Chang-Suk Chae

Lactobacillus casei potentiates induction of oral tolerance in experimental arthritis

Probiotics have been shown to exert beneficial effects on modulation of diverse diseases. However, no information is available for the effect of probiotics in the induction of oral tolerance in autoimmune diseases. The main purpose of this study was to elucidate whether Lactobacillus casei (L. casei) affect the induction of oral tolerance in experimental rheumatoid arthritis (RA). Type II collagen (CII) alone or together with L. casei was orally administered into collagen-induced arthritis (CIA) rats, and its effects on the clinical and histopathological aspects of RA were investigated. Co-administration of L. casei with CII more effectively suppressed clinical symptoms, paw swelling, lymphocyte infiltration and destruction of cartilage tissues of experimental arthritis than the rats treated with CII alone. The enhanced therapeutic efficacy was associated with an increase in anti-inflammatory cytokines (IL-10 and TGF-beta) while decreasing pro-inflammatory cytokines (IL-1beta, IL-2, IL-6, IL-12, IL-17, IFN-gamma and TNF-alpha). Co-administration of L. casei with CII more effectively suppressed CII-reactive T cell proliferation and the levels of Th1-type IgG isotypes (IgG2a and IgG2b), while up-regulating Foxp3 expression levels and the population of Foxp3(+) CD4(+) T cells. Our study provides evidence that L. casei could potentiate antigen-specific oral tolerance and suppress Th1-type immune responses of arthritic inflammation.

Interaction of Ets-1 with HDAC1 Represses IL-10 Expression in Th1 Cells

IL-10 is a multifunctional cytokine that plays a crucial role in immunity and tolerance. IL-10 is produced by diverse immune cell types, including B cells and subsets of T cells. Although Th1 produce IL-10, their expression levels are much lower than Th2 cells under conventional stimulation conditions. The potential role of E26 transformation-specific 1 (Ets-1) transcription factor as a negative regulator for Il10 gene expression in CD4+ T cells has been implicated previously. In this study, we investigated the underlying mechanism of Ets-1–mediated Il10 gene repression in Th1 cells. Compared with wild type Th1 cells, Ets-1 knockout Th1 cells expressed a significantly higher level of IL-10, which is comparable with that of wild type Th2 cells. Upregulation of IL-10 expression in Ets-1 knockout Th1 cells was accompanied by enhanced chromatin accessibility and increased recruitment of histone H3 acetylation at the Il10 regulatory regions. Reciprocally, Ets-1 deficiency significantly decreased histone deacetylase 1 (HDAC1) enrichment at the Il10 regulatory regions. Treatment with trichostatin A, an inhibitor of HDAC family, significantly increased Il10 gene expression by increasing histone H3 acetylation recruitment. We further demonstrated a physical interaction between Ets-1 and HDAC1. Coexpression of Ets-1 with HDAC1 synergistically repressed IL-10 transcription activity. In summary, our data suggest that an interaction of Ets-1 with HDAC1 represses the Il10 gene expression in Th1 cells.

Hypoxia-inducible factor-2α is an essential catabolic regulator of inflammatory rheumatoid arthritis.

Hypoxia-inducible factor-2α (HIF-2α) is sufficient to cause experimental rheumatoid arthritis and acts to regulate the functions of fibroblast-like cells from tissue surrounding joints, independent of HIF-1α.

Prophylactic Effect of Probiotics on the Development of Experimental Autoimmune Myasthenia Gravis

Probiotics are live bacteria that confer health benefits to the host physiology. Although protective role of probiotics have been reported in diverse diseases, no information is available whether probiotics can modulate neuromuscular immune disorders. We have recently demonstrated that IRT5 probiotics, a mixture of 5 probiotics, could suppress diverse experimental disorders in mice model. In this study we further investigated whether IRT5 probiotics could modulate the progression of experimental autoimmune myasthenia gravis (EAMG). Myasthenia gravis (MG) is a T cell dependent antibody mediated autoimmune disorder in which acetylcholine receptor (AChR) at the neuromuscular junction is the major auto-antigen. Oral administration of IRT5 probiotics significantly reduced clinical symptoms of EAMG such as weight loss, body trembling and grip strength. Prophylactic effect of IRT5 probiotics on EMAG is mediated by down-regulation of effector function of AChR-reactive T cells and B cells. Administration of IRT5 probiotics decreased AChR-reactive lymphocyte proliferation, anti-AChR reactive IgG levels and inflammatory cytokine levels such as IFN-γ, TNF-α, IL-6 and IL-17. Down-regulation of inflammatory mediators in AChR-reactive lymphocytes by IRT5 probiotics is mediated by the generation of regulatory dendritic cells (rDCs) that express increased levels of IL-10, TGF-β, arginase 1 and aldh1a2. Furthermore, DCs isolated from IRT5 probiotics-fed group effectively converted CD4+ T cells into CD4+Foxp3+ regulatory T cells compared with control DCs. Our data suggest that IRT5 probiotics could be applicable to modulate antibody mediated autoimmune diseases including myasthenia gravis.

