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Background: Graft-versus-host disease (GvHD) is a critical complication after allogeneic hematopoietic stem cell transplantation (HSCT). The immunosuppressants given to patients undergoing allogeneic HSCT disturb the microbiome and the host immune system, potentially leading to dysbiosis and inflammation, and may affect immune function and bone marrow transplantation. The intestinal microbiome is a target for the development of novel therapies for GvHD. Lactobacillus species are widely used supplements to induce production of antimicrobial and anti-inflammatory factors.Methods: We determined the effect of the combination of Lactobacillus acidophilus and FK506 on GvHD following major histocompatibility complex-mismatched bone marrow transplantation.Results: The combination treatment suppressed IFN-γ and IL-17-producing T cell differentiation, but increased Foxp3+Treg differentiation and IL-10 production. Also, the combination treatment and combination treated-induced Treg cells modulated the proliferation of murine alloreactive T cells in vitro. Additionally, the combination treatment upregulated Treg-related genes-Nt5e, Foxp3, Ikzf2, Nrp1 and Itgb8-in murine CD4+-T cells. The combination treatment also alleviated GvHD clinically and histopathologically by controlling the effector T cell and Treg balance in vivo. Moreover, the combination treatment decreased Th17 differentiation significantly and significantly upregulated Foxp3 and IL-10 expression in peripheral blood mononuclear cells from healthy controls and liver transplantation (LT) patients.Conclusions: Therefore, the combination of L. acidophilus and FK506 is effective and safe for patients undergoing allogeneic hematopoietic stem cell transplantation. [ABSTRACT FROM AUTHOR]

Background: Rheumatoid arthritis (RA) is a progressive systemic autoimmune disease that is characterized by infiltration of inflammatory cells into the hyperplastic synovial tissue, resulting in subsequent destruction of adjacent articular cartilage and bone. Methotrexate (MTX), the first conventional disease-modifying antirheumatic drug (DMARD), could alleviate articular damage in RA and is implicated in humoral and cellular immune responses. However, MTX has several side effects, so efficient delivery of low-dose MTX is important.Methods: To investigate the efficacy of MTX-loaded nanoparticles (MTX-NPs) against experimental model of RA, free MTX or MTX-NPs were administered as subcutaneous route to mice with collagen-induced arthritis (CIA) at 3 weeks after CII immunization. The levels of inflammatory factors in tissues were determined by immunohistochemistry, confocal microscopy, real-time PCR, and flow cytometry.Results: MTX-NPs ameliorated arthritic severity and joint destruction in collagen-induced arthritis (CIA) mice compared to free MTX-treated CIA mice. The levels of inflammatory cytokines, including interleukin (IL)-1β, tumor necrosis factor-α, and vascular endothelial growth factor, were reduced in MTX-NPs-treated mice. Number of CD4 + IL-17 + cells decreased whereas the number of CD4 + CD25 + Foxp3 + cells increased in spleens from MTX- NPs-treated CIA mice compared to MTX-treated CIA mice. The frequency of CD19 + CD25 + Foxp3 + regulatory B cells increased in ex vivo splenocytes from MTX-loaded NPs-treated CIA mice compared to MTX-treated CIA mice.Conclusion: The results suggest that MTX-loaded NPs have therapeutic potential for RA. [ABSTRACT FROM AUTHOR]

Previous studies have evaluated the roles of T and B cells in the pathogenesis of Sjögren's syndrome (SS); however, their relationships with age-dependent and metabolic abnormalities remain unclear. We examined the impacts of changes associated with aging or metabolic abnormalities on populations of T and B cells and SS disease severity. We detected increased populations of IL-17-producing T and B cells, which regulate inflammation, in the salivary glands of NOD/ShiLtJ mice. Inflammation-induced human submandibular gland cell death, determined based on p-MLKL and RIPK3 expression levels, was significantly increased by IL-17 treatment. Among IL-17-expressing cells in the salivary gland, peripheral blood, and spleen, the α4β7 (gut-homing integrin)-negative population was significantly increased in aged NOD/ShiLtJ mice. The α4β7-positive population markedly increased in the intestines of aged NOD/ShiLtJ mice following retinoic acid (RA) treatment. A significant increase in α4β7-negative IL-17-expressing cells in salivary glands may be involved in the onset and progression of SS. These results suggest the potential therapeutic utility of RA in SS treatment. [ABSTRACT FROM AUTHOR]

We examined the effect of combination therapy with metformin and tacrolimus on immune parameters including T regulatory (Treg) and type 17 helper T (Th17) cells in vitro and in vivo in mice and in liver transplantation (LT) patients. T cell proliferation and subtypes after in vitro T cell activation or allogeneic stimulation were evaluated. RNA sequencing and microarray analysis were used to evaluate differences in gene expression. Metformin and tacrolimus were administered to mice with graft-versus-host disease (GVHD) and the effects in vivo were assessed. Five LT patients were treated with metformin and the changes in Treg and Th17 cells examined. Combination therapy decreased Type 1 helper T (Th1) and Th17 cells present after in vitro T cell activation, whereas genes associated with Treg were overexpressed. During in vitro allogeneic stimulation, combination therapy increased Treg cells and decreased T cell proliferation and pro-inflammatory markers. In mice with GVHD, combination treatment decreased the clinical and pathological severity of GVHD. In LT patients, addition of metformin increased the peripheral percentage of CD4+Treg and CD8+Treg cells and decreased CD4+Th17. Our study suggests that the addition of metformin to tacrolimus may improve immunological balance by increasing Treg cells and decreasing Th17 cells. [ABSTRACT FROM AUTHOR]

Background: Myeloid-derived suppressor cells (MDSCs) play a critical role in modulating the immune response and promoting immune tolerance in models of autoimmunity and transplantation. Regulatory T cells (Tregs) exert therapeutic potential due to their immunomodulatory properties, which have been demonstrated both in vitro and in clinical trials. Cell-based therapy for acute graft-versus-host disease (aGVHD) may enable induction of donor-specific tolerance in the preclinical setting.Methods: We investigated whether the immunoregulatory activity of the combination of MDSCs and Tregs on T cell and B cell subset and alloreactive T cell response. We evaluated the therapeutic effects of combined cell therapy for a murine aGVHD model following MHC-mismatched bone marrow transplantation. We compared histologic analysis from the target tissues of each groups were and immune cell population by flow cytometric analysis.Results: We report a novel approach to inducing immune tolerance using a combination of donor-derived MDSCs and Tregs. The combined cell-therapy modulated in vitro the proliferation of alloreactive T cells and the Treg/Th17 balance in mice and human system. Systemic infusion of MDSCs and Tregs ameliorated serverity and inflammation of aGVHD mouse model by reducing the populations of proinflammatory Th1/Th17 cells and the expression of proinflammatory cytokines in target tissue. The combined therapy promoted the differentiation of allogeneic T cells toward Foxp3 + Tregs and IL-10-producing regulatory B cells. The combination treatment control also activated human T and B cell subset.Conclusions: Therefore, the combination of MDSCs and Tregs has immunomodulatory activity and induces immune tolerance to prevent of aGVHD severity. This could lead to the development of new clinical approaches to the prevent aGVHD. [ABSTRACT FROM AUTHOR]

