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ObjectiveRegulatory T cells (Tregs) play a pivotal role in tumor immune evasion, and strategies to overcome their immunosuppressive activity are urgently needed. This study investigates the immunomodulatory effects of microwave ablation (MWA) on TNFRSF4+ Tregs, focusing on the OX40L/TNFRSF4 signaling axis as a potential therapeutic target.MethodsTNFRSF4+ Tregs were isolated from C57BL/6 mice and subjected to MWA-mimetic thermal stress. In vitro functional assays and in vivo LLC xenograft models were employed, with OX40 agonist intervention. Molecular mechanisms were analyzed via RT-qPCR, Western blot, and immunohistochemistry. The balance of tumor-infiltrating immune cells was quantified by multi-color flow cytometry.ResultsMWA induced three key effects: (1) Phenotypic shift: decreased CTLA-4+ (P<0.0001) Treg subsets, but increased OX40L+ (P<0.01) in LLC cells; (2) Functional impairment: reduced Treg-mediated support for LLC proliferation, migration, and invasion; (3) Enhanced CD8+ T cell cytotoxicity. In vivo, MWA reshaped the tumor microenvironment by significantly increasing the intratumoral CD8+/Treg ratio (P<0.001), indicating a shift toward an anti-tumor inflammatory state. Mechanistically, MWA suppressed NF-¦ÊB/I¦ÊB¦Á/TRAF6 signaling, and these effects were amplified by an OX40 agonist, suggesting the pathway is potentially OX40L-dependent.ConclusionThis study demonstrates that MWA disrupts Treg immunosuppression, likely by activating OX40L/TNFRSF4 signaling, and favorably alters the balance of effector to suppressor cells, providing a novel rationale for combining thermal ablation with OX40-targeted immunotherapies in cancer treatment.

Although FOXP3+ regulatory T cells (Treg) depend on IL-2 produced by other cells for their survival and function, the levels of IL-2 in inflamed tissue are low, making it unclear how Treg access this critical resource. Here, we show that Treg use heparanase (HPSE) to access IL-2 sequestered by heparan sulfate (HS) within the extracellular matrix (ECM) of inflamed central nervous system tissue. HPSE expression distinguishes human and murine Treg from conventional T cells and is regulated by the availability of IL-2. HPSE-/- Treg have impaired stability and function in vivo, including in the experimental autoimmune encephalomyelitis (EAE) mouse model of multiple sclerosis. Conversely, endowing monoclonal antibody-directed chimeric antigen receptor (mAbCAR) Treg with HPSE enhances their ability to access HS-sequestered IL-2 and their ability to suppress neuroinflammation in vivo. Together, these data identify a role for HPSE and the ECM in immune tolerance, providing new avenues for improving Treg-based therapy of autoimmunity. Regulatory T cell (Treg) maintenance and function require IL-2, yet this cytokine is only present in low levels in vivo. In this study, the authors demonstrate that that Treg use heparanase to access IL-2 bound to heparan sulfate proteoglycans in the extracellular matrix of inflamed brain tissue in mice.

Rheumatoid arthritis (RA) is characterized by an impaired articular bone immune microenvironment, which is associated with regulatory T cells (Tregs) hypofunction and osteoclasts (OCs) hyperfunction and leads to articular bone erosion and systemic bone loss. Studies have shown that Tregs slow bone loss in RA by regulating the bone resorption function of OCs and the JAK/STAT signaling pathway can regulate the immunosuppressive function of Tregs and reduce the bone erosion function of OCs. Yi Shen Juan Bi Pill (YSJB) is a classic Chinese herbal compound for the treatment of RA. However, whether YSJB regulates bone immune microenvironment homeostasis through JAK/STAT signaling pathway remains unclear. Based on in vitro OC single culture, Treg single culture and OC-Treg coculture systems, treatments were performed using drug-containing serum, AG490 and JAK2 siRNA to explore whether YSJB-containing serum regulates the homeostasis of the bone immune microenvironment through the JAK/STAT signaling pathway. In vitro, YSJB treatment decreased the number of TRAP+ cells and the areas of bone resorption and inhibited the expression of RANK, NFATc1, c-fos, JAK2, and STAT3 in both the OC single culture system and the OC-Treg coculture system. Tregs further reduced the number of TRAP+ cells and the areas of bone resorption in the coculture system. YSJB promoted the secretion of IL-10 while inhibiting the expression of JAK2 and STAT3 in Tregs. Moreover, inhibiting the expression of JAK2 with the JAK2 inhibitor AG490 and JAK2 siRNA improved the immunosuppressive functions of Treg, inhibited OC differentiation and bone resorption. Our study demonstrates that YSJB can regulate OC-mediated bone resorption and Treg-mediated bone immunity through the JAK2/STAT3 signaling pathway. This study provides a new strategy for regulating the bone immune microenvironment in RA with traditional Chinese medicine.