Human intraepithelial mast cell differentiation and effector function are directed by TGF-b signaling

Mast cells (MCs) expressing a distinctive protease phenotype (MCTs) selectively expand within the epithelium of human mucosal tissues during type 2 (T2) inflammation. While MCTs are phenotypically distinct from subepithelial MCs (MCTCs), signals driving human MCT differentiation and this subset's contribution to inflammation remain unexplored. Here, we identify TGF-b as a key driver of the MCT transcriptome in nasal polyps. We find that short-term TGF-b signaling alters MC cell surface receptor expression and partially recapitulates the in vivo MCT transcriptome, while TGF-ß signaling during MC differentiation upregulates a larger number of MCT-associated transcripts and inhibits the hallmark MCTC proteases chymase and cathepsin G at both the transcript and protein level, allowing selective in vitro differentiation of MCTs for functional study. We identify discrete differences in effector phenotype between in vitro-derived MCTs and MCTCs, with MCTs greatly enhancing pro-inflammatory lipid mediator generation while selectively modifying cytokine, chemokine, and growth factor production in response to both innate and adaptive stimuli, recapitulating functional features of their tissue-associated counterpart MC subsets. Thus, our findings support a role for TGF-ß in promoting human MCT differentiation and identify a discrete contribution of this cell type to T2 inflammation. Overall design: This study tests the effects of TGF-b1 treatment on mature human peripheral blood-derived mast cells at two timepoints (24h, 6d) versus the effects of differentiating mast cells in the presence or absence of TGF-b1.