Single-cell RNA sequencing highlights the heterogeneity of neutrophils in granulomatosis with polyangiitis
Shuning Guo, Yangzhong Zhou, Dazhi Pang, Guanqiao Li, Yulin Lyu, Xinyuan Xu, Jing Li, Xinping TianAbstract
Background and Objectives
Granulomatosis with polyangiitis (GPA), a clinical phenotype of anti-neutrophil cytoplasmic antibody-associated vasculitis, is characterized by necrotizing inflammation of blood vessels. While neutrophil dysregulation is central to its pathogenesis, its cellular heterogeneity and dynamic functional states remain poorly defined. To better understand the transcriptomic profile of neutrophils, we applied single-cell RNA sequencing (scRNA-seq) to peripheral blood samples from GPA patients and healthy controls (HCs).
Methods
Peripheral blood samples were collected from three patients with active GPA and three HCs, followed by sequencing using the BD Rhapsody platform (Becton, Dickinson and Company). In a subsequent validation cohort, flow cytometry and quantitative real-time polymerase chain reaction were used to confirm specific neutrophil subtypes and explore their potential clinical significance.
Results
Seven neutrophil clusters with distinct transcriptomic and functional profiles were identified. CXCL8-high and TXNIP-high subsets, predominant in HCs and GPA patients respectively, showed inflammatory profiles. CXCL8-high subsets also expressed immunoregulatory molecules to restrain inflammation. MMP8-high neutrophils showed enhanced antimicrobial activity, whereas IFIT1-high cells were linked to antiviral responses, suggesting that prior infections may contribute to anti-neutrophil cytoplasmic antibody-associated vasculitis (AAV) pathogenesis. PLA2G7-high neutrophils, identified as terminally differentiated by pseudotime analysis, were associated with disease relapse and renal involvement, indicating potential as AAV biomarkers. Transcriptional profiles of peripheral T and natural killer (NK) cells were also characterized, along with the analysis of systemic immune cell-cell communication networks.
Conclusion
Our study enhances the understanding of the dynamic functional roles of neutrophils in GPA and may facilitate the development of therapeutic strategies aimed at selectively targeting disease-specific neutrophil subsets.