森田 薫

TGF-β is a pleotropic cytokine involved in various biological processes. Of the three isoforms of TGF-β, TGF-β1 has long been recognized as an important inhibitory cytokine in the immune system and has been reported to inhibit B cell function in both mice and humans. Recently, it has been suggested that TGF-β3 may play an important role in the regulation of immune system in mice. Murine CD4+CD25-LAG3+ regulatory T cells suppress B cell function through the production of TGF-β3, and it has been reported that TGF-β3 is therapeutic in a mouse model of systemic lupus erythematosus. The effect of TGF-β3 on human B cells has not been reported, and we herein examined the effect of TGF-β3 on human B cells. TGF-β3 suppressed B cell survival, proliferation, differentiation into plasmablasts, and antibody secretion. Although the suppression of human B cells by TGF-β1 has long been recognized, the precise mechanism for the suppression of B cell function by TGF-β1 remains elusive; therefore, we examined the effect of TGF-β1 and β3 on pathways important in B cell activation and differentiation. TGF-β1 and TGF-β3 inhibited some of the key molecules of the cell cycle, as well as transcription factors important in B cell differentiation into antibody secreting cells such as IRF4, Blimp-1, and XBP1. TGF-β1 and β3 also inhibited B cell receptor signaling. Our results suggest that TGF-β3 modifying therapy might be therapeutic in autoimmune diseases with B cell dysregulation in humans.

Systemic lupus erythematosus (SLE) is a prototypical autoimmune disease characterized by multiorgan inflammation induced by autoantibodies. Early growth response gene 2 (Egr2), a transcription factor essential for T-cell anergy induction, controls systemic autoimmunity in mice and humans. We have previously identified a subpopulation of CD4+ regulatory T cells, CD4+CD25-LAG3+ cells, that characteristically express both Egr2 and LAG3 and control mice model of lupus via TGF-β3 production. However, due to the mild phenotype of lymphocyte-specific Egr2-deficient mice, the presence of an additional regulator has been speculated. Here, we show that Egr2 and Egr3 expressed in T cells cooperatively prevent humoral immune responses by supporting TGF-β3 secretion. T cell-specific Egr2/Egr3 double-deficient (Egr2/3DKO) mice spontaneously developed an early onset lupus-like disease that was more severe than in T cell-specific Egr2-deficient mice. In accordance with the observation that CD4+CD25-LAG3+ cells from Egr2/3DKO mice completely lost the capacity to produce TGF-β3, the excessive germinal center reaction in Egr2/3DKO mice was suppressed by the adoptive transfer of WT CD4+CD25-LAG3+ cells or treatment with a TGF-β3-expressing vector. Intriguingly, latent TGF-β binding protein (Ltbp)3 expression maintained by Egr2 and Egr3 was required for TGF-β3 production from CD4+CD25-LAG3+ cells. Because Egr2 and Egr3 did not demonstrate cell intrinsic suppression of the development of follicular helper T cells, Egr2- and Egr3-dependent TGF-β3 production by CD4+CD25-LAG3+ cells is critical for controlling excessive B-cell responses. The unique attributes of Egr2/Egr3 in T cells may provide an opportunity for developing novel therapeutics for autoantibody-mediated diseases including SLE.

SLE is an autoimmune disease characterized by multiple organ damage mediated by autoantibodies and autoreactive T cells. Approaches utilizing genetically engineered mice as well as genome-wide association studies have identified a number of lupus-related genes. Recently, early growth response gene 2 (Egr2) and Egr3 have emerged as regulatory molecules that suppress excessive immune responses. Mice deficient for Egr2 and Egr3 develop a lupus-like disease with dysregulated activation of effector T cells. Furthermore, Egr2 and Egr3 confer suppressive activity to CD4+ T cells and regulate the production of inhibitory cytokines such as IL-10 and TGF-β1. These findings may have implications for a wide range of immune-related pathologies and suggest the possibility that efforts exploiting Egr2 and Egr3 could aid in the development of therapeutic applications. This review summarizes the recent advances regarding the roles of Egr2 and Egr3 on T cells in the control of autoimmunity.

TGF-β family members are multipotent cytokines that are involved in many cellular processes, including cell differentiation, organ development, wound healing and immune regulation. TGF-β has pleiotropic effects on adaptive immunity, especially in the regulation of CD4(+) T cell and B cell responses. Furthermore, identification of CD4(+) T cell subsets that produce TGF-β3 revealed unexpected roles of TGF-β3 in the control of adaptive immunity. In contrast to TGF-β1, which induces extensive fibrosis, TGF-β3 induces non-scarring wound healing and counteracts tissue fibrosis. Recent progress in the understanding of the activation mechanism of TGF-β may enable us to develop novel biologic therapies based on advanced protein engineering.

