Our previous findings demonstrated that naïve B cells elicit suppressive CD4+ regulatory T (Treg) cells, named as Treg-of-B cells. However, the capability of antigen-specific B cells in that process remains unclear. Using ovalbumin (OVA) as a model antigen, the present study showed that B cells from OVA-immunised mice decreased that ability. Instead, OVA-activated OVA-specific (OB1) B cells induced effector-like T-of-OB1 cells without regulatory function. Phenotypically, Treg-of-B cells reduced the production of interferon (IFN)-γ, interleukin (IL)-17 and IL-2 and expressed CD62L, PD1 and endothelial cell adhesion molecule 1 (PECAM1). Functionally, adoptive transfer of Treg-of-B cells significantly attenuated Th1 cell-mediated delayed-type hypersensitivity (DTH) responses and inhibited IFN-γ-producing Th1 cells, while T-of-OB1 cells did not. Mechanistically, activated antigen-specific B cells increased the expression of costimulatory molecules and promoted higher T cell activation, contributing to effector T cell phenotype. Conversely, Treg-of-B cells exhibited lower T cell activation, possibly mediated through the expression of PECAM1, Dusp2, Dusp5, Ptpn7, Ptpn22 and Ms4a4b. These findings suggest that non-antigen-specific B cells elicit CD4+ Treg cells, potentially via attenuating T cell activation, whereas that capacity is absent in antigen-specific B cells. This distinction underscores the critical role of B cell antigen specificity in immune regulation and inflammation.
© 2025 John Wiley & Sons Ltd.

Gain-of-function mutations in the dsDNA sensing adaptor STING lead to a severe autoinflammatory syndrome known as STING-associated vasculopathy with onset in infancy (SAVI). Patients with SAVI develop interstitial lung disease (ILD) and produce autoantibodies that are commonly associated with systemic autoimmune diseases. Mice expressing the most common SAVI mutation, STING V154M (VM), similarly develop ILD but exhibit severe T and B cell lymphopenia and low serum Ig titers, and they lack autoantibodies. Importantly, lethally irradiated VM hosts reconstituted with WT stem cells (WT→VM) still develop ILD. In this study, we find that WT→VM chimeras had restored B cell function, produced autoantibodies, and thereby recapitulated the loss of tolerance seen in patients with SAVI. Lymphocytes derived from both WT and BCR or TCR transgenic (Tg) donors accumulated in the extravascular lung tissue of WT+Tg→VM mixed chimeras, but lymphocyte activation and germinal center formation required WT cells with a diverse repertoire. Furthermore, when T cells isolated from the WT→VM chimeras were adoptively transferred to naive Rag1-deficient secondary hosts, they trafficked to the lung and recruited neutrophils. Overall, these findings indicated that VM expression by radioresistant cells promoted the activation of autoreactive B cells and T cells that then differentiated into potentially pathogenic effector subsets.

Peyer's patches, splenic, and peritoneal B cells induce regulatory T (Treg-of-B) cells in vitro without exogenous chemicals and cytokines. Treg-of-B cells exert suppressive function both in vitro and in vivo. Here, we demonstrate experimental procedures for the generation and characterization of Treg-of-B cells. We describe steps for isolating B220+ B cells, isolating CD4+CD25- T cells, inducing Treg-of-B cells, identifying Treg-of-B cells by flow cytometry, cytokine analyzing by ELISA, and functional testing by suppression assay. For complete details on the use and execution of this protocol, please refer to Chien et al.1 and Chu et al.2.
Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.

The Bursa of Fabricius, an avian unique humoral immune organ, is instrumental to B cell development. Bursal-derived peptide BP9 fosters B-cell development and formation. Yet, the exact mechanism wherein BP9 impacts B cell differentiation and antigenic presentation remains undefined. In this paper, B cell activation and differentiation in the spleen cells from mice immunized with the AIV vaccine and BP9 were detected following flow cytometry (FCM) analysis. Furthermore, the molecular mechanism of BP9 in B cell differentiation in vivo was investigated with RNA sequencing technology. To verify the potential functional mechanism of BP9 in the antigenic presentation process, the transcriptome molecular basis of chicken macrophages stimulated by BP9 was measured via high-throughput sequencing technology. The results proved that when given in experimental dosages, BP9 notably accelerated total B cells, and enhanced B-cell differentiation and plasma cell production. The gene expression profiles of B cells from mice immunized with 0.01 mg/mL BP9 and AIV vaccine disclosed that 0.01 mg/mL BP9 initiated the enrichment of several biological functions and significantly stimulated key B-cell pathways in immunized mice. Crucially, a total of 4093 differentially expressed genes were identified in B cells with BP9 stimulation, including 943 upregulated genes and 3150 downregulated genes. Additionally, BP9 induced various cytokine productions in the chicken macrophage HD11 cells and activated 9 upregulated and 20 downregulated differential miRNAs, which were involved in various signal and biological processes. Furthermore, BP9 stimulated the activation of multiple transcription factors in HD11 cells, which was related to antigen presentation processes. In summary, these results suggested that BP9 might promote B cell differentiation and induce antigen presentation, which might provide the valuable insights into the mechanism of B cell differentiation upon bursal-derived immunomodulating peptide stimulation and provide a solid experimental groundwork for enhancing vaccine-induced immunity.

Intracellular surveillance for systemic microbial components during homeostasis and infections governs host physiology and immunity. However, a long-standing question is how circulating microbial ligands become accessible to intracellular receptors. Here we show a role for host-derived extracellular vesicles (EVs) in this process; human and murine plasma-derived and cell culture-derived EVs have an intrinsic capacity to bind bacterial lipopolysaccharide (LPS). Remarkably, circulating host EVs capture blood-borne LPS in vivo, and the LPS-laden EVs confer cytosolic access for LPS, triggering non-canonical inflammasome activation of gasdermin D and pyroptosis. Mechanistically, the interaction between the lipid bilayer of EVs and the lipid A of LPS underlies EV capture of LPS, and the intracellular transfer of LPS by EVs is mediated by CD14. Overall, this study demonstrates that EVs capture and escort systemic LPS to the cytosol licensing inflammasome responses, uncovering EVs as a previously unrecognized link between systemic microbial ligands and intracellular surveillance.
© 2023. The Author(s), under exclusive licence to Springer Nature Limited.