To identify the dynamics of neutrophil autoimmunity, here we focus on anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis and perform single-cell transcriptome and surface proteome analyses on peripheral white blood cells from patients with new-onset microscopic polyangiitis (MPA). Compared with controls, two neutrophil populations, immature neutrophils and neutrophils with type II interferon signature genes (Neu_T2ISG), are increased in patients with MPA. Trajectory and cell-cell interaction analyses identify Neu_T2ISG as a subset that differentiates from mature neutrophils upon stimulation with IFN-γ and TNF, which synergize to induce myeloperoxidase and Fcγ receptors expression on the neutrophil cell surface and promote 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. A larger cohort analysis shows that serum IFN-γ levels at disease onset correlate with susceptibility to disease relapse. Our findings thus identify neutrophil diversity at the single cell level and implicate a biomarker for predicting relapse in small vessel vasculitis.
© 2025. The Author(s).

Abstract Elevated anti-citrullinated protein antibodies (ACPA) levels in the peripheral blood are associated with an increased risk for developing rheumatoid arthritis (RA). Currently, no treatments are available that prevent progression to RA in these at-risk individuals. In addition, diverse pathogenic mechanisms underlying a common clinical phenotype in RA complicate therapy as no single agent is universally effective. We propose that a unifying set of transcription factor and their downstream pathways regulate a pro-inflammatory cell communication network, and that this network allows multiple cell types to serve as pathogenic drivers in at-risk individuals and in early RA. To test this hypothesis, we identified ACPA-positive at-risk individuals, patients with early ACPA-positive RA and matched controls. We measured single cell chromatin accessibility and transcriptomic profiles from their peripheral blood mononuclear cells. The datasets were then integrated to define key TF, as well as TF-regulated targets and pathways. A distinctive TF signature was enriched in early RA and at-risk individuals that involved key pathogenic mechanisms in RA, including SUMOylation, RUNX2, YAP1, NOTCH3, and β-Catenin Pathways. Interestingly, this signature was identified in multiple cell types, including T cells, B cells, and monocytes, and the pattern of cell type involvement varied among the at-risk and early RA participants, supporting our hypothesis. Similar patterns of individualized gene expression patterns and cell types were confirmed in single cell studies of RA synovium. Cell communication analysis provided biological validation that diverse lineages can deliver the same core set of pro-inflammatory mediators to receiver cells in vivo that subsequently orchestrate rheumatoid inflammation. These cell-type-specific signature pathways could explain the personalized pathogenesis of RA and contribute to the diversity of clinical responses to targeted therapies. Furthermore, these data could provide opportunities for stratifying individuals at-risk for RA, and selecting therapies tailored for prevention or treatment of RA. Overall, this study supports a new paradigm to understand how a common clinical phenotype could arise from diverse pathogenic mechanisms and demonstrates the relevance of peripheral blood cells to synovial disease.

Elevated anti-citrullinated protein antibodies (ACPA) levels in the peripheral blood are associated with an increased risk for developing rheumatoid arthritis (RA). Currently, no treatments are available that prevent progression to RA in these at-risk individuals. In addition, diverse pathogenic mechanisms underlying a common clinical phenotype in RA complicate therapy as no single agent is universally effective. We propose that a unifying set of transcription factor and their downstream pathways regulate a pro-inflammatory cell communication network, and that this network allows multiple cell types to serve as pathogenic drivers in at-risk individuals and in early RA. To test this hypothesis, we identified ACPA-positive at-risk individuals, patients with early ACPA-positive RA and matched controls. We measured single cell chromatin accessibility and transcriptomic profiles from their peripheral blood mononuclear cells. The datasets were then integrated to define key TF, as well as TF-regulated targets and pathways. A distinctive TF signature was enriched in early RA and at-risk individuals that involved key pathogenic mechanisms in RA, including SUMOylation, RUNX2, YAP1, NOTCH3, and β-Catenin Pathways. Interestingly, this signature was identified in multiple cell types, including T cells, B cells, and monocytes, and the pattern of cell type involvement varied among the at-risk and early RA participants, supporting our hypothesis. Similar patterns of individualized gene expression patterns and cell types were confirmed in single cell studies of RA synovium. Cell communication analysis revealed that the lineages displaying this RA TF signature deliver a common set of pro-inflammatory mediators to receiver cells that subsequently orchestrate rheumatoid inflammation. These cell-type-specific signature pathways could explain the personalized pathogenesis of RA and contribute to the diversity of clinical responses to targeted therapies. Furthermore, these data could provide opportunities for stratifying individuals at-risk for RA, and selecting therapies tailored for prevention or treatment of RA. Overall, this study supports a new paradigm to understand how a common clinical phenotype could arise from diverse pathogenic mechanisms and demonstrates the relevance of peripheral blood cells to synovial disease.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies, presenting a persisting global health burden. Neutrophils have a double-edged role in tumor progression exhibiting both pro-tumor and anti-tumor functions. CD71, also known as transferrin receptor 1, performs a critical role in cellular iron uptake and is highly expressed on proliferating cells, and especially on activated immune cells. CD71 is known to be elevated in various types of solid cancers and is associated with poor prognosis, however, the expression of CD71 on neutrophils in PDAC and its potential clinical impact is still unknown. Therefore, we analyzed CD71 on circulating neutrophils in PDAC and clinical control patients and found a significant increased expression in PDAC patients. High expression of CD71 on neutrophils in PDAC patients was associated with reduced outcome compared to low expression. CD71 on neutrophils correlated positively with the levels of proinflammatory cytokines IL-6, IFN-γ, and growth factor ligands CD40-L, and BAFF in plasma of PDAC patients. Finally, we have demonstrated that high expression of CD71 on neutrophils was also associated with an increased expression of CD39 and CD25 on circulating T-cells. Based on our findings, we hypothesize that CD71 on neutrophils is associated with tumor progression in PDAC. Further studies are required to investigate the distinct functionality of CD71 expressing neutrophils and their potential clinical application.
© 2024. The Author(s).

Single-cell assay for transposase-accessible chromatin using sequencing (scATAC-seq) is being increasingly used to study gene regulation. However, major analytical gaps limit its utility in studying gene regulatory programs in complex diseases. In response, MOCHA (Model-based single cell Open CHromatin Analysis) presents major advances over existing analysis tools, including: 1) improving identification of sample-specific open chromatin, 2) statistical modeling of technical drop-out with zero-inflated methods, 3) mitigation of false positives in single cell analysis, 4) identification of alternative transcription-starting-site regulation, and 5) modules for inferring temporal gene regulatory networks from longitudinal data. These advances, in addition to open chromatin analyses, provide a robust framework after quality control and cell labeling to study gene regulatory programs in human disease. We benchmark MOCHA with four state-of-the-art tools to demonstrate its advances. We also construct cross-sectional and longitudinal gene regulatory networks, identifying potential mechanisms of COVID-19 response. MOCHA provides researchers with a robust analytical tool for functional genomic inference from scATAC-seq data.
© 2024. The Author(s).