D3.395 - Analysis of single cell multiome data (transcriptomics + ATAC) from human nasal polyp mast cells

Mast cells (MCs) are long-lived, granulated, tissue resident immune cells that respond rapidly to environmental insults through the release of biologically active mediators. Human MCs are traditionally classified into two major subtypes based on their granular content and tissue localisation: mucosal mast cells containing tryptase alone (MCTs) and connective tissue mast cells containing tryptase, chymase, and carboxypeptidase A3 (MCTCs). However, there is now growing evidence of vast heterogeneity that exists beyond classically defined MC subgroups. In this study, we aim to develop a protocol to study MC heterogeneity at the single cell epigenetic and transcriptomic levels. 

Nasal polyp samples were collected from patients with chronic rhinosinusitis with nasal polyps (CRSwNP) undergoing surgery, digested and MCs were sorted using flow cytometry. Nuclei were isolated from live polyp MCs and LAD2 cells and processed using Chromium Single Cell Multiome Assay for Transposase Accessible Chromatin (ATAC) and Gene Expression (GEX) (10X Genomics).

Good quality data were obtained from LAD2 and polyp MCs allowing for multiome analysis. Single cell GEX analysis showed the expression of MC characteristic genes such as KIT, TPSAB1 and TPSB2. LAD2 and polyp MCs expressed different sets of MC markers. Genes associated with skin MCs (CMA1, MRGPRX2) were present in LAD2 but not in polyp MCs which predominantly expressed tryptase (TPSB2) and carboxypeptidase A3 (CPA3). Higher heterogeneity (number of clusters) was observed in GEX analysis than in ATAC analysis in LAD2 and polyp MCs. Polyp MCs showed lower heterogeneity than LAD2 cells in both ATAC and GEX analyses.

We have developed a protocol for single cell multiome analysis of MCs allowing for the characterisation of MC subpopulations at the epigenetic and transcriptomic levels.