Background: Fibrosis is the formation of excess connective tissue in an organ or tissue during a reparative or reactive process. Graft-versus-host disease (GvHD) is a medical complication of allogeneic tissue transplantation with transplanted donor T cell-mediated inflammatory response; it is characterized by a severe immune response with fibrosis in the final stage of the inflammatory process. T helper 17 cells play a critical role in the pathogenesis of GvHD. Fingolimod (FTY720), an analogue of sphingosine-1-phosphate (S1P), is an effective immunosuppressive agent in experimental transplantation models.Methods: In this study, we evaluated the effects of FTY720 as a treatment for an animal GvHD model with inflammation and fibrosis. The splenocytes, lymph nodes, blood, tissues from Syngeneic mice and GvHD-induced mice treated vehicle or FTY720 were compared using flow cytometry, hematological analyses, histologic analyses.Results: FTY720 reduced clinical scores based on the following five clinical parameters: weight loss, posture, activity, fur texture, and skin integrity. FACS data showed that T lymphocyte numbers increased in mesenteric lymph nodes and decreased in splenocytes of FTY720-treated mice. Tissue analysis showed that FTY720 reduced skin, intestinal inflammation, and fibrotic markers. FTY720 dramatically decreased α-smooth muscle actin, connective tissue growth factor, and fibronectin protein levels in keloid skin fibroblasts.Conclusions: Thus, FTY720 suppressed migration of pathogenic T cells to target organs, reducing inflammation. FTY720 also inhibited fibrogenesis marker expression in vitro and in vivo. Together, these results suggest that FTY720 prevents GvHD progression via immunosuppression of TH17 and simultaneously acts an anti-fibrotic agent. [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis (RA) is a chronic inflammatory polyarthritis characterized by progressive joint destruction. IL-17-producing CD4+ T (Th17) cells play pivotal roles in RA development and progression. Retinoic acid receptor-related orphan receptor alpha (RORα) is a negative regulator of inflammatory responses, whereas RORγt, another member of the ROR family, is a Th17 lineage-specific transcription factor. Here, we investigated the immunoregulatory potential of RORα in collagen-induced arthritis (CIA) mice, an experimental model of RA. Cholesterol sulfate (CS) or SR1078, a ligand of RORα, inhibited RORγt expression and Th17 differentiation in vitro. In addition, fortification of RORα in T cells inhibited the expression levels of glycolysis-associated genes. We found that RORα overexpression in CIA mice attenuated the clinical and histological severities of inflammatory arthritis. The anti-arthritic effect of RORα was associated with suppressed Th17 differentiation and attenuated mTOR-STAT3 signaling in T cells. Furthermore, altered RORα activity could directly affect osteoclastogenesis implicated in progressive bone destruction in human RA. Our findings defined a critical role of RORα in the pathogenesis of RA. These data suggest that RORα may have novel therapeutic uses in the treatment of RA. [ABSTRACT FROM AUTHOR]

Optimizing Treg function and improving Treg stability are attractive treatment strategies for treating autoimmune rheumatoid arthritis (RA). However, the limited number of circulating Tregs and questions about the functional stability of in vitro -expanded Tregs are potential limitations of Treg-based cell therapy. The aim of this study was to analyze the regulatory effect of daurinol, a catalytic inhibitor of topoisomerase IIα, on Th cell differentiation and to evaluate their therapeutic potential in a preclinical experimental model of RA. We investigated the effect of daurinol on T cell differentiation by flow cytometry. Foxp3 stability and methylation were analyzed by suppression assays and bisulfite pyrosequencing. Daurinol was treated in the collagen-induced arthritis (CIA) model, and the effects in vivo were determined. We found that daurinol can promote Treg differentiation and reciprocally inhibit Th17 differentiation. This Treg-inducing property of daurinol was associated with decreased activity of Akt–mTOR and reciprocally increased activity of neuropilin-1 (Nrp1)–PTEN. Daurinol treatment inhibited aerobic glycolysis in Th17 conditions, indicating the metabolic changes by daurinol. We found that the daurinol increase the Treg stability was achieved by Foxp3 hypomethylation. In vivo daurinol treatment in CIA mice reduced the clinical arthritis severity and histological inflammation. The Treg population frequency increased and the Th17 cells decreased in the spleens of arthritis mice treated with daurinol. These results showed the anti-arthritic and immunoregulating properties of daurinol is achieved by increased differentiation and stabilization of Tregs. Our study provides first evidence for daurinol as a treatment for RA. [ABSTRACT FROM AUTHOR]

This study aimed to evaluate the therapeutic effect of fraxinellone on inflammatory arthritis and identify the underlying mechanisms. Fraxinellone (7.5 mg/kg) or a vehicle control was injected into mice with collagen-induced arthritis (CIA). The severity of arthritis was evaluated clinically and histologically. The differentiation of CD4+ T cells and CD19+ B cells was investigated in the presence of fraxinellone. Osteoclastogenesis after fraxinellone treatment was evaluated by staining with tartrate-resistant acid phosphatase (TRAP) and by measuring the mRNA levels of osteoclastogenesis-related genes. Fraxinellone attenuated the clinical and histologic features of inflammatory arthritis in CIA mice. Fraxinellone suppressed the production of interleukin-17 and the expression of RAR-related orphan receptor g t and phospho-signal transducer and activator of transcription 3 in CD4+ T cells. CD19+ B cells showed lower expression of activation-induced cytidine deaminase and B lymphocyte-induced maturation protein-1 after treatment with fraxinellone. The formation of TRAP-positive cells and the expression of osteoclastogenesis-related markers were reduced in the presence of fraxinellone. Inhibition of interleukin-17 and osteoclastogenesis was also observed in experiments using human peripheral mononuclear cells. Fraxinellone alleviated synovial inflammation and osteoclastogenesis in mice. The therapeutic effect of fraxinellone was associated with the inhibition of cellular differentiation and activation. The data suggests that fraxinellone could be a novel treatment for inflammatory arthritis, including rheumatoid arthritis. [ABSTRACT FROM AUTHOR]

Background: The aim of this study was to investigate the protective effect of 1α,25-dihydroxyvitamin D3 [1,25(OH)2D3] on effector CD4+ T cells or on inflammatory cytokine-induced injury in human renal proximal tubular epithelial cells (HRPTEpiC). Methods: First, we investigated the effect of 1,25(OH)2D3 on CD4+ T cell proliferation. Second, we examined the effect of 1,25(OH)2D3 on inflammatory cytokine secretion or fibrosis in HRPTEpiC induced by inflammatory cytokines or activated CD4+ T cells using ELISA and real-time PCR. Lastly, we compared urine inflammatory-cytokine (IL-6, IL-8) or KIM-1 levels in kidney transplant recipients low serum 25-hydroxyvitamin D (25(OH)D) group (< 20 ng/mL) (n = 40) and normal 25(OH)D group (n = 50). Results: Pre-incubation with 1,25(OH)2D3 significantly reduced the percentages of Th1 and Th17 cells compared to that of Th0 condition (P < 0.05 for each). In contrast, 1,25(OH)2D3 increased the proportion of Th2 and Treg cells in a dose-dependent manner (P < 0.05 for each). Treatment of HRPTEpiC with inflammatory cytokines (TNF-α, IL-17, and TGF-β) or effector CD4+ T cells resulted in increased production of IL-6, IL-8, or KIM-1 from HRPTEpiC in a dose-dependent manner. However, treatment with 1,25(OH)2D3 significantly reduced the level of these cytokines (P < 0.05 for all). Western blot analysis demonstrated that the mTOR/STAT3/ERK pathway was downregulated by 1,25(OH)2D3 in HRPTEpiC. Furthermore, the concentrations of urine IL-6/creatinine (P < 0.05) and Kim-1/creatinine (P < 0.05) were higher in the low 25(OH)D group than in the normal 25(OH)D group in kidney transplant recipients. Conclusion: The results of this study suggests that vitamin D may have a significant role in the regulation of inflammation in allograft tissue in kidney transplant recipients. Trial registration: All participants provided written informed consent in accordance with the Declaration of Helsinki. This study was approved by the Institutional Review Board of Seoul St. Mary’s Hospital (). [ABSTRACT FROM AUTHOR]

Background: Leflunomide is a low-molecular-weight compound that is widely used in the treatment of rheumatoid arthritis. Although leflunomide is thought to act through the inhibition of the de novo pyrimidine synthesis, the molecular mechanism of the drug remains largely unknown. We investigated the antiarthritis effects and mechanisms of action of the active metabolite of leflunomide, A77 1726, in interleukin-1 receptor antagonist-knockout (IL-1Ra-KO) mice.Methods: 14- to 15-week-old male IL-1Ra-KO mice were treated with 10 or 30 mg/kg A77 1726 via intraperitoneal injection three times per week for 6 weeks. The effects of A77 1726 on arthritis severities were assessed by clinical scoring and histological analysis. The serum concentrations of IL-1β, tumor necrosis factor-α (TNF-α), and malondialdehyde were measured by enzyme-linked immunosorbent assay. Histologic analysis of the joints was performed using Safranin O, and immunohistochemical staining. The frequencies of interleukin-17-producing CD4+ T (Th17) cells were analyzed by flow cytometry. Heme oxygenase-1 (HO-1) expression in splenic CD4+ T cells isolated from A77 1726-treated arthritis mice were assessed by western blotting.Results: A77 1726 treatment induced heme oxygenase-1 (HO-1) in Jurkat cells and primary mouse T cells. Interestingly, A77 1726 inhibited Th17 cell differentiation. In vivo, A77 1726 reduced the clinical arthritis severity of histological inflammation and cartilage destruction. The joints isolated from A77 1726-treated mice showed decreased expression of inducible nitric oxide synthase, nitrotyrosine, TNF-α, and IL-1β. The serum levels of TNF-α, IL-1β, and malondialdehyde were also decreased in A77 1726-treated mice. Whereas the number of Th17 cells in spleens was decreased in A77 1726-treated arthritis mice, a significant increase in the number of Treg cells in spleens was observed. Interestingly, HO-1 expression was significantly higher in splenic CD4+ T cells isolated from A77 1726-treated mice compared with those from vehicle-treated mice, whereas HO-1 expression of splenic non-CD4+ T cells did not differ between groups.Conclusion: The inhibitory effects of A77 1726 on joint inflammation and oxidative stress in autoimmune arthritis may be associated with HO-1 induction in CD4+ T cells. [ABSTRACT FROM AUTHOR]

