Helicobacter pylori and its secreted immunomodulator VacA protect against anaphylaxis in experimental models of food allergy.

BACKGROUND: Food allergy is an increasingly common health problem in Western populations. Epidemiological studies have suggested both positive and negative associations between food allergy and infection with the gastric bacterium Helicobacter pylori. OBJECTIVE: The objective of this work was to investigate whether experimental infection with H. pylori, or prophylactic treatment with H. pylori-derived immunomodulatory molecules, affects the onset and severity of food allergy, either positively or negatively. METHODS: We infected neonatal C57BL/6 or C3H mice with H. pylori or treated animals with H. pylori components (bacterial lysate or the immunomodulator VacA) and subsequently subjected them to four different protocols for food allergy induction, using either ovalbumin or peanut extract as allergens for sensitization and challenge. Readouts included anaphylaxis scoring, quantification of allergen-specific serum IgE and IgG1 and of the mast cell protease MCPT1, as well as splenic T-helper-2 cell-derived cytokine production. Mesenteric lymph node CD4+ FoxP3+ regulatory T cells were subjected to flow cytometric quantification and sorting followed by qRT-PCR, and to DNA methylation analyses of the Treg-specific demethylated region (TSDR) within the FOXP3 locus. RESULTS: Mice that had been infected with H. pylori or treated with H. pylori-derived immunomodulators showed reduced anaphylaxis upon allergen sensitization and challenge, irrespective of the allergen used. Most of the immunologic assays confirmed a protective effect of H. pylori. CD4+ FoxP3+ T cells were more abundant in protected mice and exhibited a stable Treg phenotype characterized by FOXP3 TSDR demethylation. CONCLUSIONS AND CLINICAL RELEVANCE: Helicobacter pylori confers protection against the anaphylaxis associated with ovalbumin and peanut allergy and affects the epigenome of T cells, thereby promoting stable Treg differentiation and functionality. Prophylactic treatment with H. pylori-derived immunomodulators appears to be a promising strategy for food allergy prevention.

Gut memories do not fade: epigenetic regulation of lasting gut homing receptor expression in CD4+ memory T cells.

The concept of a "topographical memory" in lymphocytes implies a stable expression of homing receptors mediating trafficking of lymphocytes back to the tissue of initial activation. However, a significant plasticity of the gut-homing receptor α4ß7 was found in CD8+ T cells, questioning the concept. We now demonstrate that α4ß7 expression in murine CD4+ memory T cells is, in contrast, imprinted and remains stable in the absence of the inducing factor retinoic acid (RA) or other stimuli from mucosal environments. Repetitive rounds of RA treatment enhanced the stability of de novo induced α4ß7. A novel enhancer element in the murine Itga4 locus was identified that showed, correlating to stability, selective DNA demethylation in mucosa-seeking memory cells and methylation-dependent transcriptional activity in a reporter gene assay. This implies that epigenetic mechanisms contribute to the stabilization of α4ß7 expression. Analogous DNA methylation patterns could be observed in the human ITGA4 locus, suggesting that its epigenetic regulation is conserved between mice and men. These data prove that mucosa-specific homing mediated by α4ß7 is imprinted in CD4+ memory T cells, reinstating the validity of the concept of "topographical memory" for mucosal tissues, and imply a critical role of epigenetic mechanisms.

Prevention of Allograft Rejection by Use of Regulatory T Cells With an MHC-Specific Chimeric Antigen Receptor.

CD4+ CD25high FOXP3+ regulatory T cells (Tregs) are involved in graft-specific tolerance after solid organ transplantation. However, adoptive transfer of polyspecific Tregs alone is insufficient to prevent graft rejection even in rodent models, indicating that graft-specific Tregs are required. We developed a highly specific chimeric antigen receptor that recognizes the HLA molecule A*02 (referred to as A2-CAR). Transduction into natural regulatory T cells (nTregs) changes the specificity of the nTregs without alteration of their regulatory phenotype and epigenetic stability. Activation of nTregs via the A2-CAR induced proliferation and enhanced the suppressor function of modified nTregs. Compared with nTregs, A2-CAR Tregs exhibited superior control of strong allospecific immune responses in vitro and in humanized mouse models. A2-CAR Tregs completely prevented rejection of allogeneic target cells and tissues in immune reconstituted humanized mice in the absence of any immunosuppression. Therefore, these modified cells have great potential for incorporation into clinical trials of Treg-supported weaning after allogeneic transplantation.

