Ovalbumin-Derived Peptides Activate Retinoic Acid Signalling Pathways and Induce Regulatory Responses Through Toll-Like Receptor Interactions.
ABSTRACT: This study investigates the potential of a hydrolysate of ovalbumin with pepsin (OP) to preclude Th2-type immunity by the enhancement of tolerogenic dendritic cells (DCs) and regulatory T (Treg) cells. Through Toll-like receptor (TLR) stimulation, OP enhances the retinoic acid pathway on DCs by means of the induction of aldehyde dehydrogenase enzymes and transforming growth factor beta (TGF-?), and it confers upon DC the ability to upregulate interleukin 10 (IL-10) as well as other tolerance-promoting mediators downstream of TRL signalling, such as IL-27, IL-33, Notch ligands, OX40L, and the transcription factors IRF4 and IRF8. OP-conditioned DCs induce the expansion of Foxp3+ and Tr1 cells in co-culture with CD4+ T cells. Furthermore, OP directly conditions CD4+ T cells from naïve mice, without the mediation of DCs, to express aldehyde dehydrogenase (ALDH) enzymes and, in the presence of the Th2 cytokine IL-4 and exogenous TGF-?, it enhances Foxp3 expression. It is noteworthy that, on CD4+ T cells isolated from egg-allergic mice, OP significantly enriches the levels of Foxp3+ and Foxp3+ ROR?t+ CD4+ T cells. In conclusion, we show that food peptides may work, analogously to microbial-driven signals, through TLRs, to promote a tolerogenic phenotype on cells of the innate and adaptive immune system, a property that is further enhanced in the context of a Th2 cytokine-rich environment.
Project description:Interplay between Foxp3(+) regulatory T cells (Treg) and dendritic cells (DCs) maintains immunologic tolerance, but the effects of each cell on the other are not well understood. We report that polyclonal CD4(+)Foxp3(+) Treg cells induced ex vivo with transforming growth factor beta (TGF?) (iTreg) suppress a lupus-like chronic graft-versus-host disease by preventing the expansion of immunogenic DCs and inducing protective DCs that generate additional recipient CD4(+)Foxp3(+) cells. The protective effects of the transferred iTreg cells required both interleukin (IL)-10 and TGF?, but the tolerogenic effects of the iTreg on DCs, and the immunosuppressive effects of these DCs were exclusively TGF?-dependent. The iTreg were unable to tolerize Tgfbr2-deficient DCs. These results support the essential role of DCs in 'infectious tolerance' and emphasize the central role of TGF? in protective iTreg/DC interactions in vivo.
Project description: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, cisplatin-treated DCs showed markedly increased IL-10 production through activation of the p38 MAPK and NF-?B signaling pathways without altering the levels of TNF-? 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 agonist-induced 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.
Project description:Tolerogenic dendritic cells (tolDCs) are central players in the maintenance of immune tolerance and thereby have been identified as the most favourable candidates for cell therapy of autoimmune diseases. We have recently shown that excretory-secretory products (ES L1) released by Trichinella spiralis larvae induce stable human tolDCs in vitro via Toll-like receptor 2 (TLR2) and TLR4. However, engagement of these receptors did not fully explain the tolerogenic profile of DCs. Here, we observed for the first time that dendritic cell-specific ICAM-3 grabbing non-integrin (DC-SIGN) interacts with highly glycosylated ES L1 and contributes to the generation of ES L1-induced tolDCs. Blocking DC-SIGN interfered with the ES L1-induced higher expression of CD40 and CCR7 and the production of IL-10 and TGF-? by DCs. The cooperation of TLR2, TLR4 and DC-SIGN receptors is of importance for the capacity of DCs to prime T cell response toward Th2 and to induce expansion of CD4+CD25+Foxp3+ T cells, as well as for the production of IL-10 and TGF-? by these cells. Overall, these results indicate that induction of tolDCs by ES L1 involves engagement of multiple pattern recognition receptors namely, TLR2, TLR4 and DC-SIGN.
