Abstract
Single cell analysis in autoimmune disease has largely focused on mononuclear cells and diseased tissues, leaving the diversity of neutrophils poorly understood. To identify the dynamics of neutrophil autoimmunity, we focus on an autoimmune disease characterized by abnormal neutrophil activation, anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis. Single-cell transcriptome and surface proteome analyses are performed on 179,664 peripheral white blood cells from treatment-naïve patients with new-onset microscopic polyangiitis (MPA) and healthy donors. Neutrophils are classified into seven subsets, two of which are significantly increased in MPA patients: immature neutrophils (Neu_Immature) and neutrophils characterized by type II interferon signature genes (Neu_T2ISG). The Neu_T2ISG subset shows increased expression of interferon (IFN)-γ response genes, many of which are identified as differentially expressed genes in neutrophils from MPA patients compared to those from healthy donors. Trajectory and cell–cell interaction analyses identify Neu_T2ISG as a uniquely primed subset that differentiates from mature neutrophils upon stimulation with IFN-γ and tumor necrosis factor (TNF)-α. The combined effect of these cytokines simultaneously exposes myeloperoxidase and Fcγ receptors on the neutrophil cell surface and promotes ANCA–induced neutrophil extracellular trap formation. Case-by-case analysis indicates that patients with a high proportion of the Neu_T2ISG subset are associated with persistent vasculitis symptoms. In a larger cohort using stored sera, serum IFN-γ levels at disease onset significantly correlate with susceptibility to disease relapse. Our findings identify neutrophil diversity at the single cell level and bridge the data to a clinically applicable biomarker for predicting relapse in small vessel vasculitis.