Background: Spondyloarthritis (SpA) usually manifests as arthritis of the axial and peripheral joints but can also result in extra-articular manifestations such as inflammatory bowel disease. Proinflammatory cytokine interleukin-17 (IL-17) plays a crucial role in the pathogenesis of SpA. Rebamipide inhibits signal transducer and activator of transcription 3 that controls IL-17 production and Th17 cell differentiation. This study examined the effect of rebamipide on SpA development.Methods: SKG ZAP-70(W163C) mice were immunized with curdlan to induce SpA features. The mice were then intraperitoneally injected with rebamipide or vehicle 3 times a week for 14 weeks and their clinical scores were evaluated. Histological scores of the paw and spine and the length of the gut were measured at sacrifice. Immunohistochemical staining of IL-17 and tumor necrosis factor-α (TNF-α) was performed using tissue samples isolated from the axial joints, peripheral joints, and gut. Spleen tissue samples were isolated from both rebamipide- or vehicle-treated mice with SpA at 14 weeks after curdlan injection to determine the effect of rebamipide on Th17 and regulatory T (Treg) cell differentiation.Results: Rebamipide decreased the clinical and histological scores of the peripheral joints. The total length of the gut was preserved in rebamipide-treated mice. IL-17 and TNF-α expression in the spine, peripheral joints, and gut was lower in rebamipide-treated mice than in control mice. Th17 cell differentiation was suppressed whereas Treg cell differentiation was upregulated in the spleen of rebamipide-treated mice.Conclusion: Rebamipide exerted beneficial effects in mice with SpA by preventing peripheral arthritis and intestinal inflammation and by regulating Th17/Treg cell imbalance, suggesting that it can be used as a potential therapeutic agent for treating arthritis to SpA patients. [ABSTRACT FROM AUTHOR]

Background: Systemic lupus erythematosus (SLE) is an autoimmune-mediated chronic inflammatory disease. Half of patients with SLE suffer from lupus nephritis, which is major cause of death in SLE. TNF-like weak inducer of apoptosis (TWEAK)/fibroblast growth factor-inducible 14 (Fn14) interactions mediate inflammatory responses that are linked to the pathogenesis of lupus nephritis. Blocking of the TWEAK/Fn14 pathway by Fn14-Fc was performed in a SLE mouse model and the likely therapeutic mechanisms were investigated.Methods: To investigate the impact of TWEAK on B cell differentiation in SLE, the levels of AID, Blimp-1, and IRF4 messenger RNA were measured in CD19(+) B cells extracted from the spleens of sanroque mice and cultured with TWEAK. To identify the therapeutic effects of Fn14-Fc in SLE, sanroque mice were treated with Fn14-Fc or a control-Fc for 3 weeks. Immunoglobulin (Ig) G, IgG1, IgG2a, and anti-dsDNA antibody (Ab) levels were measured in the sera of each group. Spleens from each group were stained with antibodies against CD4, B220, GL-7, CD138, and PD-1. Kidneys were stained with hematoxylin and eosin (H&E) and periodic acid-Schiff (PAS).Results: Administration of TWEAK increased the mRNA levels of AID, Blimp-1, and IRF4. Treatment with Fn14-Fc suppressed levels of IgG, IgG1, IgG2a, and anti-dsDNA Ab in sera and reduced numbers of B, plasma, and follicular helper T (Tfh) cells in spleens of sanroque mice. In addition, renal protective effects of Fn14-Fc were shown.Conclusion: Fn14-Fc had beneficial effects in a SLE mouse model by repressing B cells, plasma cells, Tfh, and renal damage. This suggested that Fn14-Fc represents a potential therapeutic agent for SLE. [ABSTRACT FROM AUTHOR]

Sirolimus ( SRL) is a promising alternative to calcineurin inhibitors, such as tacrolimus ( TAC), in kidney transplant recipients ( KTRs), but the immunological benefits of conversion from calcineurin inhibitors to SRL are not fully investigated. In the present study, we evaluated the effect of conversion from TAC to SRL on the T helper type 17/regulatory T (Th17/Treg) axis in three separate studies. First, the effect of SRL on the Th17/Treg axis was evaluated in vitro using peripheral blood mononuclear cells ( PBMCs). Second, the effect of conversion from TAC to SRL on the Th17/Treg axis was studied in KTRs. Finally, the effect of SRL on CD8+ Treg cells was evaluated. In vitro analysis of PBMCs isolated from KTRs showed that SRL suppressed Th17 cell differentiation but TAC did not. Conversion from TAC to SRL markedly decreased the number of effector memory CD8+ T cells and significantly increased the proportion of CD4+ and CD8+ Treg cells compared with TAC in KTRs. SRL treatment induced the CD8+ Treg cells, and these cells inhibited the proliferation of allogeneic CD4+ T cells and Th17 cells. In conclusion, conversion from TAC to SRL favourably regulates Th17 and Treg cell differentiation in KTRs. These findings provide a rationale for conversion from TAC to SRL in KTRs. [ABSTRACT FROM AUTHOR]

Systemic lupus erythematosus (SLE) is an autoimmune disease in which abnormal immune responses are mediated by tissue-binding autoantibodies and immune complex deposition. Because most SLE patients are women of child-bearing age, oestrogen has been suggested to play an important role in SLE pathogenesis. One proposed role is to induce B-cell activation, culminating in increased autoantibody production. Interleukin- 21 (IL-21) has been shown to be crucial in the differentiation of activated B cells into plasma cells. We therefore hypothesized that oestrogen up-regulates IL-21 production and induces subsequent B-cell activation in SLE patients. Peripheral blood was obtained from 22 SLE patients and 16 healthy controls. Expression levels of IL-21 and its receptor in serum, peripheral blood mononuclear cells, and CD4+ T cells were higher in SLE patients than in healthy controls. Exposure of CD4+ T cells from SLE patients to 17β-oestradiol led to a dose- and time-dependent increase in IL-21 expression, which was abolished in the presence of mitogen-activated protein kinase (MAPK) (MAPK kinase, p38, Jun N-terminal kinase) inhibitors. B cells from healthy controls showed increased antibody production when they were co-cultured with oestrogen-treated CD4+ T cells from SLE patients. Treatment with IL-21 antibody abrogated the increased antibody production of the co-culture systems. This study revealed the association between oestrogen and IL-21 in SLE patients. Oestrogen up-regulates IL-21 expression of CD4+ T cells via MAPK-dependent pathways in SLE patients, which in turn induces increased antibody production by B cells. [ABSTRACT FROM AUTHOR]

Objective. Intravenous immunoglobulin (IVIG) is used as a therapeutic agent in various autoimmune diseases. The aims of this study were to investigate the therapeutic effects of IVIG on collagen-induced arthritis (CIA) and identify the mechanism responsible for any therapeutic effects. Methods. IVIG was administered to mice with CIA, and the in vivo effects were determined. Th17 and Treg cell frequencies were analyzed by flow cytometry, and cytokine levels in the supernatant were measured by enzyme-linked immunosorbent assay. Subpopulations of T cells and B cells in the spleen were assessed by confocal microscopy. Results. The arthritis severity score and incidence of arthritis were lower in mice treated with IVIG compared with untreated mice. Histopathologic analysis showed less joint damage in mice treated with IVIG. The expression of proinflammatory cytokines, specific type II collagen antibodies, and osteoclast markers was significantly reduced in mice treated with IVIG. Administration of IVIG induced increased FoxP3 expression and inhibited Th17 cell development. The number of FoxP3+ Treg cells was increased, and the number of Th17 cells was decreased in the spleens of mice treated with IVIG. The number of FoxP3+ follicular helper T cells was increased, and subsequent maturation of germinal center B cells was inhibited by IVIG. In addition, IVIG up-regulated interleukin-10 (IL-10) and Fcγ receptor IIB expression. The treatment effects of IVIG on arthritis were lost in IL-10-knockout mice. Conclusion. These results showed that IVIG has therapeutic effects by modulating CD4+ T cell differentiation. The therapeutic effects of IVIG are dependent on IL-10. [ABSTRACT FROM AUTHOR]