Project description:Earlier, we have shown that GM-CSF-exposed CD8α- DCs that express low levels of pro-inflammatory cytokines IL-12 and IL-1β can induce Foxp3+ Tregs leading to suppression of autoimmunity. Here, we examined the differential effects of IL-12 and IL-1β on Foxp3 expression in T cells when activated in the presence and absence of DCs. Exogenous IL-12 abolished, but IL-1β enhanced, the ability of GM-CSF-exposed tolerogenic DCs to promote Foxp3 expression. Pre-exposure of DCs to IL-1β and IL-12 had only a modest effect on Foxp3- expressing T cells; however, T cells activated in the absence of DCs but in the presence of IL-1β or IL-12 showed highly significant increase and decrease in Foxp3+ T cell frequencies respectively suggesting direct effects of these cytokines on T cells and a role for IL-1β in promoting Foxp3 expression. Importantly, purified CD4+CD25+ cells showed a significantly higher ability to maintain Foxp3 expression when activated in the presence of IL-1β. Further analyses showed that the ability of IL-1β to maintain Foxp3 expression in CD25+ T cells was dependent on TGF-β1 and IL-2 expression in Foxp3+Tregs and CD25- effectors T cells respectively. Exposure of CD4+CD25+ T cells to IL-1β enhanced their ability to suppress effector T cell response in vitro and ongoing experimental autoimmune thyroidits in vivo. These results show that IL-1β can help enhance/maintain Tregs, which may play an important role in maintaining peripheral tolerance during inflammation to prevent and/or suppress autoimmunity.
Project description:Trichinella spiralis, as well as its muscle larvae excretory-secretory products (ES L1), given either alone or via dendritic cells (DCs), induce a tolerogenic immune microenvironment in inbred rodents and successfully ameliorate experimental autoimmune encephalomyelitis. ES L1 directs the immunological balance away from T helper (Th)1, toward Th2 and regulatory responses by modulating DCs phenotype. The ultimate goal of our work is to find out if it is possible to translate knowledge obtained in animal model to humans and to generate human tolerogenic DCs suitable for therapy of autoimmune diseases through stimulation with ES L1. Here, the impact of ES L1 on the activation of human monocyte-derived DCs is explored for the first time. Under the influence of ES L1, DCs acquired tolerogenic (semi-matured) phenotype, characterized by low expression of HLA-DR, CD83, and CD86 as well as moderate expression of CD40, along with the unchanged production of interleukin (IL)-12 and elevated production of IL-10 and transforming growth factor (TGF)-?, compared to controls. The interaction with DCs involved toll-like receptors (TLR) 2 and 4, and this interaction was mainly responsible for the phenotypic and functional properties of ES L1-treated DCs. Importantly, ES L1 potentiated Th2 polarizing capacity of DCs, and impaired their allo-stimulatory and Th1/Th17 polarizing properties. Moreover, ES L1-treated DCs promoted the expansion of IL-10- and TGF-?- producing CD4+CD25hiFoxp3hi T cells in indolamine 2, 3 dioxygenase (IDO)-1-dependent manner and increased the suppressive potential of the primed T cell population. ES L1-treated DCs retained the tolerogenic properties, even after the challenge with different pro-inflammatory stimuli, including those acting via TLR3 and, especially TLR4. These results suggest that the induction of tolerogenic properties of DCs through stimulation with ES L1 could represent an innovative approach for the preparation of tolerogenic DC for treatment of inflammatory and autoimmune disorders.
