Raw spectral flow cytometry data for "Deep Phenotyping Identifies a Pathogenic Axis of CD20+ DN B cells, Monocyte-Derived Dendritic Cells and CD27-T cells in Antiphospholipid Syndrome"

Antiphospholipid Syndrome (APS) is a systemic autoimmune disease defined by pathogenic anti-phospholipid antibodies and a high risk of vascular thrombosis. B-cell dysfunction is a known driver; however, the specific cellular networks and non-classical maturation pathways that sustain disease progression remain poorly understood. We used high-dimensional 25-color spectral flow cytometry and unsupervised machine learning algorithms, including Tracking Responders Expanding (T-REX), to characterize the peripheral blood immune landscape in APS patients (n=45) and healthy controls (n=14). These findings were correlated with serum levels of 46 cytokines and chemokines to identify systemic inflammatory axes. High-dimensional analysis revealed remodeling of the B-cell compartment, marked by a significant expansion of activated Double Negative (DN CD20+ IgD- CD27-) B cells expressing CD38, HLA-DR, and CD11c across all APS subgroups. This unconventional subset likely emerges via an extra-follicular maturation pathway. Within the T-cell compartment, we identified a global shift toward fully differentiated effector states, including increased CD4+CD27- and CD8+ T EMRA cells. Notably, T-REX identified a core myeloid signature of activated, migratory monocyte-derived dendritic cells (MoDCs) (CD14+ CCR7+ HLA-DR+ CD38+) that functioned as a central immunological node, correlating strongly with both CD20+DN B-cell expansion and highly differentiated CD8+ T cells. Serum analysis further linked this cellular axis to a pro-inflammatory milieu. Our study identifies a convergent pathogenic axis in APS, linking activated MoDCs to dysregulated B-cell differentiation and systemic CD8 T-cell activation. These findings highlight the CD20+DN B-cell and moDCs signatures as potential biomarkers for disease activity.