Background/Purpose: We recently identified a human B cell population, which is naturally autoreactive and tolerized by functional anergy. This population was naïve, fully mature, negative for surface IgM and expressing only IgD BCR (BND cells: naïve, D-only). The BND cells have antibody variable region genes in germline configuration and a significant proportion of anergic cells exhibit natural reactivity to antinuclear antigens and dsDNA. Break of B cell tolerance leading to presence of autoantibodies in systemic autoimmune rheumatic diseases may cause the initiation and perpetuation of autoimmune processes. The aim of this study was therefore to investigate if anergy in BND cells could be overcome by T cell mediated signals, as has been reported in murine systems. Furthermore, investigate the molecular mechanisms underlying this process, and determine whether autoreactive BND cells are tolerized in Systemic Lupus Erythematosus  (SLE) patients.

Methods: Untouched naive and BND human B cells from healthy blood donors or SLE patients were purified and cultured in the presence of T cell help (e.g. IL-4 and CD40L) followed by functional assays such as Ca2+-flux, phospho-flow (pSyk, pLyn) and standard flow cytometry measurements of intracellular and surface B cell markers. Finally, CD45 phosphatase activity was measured utilizing a novel fluorescent peptide-based assay.

Results: Here we report that T helper signals reverse the state of anergy allowing BND cells to fully respond to antigenic stimulation by restoring signaling through the B cell receptor. The mechanism was dependent on increased activity of the tyrosine phosphatase CD45, and CD45-dependent activation of Lyn and Syk kinases. CD45 phosphatase activity was upregulated by T cell help both in BND and naïve B cells, suggesting that CD45 is a key regulator of B cell receptor signaling thresholds mediated by T cell help. Furthermore, we found that BND cells obtained from SLE patients exhibited increased CD45 activity and B cell receptor signaling capacity, thus being less tolerized,than BND cells from healthy controls.

Conclusion: We discovered that T helper signals regulate CD45 phosphatase activity in B cells, and that this lead to increased B cell receptor signaling and recovered ability of tolerized BND cells to respond to antigen. Strikingly, we found that BND cells obtained from SLE patients were less tolerized than BND cells from healthy controls. This raises the possibility that BND cells could represent precursors of autoantibody-secreting plasma cells and suggests a role for these autorective B cells in contributing to autoimmunity if not properly controlled. Our data provide new insight in the break of humoral immune tolerance with possible diagnostic and therapeutic implications in patients with SLE and other systemic autoimmune and rheumatic diseases.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/incomplete-tolerance-in-anergic-b-cells-in-sle-t-helper-signals-restore-b-cell-receptor-signaling-in-autoreactive-anergic-b-cells-by-upregulating-cd45-phosphatase-activity/