Project description:Immature or semi-mature dendritic cells (DCs) represent tolerogenic maturation stages that can convert naive T cells into Foxp3+ induced regulatory T cells (iTreg). Here we found that murine bone marrow-derived DCs (BM-DCs) treated with cholera toxin (CT) matured by up-regulating MHC-II and costimulatory molecules using either high or low doses of CT (CThi, CTlo) or with cAMP, a known mediator CT signals. However, all three conditions also induced mRNA of both isoforms of the tolerogenic molecule cytotoxic T lymphocyte antigen 2 (CTLA-2α and CTLA-2β). Only DCs matured under CThi conditions secreted IL-1β, IL-6 and IL-23 leading to the instruction of Th17 cell polarization. In contrast, CTlo- or cAMP-DCs resembled semi-mature DCs and enhanced TGF-β-dependent Foxp3+ iTreg conversion. iTreg conversion could be reduced using siRNA blocking of CTLA-2 and reversely, addition of recombinant CTLA-2α increased iTreg conversion in vitro. Injection of CTlo- or cAMP-DCs exerted MOG peptide-specific protective effects in experimental autoimmune encephalomyelitis (EAE) by inducing Foxp3+ Tregs and reducing Th17 responses. Together, we identified CTLA-2 production by DCs as a novel tolerogenic mediator of TGF-β-mediated iTreg induction in vitro and in vivo. The CT-induced and cAMP-mediated up-regulation of CTLA-2 also may point to a novel immune evasion mechanism of Vibrio cholerae.
Project description:Asthma is a common chronic inflammatory disease, which is characterized by airway hyperresponsiveness (AHR), high serum levels of immunoglobulin (Ig)E, and recruitment of various inflammatory cells such as eosinophils and lymphocytes. Korean traditional fermented foods have been reported to exert beneficial effects against allergic diseases such as asthma and atopic dermatitis. In this study, we investigated whether Staphylococcus succinus strain 14BME20 (14BME20) isolated from doenjang, a traditional high-salt-fermented soybean food of Korea, exerts suppressive effects on allergic airway inflammation in a murine model. Mice were orally administered with 14BME20, then sensitized and challenged with ovalbumin as an allergen. Administration of the 14BME20 significantly suppressed AHR and influx of inflammatory cells into the lungs and reduced serum IgE levels. Moreover, the proportion of T helper type 2 (Th2) cells and the production of Th2 cytokines were decreased in 14BME20-treated mice, whereas dendritic cells (DCs) with tolerogenic characteristics were increased. In contrast, oral administration of 14BME20 increased the proportion of CD4+CD25+Foxp3+ regulatory T (Treg) cells and the level of interleukin (IL)-10 in 14BME20-treated mice. Furthermore, 14BME20 induced maturation of tolerogenic DCs, and 14BME20-treated DCs increased Treg cell population in a co-culture system of DCs and CD4+ T cells. The addition of a neutralizing anti-IL-10 mAb to the culture of cells that had been treated with 14BME20 decreased the enhanced Treg cell population, thereby indicating that 14BME20-treated DCs increase Treg cell population via DC-derived IL-10. These results demonstrate that oral administration of 14BME20 suppresses airway inflammation by enhancing Treg responses and suggest that the 14BME20 isolated from doenjang may be a therapeutic agent for allergic asthma.
Project description:Protosappanin A (PrA), an immunosuppressive ingredient of the medicinal herb Caesalpinia sappan L, prolongs heart allograft survival in rats, possibly by impairing the function of antigen-presenting cells (APCs). We examined the effects of PrA on the maturation and function of dendritic cells (DCs), a potent class of APCs, and the downstream cell-cell and intracellular signaling pathways mediating the immunosuppressive activity of PrA. PrA inhibited LPS-stimulated maturation of Wistar rat DCs in vitro as reflected by reduced expression of costimulatory molecules (CD80 and CD86) and reduced expression of TLR4 and NF-κB, two critical signaling components for antigen recognition. PrA also enhanced the release of IL-10 and decreased the release of IL-12 from DCs, but had no effect on the production of TGF-ß. In mixed cultures, Wistar DCs pretreated with PrA impaired the proliferation of Sprague Dawley (SD) rat T cells while promoting the expansion of SD rat CD4(+)CD25(+) regulatory T cells (Tregs). Both oral PrA treatment and infusion of PrA-pretreated Wistar DCs prolonged cardiac allograft survival (Wistar donor, SD recipient) and expanded recipient CD4(+)CD25(+)Foxp3(+) Tregs. Donor spleen cells, but not spleen cells from a third rat strain (DA), supported the expansion of recipient CD4(+)CD25(+)Foxp3(+) Tregs and suppressed recipient T cell proliferation. We conclude that PrA triggers a tolerogenic state in DCs that allows for the induction of alloantigen-specific Tregs and the suppression of allograft rejection in vivo.