Suppression of experimental myasthenia gravis by a B-cell epitope-free recombinant acetylcholine receptor

Myasthenia gravis (MG) and experimental autoimmune MG (EAMG) are antibody-mediated autoimmune diseases in which the nicotinic acetylcholine receptor (AChR) is the major autoantigen. Previously we have revealed that oral treatment with the less native recombinant fragment of the extracellular domain of the human AChR (Halpha1-205) suppressed ongoing EAMG, whereas the more native recombinant Trx-Halpha1-210 exacerbated EAMG. In this study, we speculated on the role of B-cell epitopes in oral tolerogens for the induction of oral tolerance in EAMG. We developed a B-cell epitope-free AChR fragment (BF-AChR) by removing two major B-cell epitopes (67-76 and 129-145) from Trx-Halpha1-210. BF-AChR exhibited a poor response to EAMG sera and to AChR-specific B- and T-cells while its parent fragment, Trx-Halpha1-210, showed much higher reactivity. Oral administration of BF-AChR ameliorated the symptoms in ongoing myasthenic rats accompanied by a significant decrease in AChR-specific humoral and Th1 cellular responses. The underlying mechanism for BF-AChR-induced oral tolerance was mediated by a shift from Th1 to regulatory T-cell (IL-10(+), CD4(+) TGF-beta(+) or Foxp3(+)) responses. This shift was assessed by changes in the cytokine profile and a deviation in the anti-AChR IgG isotypes from IgG2a/IgG2b to IgG1. Our results suggest that the removal of pathogenic B-cell epitopes from AChR fragments increases tolerogenicity by reducing the activation and proliferation of autoreactive B- and T-cells. Collectively, careful consideration of the immunogenicity of a tolerogen is necessary to induce successful oral tolerance in autoimmune disorders.

NFAT1 and JunB Cooperatively Regulate IL-31 Gene Expression in CD4+ T Cells in Health and Disease

IL-31 is a key mediator of itching in atopic dermatitis (AD) and is preferentially produced by activated CD4+ T cells and Th2 cells. Although pathophysiological functions of IL-31 have been suggested in diverse immune disorders, the molecular events underlying IL-31 gene regulation are still unclear. In this study we identified the transcription start site and functional promoter involved in IL-31 gene regulation in mouse CD4+ T cells. TCR stimulation–dependent IL-31 expression was found to be closely linked with in vivo binding of NFAT1 and JunB to the IL-31 promoter. Although NFAT1 alone enhanced IL-31 promoter activity, it was further enhanced in the presence of JunB. Conversely, knockdown of either NFAT1 or JunB resulted in reduced IL-31 expression. NFAT1-deficient CD4+ T cells showed a significant defect in IL-31 expression compared with wild-type CD4+ T cells. In agreement with these findings, mice subjected to atopic conditions showed much higher levels of IL-31, which were closely correlated with a significant increase in the number of infiltrated NFAT1+CD4+ T cells into the AD ears. Amelioration of AD progression by cyclosporin A treatment was well correlated with downregulation of IL-31 expressions in CD4+ T cells and total ear residual cells. In summary, our results suggest a functional cooperation between NFAT1 and JunB in mediating IL-31 gene expression in CD4+ T cells and indicate that interference with this interaction or their activity has the potential of reducing IL-31–mediated AD symptoms.

