven by the recruitment of MCps exhibiting a robust proliferative capacity, and that the proliferation and differentiation of these recruited MCps towards the histochemically recognized MCT and MCTC subsets are driven by key signals secreted by airway structural cells. A related hypothesis is that the resulting intraepithelial and subepithelial MC phenotypes are maintained through a network of transcription factors regulated by these tissue stromal-derived signals, and that signal nucleotide polymorphisms in these transcription factors increase risk of developing airway disease. Aim 1 of this proposal evaluates MCp concentration in human sinonasal tissue across disease endotypes, determining the relationship between recruited MCp and the proliferative population of MCs we previously identified in nasal polyposis. Aim 2 of this proposal tests the impact of a series of candidate stromal-derived ligands on directing MCp proliferation and differentiation using CRISPR/Cas9 gene editing. Aim 3 probes the transcription factor network underlying MC polarization via shRNA knockdown and evaluates the impact of key asthma- associated single nucleotide polymorphisms in one such transcription factor on intraepithelial MC differentiation. Completion of these aims will greatly expand our understanding of the pathways through which MCs are capable of influencing tissue inflammation and the mechanism(s) through which these pathways are regulated by their tissue microenvironment.