Objective The small molecule halofuginone has been shown to inhibit fibrosis, angiogenesis, and tumor progression. This study was undertaken to evaluate the effects of halofuginone in preventing autoimmune arthritis in mice. Methods The effects of halofuginone on joint diseases were assessed by clinical scoring and histologic analysis. Protein expression levels were confirmed by immunohistochemistry, enzyme-linked immunosorbent assay, flow cytometry, and/or Western blotting. The expression levels of messenger RNA (mRNA) for various molecules were determined by real-time polymerase chain reaction (PCR). Proliferation of osteoclast precursors was assessed by bromodeoxyuridine uptake. Osteoclast differentiation and activity were determined by quantifying tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells and area of resorbed bone. Results Treatment with halofuginone suppressed the development of autoimmune arthritis and reciprocally regulated Th17 cells and FoxP3+ Treg cells. These effects of halofuginone on Th17 differentiation involved increased signaling of ERK and reduction of STAT-3 and NF-ATc1 expression. Furthermore, halofuginone induced the expression of indoleamine 2,3-dioxygenase (IDO) in dendritic cells, leading to reduced production of Th17 cells. In addition, halofuginone prevented the formation and activity of osteoclasts through suppression of transcription factors, such as activator protein 1 and NF-ATc1, and inhibited cell cycle arrest by the committed osteoclast precursors via expression of Ccnd1 encoding cyclin D1. Conclusion Taken together, our results suggest that halofuginone is a promising therapeutic agent for the treatment of Th17 cell-mediated inflammatory diseases and bone diseases. [ABSTRACT FROM AUTHOR]

Objective Stromal cell-derived factor 1 (SDF-1) is a chemokine that is involved in the bone-destructive process in rheumatoid arthritis (RA) and bony metastasis in malignancy. This study was undertaken to determine the role and mechanism of SDF-1 in RA-associated osteoclastogenesis. Methods The expression of SDF-1, tumor necrosis factor α (TNFα), and RANKL in RA synovial tissue was analyzed using confocal microscopy. After synovial fibroblasts and CD4+ T cells were treated with SDF-1, RANKL messenger RNA expression was determined by real-time and reverse transcription polymerase chain reaction. Osteoclastogenesis was assessed by counting tartrate-resistant acid phosphatase-positive multinucleated cells in CD14+ monocytes cultured with SDF-1 in the presence of anticytokine antibodies or signal inhibitors and in monocytes cocultured with SDF-1-pretreated synovial fibroblasts and CD4+ T cells. Results RANKL, TNFα, and SDF-1 were coexpressed in the lining and sublining of RA synovium. SDF-1 stimulated RANKL expression in RA synovial fibroblasts and CD4+ T cells, and TNFα inhibition reduced this stimulation. When monocytes isolated from human peripheral blood were cultured with SDF-1, they were differentiated into osteoclasts in the absence of RANKL. Monocytes were also differentiated into osteoclasts when they were cocultured with SDF-1-pretreated synovial fibroblasts or CD4+T cells; however, this osteoclastogenesis was reduced by TNFα inhibition. Conclusion Our findings indicate that SDF-1 induces osteoclastogenesis directly and indirectly via up-regulating RANKL expression in RA synovial fibroblasts and CD4+ T cells, and that this is mediated by TNFα. The axis of SDF-1 and RANKL is a potential therapeutic target for RA-associated bone destruction. [ABSTRACT FROM AUTHOR]

Objective STAT-3 is a key transcriptional factor in the interleukin-6 (IL-6)-mediated differentiation of Th17 cells. Because Th17 is believed to be a central player in rheumatoid arthritis (RA), we sought to evaluate whether an endogenous inhibitor of the STAT3 gene, GRIM-19 (gene associated with retinoid-interferon-induced mortality 19), could attenuate the progression and severity of murine collagen-induced arthritis (CIA) through suppression of Th17 cells and, reciprocally, could increase expression of Treg cells. Methods Overexpression of GRIM-19 was produced either by intravenous/intramuscular administration of a GRIM-19 overexpression vector in DBA1/J mice or by development of GRIM-19-transgenic (Tg) mice on a C57BL/6 background. Clinical signs were scored for arthritis severity, and mouse splenocytes, serum, and joint tissue were obtained for immunostaining and histologic analyses. Results The numbers of CD4+IL-17+ cells and CD4+pSTAT3+ cells were decreased, while the numbers of CD4+CD25+Foxp3+ cells and CD4+pSTAT5+ cells were increased, in both GRIM-19 vector-transfected and GRIM-19-Tg mice. Administration of the GRIM-19 overexpression vector into mice with CIA markedly suppressed the clinical and histologic signs of arthritis in the affected joints. Similarly, when CIA was induced in GRIM-19-Tg mice, the arthritis phenotype was markedly attenuated and the expression of inflammatory cytokines (IL-1β, IL-6, tumor necrosis factor α, and IL-17) in the arthritic joints was also significantly reduced. Moreover, bone marrow-derived monocyte/macrophages obtained from GRIM-19-Tg mice showed attenuated RANKL-induced osteoclastogenesis in vitro. Conclusion GRIM-19 improved the clinical and histologic features of CIA and also inhibited osteoclast formation. These findings suggest that GRIM-19 may be a novel treatment agent for RA. [ABSTRACT FROM AUTHOR]

Mesenchymal stromal cells (MSCs) are seen as an ideal source of cells to induce graft acceptance; however, some reports have shown that MSCs can be immunogenic rather than immunosuppressive. We speculate that the immunomodulatory effects of regulatory T cells (Tregs) can aid the maintenance of immunoregulatory functions of MSCs, and that a combinatorial approach to cell therapy can have synergistic immunomodulatory effects on allograft rejection. After preconditioning with Fludarabine, followed by total body irradiation and anti-asialo-GM-1(ASGM-1), tail skin grafts from C57BL/6 (H-2kb) mice were grafted onto the lateral thoracic wall of BALB/c (H-2kd) mice. Group A mice (control group, n = 9) did not receive any further treatment after preconditioning, whereas groups B and C (n = 9) received cell therapy with MSCs or Tregs, respectively, on days −1, +6 and +13 relative to the skin transplantation. Group D (n = 10) received cell therapy with MSCs and Tregs on days −1, +6 and +13. Cell suspensions were obtained from the spleens of five randomly chosen mice from each group on day +7, and the immunomodulatory effects of the cell therapy were evaluated by flow cytometry and real-time PCR. Our results show that allograft survival was significantly longer in group D compared to the control group (group A). Flow cytometric analysis and real-time PCR for splenocytes revealed that the Th2 subpopulation in group D increased significantly compared to the group B. Also, the expression of Foxp3 and STAT 5 increased significantly in group D compared to the conventional cell therapy groups (B and C). Taken together, these data suggest that a combined cell therapy approach with MSCs and Tregs has a synergistic effect on immunoregulatory function in vivo, and might provide a novel strategy for improving survival in allograft transplantation. [ABSTRACT FROM AUTHOR]

Background: In this study we examined the in vivo and in vitro effects and mechanisms of action of curcumin on the development of acute graft-versus-host disease (GVHD) using a murine model. Methodology/Principal Findings: Mixed lymphocyte reactions were used to determine the in vitro effects of curcumin. Treatment with curcumin attenuated alloreactive T cell proliferation and inhibited the production of interferon (IFN)-γ and interleukin (IL)-17. In a murine acute GVHD model, transplantation of curcumin-treated allogeneic splenocytes into irradiated recipient mice significantly reduced the clinical severity scores of acute GVHD manifested in the liver, skin, colon and lung as compared with animals receiving vehicle-treated splenocytes. c-Fos and c-Jun expression levels in the skin and intestine, which are major target organs, were analyzed using immunohistochemical staining. Expression of both proteins was reduced in epithelial tissues of skin and intestine from curcumin-treated GVHD animals. The IFN-γ-expressing CD4+ splenocytes and IFN-γ-expressing lymph node cells were dramatically decreased in curcumin-treated mice. In contrast, CD4+Foxp3+ splenocytes were increased in the curcumin-treated acute GVHD animals. Flow cytometric analysis revealed that animals transplanted with curcumin-treated allogeneic splenocytes showed increased populations of CD4+ regulatory T cells (Tregs) as well as CD8+ Treg cells, compared to animals administered vehicle-treated splenocytes. Curcumin-treated acute GVHD animals could have a change in B cell subpopulations. Conclusion/Significance: In the present study, we investigated the efficacy and mechanism of action of curcumin treatment against acute GVHD. The acute GVHD mice administered with curcumin-treated splenocytes showed significantly reduced severity of acute GVHD. Curcumin exerted in vivo preventive effects on acute GVHD by reciprocal regulation of T helper 1 (Th1) and Treg (both CD4+ and CD8+ Treg) cell lineages as well as B cell homeostasis. [ABSTRACT FROM AUTHOR]

