Inflammation-induced Id2 promotes plasticity in regulatory T cells

T(H)17 cells originating from regulatory T (T-reg) cells upon loss of the T-reg-specific transcription factor Foxp3 accumulate in sites of inflammation and aggravate autoimmune diseases. Whether an active mechanism drives the generation of these pathogenic 'ex-Foxp3 T(H)17' cells, remains unclear. Here we show that pro-inflammatory cytokines enhance the expression of transcription regulator Id2, which mediates cellular plasticity of T-reg into 'ex-Foxp3' T(H)17 cells. Expression of Id2 in in vitro differentiated iT(reg) cells reduces the expression of Foxp3 by sequestration of the transcription activator E2A, leading to the induction of T(H)17-related cytokines. T-reg-specific ectopic expression of Id2 in mice significantly reduces the T-reg compartment and causes immune dysregulation. Cellular fate-mapping experiments reveal enhanced T-reg plasticity compared to wild-type, resulting in exacerbated experimental autoimmune encephalomyelitis pathogenesis or enhanced anti-tumor immunity. Our findings suggest that controlling Id2 expression may provide a novel approach for effective T-reg cell immunotherapies for both autoimmunity and cancer.11sciescopu

Immunomodulatory effect of water soluble extract separated from mycelium of Phellinus linteus on experimental atopic dermatitis

Background: Complementary and alternative medicine (CAM) is becoming a popular treatment for modulating diverse immune disorders. Phellinus linteus (P. linteus) as one of the CAMs has been used to modulate cancers, inflammation and allergic activities. However, little evidence has been shown about its underlying mechanism of action by which it exerts a beneficial role in dermatological disease in vivo. In this study, we examined the immunomodulatory effects of P. linteus on experimental atopic dermatitis (AD) and elucidated its action mechanism. Methods: The immunomodulatory effect of total extract of P. linteus on IgE production by human myeloma U266B1 cells was measured by ELISA. To further identify the effective components, P. linteus was fractionated into methanol soluble, water soluble and boiling water soluble extracts. Each extract was treated to U266B1 cells and primary B cells to compare their inhibitory effects on IgE secretion. To test the in vivo efficacy, experimental atopic dermatitis (AD) was established by alternative treatment of DNCB and house dust mite extract into BALB/c mice. Water soluble extract of P. linteus (WA) or ceramide as a positive control were topically applied to ears of atopic mouse every day for 2 weeks and progression of the disease was estimated by the following criteria: (a) ear thickness, clinical score, (b) serum total IgE, IgG and mite specific IgE level by ELSIA, (c) histological examination of ear tissue by H&E staining and (d) cytokine profile of total ear cells and CD4+ T cells by real time PCR and ELSIA. Results: Treatment of total extracts of P. linteus to U266B1 inhibited IgE secretion. Among the diverse extracts of P. linteus, water soluble extract of P. linteus (WA) significantly reduced the IgE production in primary B cells and B cell line U266B1. Moreover, treatment of WA reduced AD symptoms such as ear swelling, erythema, and dryness and decreased recruitment of lymphocyte into the inflamed site. Interestingly WA treatment significantly reduced IgE level without affecting IgG levels and also down-regulated the levels of pathogenic cytokines (IL-4, IL-13, IL-12 and IFN-γ) and chemokines (CCL17 and CCL22) involved in AD development. Conclusions: Our study indicates that protective effect of water soluble extract of P. linteus in atopic dermatitis is mediated by inhibiting IgE production and expression of AD associated pathogenic cytokines as well as chemokines, suggesting the beneficial effect of P. linteus to modulate allergic skin disease

IRT5 Probiotics Changes Immune Modulatory Protein Expression in the Extraorbital Lacrimal Glands of an Autoimmune Dry Eye Mouse Model

PURPOSE. While the association between the gut microbiome and the immune system has been studied in autoimmune disorders, little is known about ocular disease. Previously we reported that IRT5, a mixture of five probiotic strains, could suppress autoimmune dry eye. In this study, we investigated the mechanism by which IRT5 performs its immunomodulatory function in a mouse model of autoimmune dry eye. METHODS. NOD.B10.H2b mice were used as an autoimmune dry eye model. Either IRT5 or PBS was gavaged orally for 3 weeks, with or without 5 days of antibiotic pretreatment. The effects on clinical features, extraorbital lacrimal gland and spleen proteins, and fecal microbiota were analyzed. RESULTS. The ocular staining score was lower, and tear secretion was higher, in the IRT5-treated groups than in the PBS-treated groups. After IRT5 treatment, the downregulated lacrimal gland proteins were enriched in the biological processes of defense response and immune system process. The relative abundances of 33 operational taxonomic units were higher, and 53 were lower, in the feces of the IRT5-treated groups than in those of the PBS-treated groups. IRT5 administration without antibiotic pretreatment also showed immunomodulatory functions with increases in the Lactobacillus helveticus group and Lactobacillus hamsteri. Additional proteomic assays revealed a decrease of proteins related to antigen-presenting processes in the CD11b(+) and CD11c(+) cells of spleen in the IRT5-treated groups. CONCLUSIONS. Changes in the gut microbiome after IRT5 treatment improved clinical manifestations in the autoimmune dry eye model via the downregulation of antigen-presenting processes in immune networks.11Ysciescopu

