3c), and it has been difficult to define the sources of TGF-1 that are relevant to immune suppression. T cells (Th cells) that are functionally specialized to provide help to B cells, allowing the formation of GC and the subsequent long-lived plasma cell differentiation. Therefore, regulation of the quality and quantity of TFH cells and memory B-cell populations in GC (GCB) is important to prevent immunopathology. CD4+CD25+ Treg (CD25+ Treg) that express Foxp3 play the key roles in the Col6a3 maintenance of self-tolerance and suppress the activation of conventional T cells and dendritic cells3. Moreover, accumulating evidence indicates the essential role of CD25+ Treg, including CD4+CD25+CXCR5+ follicular Treg2 and CD4+CD25+CD69? Treg4, in the regulation of humoral immunity. These observations highlight the protective role of CD25+ Treg in systemic autoimmunity; however, the disease induced by the absence of functional CD25+ Treg is quite different from SLE1,5. Moreover, a role for CD25+ Treg in SLE has not been clearly established6. Recent advances in understanding of CD8+ Treg have underscored the importance of Qa-1-restricted CD8+ Treg for the maintenance of B-cell tolerance. Mice with functional impairment in CD8+ Treg exhibit a lupus-like disease with a significant increase in TFH7. The development of systemic autoimmunity in B6.mutant mice is associated with a pronounced defect in CD8+ Treg activity8. Nevertheless, the actual contribution of CD8+ Treg to the regulation of human autoimmunity remains unclear. Early growth response gene 2 (Egr2), a zinc-finger transcription factor, plays a critical role in hindbrain development and myelination of the peripheral nervous system9. In T cells, Egr2 is important for the maintenance of T-cell anergy by negatively regulating T-cell activation10. The involvement of Egr2 in the control of systemic autoimmunity was first suggested by the observation that lymphocyte-specific Egr2-deficient mice develop a lupus-like disease with no impact on the development of Foxp3-expressing CD25+ Treg11. Moreover, mice deficient for both Egr2 and Egr3 in B and T cells present lethal and early-onset systemic autoimmunity, suggesting a synergistic role for Egr2 and Egr3 in controlling B-cell tolerance12. We and our collaborators have shown that polymorphisms in influence SLE susceptibility in humans13. We have previously identified Egr2-controlled CD4+CD25?LAG3+ Treg (LAG3+ Treg)14. LAG3 is a CD4-related molecule that binds to MHC class II, and the binding induces immunoreceptor tyrosine-based activation motif (ITAM)-mediated inhibitory signalling15. Approximately 2% of the CD4+CD25? T-cell population in the spleen express LAG3. These LAG3+ Treg produce high levels of interleukin (IL)-10 and are suppressive in a murine model of colitis in an IL-10-dependent manner. Unlike CD25+ Treg, high-affinity interactions with selecting peptide/MHC ligands expressed in the thymus do not induce the development of LAG3+ Treg. Recently, Gagliani lupus-prone mice, adoptive transfer of LAG3+ Treg from MRL/+ mice suppresses the progression of lupus in a TGF-3-dependent manner. Expression of both Fas and Egr2 by LAG3+ Treg is necessary for TGF-3 production and for the suppression of humoral immunity. These results clarify the mechanisms underlying LAG3+ Azelaic acid Treg-mediated B-cell regulation. Results Egr2 mediates control of humoral immunity by LAG3+ Treg To clarify the role of Egr2 in Azelaic acid T cells, we generated T-cell-specific Egr2 conditional knockout (CKO) mice (B-cell antibody production and the development of TFH and GCB (Fig. 1e,f). Thus, the expression of Egr2 on LAG3+ Treg is necessary for the suppression of B-cell responses. In transgenic mice that express green fluorescent protein (GFP) under the control of the Egr2 promoter (Egr2-GFP mice; Supplementary Fig. 3a), the expression of GFP in CD4+ T cells correlated with Egr2 protein expression (Supplementary Fig. 3b). The importance of Egr2 was confirmed by the observation that CD4+CD25?Egr2-GFP+ cells from Egr2-GFP mice also exhibited B-cell-suppressive activity B-cell suppression by LAG3+ Treg. Each T-cell subset stimulated with anti-CD3 mAb was co-cultured with stimulated B cells. (c) Live B220+ B cells were quantified with AnnexinV/PI staining 72?h after anti-IgM stimulation (NP-specific antibody responses. C57BL/6 (B6) B cells and OT-II CD4+CD25?LAG3? Th cells were injected into Rag1KO mice in combination with or without LAG3+ Treg from B6 mice 1 day before the immunization with NP-OVA/alum, and Azelaic acid given a booster.