Autoantibodies induce various autoimmune diseases, including systemic lupus erythematosus (SLE). We previously described that CD4(+)CD25(-)LAG3(+) regulatory T cells (LAG3(+) Treg) are regulated by Egr2, a zinc-finger transcription factor required for the induction of T-cell anergy. We herein demonstrate that LAG3(+) Treg produce high amounts of TGF-β3 in an Egr2- and Fas-dependent manner. LAG3(+) Treg require TGF-β3 to suppress B-cell responses in a murine model of lupus. Moreover, TGF-β3- and LAG3(+) Treg-mediated suppression requires PD-1 expression on B cells. We also show that TGF-β3-expressing human LAG3(+) Treg suppress antibody production and that SLE patients exhibit decreased frequencies of LAG3(+) Treg. These results clarify the mechanism of B-cell regulation and suggest therapeutic strategies.

多くの自己免疫疾患において自己抗体が産生される事が知られている．この自己抗体産生がどのような生理的な抑制機構により制御されているか，は未だよく分かっていない．免疫応答に多様な作用をもつサイトカインの中で，B細胞を抑制するものとしてはTGF-β1が知られている．TGF-β1はCD4陽性CD25陽性Foxp3陽性制御性T細胞（CD25Treg）が産生することが知られているがその量は限定されており，CD25TregによるB細胞抑制能にもTGF-β1あまり関与していない．我々はマウスのCD4陽性CD25陰性LAG3陽性制御性T細胞（LAG3Treg）がTGF-β3産生により，B細胞による抗体産生を抑制することを見出した．TGF-β3はこれまで発生における重要性は知られていたが，免疫抑制能については知られていなかった．ヒトのCD4陽性CD25陰性LAG3陽性細胞もB細胞機能を抑制しTGF-β3を発現するが，この細胞は全身性エリテマトーデスや活動性の関節リウマチ症例では減少している．さらに最近，ヒトTGF-β3がヒトB細胞に対して強い抑制能を発揮する事を確認している．ここでは，このような生理的な免疫抑制機構による自己抗体産生抑制の可能性について，議論してみたい．

【目的】Transforming growth factor beta（TGFβ）は多彩な機能を持つサイトカインであり，TGFβ1は免疫において抑制能を発揮することが知られている．一方，アミノ酸の異なるisoformであるTGFβ3の免疫系における機能はあまり分かっていない．我々は，マウスにおいて，CD4+CD25-LAG3+制御性T細胞がTGFβ3を分泌し，マウスB細胞の増殖・抗体産生を抑制することを報告した．今回我々は，TGFβ3のヒトB細胞に対する作用を検討した．【方法】ヒト末梢血B細胞を用いて，種々の刺激条件下で，TGFβ3によるアポトーシス，増殖，抗体産生，形質細胞への分化を検討した．また，TGFβ3存在下で刺激したB細胞においてRNA-seqによる発現解析を行い，B細胞の活性化や抗体産生に関わるパスウェイへの影響を検討した．【結果】T細胞依存性・非依存性の両方の系において，TGFβ3は，ヒトB細胞のアポトーシスを誘導し，増殖・抗体産生を抑制した．また，TGFβ3は，形質細胞への分化に重要なPRDM1やXBP1の発現を抑制し，形質細胞への分化を抑制した．【結論】TGFβ3は，ヒトB細胞の活性化を抑制し，TGFβ3の治療応用は全身性エリテマトーデスなど自己抗体を伴う自己免疫疾患に有用である可能性が示唆された．

Transforming growth factor-betas (TGF-βs) are multifunctional cytokines that have been implicated in the regulation of a broad range of biological processes, including cell proliferation, cell survival, and cell differentiation. The three isoforms identified in mammals, TGF-β1, TGF-β2, and TGF-β3, have high sequence homology, bind to the same receptors, and show similar biological functions in many in vitro studies. However, analysis of the in vivo functions of the three isoforms and mice deficient for each cytokine reveals striking differences, illustrating their unique biological importance and functional non-redundancy. Although increasing evidence suggests that TGF-β1 and, to a lesser extent, TGF-β2 play an integral role in maintaining immune tolerance, the immunological role of TGF-β3 has not been carefully evaluated. Recent studies have focused on the multifunctional role of TGF-β3. In this review, we provide an overview of the role of TGF-β3 in immunity, with comparison to TGF-β1 and -β2.

Autoantibodies are associated with various autoimmune diseases. Systemic lupus erythematosus (SLE) is an autoantibody-associated autoimmune disease which affects multiple organs. Although both genetic and environmental factors are implicated in lupus pathogenesis, the etiology of the disease remains elusive. The discovery of CD4(+)CD25(+) regulatory T cells (Tregs) that characteristically express forkhead box p3 (Foxp3) gene have greatly advanced our understanding of immune systems in autoimmune diseases. CD4(+) Tregs can be classified into two main populations: thymus-derived naturally occurring Tregs (nTregs) and induced Tregs (iTregs) generated from CD4(+)CD25(-) precursors in the periphery. Recently, accumulating evidence suggests that these Tregs play important roles in the regulation of humoral immune responses. In this review, we discuss recent findings on the role of Tregs in SLE.