Enhanced Chromatin Accessibility and Recruitment of JUNB Mediate the Sustained IL-4 Expression in NFAT1 Deficient T Helper 2 Cells

Nuclear factor of activated T cells (NFAT) is a family of transcription factors composed of five proteins. Among them, NFAT1 is a predominant NFAT protein in CD4+ T cells. NFAT1 positively regulates transcription of a large number of inducible cytokine genes including IL-2, IL-4, IL-5 and other cytokines. However, disruption of NFAT1 results in an unexpected increase of IL-4. In this study, we have investigated the role of NFAT1 in regulation of IL-4 gene expression in T helper 2 cells (Th2) from an epigenetic viewpoint. NFAT1 deficient Th2 cells showed a sustained IL-4 expression while wild type (WT) cells reduced its expression. We tested whether epigenetic maintenance and changes in the chromatin architecture of IL-4 promoter locus play a role in differential IL-4 transcription between in WT and NFAT1 deficient Th2 cells. Compared with WT, NFAT1 deficient CD4+ Th2 cells exhibited enhanced chromatin accessibility with permissive histone modification and DNA demethylation in the IL-4 promoter region. Transcription factors bound to IL-4 promoter region in the absence of NFAT1 were identified by Micro-LC/LC-MS/MS analysis. Among the candidates, preferential recruitment of JUNB to the IL-4 promoter was confirmed by chromatin immunoprecipitation analysis. Overexpression of JUNB together with SATB1 synergistically upregulated IL-4 promoter activity, while knockdown JUNB significantly reduced IL-4 expression. Our results suggest that the prolonged IL-4 expression in NFAT1 deficient Th2 cells is mediated by preferential binding of JUNB/SATB1 to the IL-4 promoter with permissive chromatin architecture.

Transcription factor NFAT1 controls allergic contact hypersensitivity through regulation of activation induced cell death program

Allergic contact hypersensitivity (CHS) is an inflammatory skin disease mediated by allergen specific T cells. In this study, we investigated the role of transcription factor NFAT1 in the pathogenesis of contact hypersensitivity. NFAT1 knock out (KO) mice spontaneously developed CHS-like skin inflammation in old age. Healthy young NFAT1 KO mice displayed enhanced susceptibility to hapten-induced CHS. Both CD4+ and CD8+ T cells from NFAT1 KO mice displayed hyper-activated properties and produced significantly enhanced levels of inflammatory T helper 1(Th1)/Th17 type cytokines. NFAT1 KO T cells were more resistant to activation induced cell death (AICD), and regulatory T cells derived from these mice showed a partial defect in their suppressor activity. NFAT1 KO T cells displayed a reduced expression of apoptosis associated BCL-2/BH3 family members. Ectopic expression of NFAT1 restored the AICD defect in NFAT1 KO T cells and increased AICD in normal T cells. Recipient Rag2−/− mice transferred with NFAT1 KO T cells showed more severe CHS sensitivity due to a defect in activation induced hapten-reactive T cell apoptosis. Collectively, our results suggest the NFAT1 plays a pivotal role as a genetic switch in CD4+/CD8+ T cell tolerance by regulating AICD process in the T cell mediated skin inflammation.

NFAT1 Regulates Systemic Autoimmunity through the Modulation of a Dendritic Cell Property

The transcription factor NFAT1 plays a pivotal role in the homeostasis of T lymphocytes. However, its functional importance in non-CD4+ T cells, especially in systemic immune disorders, is largely unknown. In this study, we report that NFAT1 regulates dendritic cell (DC) tolerance and suppresses systemic autoimmunity using the experimental autoimmune myasthenia gravis (EAMG) as a model. Myasthenia gravis and EAMG are T cell–dependent, Ab-mediated autoimmune disorders in which the acetylcholine receptor is the major autoantigen. NFAT1-knockout mice showed higher susceptibility to EAMG development with enhanced Th1/Th17 cell responses. NFAT1 deficiency led to a phenotypic alteration of DCs that show hyperactivation of NF-κB–mediated signaling pathways and enhanced binding of NF-κB (p50) to the promoters of IL-6 and IL-12. As a result, NFAT1-knockout DCs produced much higher levels of proinflammatory cytokines such as IL-1β, IL-6, IL-12, and TNF-α, which preferentially induce Th1/Th17 cell differentiation. Our data suggest that NFAT1 may limit the hyperactivation of the NF-κB–mediated proinflammatory response in DCs and suppress autoimmunity by serving as a key regulator of DC tolerance.