Objective To investigate the connection between p53 and interleukin-17-producing Th17 cell/Treg cell balance in rheumatoid arthritis (RA). Methods Th17 cell and Treg cell frequencies were analyzed by flow cytometry, and cytokine levels in the supernatant were determined using enzyme-linked immunosorbent assays. The expression of transcription factors was analyzed by immunostaining and Western blotting, and the interactions between p53 and STAT-3 or STAT-5 were determined by immunoprecipitation-Western blot analysis. A p53 agonist was administered in the collagen-induced arthritis (CIA) model, and the effects in vivo were determined. Results CD4+ T cells from p53-/- mice decreased the activity of STAT-5, lowered the level of phosphorylated STAT-5, and compromised Treg cell differentiation. The protein p53 bound STAT-5 directly, and this interaction was enhanced with increasing p53 activity. Under inflammatory conditions, p53 suppressed Th17 cell differentiation and skewed T cells toward Treg cell differentiation through the activation of STAT-5 signaling cascades. In mice with CIA, injection of a p53 overexpression vector or an antagonist of Mdm2 had the effect of controlling arthritis development in vivo. The regulatory effect of p53 was recapitulated in the cells of RA patients, with more pronounced suppression due to the repressed status of p53 in RA. Conclusion We demonstrated a link between p53-mediated and STAT-mediated regulation of Th17 cells/Treg cells in RA. Our results suggest that factors involved in this pathway might constitute novel therapeutic targets for the treatment of RA. [ABSTRACT FROM AUTHOR]

Objective To investigate the impact of STAT-3-mediated regulation on Th17 differentiation in patients with rheumatoid arthritis (RA). Methods CD4+ T cells isolated from peripheral blood (PB) and synovial fluid (SF) were stimulated to differentiate into Th17 cells or Treg cells. The activity of STAT-3 was knocked down by transfecting CD4+ T cells with small interfering RNA (siRNA). After 3 days in culture, the proportions of Th17 cells and Treg cells were measured by flow cytometry, and the production of interleukin-17 (IL-17) was measured by reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay. Results The levels of IL-17, IL-6, IL-23, IL-1, and tumor necrosis factor α were significantly higher in RA SF and synovial tissue than in SF and synovial tissue from osteoarthritis patients. In RA synovial tissue, the expression of STAT-3 increased in proportion to the severity of synovitis, as shown by stromal cellularity, intimal hyperplasia, and inflammatory infiltration. The degree of Th17 differentiation was highest in RA SF, followed by RA PB, and lowest in normal subjects. In CD4+ T cells, transfection with STAT-3 siRNA prevented Th17 differentiation of mononuclear cells from RA PB and SF but increased the proportion of Treg cells. In contrast, inhibition of STAT-5, the transcription factor for Treg cells, increased the proportion of Th17 cells and reduced that of Treg cells. Conclusion Our findings indicate that modulation of STAT-3 in CD4+ T cells affects the differentiation of Th17 cells and Treg cells in patients with RA. This role of STAT-3 in RA synovial T cells may provide a new therapeutic target for the management of RA. [ABSTRACT FROM AUTHOR]

Maintaining an appropriate balance between subsets of CD4+ helper T cells and T regulatory cells (Tregs) is a critical process in immune homeostasis and a protective mechanism against autoimmunity and inflammation. To identify the role of vitamin A-related compounds, we investigated the regulation of interleukin (IL)-17-producing helper T cells (Th17 cells) and Tregs treated with all-trans-retinal (retinal). CD4+T cells or total cells from the spleens of C57BL/6 mice were stimulated under Treg-polarizing (anti-CD3/CD28 and TGF-β) or Th17-polarizing (anti-CD3/CD28, TGF-β, and IL-6) conditions in the presence or absence of retinal. To analyze their suppressive abilities, retinal-induced Tregs or TGF-β-induced Tregs were co-cultured with responder T cells. Collagen-induced arthritis (CIA) was established in interferon (IFN)-γ knockout mice. On day 13, retinal-induced Tregs were adoptively transferred to mice with established CIA after second immunizations. Compared with TGF-β-induced Treg cells, retinal-induced Tregs showed increased Foxp3 expression and mediated stronger suppressive activity. Under Th17-polarizing conditions, retinal inhibited the production of IL-17 and increased the expression of Foxp3.Retinal-induced Tregs showed therapeutic effects in IFN-γ knockout CIA mice. Thus, we demonstrated that retinal reciprocally regulates Foxp3+ Tregs and Th17 cells. These findings suggest that retinal, a vitamin A metabolite, can regulate the balance between pro- and anti-inflammatory immunity. A better understanding of the manipulation of Foxp3 and Tregs may enable the application of this tremendous therapeutic potential in various autoimmune diseases. [ABSTRACT FROM AUTHOR]

Abstract: Cytotoxic T lymphocyte antigen-4 (CTLA4) and IgG fusion protein, CTLA4-Ig, is a therapeutic agent used for rheumatoid arthritis. It binds B7 molecules on dendritic cells (DCs) and thereby blocks B7/CD28 costimulatory interaction and inhibits effective T cell proliferation. However, the effect of CTLA4-Ig on the regulatory T cell (Treg) is still not known. In this study, we investigated the influence of CTLA4-Ig on the CD4+CD25+Foxp3+ Treg population in collagen-induced arthritis (CIA) mouse model. CTLA4-Ig suppressed CIA and increased the CD4+CD25+Foxp3+ Treg population in joint and spleen. When CD11c + DCs and CD4+T cells from CIA mice were cultured with anti-CD3, CTLA4-Ig increased the CD4+CD25 + Foxp3+ Treg population in a TGF-β-dependent manner. When CD11c + DCs from CIA mice were treated with CTLA4-Ig and adoptively transferred into CIA-induced mice, arthritis did not develop in association with the increase in CD4+CD25+Foxp3+ Treg population. However, in CTLA4-Ig-untreated DC-transferred CIA mice, arthritis developed and then rapidly progressed. Our study demonstrated that CTLA4-Ig suppressed CIA by modifying DCs from CIA mice into tolerogenic DCs to increase the CD4+CD25+Foxp3+ Treg population and this seems to be the new immune regulatory mechanism of CTLA4-Ig. [Copyright &y& Elsevier]

Abstract: To examine the effects of nuclear factor kappa B (NF-κB) inhibition on the secretion of macrophage migration inhibitory factor (MIF) in human CD4+ T cells. Isolated human CD4+ T cells were cultured for 24h with pharmacological inhibitors of NF-κB including parthenolide, pyrrolidine dithiocarbamate, BAY 11-7082, gliotoxin, oridonin, andrographolide, and NF-κB shRNA. MIF concentration was measured by intracellular flow cytometry, enzyme-linked immunosorbent assay, and real-time polymerase chain reaction. The intracellular concentrations O2 −, H2O2, and glutathione were measured using the oxidation-sensitive fluorescent dyes dihydroethidium, dichlorodihydrofluorescein diacetate, and monochlorobimane, respectively. The amount of phosphorylated c-Jun was measured by Western blotting. Treatment of CD4+ T cells with NF-κB inhibitors significantly increased MIF concentration in culture supernatants, MIF gene expression, and O2 − production, and decreased the intracellular concentrations of MIF, H2O2, and glutathione. Treatment with LY294002 (PI3K inhibitor) and SP600125 (JNK inhibitor) suppressed NF-κB inhibitor induced MIF mRNA expression and MIF secretion. LY294002 and SP600125 inhibited the parthenolide-induced phosphorylation of c-Jun. Treatment with H2O2 decreased the amount of intracellular MIF protein and increased MIF concentration in the culture supernatant. N-acetylcysteine, an antioxidant precursor of glutathione, inhibited the parthenolide-induced and H2O2-induced secretion of MIF. These results indicate that pharmacological inhibition of NF-κB causes the release of MIF through de novo synthesis of MIF and the secretion of preformed MIF in CD4+ T cells through the production of reactive oxygen species. [Copyright &y& Elsevier]

