Heterogeneity and Clinical Relevance of Group 2 Innate Lymphoid Cells Subsets in Nasal Polyps

ABSTRACTBackground Chronic rhinosinusitis with nasal polyps (CRSwNP) is characterized by type 2 (T2) inflammation with elevated IL-5 and IL-13. Although group 2 innate lymphoid cells (ILC2s) drive T2 inflammation, their subset diversity and clinical relevance in nasal polyps (NPs) remain unclear. Objective To investigate cellular sources of T2 cytokines, subset heterogeneity of ILC2s and relationship between ILC2 subsets and clinical severity in CRSwNP. Methods ILC2s and CD4+ T cells were isolated from NP tissue and analyzed for cytokine production. ILC2s from six NP and four peripheral blood (PB) samples underwent single-cell RNA sequencing. Differential gene expression, subset heterogeneity, and pseudotime trajectory analyses were performed. Flow cytometry validated ILC2 subsets and associations with clinical parameters. Results Under ex vivo conditions, NP-derived ILC2s produced higher IL-5 and IL-13 levels than Th2 cells. NP-derived ILC2s showed downregulation of early developmental and trafficking genes (e.g. CD48, S1PR1) and upregulation of T2-associated cytokines (e.g. IL5, IL13), chemokines (e.g. XCL1, CXCL8), remodeling factors (e.g. AREG, TNFSF14), and METRNL. Four ILC2 subsets reflecting activation states were identified: migratory (tissue-homing with less activation), transitional (intermediate activation), inflammatory (high activation and T2 cytokines), and exhausted-like (expression of inhibitory receptors: e.g. TIGIT). The combined number of T2 cytokine-enriched subsets (transitional, inflammatory, and exhausted-like ILC2s) correlated with Lund-Mackay CT scores. Inflammatory and exhausted-like ILC2s were associated with clinical symptom severity. Conclusion ILC2s are one of the important effector populations driving local T2 inflammation in NPs and include four subsets with specific clinical associations. Subset-level ILC2 profiling may clarify CRSwNP pathophysiology and inform clinical stratification.