Structural features and immunological perception of the cell surface glycans of Lactobacillus plantarum: a novel rhamnose-rich polysaccharide and teichoic acids

Abstract The capsular material from Lactobacillus plantarum IMB19, an isolate from fermented vegetables, has been analyzed and our results demonstrate that most of the coat of this bacterium consists of glycerol- and ribitol-type teichoic acids, further decorated with other substituents (α-glucose and alanine), and of a capsular polysaccharide (CPS) with a linear nonasaccharide repeating unit, rich in rhamnose, interconnected to the next via a phosphodiester bridge. Stimulation of immune cells with the total capsular material resulted in the enhancement of immunostimulatory (IFNγ, TNF-α, IL-6 and IL-12) or immuno-regulatory (IL-10) cytokines in an in vitro splenocyte culture system. The capsular polysaccharide, and not the teichoic acids mixture, was responsible for the IFNγ production. Indeed, a significant increase of IFNγ along with other inflammatory cytokines, and a dose response in IFNγ expression with an EC50 of 3.16 μM was found for CPS, disclosing that this polysaccharide is a potent immunostimulatory molecule

Cisplatin induces tolerogenic dendritic cells in response to TLR agonists via the abundant production of IL-10, therby promoting Th2- and Tr1-biased T-cell immunity

Although many advantageous roles of cisplatin (cis-diamminedichloroplatinum (II), CDDP) have been reported in cancer therapy, the immunomodulatory roles of cisplatin in the phenotypic and functional alterations of dendritic cells (DCs) are poorly understood. Here, we investigated the effect of cisplatin on the functionality of DCs and the changes in signaling pathways activated upon toll-like receptor (TLR) stimulation. Cisplatin-treated DCs down-regulated the expression of cell surface molecules (CD80, CD86, MHC class I and II) and up-regulated endocytic capacity in a dose-dependent manner. Upon stimulation with various TLR agonists, cisplatintreated DCs showed markedly increased IL-10 production through activation of the p38 MAPK and NF-κB signaling pathways without altering the levels of TNF-α and IL- 12p70, indicating the cisplatin-mediated induction of tolerogenic DCs. This effect was dependent on the production of IL-10 from DCs, as neither DCs isolated from IL-10- /- mice nor IL-10-neutralized DCs generated tolerogenic DCs. Interestingly, DCs that were co-treated with cisplatin and lipopolysaccharide (LPS) exhibited a decreased immunostimulatory capacity for inducing the proliferation of Th1- and Th17-type T cells; instead, these DCs contributed to Th2-type T cell immunity. Furthermore, in vitro and in vivo investigations revealed a unique T cell population, IL-10-producing CD3+CD4+LAG-3+CD49b+CD25-Foxp3- Tr1 cells, that was significantly increased without altering the Foxp3+ regulatory T cell population. Taken together, our results suggest that cisplatin induces immune-suppressive tolerogenic DCs in TLR agonistinduced inflammatory conditions via abundant IL-10 production, thereby skewing Th cell differentiation towards Th2 and Tr1 cells. This relationship may provide cancer cells with an opportunity to evade the immune system.1231sciescopu

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

Abalone visceral extract inhibit tumor growth and metastasis by modulating Cox-2 levels and CD8+ T cell activity