A 75-year-old woman was admitted to our department because of backache and multiple joint pain. Serum C-reactive protein (CRP) level was 6.8 mg/dL, and serum rheumatoid factor and anti-citrullinated peptide antibody were negative. Magnetic resonance imaging (MRI) showed bone edema and synovitis of the sacroiliac joints. (18)Fluorodeoxyglucose-positron emission tomography ((18)FDG-PET) showed increased uptakes in the bilateral elbow, shoulder, sternoclavicular, hip, and sacroiliac joints. In addition, increased uptakes were also observed in the bilateral cervical and subclavian arteries, and the thoracic aorta. Moreover, inflammation of the vascular walls and an aneurism in the right subclavian artery were observed on MRI. The patient's HLA type was HLA-B48 and B60. According to the Assessment of Spondyloarthritis (ASAS) classification criteria, peripheral spondyloarthropathy (SpA) was also diagnosed. Although the diagnostic criteria of Takayasu aortitis or giant cell aortitis were not fulfilled, we thought that active aortitis was also involved, and high-dose prednisolone was started. The patient's symptoms were diminished immediately, and CRP levels returned to normal. Although the etiology of the aortitis was not certain, this is the first report of late-onset peripheral SpA with aortitis, diagnosed by (18)FDG-PET and MRI. We recommended that it is important to evaluate the aortic involvement in late-onset SpA patients, when elevated systemic inflammation is observed.

TGF-β1 is an important anti-inflammatory cytokine, and several regulatory T cell (Treg) subsets including CD4(+)CD25(+)Foxp3(+) Tregs and Th3 cells have been reported to exert regulatory activity via the production of TGF-β1. However, it has not yet been elucidated which transcription factor is involved in TGF-β1 transcription. Early growth response 3 (Egr-3) is a zinc-finger transcription factor that creates and maintains T cell anergy. In this study, we found that Egr-3 induces the expression of TGF-β1 in both murine and human CD4(+) T cells. Egr-3 overexpression in murine CD4(+) T cells induced the production of TGF-β1 and enhanced the phosphorylation of STAT3, which is associated with TGF-β1 transcription. Moreover, Egr-3 conferred Ag-specific regulatory activity on murine CD4(+) T cells. In collagen-induced arthritis and delayed-type hypersensitivity model mice, Egr-3-transduced CD4(+) T cells exhibited significant regulatory activity in vivo. In particular, the suppression of delayed-type hypersensitivity depended on TGF-β1. In human tonsils, we found that CD4(+)CD25(-)CD45RO(-)lymphocyte activation gene 3 (LAG3)(-) T cells express membrane-bound TGF-β1 in an EGR3-dependent manner. Gene-expression analysis revealed that CD4(+)CD25(-)CD45RO(-)LAG3(-) T cells are quite different from conventional CD4(+)CD25(+)Foxp3(+) Tregs. Intriguingly, the CD4(+)CD25(-)CD45RO(-)LAG3(-) T cells suppressed graft-versus-host disease in immunodeficient mice transplanted with human PBMCs. Our results suggest that Egr-3 is a transcription factor associated with TGF-β1 expression and in vivo regulatory activity in both mice and humans.

A 69-year-old man with sensorineural hearing loss and iritis was diagnosed with atypical Cogan's syndrome. He had several systematic manifestations: aortitis, meningitis, panniculitis and seronegative arthritis. Remission induced by treatment with high doses of prednisolone was followed by relapse within 1 year. Although his condition was resistant to various immunosuppressive drugs, including methotrexate, cyclosporine, azathioprine and adalimumab, his symptoms, inflammatory response and quality of life measures were successfully improved by tocilizumab, a humanized anti-interleukin-6 receptor antibody.

制御性T細胞（Treg）は自己免疫応答に対する重要な抑制機構であり，CD4陽性CD25陽性Foxp3陽性Treg，Tr-1，CD8陽性制御性T細胞等が知られている．我々は近年IL-10を大量に産生し転写因子Egr2を発現するCD4陽性CD25陰性LAG3陽性Treg（LAG3 Treg）を新たなTregサブセットとしてマウスとヒトで同定した．興味深いことにマウスLAG3TregはSLEモデルマウスMRL/lprマウスの自己抗体産生と腎炎進行を抑制し，試験管内および生体内でのヘルパーT細胞とB細胞による抗体産生をPD-1-PD-L1およびFas-FasL依存性に抑制した．試験管内での抗CD40抗体によるT細胞非存在下でのB細胞の分裂および抗体産生はLAG3Tregにより抑制され，またFasL欠損B細胞を生体内に移入するとLAG3Tregは抗体産生を抑制できなかったことから，LAG3TregはB細胞を直接抑制すると考えられた．ヒトでも扁桃腺と末梢血にLAG3Tregが存在し，試験管内で細胞接触依存性に強力な抗体産生抑制能を発揮した．関節リウマチ（RA）や全身性エリテマトーデス（SLE）患者末梢血ではLGA3Tregが減少していた．現在LAG3TregによるB細胞抑制機構と各種免疫抑制治療によるLAG3Tregへの修飾を解析中である．<br>