Sublethal DNA Damage Switches Off B-Cell Effector Programs in an RA-FLS- PBMC Co-culture

Rheumatoid arthritis (RA) features lymphocyte-driven inflammation in which B cells, alongside T cells, play key effector roles (autoantibody production, antigen presentation, cytokines, or chemokines). Within B cells, during normal diversification, activation-induced cytidine deaminase (AID) introduces targeted DNA lesions in immunoglobulin loci (class-switch recombination/somatic hypermutation), creating a potential vulnerability to sublethal genotoxic stress. T cells also contribute to RA pathogenesis through cytokine production and cell-mediated responses, and are exposed to similar genotoxic stressors in the inflamed joint environment. Given this, we asked whether a single, sublethal insult can modulate lymphocyte effector function without overt cytotoxicity.Peripheral blood mononuclear cells from healthy donors were co-cultured with RA fibroblast-like synoviocytes and exposed once to an IC₂₀ or IC₅₀ concentration of γ-irradiation (γ-IR), hydrogen peroxide (H₂O₂), or the oxazaphosphorine metabolite 4-hydroperoxyifosfamide (4-OOH IFA). Viability, γ-H2AX kinetics, cell cycle status, cytokine and immunoglobulin secretion, and a 28-gene damage response/differentiation panel were quantified at either 24 hours or 5 days.Together, the data indicate that a single, carefully titrated low-concentration genotoxic hit can selectively suppress lymphocyte effector programs, with B cells being more durably affected than T cells. At 2 Gy, overall cell viability remained above 80%, whereas IL-10 expression declined by approximately 70%, demonstrating functional silencing in the absence of substantial cytotoxicity. Targeting this vulnerability may selectively dampen pathogenic B cell activity in RA while sparing overall immune viability and T-cell competence.