Abstract: The receptor activator of nuclear factor κB ligand (RANKL) is an osteoclastogenic mediator, which is mainly expressed by stromal cells and osteoblast. However, T cells can also be an important provider for RANKL in special condition such as autoimmune arthritis. We examined the RANKL expression of hyporesponsive CD4+ T cells induced by oral feeding with type II collagen in collagen-induced arthritis (CIA) mice. The potential of RANKL expression in CD4+ T cells was downregulated in tolerance, as compared with CIA. One of possible explanations for this phenomenon is that CII-specific T cell activation was intrinsically impaired in oral tolerance, which caused suppression of RANKL expression of CD4+ T cells. We also investigated the extrinsic role of cytokine in this process. IL-17, well-known pro-inflammatory cytokine was upregulated in CIA and downregulated in tolerance. IL-17 had a potential to stimulate T cells to express RANKL in dose-dependent manner. IL-17-associated RANKL expression of CD4+ T cells was downregulated in oral tolerance, suggesting that the induction of tolerance ameliorates IL-17-induced RANKL expression of T cells in murine CIA. We also discovered that CIA – T cells could enhance osteoclastogenesis but not oral tolerance – T cells. Oral tolerance might be promising therapeutic option in viewpoints of modulating autoreactivity of CII which can induce not only IL-17 production but also RANKL expression in CD4+ T cells. [Copyright &y& Elsevier]

Background: To evaluate the immunomodulatory effect of Lactobacillus sakei in a mouse model of rheumatoid arthritis (RA) and in human immune cells.Methods: We evaluated whether L. sakei reduced the severity of collagen-induced arthritis (CIA) and modulated interleukin (IL)-17 and IL-10 levels, as well as whether it affected the differentiation of CD4+ T cells and regulatory B cells. We evaluated osteoclastogenesis after culturing bone marrow-derived mononuclear cells with L. sakei.Results: The differentiation of T helper 17 cells and the serum level of IL-17 were suppressed by L. sakei in both human peripheral blood mononuclear cells and mouse splenocytes. The serum level of IL-10 was significantly increased in the L. sakei-treated group, whereas the regulatory T cell population was unchanged. The population of regulatory B cells significantly increased the in L. sakei-treated group. Oral administration of L. sakei reduced the arthritis incidence and score in mice with CIA. Finally, osteoclastogenesis and the mRNA levels of osteoclast-related genes were suppressed in the L. sakei-treated group.Conclusion: L. sakei exerted an anti-inflammatory effect in an animal model of RA, regulated Th17 and regulatory B cell differentiation, and suppressed osteoclastogenesis. Our findings suggest that L. sakei has therapeutic potential for RA. [ABSTRACT FROM AUTHOR]

Background: Spondyloarthritis (SpA) is chronic inflammatory arthritis, and interleukin (IL)-17 is crucial in SpA pathogenesis. Type 17 helper T (Th17) cells are one of major IL-17-secreting cells. Signal transducer and activator of transcription (STAT)-3 signaling induces Th17 differentiation. This study investigated the effects of protein inhibitor of activated STAT3 (PIAS3) on SpA pathogenesis. Curdlan was injected into SKG ZAP-70W163C mice for SpA induction.Methods: The PIAS3 or Mock vector was inserted into mice for 10 weeks. Clinical and histologic scores of the paw, spine, and gut were evaluated. The expression of IL-17, tumor necrosis factor-α (TNF-α), STAT3, and bone morphogenic protein (BMP) was measured. Confocal microscopy and flow cytometry were used to assess Th cell differentiation.Results: PIAS3 significantly diminished the histologic scores of the paw and gut. PIAS3-treated mice displayed decreased expression of IL-17, TNF-α, and STAT3 in the paw, spine, and gut. BMP-2/4 expression was lower in the spines of PIAS3-treated mice. Th cell differentiation was polarized toward the upregulation of regulatory T cells (Tregs) and the downregulation of Th17 in PIAS3-treated mice.Conclusion: PIAS3 had beneficial effects in mice with SpA by reducing peripheral arthritis and gut inflammation. Pro-inflammatory cytokines and Th17/Treg differentiation were controlled by PIAS3. In addition, BMPs were decreased in the spines of PIAS3-treated mice. These findings suggest that PIAS3 could have therapeutic benefits in patients with SpA. [ABSTRACT FROM AUTHOR]

• A3373, a novel chemical inhibitor of PLD1, inhibits LPS-induced TNF-α and IL-6 production in mouse splenocytes. • A3373 reduces the production of IL-17 and IgG in murine splenocytes in vitro. • A3373 ameliorates autoimmune arthritis in vivo. • A3373 prevents systemic bone demineralization. • A3373 suppresses osteoclastogenesis in vitro. Phospholipase D1 (PLD1), which catalyzes the hydrolysis of phosphatidylcholine to phosphatidic acid and choline, plays multiple roles in inflammation. We investigated the therapeutic effects of the newly developed PLD1 inhibitors A2998, A3000, and A3773 in vitro and in vivo rheumatoid arthritis (RA) model. A3373 reduced the levels of LPS-induced TNF-α, IL-6, and IgG in murine splenocytes in vitro. A3373 also decreased the levels of IFN-γ and IL-17 and the frequencies of Th1, Th17 cells and germinal-center B cells, in splenocytes in vitro. A3373 ameliorated the severity of collagen-induced arthritis (CIA) and suppressed infiltration of inflammatory cells into the joint tissues of mice with CIA compared with vehicle-treated mice. Moreover, A3373 prevented systemic bone demineralization in mice with CIA and suppressed osteoclast differentiation and the mRNA levels of osteoclastogenesis markers in vitro. These results suggest that A3373 has therapeutic potential for RA. [ABSTRACT FROM AUTHOR]

• IL-21 induces the expression of the fibrotic factor, Col1a1, by enhancing STAT3 phosphorylation following IL-21 administration in dermal fibroblasts. • IL-21 deficiency ameliorated the development and progression of SSc. • IL-21 deficiency reduced fibrosis in the skin and lung tissues by downregulating STAT3 activity and Th1, Th2, and Th17 cells. • IL-21 is a crucial molecule in SSc disease and could be a therapeutic target for SSc treatment. Systemic sclerosis (SSc) is an autoimmune disease characterized by vasculopathy, abnormal inflammation, and fibrosis of the skin and internal organs, notably the skin and lungs, significantly impairing quality of life. There is currently no cure for SSc, and its etiology remains largely unknown, presenting a primary barrier to effective treatment. We investigated the role of interleukin-21 (IL-21) in the pathogenesis of SSc. We assessed the expression levels of fibrosis-related genes in human dermal fibroblasts exposed to IL-21 and TGF beta. We also induced SSc in wild-type C57BL/6 mice and IL-21 knockout (KO) mice with a C57BL/6 background using bleomycin (Bleomycin). Histological analyses were conducted on skin and lung tissues from these mice. The distribution and expression levels of fibrosis-related proteins in the tissues were examined via immunohistochemistry and quantitative real-time PCR. Furthermore, we measured the frequency of Th1, Th2, and Th17 cells among splenocytes through flow cytometry. IL-21 activation led to STAT3 phosphorylation more than TGF beta in dermal fibroblasts. In IL-21 KO mice with BLM-induced SSc, skin thickness and lung fibrosis were reduced. The absence of IL-21 in these mice resulted in suppressed expression of fibrosis-related genes, including Col1a1, Col1a2, Col3a1, CTGF, α-SMA, STAT3 , and TGFβ , in the skin and lungs. It also led to a decreased frequency of Th1, Th2, and Th17 cells, as well as a lower Th17/Treg ratio among splenocytes, factors known to contribute to the development of SSc. IL-21 contributes to the development of SSc by promoting the expression of fibrosis-related genes and modulating the levels of CD4+ T cells. [ABSTRACT FROM AUTHOR]

Vimentin contributes to the positioning and function of organelles, cell migration, adhesion, and division. However, secreted vimentin accumulates on the cell surface (Mor-Vaknin et al., 2003; Ramos et al., 2020 [ 1 , 2 ]) where it acts as a coreceptor for viral infection and as an autoantigen in inflammatory and autoimmune diseases. The roles of vimentin in Th17 cells were examined in mice with knockdown of vimentin. We also examined whether STAT3 is required for vimentin expression. Vimentin expression was significantly increased in Th17 cells through STAT3 activation, and vimentin+ IL-17+ T cells were markedly increased in the joint and spleen tissues of CIA mice. The arthritis score and expression levels of proinflammatory cytokines were significantly decreased in CIA mice treated with vimentin shRNA vector. In this study, we demonstrated that vimentin is significantly expressed in Th17 cells through STAT3 activation. Our results provide new insights into the role of vimentin in Th17 cells and the complex pathogenesis of RA. [ABSTRACT FROM AUTHOR]

