BATF-overexpressing Treg cell infusion attenuates experimental osteoarthritis by enhancing Treg immunoregulatory function

Osteoarthritis (OA) is a whole-joint degenerative disease characterized by progressive cartilage damage and persistent local immune dysregulation. Regulatory T cells (Tregs) contribute to immune homeostasis, but their functional heterogeneity and the molecular mechanisms governing their activity in OA remain incompletely understood. This study aimed to characterize Treg-cell heterogeneity in joint-associated tissues, investigate the role of basic leucine zipper ATF-like transcription factor (BATF) in regulating Treg function, and evaluate whether BATF overexpression enhances the therapeutic activity of adoptively transferred Treg cells in experimental OA. Public bulk transcriptomic and single-cell RNA-sequencing datasets were analyzed to characterize disease-associated immune alterations, chondrocyte states, and synovial Treg-cell heterogeneity. BATF was overexpressed or knocked out in mouse Treg cells, followed by analyses of proliferation, IL-10 and TGF-β1 secretion, and suppression of Th17-cell proliferation. ChIP-qPCR, dual-luciferase reporter assays, qRT-PCR, and Western blotting were used to examine the potential BATF-IL-2RB-JAK1/STAT5 regulatory relationship. FOXP3/BATF double immunofluorescence was performed to assess BATF-positive Treg cells in control and OA joint tissues. The therapeutic effects of BATF-overexpressing Treg cells were evaluated in papain- and DMM-induced OA models. An empty-vector-transfected Treg-cell group was included in the papain-induced model to distinguish the effects of BATF overexpression from those of Treg-cell transfer itself. Transcriptomic analyses identified disease-associated inflammatory alterations and impaired Treg-related immune features, while single-cell analysis revealed functionally heterogeneous synovial Treg populations. BATF-high Treg cells displayed stronger immunosuppressive transcriptional characteristics. FOXP3/BATF double immunofluorescence showed a reduced abundance of FOXP3⁺BATF⁺ Treg cells in OA joint tissues compared with controls. In vitro, BATF overexpression enhanced Treg-cell proliferation, increased IL-10 and TGF-β1 secretion, and strengthened suppression of Th17-cell proliferation, whereas BATF knockout produced opposite effects. ChIP-qPCR, promoter-reporter assays, and signaling analyses supported an association between BATF-mediated IL-2RB transcription and JAK1-STAT5 activation. In vivo, BATF-overexpressing Treg-cell transfer attenuated cartilage degeneration, preserved Collagen II and Aggrecan expression, and improved the local inflammatory cytokine profile. In the papain-induced OA model, these effects were greater than those observed following NC-Treg transfer. BATF is associated with an enhanced immunosuppressive phenotype of Treg cells and may contribute to their functional regulation partly through the IL-2RB/JAK1-STAT5 signaling axis. BATF overexpression enhanced the therapeutic activity of adoptively transferred Treg cells in experimental OA, supporting further investigation of functionally engineered Treg cells as a potential immune-modulatory strategy for OA. Further studies are required to validate the underlying mechanism in vivo and thoroughly evaluate potential side effects, off-target tissue distribution, long-term phenotypic stability, and the translational safety of this engineered cell therapy.