Hejian Zou

Cutting Edge: All-Trans Retinoic Acid Sustains the Stability and Function of Natural Regulatory T Cells in an Inflammatory Milieu

Recent studies have demonstrated that plasticity of naturally occurring CD4+Foxp3+ regulatory T cells (nTregs) may account for their inability to control chronic inflammation in established autoimmune diseases. All-trans retinoic acid (atRA), the active derivative of vitamin A, has been demonstrated to promote Foxp3+ Treg differentiation and suppress Th17 development. In this study, we report a vital role of atRA in sustaining the stability and functionality of nTregs in the presence of IL-6. We found that nTregs treated with atRA were resistant to Th17 and other Th cell conversion and maintained Foxp3 expression and suppressive activity in the presence of IL-6 in vitro. atRA decreased IL-6R expression and signaling by nTregs. Of interest, adoptive transfer of nTregs even from arthritic mice treated with atRA suppressed progression of established collagen-induced arthritis. We suggest that nTregs treated with atRA may represent a novel treatment strategy to control established chronic immune-mediated inflammatory diseases.

Activation of Sirt1 by Resveratrol Inhibits TNF-α Induced Inflammation in Fibroblasts

Inflammation is one of main mechanisms of autoimmune disorders and a common feature of most diseases. Appropriate suppression of inflammation is a key resolution to treat the diseases. Sirtuin1 (Sirt1) has been shown to play a role in regulation of inflammation. Resveratrol, a potent Sirt1 activator, has anti-inflammation property. However, the detailed mechanism is not fully understood. In this study, we investigated the anti-inflammation role of Sirt1 in NIH/3T3 fibroblast cell line. Upregulation of matrix metalloproteinases 9 (MMP-9), interleukin-1beta (IL-1β), IL-6 and inducible nitric oxide synthase (iNOS) were induced by tumor necrosis factor alpha (TNF-α) in 3T3 cells and resveratrol suppressed overexpression of these pro-inflammatory molecules in a dose-dependent manner. Knockdown of Sirt1 by RNA interference caused 3T3 cells susceptible to TNF-α stimulation and diminished anti-inflammatory effect of resveratrol. We also explored potential anti-inflammatory mechanisms of resveratrol. Resveratrol reduced NF-κB subunit RelA/p65 acetylation, which is notably Sirt1 dependent. Resveratrol also attenuated phosphorylation of mammalian target of rapamycin (mTOR) and S6 ribosomal protein (S6RP) while ameliorating inflammation. Our data demonstrate that resveratrol inhibits TNF-α-induced inflammation via Sirt1. It suggests that Sirt1 is an efficient target for regulation of inflammation. This study provides insight on treatment of inflammation-related diseases.

Antigen-specific transforming growth factor β-induced Treg cells, but not natural Treg cells, ameliorate autoimmune arthritis in mice by shifting the Th17/Treg cell balance from Th17 predominance to Treg cell predominance.

OBJECTIVE Transferred CD4+CD25+FoxP3+ Treg cells can prevent autoimmune disease, but generally fail to ameliorate established disease. This study was undertaken to compare the effects of antigen-specific Treg cells induced with interleukin-2 (IL-2) and transforming growth factor β (TGFβ) ex vivo (induced Treg [iTreg] cells) to the effects of equivalent expanded thymus-derived natural Treg (nTreg) cells on established collagen-induced arthritis (CIA). METHODS CIA was induced in DBA/1 mice by immunization with type II collagen (CII), and before or shortly after immunization, mice were treated with iTreg or nTreg cells that were generated or expanded in vitro. Clinical scores were determined. Inflammatory responses were determined by measuring the levels of anti-CII antibody in the serum and examining the histologic features of the mouse joints. The Th1/Th17-mediated autoreactive response was evaluated by determining the cytokine profile of the draining lymph node (LN) cells of the mice by flow cytometry. RESULTS Following transfer, nTreg cells exhibited decreased FoxP3 and Bcl-2 expression and decreased suppressive activity, and many converted to Th17 cells. In contrast, transferred iTreg cells were more numerous, retained FoxP3 expression and their suppressive activity in the presence of IL-6, and were resistant to Th17 conversion. Notably, 10 days after the transfer of donor iTreg cells, predominance was shifted from Th17 cells to Treg cells in the draining LNs of recipient mice. CONCLUSION These findings provide evidence that transferred TGFβ-induced iTreg cells are more stable and functional than nTreg cells in mice with established autoimmunity. Moreover, iTreg cells can have tolerogenic effects even in the presence of ongoing inflammation. The therapeutic potential of human iTreg cells in subjects with chronic, immune-mediated inflammatory diseases should be investigated.