Abstract: Grape seed proanthocyanidin extract (GSPE), which is the antioxidant derived from grape seeds, has been reported to possess a variety of potent properties. We have previously shown that GSPE attenuates collagen-induced arthritis. However the mechanism by which GSPE regulates the immune response remains unclear, although it may involve effects on the regulation of pathogenic T cells in autoimmune arthritis. To clarify this issue, we have assessed the effects of GSPE on differential regulation of Th17 and regulatory T (Treg) cells subsets in vitro in mouse and human CD4+ T cells. We observed that GSPE decreased the frequency of IL-17+CD4+Th17 cells and increased induction of CD4+CD25+forkhead box protein 3 (Foxp3)+ Treg cells. In vivo, GSPE effectively attenuated clinical symptoms of established collagen-induced arthritis in mice with concomitant suppression of IL-17 production and enhancement of Foxp3 expression (type II collagen-reactive Treg cells) in CD4+ T cells of joints and splenocytes. The presence of GSPE decreased the levels of IL-21, IL-22, IL-26 and IL-17 production by human CD4+ T cells in a STAT3-dependent manner. In contrast, GSPE induces Foxp3+ Treg cells in humans. Our results suggest that GSPE possesses a reciprocal control over IL-17 and Foxp3. By potently regulating inflammatory T cell differentiation, GSPE may serve as a possible novel therapeutic agent for inflammatory and autoimmune diseases, including rheumatoid arthritis. [ABSTRACT FROM AUTHOR]

Highlights • Cucurbitacine E (CuE) repressed alloreactive T cell response in mouse splenocytes and human PBMCs. • CuE suppressed IL-17 expression through suppression of STAT3 phosphorylation. • CuE inhibited development of acute GvHD • CuE mitigated inflammation in intestines and fibrotic skin in aGvHD Abstract Cucurbitacin E (CuE) is a biochemical compound found in plants that are members of the family CuE has been studied for its roles in anti-inflammation and the inhibition of angiogenesis as well as for its properties as an antioxidant. CuE is a new agent that was identified as a selective inhibitor of the signal transducer and activator of transcription 3 (STAT3)-related pathway. STAT3, a pivotal transcription factor for Th17 differentiation, is critical for T cell alloactivation in acute graft-versus-host disease (aGvHD). We investigated whether CuE attenuates the development of aGvHD through the suppression of Th17 cells. The alloreactive proliferation of mouse and human T cells was reduced by CuE treatment. CuE also decreased pro-inflammatory cytokines, such as IL-17 and IFN-γ, in alloreactive T cells. STAT3-responsive and IL-17A-promoter activities were also suppressed by CuE treatment, confirming that activated STAT3 was decreased by CuE treatment. To construct an aGvHD-induced mouse line, splenocytes and bone marrow cells from C57BL/6 mice were transplanted into BALB/c mice with complete mis-matched major histocompatibility complex molecules. CuE was administered to aGvHD animals 3 days per week via intraperitoneal injection. CuE attenuated the severity of aGvHD disease-related scores compared to the vehicle group. CuE inhibited skin inflammation and fibrosis, as evidenced by the expression of α-Sma and Col-I in aGvHD mice compared to the vehicle group. Additionally, aGvHD mice treated with CuE showed improved histopathological features in the small and large intestines, whereas the vehicle group showed collapsed villi in the small intestine and cryptic structures in the large intestine. We also observed a marked reduction of pro-inflammatory cytokines in the intestinal tissue. Collectively, our data suggest that CuE could serve as a therapeutic agent for patients with aGvHD. [ABSTRACT FROM AUTHOR]

Coenzyme Q10 (CoQ10) is a powerful antioxidant substance synthesized in the body. The current study aimed to determine whether CoQ10 suppresses inflammation and inhibits p-STAT3 expression in an experimental colitis mouse model. The mice were orally fed with CoQ10 once a day for 13 days. Histological analysis of the colons was performed by immunohistochemistry. Expression of IL-17, FOXP3, p53, AMPK, and mTOR and activation of p-STAT3 and p-STAT5 in lymph node and spleen tissues were detected by confocal microscopy of stained tissue sections. The relative mRNA expression was measured with real-time PCR, and protein levels were examined by western blot. CoQ10 reduced the disease activity index score and the colon histological score. It also reduced inflammatory mediators in the colon and increased the colon length. The expression of IL-17 and p-STAT3 was decreased with CoQ10 treatment. In contrast, CoQ10 treatment increased the expression of p-AMPK and FOXP3. Expression of anti-inflammatory cytokines was shown to increase in colitis mice treated with CoQ10. These results suggested that CoQ10 may reduce the severity of colitis and suppress inflammation through the inhibition of p-STAT3 and IL-17. These results support the use of CoQ10 as a potential targeted therapy for the treatment of colitis. [ABSTRACT FROM AUTHOR]

IL-12p40 homodimer, a natural antagonist of IL-12 and IL-23, performs an important role in the expression of proinflammatory cytokines that is essential for Th1 and Th17 immune responses. Here, we reveal the therapeutic and immunosuppressive effect of the IL-12p40 subunit ((p40) 2 -Fc) in an experimental autoimmune arthritis model. We hypothesized that (p40) 2 -Fc may reduce the inflammatory response and the activation of T cells. In this study, we intraperitoneally injected (p40) 2 -Fc into collagen induced arthritis (CIA) mice to identify whether (p40) 2 -Fc attenuates CIA severity. (p40) 2 -Fc reduced the development of CIA, joint inflammation and cartilage destruction. (p40) 2 -Fc also significantly decreased the concentration of serum immunoglobulin as well as the number of T cells and C II specific T cells. In addition, osteoclastogenesis in (p40) 2 -Fc treated mice was down-regulated compared to the mice treated with (p40) 2 -Fc control. We observed that (p40) 2 -Fc treatment alleviates arthritis in mice with CIA, reducing inflammation and osteoclast differentiation. These findings suggest that (p40) 2 -Fc can be a potential therapeutic approach for autoimmune arthritis. [ABSTRACT FROM AUTHOR]

Introduction IL-1β signalling has a critical role in the pathogenesis of various types of inflammatory arthritis including rheumatoid arthritis (RA). We aimed to investigate the therapeutic effects of human IL-1 receptor antagonist with Fc fragment (hIL-1Ra-Fc) on autoimmune arthritis and to identify the possible mechanisms by which hIL-1RA-Fc exerts anti-arthritic effects in a murine model of RA and patients with rheumatoid arthritis. Methods Collagen-induced arthritis (CIA) murine model was established in DBA/1J mice. The levels of various cytokines were determined by using enzyme-linked immunosorbent assay. The mouse joints were assessed for clinical arthritis score and histologic features. Th17 cells and Treg cells were stained by using antibodies specific for CD4, IL-17, CD25, and FoxP3. Osteoclastogenesis was determined by TRAP staining and real-time PCR. Results hIL-1RA-Fc reduced the arthritis incidence, histological inflammation and cartilage score in the CIA model. The expression of proinflammatory cytokines, VEGF and RANK, was reduced in the affected joint of hIL-1Ra-Fc-treated mice. hIL-1Ra-Fc-treated mice showed decreased number of Th17 cells with increased number of Treg cells in spleens. hIL-1Ra-Fc reduced Th17 cell differentiation by inactivation of STAT3 signalling, and reciprocally induced Treg cell differentiation through STAT5 signalling. In addition, the expression of IL-17, TNF-α, RANKL, and VEGF was decreased, while Foxp3 gene expression was increased in PBMCs of RA patients after administration of hIL-1Ra-Fc. Conclusion The anti-arthritis effects of hIL-1RA-Fc are associated with regulation of balance between Th17 cells and Treg cells and suppression of osteoclastogenesis and angiogenesis in the affected joints. [ABSTRACT FROM AUTHOR]

