Phosphatidylcholine-Specific B Cells Are Enriched Among Atypical CD11c high and CD21 low Memory B Cells in Antiphospholipid Syndrome

Eduard Tim Nitschke^1,2*Van Duc Dang^1,2,3Hector Rincon-Arevalo^1,2,4,5Franziska Szelinski^1,2Jacob Ritter^1,2Eva Schrezenmeier^2,5Tobias Alexander^1,2Tuan Anh Le^1,2Yidan Chen^1,2Annika Wiedemann^1,2Jose Bernardino González-González⁶Andreia C Lino²Ana-Luisa Stefanski^1,2Thomas Dörner^1,2

• ¹Charité – Universitätsmedizin Berlin, corporate member of Freie Universität Berlin and Humboldt- Universität zu Berlin, Department of Rheumatology and Clinical Immunology, Charitéplatz 1, 10117 Berlin, Berlin, Germany
• ²German Rheumatism Research Center (DRFZ), Berlin, Baden-Württemberg, Germany
• ³VNU University of Science, Vietnam National University, Faculty of Biology, Hanoi, Vietnam, Hanoi, Vietnam
• ⁴Universidad de Antioquia UdeA, Grupo de Inmunología Celular e Inmunogenética, Facultad de Medicina, Instituto de Investigaciones Médicas, Medellín, Colombia, Medellín, Colombia
• ⁵Charité – Universitätsmedizin Berlin, corporate member of Freie Universität Berlin and Humboldt- Universität zu Berlin, Department of Nephrology and Medical Intensive Care, Charitéplatz 1, 10117, Berlin, Germany
• ⁶Labor Berlin Charité Vivantes GmbH, Berlin, Berlin, Germany

Background: Patients with antiphospholipid syndrome (APS) carry an increased risk of thrombosis and adverse pregnancy outcomes due to the presence of antiphospholipid autoantibodies (aPL). However, the pathogenesis of the disease remains incompletely understood regarding various aPL and the role of autoreactive B cells as precursors of antibodysecreting plasma cells (PC).Objective: To assess B-cell dysregulation in APS, with a focus on the distribution of B cell subsets and phosphatidylcholine (PtC)-specific cells.We used flow cytometry to study B cell subsets in peripheral blood mononuclear cells (PBMCs) from 20 healthy controls (HCs), 21 patients with primary APS (pAPS), and 16 patients with secondary APS (sAPS). A novel fluorescent liposome-based method was used to identify PtC-specific B cells in these subsets. Data were analyzed using manual gating and unsupervised clustering. We quantified aPtC antibody serum levels using ELISA and conducted correlation analyses between PtC-specific B cell subsets and aPL titers. Results: Patients with pAPS and sAPS exhibited significantly increased frequencies of atypical CD21 low and CD11c high B cells, including PtC-specific B cells. Notably, both total and unswitched memory PtC-specific B cells were elevated in pAPS patients and correlated with aPL antibody titers. Unsupervised clustering further highlighted the increased frequencies of PtC-specific CD21 low CD11c high unswitched and switched memory B cells in both pAPS and sAPS.The enrichment of PtC-specific B cells among CD21 low and CD11c high atypical memory subsets, along with their correlation with aPL serum levels, suggest a linkage between these atypical memory B cell subsets and autoantibody producing cells in APS.