Foxp3(+) T cells expressing RORγt represent a stable regulatory T-cell effector lineage with enhanced suppressive capacity during intestinal inflammation.

Foxp3 (forkhead box P3 transcription factor)-expressing regulatory T cells (Tregs) are essential for immunological tolerance, best illustrated by uncontrolled effector T-cell responses and autoimmunity upon loss of Foxp3 expression. Tregs can adopt specific effector phenotypes upon activation, reflecting the diversity of functional demands in the different tissues of the body. Here, we report that Foxp3(+)CD4(+) T cells coexpressing retinoic acid-related orphan receptor-γt (RORγt), the master transcription factor for T helper type 17 (Th17) cells, represent a stable effector Treg lineage. Transcriptomic and epigenetic profiling revealed that Foxp3(+)RORγt(+) T cells display signatures of both Tregs and Th17 cells, although the degree of similarity was higher to Foxp3(+)RORγt(-) Tregs than to Foxp3(-)RORγt(+) T cells. Importantly, Foxp3(+)RORγt(+) T cells were significantly demethylated at Treg-specific epigenetic signature genes such as Foxp3, Ctla-4, Gitr, Eos, and Helios, suggesting that these cells have a stable regulatory rather than inflammatory function. Indeed, adoptive transfer of Foxp3(+)RORγt(+) T cells in the T-cell transfer colitis model confirmed their Treg function and lineage stability in vivo, and revealed an enhanced suppressive capacity as compared with Foxp3(+)RORγt(-) Tregs. Thus, our data suggest that RORγt expression in Tregs contributes to an optimal suppressive capacity during gut-specific immune responses, rendering Foxp3(+)RORγt(+) T cells as an important effector Treg subset in the intestinal system.

CD11b(+)Ly6C(++)Ly6G(-) Cells with Suppressive Activity Towards T Cells Accumulate in Lungs of Influenza a Virus-Infected Mice.

Influenza A virus (IAV) infection causes an acute respiratory disease characterized by a strong inflammatory immune response and severe immunopathology. Proinflammatory mechanisms are well described in the murine IAV infection model, but less is known about the mechanisms leading to the resolution of inflammation. Here, we analyzed the contribution of CD11b(+)Ly6C(++)Ly6G(-) cells to this process. An accumulation of CD11b(+)Ly6C(++)Ly6G(-) cells within the lungs was observed during the course of IAV infection. Phenotypic characterization of these CD11b(+)Ly6C(++)Ly6G(-) cells by flow cytometry and RNA-Seq revealed an activated phenotype showing both pro- and anti-inflammatory features, including the expression of inducible nitric oxide synthase (iNOS) by a fraction of cells in an IFN-γ-dependent manner. Moreover, CD11b(+)Ly6C(++)Ly6G(-) cells isolated from lungs of IAV-infected animals displayed suppressive activity when tested in vitro, and iNOS inhibitors could abrogate this suppressive activity. Collectively, our data suggest that during IAV infection, CD11b(+)Ly6C(++)Ly6G(-) cells acquire immunoregulatory function, which might contribute to the prevention of pathology during this life-threatening disease.

Foxp3(+) regulatory T cells are expanded in severe atopic dermatitis patients.

Regulatory T cells (Tregs) are known to play critical roles in homeostasis and immune responses in the skin. Whether Treg frequencies are altered in atopic dermatitis (AD) patients has been addressed by several studies, leading to conflicting results. The detection of Tregs by FOXP3 expression may lead to false-positive results as activated T cells without regulatory function may transiently upregulate this transcription factor. In contrast, measurement of the DNA methylation status of a region within the FOXP3 locus that is selectively demethylated only in bona fide Tregs (Treg-specific demethylated region, TSDR) represents a reliable method to quantify Tregs. Here, we measured circulating Treg frequencies of adult patients and detected a positive correlation with disease severity. Subsequent surface marker analysis revealed higher frequencies of CD45RA(+) CCR7(-) tissue-homing Tregs in the patient group with a tendency of reduced expression of CD39 compared with healthy donors, a marker for the highly suppressive TREM subtype.

The intestinal micro-environment imprints stromal cells to promote efficient Treg induction in gut-draining lymph nodes.