The objective of this study was to determine whether coenzyme Q10 (CoQ10) can reduce the severity of graft versus host disease (GVHD) by inducing regulatory T cells (Tregs). CoQ10 or vehicle was orally administrated once a day for 22 days to mice with GVHD. We measured the alloresponse of the T cells and the GVHD clinical scores. Real-time polymerase chain reaction was used to examine messenger RNA (mRNA) level. Flow cytometry and enzyme-linked immunosorbent assay were used to evaluate protein expression. CoQ10 reduced the T-cell alloresponse and the expression of interferon (IFN)- γ and interleukin (IL)-17. The severity of GVHD and gene expressions of IL-6 and tumor necrosis factor (TNF)- α decreased with CoQ10 treatment. Furthermore, CoQ10 promoted weight gain and survival in GVHD mice. Flow cytometry revealed that CoQ10 dose dependently induced Treg differentiation, but FK506, an immunosuppressive drug, decreased Treg differentiation dose dependently. In conclusion, CoQ10 downregulates the alloreactivity of T cells and reduces GVHD severity, enhancing the differentiation of Tregs. [ABSTRACT FROM AUTHOR]

Establishment of mixed chimerism is an ideal approach to induce donor-specific tolerance while expanding its potential in various clinical settings. Despite the developments in partial conditioning regimens, improvements are still needed in reducing toxicity and bone marrow transplantation-related complications. Recently, cell-based therapies, including mesenchymal stem cells (MSCs), have been incorporated in establishing noncytoreductive mixed chimerism protocols; however, its efficacy is only partial and shows reversed immunosuppressive properties. This study demonstrates a novel approach to induce mixed chimerism and tolerance through combinatory cell-based immune modulation (CCIM) of MSCs and regulatory T cells (Tregs). We hypothesize that the interaction between these cells may lead to greater inhibition of host immune responses. Compared with single cell therapy, CCIM induced a higher engraftment rate and robust donor-specific tolerance to skin allografts across full major histocompatibility complex barriers. These regulatory effects were associated with inhibition of natural killer cell cytotoxic activity, CD4+IL-17+ cells, memory B cells, plasma cells, and immunoglobulin production levels along with increased frequencies of CD4+Foxp3+ cells, IL-10-producing mature B cells, and myeloid-derived suppressor cells. Furthermore, CCIM was able to regulate mortality in a graft-versus-host disease model through reciprocal regulation of Treg/Th17. Taken together, we suggest CCIM as a clinically applicable strategy for facilitating the induction of mixed chimerism and permanent tolerance. [ABSTRACT FROM AUTHOR]

Highlights: [•] PIAS3 overexpression suppressed Graft-versus-host disease models. [•] PIAS3 reciprocally regulated Th17/Treg balance via STAT3 targeting. [•] PIAS3 induced CD4+Foxp3+ Treg and CD8+Foxp3+ Treg in GVHD. [•] PIAS3 inhibit plasma B cell and IgG producing cell. [ABSTRACT FROM AUTHOR]

Highlights: [•] EGCG has a therapeutic effect on both obese (DIO) mice and obese CIA mice models. [•] EGCG suppress inflammatory mediators and increases regulatory components in the obese CIA mice. [•] EGCG increases the frequency of Foxp3+ Treg cells and decreases the frequency of Th17 cells in obese CIA mice. [Copyright &y& Elsevier]

Abstract: Retinoids (e.g., vitamin A and its derivatives) can regulate immune responses. The aim of this study was to determine whether all-trans retinaldehyde (retinal), a vitamin A derivative, can inhibit inflammatory responses and joint destruction in DBA/1J mice with collagen-induced arthritis (CIA). The arthritis score and incidence of arthritis were lower in mice treated with retinal compared to those treated with cottonseed oil. Histopathologic evidence of joint damage was lower in mice treated with retinal, corresponding with a reduction in the infiltration of immune cells in mice treated with retinal type II collagen (CII)-stimulated spleen cells. In addition, the expression of proinflammatory cytokines, oxidative stress proteins, and osteoclast markers were significantly reduced in mice treated with retinal. In vitro, retinal induced increased Foxp3 expression and inhibited Th17 development. The proportion of Foxp3+ Treg cells was increased in the spleens of mice treated with retinal, whereas the proportion of Th17 cells was reduced. In both mice and a human culture system, tartrate-resistant acid phosphatase (TRAP) positive mononuclear cells and multinucleated cells were significantly reduced after treatment with retinal. The expression of osteoclast differentiation markers was dramatically decreased upon addition of retinal. This is the first study to demonstrate the therapeutic effect of retinal on an autoimmune arthritis model in mice through reciprocal regulation of Th17 and regulatory T cells and protection of differentiation and activation of osteoclasts. Taken together, our findings indicate that retinal has profound immunoregulatory functions and potential value for the treatment of autoimmune inflammatory disorders. [Copyright &y& Elsevier]

Abstracts: Patients with end-stage renal disease (ESRD) exhibit immune dysregulation, but the precise immunological profile and the effect of hemodialysis (HD) on it has not been investigated fully. Thirty-eight ESRD patients (22 on HD and 16 in pre-dialysis) and 24 healthy volunteers were included. We compared the T cell immune profiles as in these patients. Among the effector T cell subset, the percentages of Th17 and Th2 cells were significantly higher in the ESRD group than in the healthy controls (P <0.05). The percentage of Th1 cells did not differ significantly between these groups. The percentages of Th1, Th2 and Th17 cells did not differ significantly (P >0.05) between the two subgroups within the ESRD group. The CCR4−CCR6+/CD4+ T cell percentage was also significantly higher in the ESRD group. The naïve T cell (Tnaïve) percentage was significantly lower in the ESRD group, and the difference between patients and controls was greater in the pre-dialysis patients than in the HD patients (P <0.05, for each comparison). By contrast, the percentages of central memory T cells (TCM) and effector memory T (TEM) cells were significantly higher in the ESRD group. Interleukin-17 production by TEM cells was significantly higher in the ESRD group. The severity of uremia was related negatively to the Tnaïve cell percentage but positively to the TCM and TEM cell percentages. The percentages of TEM and CD45RA+ T effector memory subsets of CD8+ T cells were significantly higher in the ESRD group (P <0.05). The result of this study showed significantly altered T cell-associated immunity and that it could not be corrected with hemodialysis. [Copyright &y& Elsevier]

Sjögren's syndrome (SS) is a systemic autoimmune disease with severe dysfunction of glandular secretory function mediated by T and B lymphocyte infiltration into the exocrine glands, including the salivary and lacrimal glands. Follicular helper T (Tfh) cells exacerbate the disease by causing B cell hyperactivity. Inhibitor of DNA binding 3 (Id3) deficiency causes activation of Tfh cells and is known to be a clinical manifestation of human SS disease. In this study, we investigated the mechanism of action of Pax3, which is reduced in SS and can interact with Id3, in NOD/ShiLtJ mice as an animal model of SS. Treatment with interleukin (IL)-21, a major cytokine secreted from Tfh cells, suppressed Pax3 and Id3 expression via STAT3 in splenic T cells in vitro. Administration of pCMV14-3xFlag PAX3 vector improved the severity of SS by reducing the number of Tfh cells in NOD/ShiLtJ mice. Application of IL-21R-Fc increased the number of Pax3- and Id3-positive cells in the salivary glands, while reducing the proportion of Tfh cells and IL-17-producing T cells in NOD/ShiLtJ mice. The salivary glands from SS patients showed decreased levels of Pax3 or Id3 expression compared with healthy controls. Our findings regarding reinforcement of the Pax3-Id3 signal pathway may facilitate the development of novel therapeutic strategies for SS. [ABSTRACT FROM AUTHOR]

Interleukin (IL) 21 plays a key role in the development of acute graft-versus-host disease (GVHD) after allogeneic bone marrow transplantation. Therapeutic manipulation of IL-21 activity may improve acute GVHD during the early-posttransplant period. We investigated the mechanisms regulating T- and B-cells during IL-21 blockade in acute GVHD. Interleukin 21 blockade enhanced regulatory T and T helper (Th) 2 cell differentiation and inhibited Th1- and Th17-derived transcription factors and cytokines as a modulator of activated T-cells. Interleukin 21 -/- cell recipients showed increased mature B- and marginal-zone B-cells, but decreased memory B-cells, germinal center formation, and plasma cells that did not lead to immunoglobulin production. B-cell activating factor (BAFF) and a proliferation-inducing ligand (APRIL) are involved in the induction and maintenance of T- and B-cell responses. We observed decreased levels of only BAFF during acute GVHD and confirmed that mammalian target of rapamycin complex 1 was reduced by the BAFF/BAFF-receptor pathway. Therefore, this study suggests that IL-21 blockade modulates activated T- and B-cell homeostasis via BAFF-pathway-mediated inhibition in acute GVHD following murine allogeneic bone marrow transplantation. [ABSTRACT FROM AUTHOR]