Single-Cell Transcriptomics Reveals CCL3+ Classical Monocyte Subset Linked to Autoimmune Pathogenesis

Objective: The diverse differentiation states of mononuclear macrophages are closely associated with the pathogenesis of autoimmune diseases. This study integrates single-cell RNA sequencing data from six autoimmune diseases to characterize shared and disease-specific alterations in mononuclear macrophages, with the aim of enhancing our understanding of the immune landscape in autoimmune diseases and refining clinical treatment strategies. Methods: We collected single-cell RNA-sequencing data of autoimmune diseases including primary Sjogren’s syndrome (pSS), Behçet’s disease (BD), juvenile dermatomyositis (JDM), rheumatoid arthritis (RA), relapsing-remitting multiple sclerosis (RRMS), and systemic lupus erythematosus (SLE). We performed scRNA-seq analysis on 350,043 peripheral blood immune cells from autoimmune diseases patients and healthy controls, followed by validations with flow cytometry, immunohistochemical staining, and immunofluorescence. Results: Fifteen mononuclear phagocyte subpopulations were clustered from peripheral blood mononuclear cells (PBMCs), we identified a new subpopulation named CCL3 + classical monocytes (cMo) that is co-amplified in multiple autoimmune diseases (BD, JDM, pSS, RRMS, SLE). The CCL3 + cMo cells are characterized by high M1-like score, exhibiting strong inflammatory characteristics and high chemotaxis toward other monocytes. In addition, CCL3 + cMo cells upregulated antigen presentation-related signaling pathways, and the cytotoxic CD8 + T or memory CD8 + T cells were strongly perturbed by their signaling crosstalk. Conclusion: This study delineates a comprehensive landscape of mononuclear phagocyte heterogeneity in autoimmune diseases and reveals CCL3 + cMo as a commonly amplified immune subset associated with multiple autoimmune diseases. These findings highlight its potential role in disease mechanisms and nominate CCL3 + cMo as a candidate therapeutic target. Keywords: autoimmune diseases, single-cell RNA sequencing, CCL3 + classical monocytes, mononuclear phagocytes, primary Sjogren’s syndrome