Project description:<h4>Background</h4>Asthma is a chronic inflammatory disorder of the airways with the proven role of Th2 cells in its pathogenesis. The role and characteristic of different subsets of CD4(+) cells is much less known.<h4>Aim</h4>The aim of the study was to analyze the incidence of different subsets of CD4(+) T cells, in particular different subsets of CD4(+) cells with the co-expression of different cytokines.<h4>Methods</h4>Twenty five stable asthmatic and twelve age-matched control subjects were recruited to the study. Bronchoscopy and bronchoalveolar lavage (BAL) were performed in all study subjects. CD4(+) T cells were isolated from BAL fluid by positive magnetic selection. After stimulation simultaneous expression of TGF-β, FoxP3, CD25, IFN-γ, IL-4, TNF-α (set 1); IL-10, FoxP3, CD25, IFN-γ, IL-4, MIP-1β (set 2); IL-17A, IL-8, IFN-γ, IL-4, MIP-1β (set 3) were measured by flow cytometry.<h4>Results</h4>The percentage of CD4(+) cells co-expressing Foxp3 and TGF-β (CD4(+)Foxp3(+)TGF-β(+) cells) was significantly lower (P = 0.03), whereas the percentage of CD4(+)IL-17(+) cells (P = 0.008), CD4(+)IL-17(+) IFN-γ(+) cells (P = 0.047) and CD4(+)IL-4(+) cells (P = 0.01) were significantly increased in asthmatics compared with that seen in healthy subjects. A significantly higher percentage of CD4(+)Foxp3(+) cells from asthma patients expressed IFN-γ (P = 0.01), IL-4 (P = 0.004) and CD25 (P = 0.04), whereas the percentage of CD4(+)IL-10(+) cells expressing Foxp3 was significantly decreased in asthmatics (P = 0.03). FEV1% predicted correlated negatively with the percentage of CD4(+)IL-17(+) cells (r = -0.33; P = 0.046) and positively with CD4(+)Foxp3(+)TGF-β(+) cells (r = 0.43; P = 0.01).<h4>Conclusions</h4>Our results suggest that in the airways of chronic asthma patients there is an imbalance between increased numbers of CD4(+)IL-17(+) cells and Th2 cells and decreased number of CD4(+)Foxp3(+)TGF-β(+).
Project description:Naturally occurring, thymic-derived Foxp3+CD25+CD4+ regulatory T cells (nTregs) are pivotal for the maintenance of self-tolerance. nTregs, however, are sparse and lack alloantigen specificity, and these properties pose challenges for their use in clinical transplantation.We established mixed leukocyte reaction (MLR) with dendritic cells (DCs) as stimulators and CD4+ T cells as responders and supplemented the MLR with IL-2 and TGF-?1 and investigated whether DCs+IL-2+TGF-?1 differentiate the polyclonal CD4+ cells into alloantigen-specific and allograft protective Tregs.We found a greater than a 10-fold increase in Foxp3+CD25+ subpopulation (P<0.01) following stimulation of BALB/c CD4+ cells with C57BL/6 (B6) CD11c+ DCs+IL-2+TGF-?1 in the MLR. Levels of TGF-?1 messenger RNA (mRNA) (P=0.01) and the ratios of TGF-?1 mRNA to granzyme B mRNA (P=0.0003) and Foxp3 mRNA to granzyme B mRNA (P<0.01) were higher in alloantigen-induced Tregs (alloTregs) compared with nTregs. alloTregs suppressed MLR at a 16:1 responder to suppressor ratio, whereas nTregs suppressed at 4:1. Suppression by alloTregs was alloantigen specific and was observed at the level of responder cells and at the level of stimulator cells. In a fully H-2-mismatched, nonlymphopenic, immunocompetent mouse islet transplantation model, alloTregs but not nTregs prolonged survival of islet allografts without any other immunosuppressive therapy (P=0.0003), and the protection was alloantigen specific.A combination of CD11c+ DCs, IL-2, and TGF-?1 may help differentiate naive, high abundant CD4+ T into alloantigen-specific and allograft protective Foxp3+Tregs.