Abstract: FR-PO830
Patient-Derived Cell Line as an In Vitro Model System for Autoantigen-Specific B Cells in Myeloperoxidase-ANCA
Session Information
- Glomerular Diseases: Inflammation and Immunology
October 25, 2024 | Location: Exhibit Hall, Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Glomerular Diseases
- 1401 Glomerular Diseases: Mechanisms, including Podocyte Biology
Authors
- Moon, Young-Hyun, The University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, North Carolina, United States
- Chen, Dhruti P., The University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, North Carolina, United States
- Taylor, Justin J., University of Virginia School of Medicine, Charlottesville, Virginia, United States
- Falk, Ronald, The University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, North Carolina, United States
- Bunch, Donna O., The University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, North Carolina, United States
Background
Autoantigen-specific B cells are important in ANCA vasculitis but are rare and hard to study. Existing myeloperoxidase (MPO)-ANCA vasculitis mouse models have been insightful although limited by their non-human nature, time intensiveness and cost. We hypothesized that a patient-derived B cell line could be modulated to differentiate into antibody-producing cells and establish an in vitro model system to inform specific targeting of human B cells.
Methods
We created Epstein-Barr virus immortalized B cell lines with PBMCs from patients with ANCA vasculitis (23 MPO, 23 PR3). Two lines were cultured in “B cell media” (IMDM, penicillin-streptomycin, 10% FBS, GlutaMax, β-ME, CD40L, CpG) with or without MPO autoantigen and IL-21. Autoantibody production was tracked longitudinally by testing supernatants for anti-MPO IgG by ELISA. Cells were stained with allophycocyanin (APC)-tagged MPO-tetramers, positively selected for APC, and analyzed by flow cytometry.
Results
Immortalized cells maintained expression of B cell markers (CD19, CD22) and 4% MPO-specificity. Cells cultured with MPO and IL-21 had more CD27high/CD22low B cells, consistent with plasmablast differentiation, in both parent (9%) and antigen-specific populations (15%) compared to cells cultured without MPO or IL-21 (5 and 10% respectively) (Fig 1a). This cell line produced ANCAs; cells cultured with MPO and IL-21 produced more autoantibody than cells grown without (Fig 1b). Cells cultured to 92 days maximally had 29% MPO-specificity.
Conclusion
We developed immortalized patient-derived B cell lines that maintain antigen-specificity in culture, with targetable B cell markers, and inducible autoantibody production. This system could be used to test B cell-depleting and autoantigen-specific therapies. In summary, an immortalized patient-derived cell line offers a viable model system of autoantigen-specific B cells in ANCA vasculitis.
Funding
- NIDDK Support