Cutting Edge: Leptin-Induced RORγt Expression in CD4+ T Cells Promotes Th17 Responses in Systemic Lupus Erythematosus

Th17 CD4+ cells promote inflammation and autoimmunity. In this study, we report that Th17 cell frequency is reduced in ob/ob mice (that are genetically deficient in the adipokine leptin) and that the administration of leptin to ob/ob mice restored Th17 cell numbers to values comparable to those found in wild-type animals. Leptin promoted Th17 responses in normal human CD4+ T cells and in mice, both in vitro and in vivo, by inducing RORγt transcription. Leptin also increased Th17 responses in (NZB × NZW)F1 lupus-prone mice, whereas its neutralization in those autoimmune-prone mice inhibited Th17 responses. Because Th17 cells play an important role in the development and maintenance of inflammation and autoimmunity, these findings envision the possibility to modulate abnormal Th17 responses via leptin manipulation, and they reiterate the link between metabolism/nutrition and susceptibility to autoimmunity.

Therapeutic potential of TGF-β-induced CD4(+) Foxp3(+) regulatory T cells in autoimmune diseases.

Foxp3+ T regulatory cell (Treg) subsets play a crucial role in the maintenance of immune homeostasis against self-antigens. The lack or dysfunction of these cells contributes to the pathogenesis and development of many autoimmune diseases. Therefore, manipulation of these cells may provide a novel therapeutic approach to treat autoimmune diseases. In this review, we provide current opinions concerning the classification, developmental, and functional characterization of Treg subsets. Particular emphasis will be focused on the therapeutic role of TGF-β-induced CD4M+ Foxp3+ cells (iTregs) in established autoimmune disease. Moreover, the similarity and diversity of iTregs and naturally occurring, thymus-derived CD4+ CD25+ Foxp3+ regulatory T cells (nTregs) will be discussed, including the finding that the pro-inflammatory cytokine IL-6 can convert nTregs to IL-17-producing cells, whereas iTregs induced by TGF-β are resistant to the effects of this cytokine. Understanding these aspects may help to determine how Tregs can be used in the treatment of autoimmune diseases.

Adipokines in Psoriatic Arthritis Patients: The Correlations with Osteoclast Precursors and Bone Erosions

Significant bone remodeling with disordered osteoclastogenesis has been implicated in the pathogenesis of psoriatic arthritis (PsA). And there is a high prevalence of the metabolic syndrome (MS) in PsA patients. Adipokines, especially leptin and adiponectin, have recently been reported to be involved in the development and regulation of some autoimmune diseases. In this study, we examined the alternation of circulating osteoclastogenesis related cytokines [tumor necrosis factor-α (TNF-α), osteoprotegerin (OPG) and receptor activator of nuclear factor-κB ligand (RANKL)] and adipokines (leptin, adiponectin, resistin, chemerin, omentin) in PsA patients, and analysed the correlations between these factors and osteoclast precursors numbers, radiographic damage scores, and disease activity index. 41 PsA patients, 20 psoriasis patients, and 24 healthy controls were recruited. Blood samples were obtained for detecting the levels of TNF-α, OPG, RANKL and the adipokines. The numbers of osteoclast precursors (OCs) in peripheral blood were assessed. Radiographs of affected joints in PsA patients were scored for erosion, joint-space narrowing, osteolysis, and new bone formation. Compared with healthy controls, patients with PsA had higher TNF-α, RANKL, OCs, leptin and omentin but lower adiponectin and chemerin. Increased serum levels of TNF-α, RANKL, leptin, and omentin were positively correlated with OCs numbers. In contrast, serum adiponectin levels were decreased in PsA patients and negatively correlated with OCs numbers. TNF-α, RANKL and leptin were positively correlated with Psoriatic Arthritis Joint Activity Index (PsAJAI). Only TNF-α was positively correlated with radiographic damage scores. Our data demonstrated that systemic expression of soluble mediators of osteoclastogenesis and adipokines were disordered in PsA. Certain adipokines were elevated in the circulation of patients with PsA and might contribute to pathogenesis of arthritis. Prospective studies will be of interest to determine the pluripotent effects of adipokines on osteoclastogenesis in chronic inflammatory rheumatic diseases. Future studies may lead to novel therapeutic strategies.

Induced T regulatory cells suppress osteoclastogenesis and bone erosion in collagen-induced arthritis better than natural T regulatory cells

Background Although natural regulatory T cells (nTregs) can suppress osteoclastogenesis, the role of TGF-β-induced CD4+Foxp3+ Tregs (iTregs) in osteoclastogenesis remains unknown. Objective To determine the effects of iTregs on osteoclastogenesis in vitro and on bone erosion in vivo in collagen-induced arthritis (CIA). Methods Osteoclastogenesis was induced in bone marrow CD11b+ cells with receptor activator of nuclear factor κ B (NF-κB) ligand (RANKL) and macrophage colony stimulating factor. Graded doses of Tregs were added to inhibit osteoclastogenesis. Transwell and antibody blockade experiments were performed to assess the roles for cell contact and soluble cytokines. NF-κB activation was determined by western blot. iTregs or nTregs were adoptively transferred to mice with CIA to assess in vivo effects on disease incidence and bone erosion, the latter determined by CT scanning. Results Both nTregs and iTregs greatly suppressed osteoclastogenesis in vitro, but only iTregs sustained this effect when interleukin-6 was present. iTregs, but not nTregs, significantly suppressed development of CIA. Bone erosions in iTregs-treated mice were diminished compared with untreated mice or nTregs-treated mice. Treatment with iTregs, but not with nTregs, dramatically decreased NF-κB p65/p50 levels in osteoclasts in vitro and p65/50 and RANKL expression by synovial tissues in vivo. Conclusion iTregs may be therapeutically beneficial in rheumatoid arthritis and related diseases associated with bone erosions.