De novo induction of Foxp3⁺ regulatory T cells (Tregs) is particularly efficient in gut-draining mesenteric and celiac lymph nodes (mLN and celLN). Here we used LN transplantations to dissect the contribution of stromal cells and environmental factors to the high Treg-inducing capacity of these LN. After transplantation into the popliteal fossa, mLN and celLN retained their high Treg-inducing capacity, whereas transplantation of skin-draining LN into the gut mesenteries did not enable efficient Treg induction. However, de novo Treg induction was abolished in the absence of dendritic cells (DC), indicating that this process depends on synergistic contributions of stromal and DC. Stromal cells themselves were influenced by environmental signals as mLN grafts taken from germ-free donors and celLN grafts taken from vitamin A-deficient donors did not show any superior Treg-inducing capacity. Collectively, our observations reveal a hitherto unrecognized role of LN stromal cells for the de novo induction of Foxp3⁺ Tregs.

Cord blood Tregs with stable FOXP3 expression are influenced by prenatal environment and associated with atopic dermatitis at the age of one year.

BACKGROUND: Regulatory T cells (Tregs) with stable FOXP3 expression are characterized by a specific demethylated region in the FOXP3 gene (Treg-specific demethylated region, TSDR). The aim of this study was to analyse the influence of prenatal factors on cord blood Treg numbers, as detected by changes in the TSDR demethylation, and the subsequent risk for allergic diseases. METHODS: Analyses were performed within the LINA study in blood samples from pregnant women (34th gestational week) and in cord blood (n = 346 mother-child pairs). Treg numbers were detected via DNA demethylation in the FOXP3 TSDR. At age 1, total and specific IgE was measured in children's blood. In addition, maternal cytokine production (Th1/Th2/Th17) was analysed. Exposure and disease outcomes were assessed by questionnaires. RESULTS: Boys had lower Treg numbers compared with girls (P < 0.001). Parental atopy history, particularly maternal hay fever and paternal asthma were related to lower Treg numbers in cord blood (adj. MR = 0.81, 95% CI = 0.68-0.97; adj. MR = 0.60, 95% CI = 0.45-0.81). Maternal cytokines (IL-13, IL-17E and IFN-γ) and maternal smoking/exposure to tobacco smoke during pregnancy were also associated with decreased cord blood Treg numbers (adj. MR = 0.89, 95% CI = 0.97-1.00). Children with lower Treg numbers at birth had a higher risk to develop atopic dermatitis (adj. OR = 1.55, 95% CI = 1.00-2.41) and sensitization to food allergens (adj. OR = 1.55, 95% CI = 1.06-2.25) during the first year of life. CONCLUSIONS: These results indicate that both genetic and environmental factors presumably influence the development of foetal Tregs. Low cord blood Treg numbers may predict early atopic dermatitis.

[To be or not to be a Treg: epigenetic regulation of Foxp3 expression in regulatory T cells]. / To be or not to be a Treg: Epigenetische Regulation der Foxp3-Expression in regulatorischen T-Zellen.

Regulatory T cells (Treg) harbor great therapeutic potential for the treatment of autoimmune diseases due to their potent suppressive capacity. The majority of these cells express the transcription factor Foxp3, which is critical for both development and function of Treg. We discuss here our recent data indicating a contribution of epigenetic regulation for the permanent expression of Foxp3 in stable Treg a finding that is of significant importance if Treg are devised for clinical applications.

Migration rules: functional properties of naive and effector/memory-like regulatory T cell subsets.

Suppressor T cells were first described in the early 1970s, but since the hypothetical soluble suppressor factor could not be identified on a molecular level and since appropriate cellular markers were lacking, the suppressor T cell concept vanished for a long time. The discovery by Sakaguchi and co-workers, that the adoptive transfer of CD25+CD4+ -depleted T cells induced several organ-specific autoimmune diseases in immunodeficient recipients, put the suppressor T cell model back into the focus of many immunologists. CD25+CD4+ T cells were named regulatory T cells (Treg) and since then have been intensively characterized by many groups. It has now been well documented in a variety of models that CD25+CD4+ Tregs, in addition to cell-intrinsic peripheral tolerance mechanisms such as anergy induction and peripheral deletion, play indispensable roles in the maintenance of natural self-tolerance, in averting autoimmune responses as well as in controlling inflammatory reactions. However, a number of fundamental questions concerning their origin, mechanism of action, and the sites of suppression remain elusive and are currently a matter of debate. Notably, the potential heterogeneity of Tregs with respect to phenotype and function deserves attention and is a major issue discussed in this review.