B cells induced regulatory T cells attenuated the classical M1 polarization of mouse bone marrow-derived macrophages

Regulatory T (Treg) cells are effective immunomodulators of adaptive and innate immune responses. Our previous studies have demonstrated that B-cell-induced CD4⁺Foxp3^- regulatory T cells, referred to as Treg-of-B cells, exert suppressive capacity, by inhibiting CD4⁺CD25^- T-cell proliferation and inflammasome activation. In the present study, Treg-of-B cells downregulated proinflammatory M1-like markers and partially induced M2-associated genes in unpolarized bone marrow-derived macrophages (BMDMs), as evidenced by RNA expression of Nos2, Arg1, Retnla, Mrc1, and Egr2. Treg-of-B cells decreased the RNA levels of Nos2, Tnfa, Cd86, and Cxcl9, and reduced the production of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and nitrite in LPS/interferon (IFN)-γ-stimulated M1-like macrophages in a dose-dependent manner. These cells also secreted Th2 cytokines, including IL-10, IL-4, and IL-13, with enhanced cytokine production observed when cocultured with macrophages. Mechanistically, Treg-of-B cells exerted their modulatory effects via both cell-cell contact and contact-dependent induction of soluble mediators, particularly Th2 cytokines. Furthermore, Treg-of-B cells promoted IκBα accumulation and suppressed RNA expression of Kruppel-like factor 4 (Klf4), thereby inhibiting NF-κB activation. These findings suggest that Treg-of-B cells regulate macrophage plasticity and might prevent excessive inflammation.