Background: Chronic rhinosinusitis with nasal polyps (CRSwNP) is characterized by type 2 (T2) inflammation. Recent studies, including our own, suggest that neutrophils are also elevated in T2 nasal polyps (NP) and that elevated neutrophils display an activated phenotype. However, the actual roles of neutrophils in NP pathogenesis in T2 CRSwNP are still largely unclear.
Objective: To reveal the roles and heterogeneity of neutrophils in NP tissue by single-cell RNA sequencing analysis.
Methods: We developed a novel microwell-based single-cell RNA sequencing assay using granulocyte-enriched samples from 5 control sinus tissues, 5 NP tissues and patient-matched peripheral blood (PB) samples. This approach allowed for examination of differential expression of genes in NP neutrophils by the Benjamini-Hochberg algorithm and predicted the overall function of NP neutrophils by pathway and Gene Ontology enrichment analyses.
Results: After performing all quality control steps, we successfully detected neutrophils. We identified 333 downregulated and 128 upregulated genes in NP neutrophils (1,151 cells) compared with all PB neutrophils (13,591 cells) (>1.5-fold, q < 0.05) and found commonly dysregulated genes in NP neutrophils compared with both all PB and control sinus tissue neutrophils (3,136 cells). Commonly downregulated genes in NP neutrophils were associated with the innate immune system, and upregulated genes were associated with nuclear factor-κB signaling, cytokine activity, and cellular response to oxygen-containing compounds. NP neutrophils displayed 4 clusters revealing potential heterogeneity of neutrophils in NP tissue.
Conclusions: Elevated neutrophils in NP tissue appear to exist in several subphenotypes that may play important pathogenic roles in CRSwNP.
Keywords: Neutrophils; chronic rhinosinusitis; heterogeneity; nasal polyps; scRNA-Seq.
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