<p>Abstract</p> <p>Background</p> <p>Abalone has long been used as a valuable food source in East Asian countries. Although the nutritional importance of abalone has been reported through <it>in vitro </it>and <it>in vivo </it>studies, there is little evidence about the potential anti-tumor effects of abalone visceral extract. The aim of the present study is to examine anti-tumor efficacy of abalone visceral extract and to elucidate its working mechanism.</p> <p>Methods</p> <p>In the present study, we used breast cancer model using BALB/c mouse-derived 4T1 mammary carcinoma and investigated the effect of abalone visceral extract on tumor development. Inhibitory effect against tumor metastasis was assessed by histopathology of lungs. Cox-2 productions by primary and secondary tumor were measured by real-time RT-PCR and immunoblotting (IB). Proliferation assay based on [³H]-thymidine incorporation and measurement of cytokines and effector molecules by RT-PCR were used to confirm tumor suppression efficacy of abalone visceral extract by modulating cytolytic CD8+ T cells. The cytotoxicity of CD8⁺T cell was compared by JAM test.</p> <p>Results</p> <p>Oral administration of abalone visceral extract reduced tumor growth (tumor volume and weight) and showed reduced metastasis as confirmed by decreased level of splenomegaly (spleen size and weight) and histological analysis of the lung metastasis (gross analysis and histological staining). Reduced expression of Cox-2 (mRNA and protein) from primary tumor and metastasized lung was also detected. In addition, treatment of abalone visceral extract increased anti-tumor activities of CD8⁺T cells by increasing the proliferation capacity and their cytolytic activity.</p> <p>Conclusions</p> <p>Our results suggest that abalone visceral extract has anti-tumor effects by suppressing tumor growth and lung metastasis through decreasing Cox-2 expression level as well as promoting proliferation and cytolytic function of CD8⁺T cells.</p

The Kinase PDK1 Is Essential for B-Cell Receptor Mediated Survival Signaling

Phosphoinositide-dependent kinase 1 (PDK1) plays an important role in integrating the T cell antigen receptor (TCR) and CD28 signals to achieve efficient NF-κB activation. PDK1 is also an important regulator of T cell development, mediating pre-TCR induced proliferation signals. However, the role of PDK1 in B cell antigen receptor (BCR) signaling and B cell development remains largely unknown. In this study we provide genetic evidence supporting the role of PDK1 in B cell survival. We found PDK1 is required for BCR mediated survival in resting B cells, likely through regulation of Foxo activation. PDK1-dependent signaling to NF-κB is not crucial to resting B cell viability. However, PDK1 is necessary for triggering NF-κB during B cell activation and is required for activated B cell survival. Together these studies demonstrate that PDK1 is essential for BCR-induced signal transduction to Foxo and NF-κB and is indispensable for both resting and activated B cell survival

Dietary Glucose Consumption Promotes RALDH Activity in Small Intestinal CD103(+)CD11b(+) Dendritic Cells

Retinal dehydrogenase (RALDH) enzymatic activities catalyze the conversion of vitamin A to its metabolite Retinoic acid (RA) in intestinal dendritic cells (DCs) and promote immunological tolerance. However, precise understanding of the exogenous factors that act as initial trigger of RALDH activity in these cells is still evolving. By using germ-free (GF) mice raised on an antigen free (AF) elemental diet, we find that certain components in diet are critically required to establish optimal RALDH expression and activity, most prominently in small intestinal CD103(+)CD11b(+) DCs (siLP-DCs) right from the beginning of their lives. Surprisingly, systematic screens using modified diets devoid of individual dietary components indicate that proteins, starch and minerals are dispensable for this activity. On the other hand, in depth comparison between subtle differences in dietary composition among different dietary regimes reveal that adequate glucose concentration in diet is a critical determinant for establishing RALDH activity specifically in siLP-DCs. Consequently, pre-treatment of siLP-DCs, and not mesenteric lymph node derived MLNDCs with glucose, results in significant enhancement in the in vitro generation of induced Regulatory T (iTreg) cells. Our findings reveal previously underappreciated role of dietary glucose concentration in establishing regulatory properties in intestinal DCs, thereby extending a potential therapeutic module against intestinal inflammation11Ysciescopu

Upregulation of Ets1 expression by NFATc2 and NFKB1/RELA promotes breast cancer cell invasiveness

Breast cancer is highly aggressive and is the leading cause of cancer-related mortality in women in developed countries. The ETS proto-oncogene 1 (Ets1) has versatile roles during the cellular processes of cancer development. It is often highly expressed in breast cancers and mediates migration and invasion of human breast cancer cells. However, underlying mechanisms of Ets1 gene expression is still ambiguous. Here, we identified a core-regulatory element (CRE) located in the Ets1 promoter region (−540/−80 bp from TSS) that contains elements responsible for associating with NFATs and NF-κBs. Compared with the less metastatic breast cancer cells, metastatic breast cancer cells (MDA-MB-231) show open chromatin configurations in the CRE, which facilitates direct binding of NFATc2 and/or NFKB1/RELA complex to trans-activate Ets1 transcription. Moreover, enhanced level of Nfatc2 and Nfkb1 positively correlated with Ets1 expression in the human breast cancer specimens. Deletion of the CRE region by CRISPR/Cas9 system resulted in significant reduction in Ets1 expression, which led to alterations of Ets1-mediated transcription programs including tumor invasiveness-related genes. Proper regulation of Ets1 gene expression by targeting the NFATc2 and NFKB1/RELA interaction could be a potential therapeutic target for Ets1-mediated metastatic breast cancer.11sciescopu