T Follicular Helper Cells Mediate Expansion of Regulatory B Cells via IL-21 in Lupus-Prone MRL/lpr Mice

T follicular helper (Tfh) cells can mediate humoral immune responses and augment autoimmunity, whereas the role of Tfh cells on regulatory B (B10) cells in autoimmunity diseases is not clear. Here, we investigated the percentages of Tfh cells and B10 cells in lupus-prone MRL/Mp-lpr/lpr (MRL/lpr) mice and examined the effects and mechanism of Tfh cell-derived interleukin-21 (IL-21) on IL-10 production during the differentiation of B10 cells. Both Tfh cells and B10 cells were expanded in spleens of MRL/lpr mice. In addition, a positive correlation between the proportions of Tfh cells and B10 cells was observed. Tfh cell-derived IL-21 from MRL/lpr mice could promote IL-10 production during the differentiation of B10 cells. Importantly, neutralization of IL-21 inhibited IL-10 production and expansion of B10 cells both in vitro and in vivo. IL-21 induced IL-10 production via activation of phosphorylated signal transduction and activator of transcription 3 (p-STAT3). Inhibition of p-STAT3 effectively blocked IL-10 production during the differentiation of B10 cells. Moreover, IL-21-induced IL-10 exerted a regulatory function by inhibiting the proliferation of T cells. These data suggest that Tfh cells not only mediate humoral immune responses and augment autoimmunity but also play a broader role in immune regulatory actions via the induction of IL-10 production.

T follicular helper cells and regulatory B cells dynamics in systemic lupus erythematosus.

T follicular helper (Tfh) cells aid effector B cells, and augment autoimmunity, whereas the role of Tfh cells on regulatory B (Breg) cells in systemic lupus erythematosus (SLE) is not known. The aim of this study is to investigate the percentage of Breg cells in SLE, and the role of Tfh cells on Breg cells. First, we demonstrated the presence of Breg cells in SLE peripheral blood mononuclear cells and in involved skins. Both the percentage of circulating Breg cells and the ability to produce interleukin-10 (IL-10) were elevated in SLE patients. The percentage of Breg cells increased during SLE flares and decreased following disease remission. Second, Tfh cell expansion was not only related to autoantibody production but also correlated with the increased percentage of Breg cells. Third, in vitro studies revealed that Tfh cell-derived IL-21 could promote IL-10 production and Breg cell differentiation. In conclusions, these data imply that SLE flares may be linked to the expansion of Tfh cells and that Breg cells are increased in a regulatory feedback manner. Thus, SLE development may be associated with the complex regulation of Tfh cells and diverse B cell subsets.

S100A9 promotes human lung fibroblast cells activation through receptor for advanced glycation end‐product‐mediated extracellular‐regulated kinase 1/2, mitogen‐activated protein‐kinase and nuclear factor‐κB‐dependent pathways

S100A9 belongs to the S100 family of calcium‐binding proteins and plays a key role in many inflammatory conditions. Recent studies have found that S100A9 was elevated significantly in the bronchoalveolar lavage fluid of idiopathic pulmonary fibrosis patients, and might be a biomarker for fibrotic interstitial lung diseases. However, the exact function of S100A9 in pulmonary fibrosis needs further studies. We performed this study to investigate the effect of S100A9 on human embryo lung fibroblast (HLF) proliferation and production of cytokines and collagen, providing new insights into the possible mechanism. S100A9 promoted proliferation of fibroblasts and up‐regulated expression of both proinflammatory cytokines interleukin (IL)‐6, IL‐8, IL‐1β and collagen type III. S100A9 also induced HLF cells to produce α‐smooth muscle actin (α‐SMA) and receptor for advanced glycation end‐product (RAGE). In addition, S100A9 caused a significant increase in extracellular‐regulated kinase (ERK)1/2 mitogen‐activated protein kinase (MAPK) phosphorylation, while the status of p38 and c‐Jun N‐terminal kinase (JNK) phosphorylation remained unchanged. Treatment of cells with S100A9 also enhanced nuclear factor kappa B (NF‐κB) activation. RAGE blocking antibody pretreatment inhibited the S100A9‐induced cell proliferation, cytokine production and pathway phosphorylation. S100A9‐mediated cell activation was suppressed significantly by ERK1/2 MAPK inhibitor and NF‐κB inhibitor. In conclusion, S100A9 promoted HLF cell growth and induced cells to secret proinflammatory cytokines and collagen through RAGE signalling and activation of ERK1/2 MAPK and NF‐κB pathways.