OX40 is a recently identified T-cell costimulatory molecule that belongs to the TNF/TNFR superfamily. OX40 can be expressed by both activated T effector cells and Foxp3⁺ Tregs. It is well known that OX40 delivers a potent costimulatory signal to T effector cells, but very little is known about the role of OX40 in regulating the suppressor properties of Foxp3⁺ Tregs and the de novo generation of new inducible Foxp3⁺ Tregs from T effector cells. In the present study, we found, by using a newly created foxp3gfp knockin model, that OX40 was dispensable for the genesis and suppressor functions of naturally arising CD4⁺Foxp3⁺ Tregs, but stimulating OX40 on the Foxp3⁺ Tregs abrogated their ability to suppress T effector cell proliferation, IFN-γ production, and T effector cell-mediated allograft rejection. OX40 costimulation did not significantly affect proliferation and survival of the naturally arising Foxp3⁺ Tregs, but profoundly inhibited Foxp3 gene expression. Importantly, OX40 costimulation to T effector cells prevented the induction of new inducible Foxp3⁺ Tregs from T effector cells. Our study identified OX40 as a key negative regulator of Foxp3⁺ Tregs and may have important clinical implications in models of transplantation and autoimmunity.

Whereas a B cell–transcriptional profile has been recorded for operationally tolerant kidney graft patients, the role that B cells have in this tolerance has not been reported. In this study, we analyzed the role of B cells from operationally tolerant patients, healthy volunteers, and kidney transplant recipients with stable graft function on T cell suppression. Proliferation, apoptosis, and type I proinflammatory cytokine production by effector CD4⁺CD25⁻ T cells were measured after anti-CD3/anti-CD28 stimulation with or without autologous B cells. We report that B cells inhibit CD4⁺CD25⁻ effector T cell response in a dose-dependent manner. This effect required B cells to interact with T-cell targets and was achieved through a granzyme B (GzmB)–dependent pathway. Tolerant recipients harbored a higher number of B cells expressing GzmB and displaying a plasma cell phenotype. Finally, GzmB⁺ B-cell number was dependent on IL-21 production, and B cells from tolerant recipients but not from other patients positively regulated both the number of IL-21⁺ T cells and IL-21 production, suggesting a feedback loop in tolerant recipients that increases excessive B cell activation and allows regulation to take place. These data provide insights into the characterization of B cell–mediated immunoregulation in clinical tolerance and show a potential regulatory effect of B cells on effector T cells in blood from patients with operationally tolerant kidney grafts.

Regulatory T (T_reg) cells are indispensable for maintaining peripheral tolerance, whereas T helper (Th)1 and Th17 cells induce inflammation and tissue destruction. Using Foxp3‐GFP knock‐in mice, we report a novel regulatory role for B cell subsets in influencing the differentiation of T_reg versus Th1/Th17 cells. Peritoneal B1 cells strongly promoted T cell proliferation and cytokine secretion when presenting nominal or allogeneic antigens, as compared to conventional follicular B (B2) cells. However, peritoneal B1 cells largely failed to convert naive Foxp3^–CD4⁺ T cells into Foxp3⁺ T_reg cells in the presence of TGF‐β and IL‐2, in marked contrast to conventional B2 cells, which excelled in T_reg conversion. Interestingly, under the same T_reg conversion conditions, peritoneal B1 cells preferentially promoted Th1 and Th17 cell differentiation. Blockade of CD86 but not CD80 costimulation markedly enhanced T_reg cell induction by B1 cells. Thus, B cell antigen presentation function is inversely correlated with de novo T_reg cell induction for these B cell subsets. Our findings suggest that B1 and B2 cell subsets play distinct roles in immune regulation by promoting reciprocal differentiation of T cell lineages.

Invasive insulitis is a destructive T cell-dependent autoimmune process directed against insulin-producing β cells that is central to the pathogenesis of type 1 diabetes mellitus (T1DM) in humans and the clinically relevant nonobese diabetic (NOD) mouse model. Few therapies have succeeded in restoring long-term, drug-free euglycemia and immune tolerance to β cells in overtly diabetic NOD mice, and none have demonstrably enabled enlargement of the functional β cell mass. Recent studies have emphasized the impact of inflammatory cytokines on the commitment of antigen-activated T cells to various effector or regulatory T cell phenotypes and insulin resistance and defective insulin signaling. Hence, we tested the hypothesis that inflammatory mechanisms trigger insulitis, insulin resistance, faulty insulin signaling, and the loss of immune tolerance to islets. We demonstrate that treatment with α1-antitrypsin (AAT), an agent that dampens inflammation, does not directly inhibit T cell activation, ablates invasive insulitis, and restores euglycemia, immune tolerance to β cells, normal insulin signaling, and insulin responsiveness in NOD mice with recent-onset T1DM through favorable changes in the inflammation milieu. Indeed, the functional mass of β cells expands in AAT-treated diabetic NOD mice.

Operationally tolerant patients (TOL) display a higher number of blood B cells and transcriptional B cell signature. As they rarely develop an allo‐immune response, they could display an abnormal B cell differentiation. We used an in vitro culture system to explore T‐dependent differentiation of B cells into plasma cells. B cell phenotype, apoptosis, proliferation, cytokine, immunoglobulin production and markers of differentiation were followed in blood of these patients. Tolerant recipients show a higher frequency of CD20⁺CD24^hiCD38^hi transitional and CD20⁺CD38^loCD24^lo naïve B cells compared to patients with stable graft function, correlating with a decreased frequency of CD20⁻CD38⁺CD138⁺ differentiated plasma cells, suggestive of abnormal B cell differentiation. B cells from TOL proliferate normally but produce more IL‐10. In addition, B cells from tolerant recipients exhibit a defective expression of factors of the end step of differentiation into plasma cells and show a higher propensity for cell death apoptosis compared to patients with stable graft function. This in vitro profile is consistent with down‐regulation of B cell differentiation genes and anti‐apoptotic B cell genes in these patients in vivo. These data suggest that a balance between B cells producing IL‐10 and a deficiency in plasma cells may encourage an environment favorable to the tolerance maintenance.