The Pioneer Transcription Factor Foxa2 Modulates T Helper Differentiation to Reduce Mouse Allergic Airway Disease.

Foxa2, a member of the Forkhead box (Fox) family of transcription factors, plays an important role in the regulation of lung function and lung tissue homeostasis. FOXA2 expression is reduced in the lung and airways epithelium of asthmatic patients and in mice absence of Foxa2 from the lung epithelium contributes to airway inflammation and goblet cell hyperplasia. Here we demonstrate a novel role for Foxa2 in the regulation of T helper differentiation and investigate its impact on lung inflammation. Conditional deletion of Foxa2 from T-cells led to increased Th2 cytokine secretion and differentiation, but decreased Th1 differentiation and IFN-γ expression in vitro. Induction of mouse allergic airway inflammation resulted in more severe disease in the conditional Foxa2 knockout than in control mice, with increased cellular infiltration to the lung, characterized by the recruitment of eosinophils and basophils, increased mucus production and increased production of Th2 cytokines and serum IgE. Thus, these experiments suggest that Foxa2 expression in T-cells is required to protect against the Th2 inflammatory response in allergic airway inflammation and that Foxa2 is important in T-cells to maintain the balance of effector cell differentiation and function in the lung.

Sonic Hedgehog signalling in the regulation of barrier tissue homeostasis and inflammation.

Epithelial barrier tissues such as the skin and airway form an essential interface between the mammalian host and its external environment. These physical barriers are crucial to prevent damage and disease from environmental insults and allergens. Failure to maintain barrier function against such risks can lead to severe inflammatory disorders, including atopic dermatitis and asthma. Here, we discuss the role of the morphogen Sonic Hedgehog in postnatal skin and lung and the impact of Shh signalling on repair, inflammation, and atopic disease in these tissues.

Regulation of Adaptive Tumor Immunity by Non-Coding RNAs.

Cancer immunology research has mainly focused on the role of protein-coding genes in regulating immune responses to tumors. However, despite more than 70% of the human genome is transcribed, less than 2% encodes proteins. Many non-coding RNAs (ncRNAs), including microRNAs (miRNAs) and long non-coding RNAs (lncRNAs), have been identified as critical regulators of immune cell development and function, suggesting that they might play important roles in orchestrating immune responses against tumors. In this review, we summarize the scientific advances on the role of ncRNAs in regulating adaptive tumor immunity, and discuss their potential therapeutic value in the context of cancer immunotherapy.

Systemic Pharmacological Smoothened Inhibition Reduces Lung T-Cell Infiltration and Ameliorates Th2 Inflammation in a Mouse Model of Allergic Airway Disease.

Allergic asthma is a common inflammatory airway disease in which Th2 immune response and inflammation are thought to be triggered by inhalation of environmental allergens. Many studies using mouse models and human tissues and genome-wide association have indicated that Sonic Hedgehog (Shh) and the Hedgehog (Hh) signaling pathway are involved in allergic asthma and that Shh is upregulated in the lung on disease induction. We used a papain-induced mouse model of allergic airway inflammation to investigate the impact of systemic pharmacological inhibition of the Hh signal transduction molecule smoothened on allergic airway disease induction and severity. Smoothened-inhibitor treatment reduced the induction of Shh, IL-4, and IL-13 in the lung and decreased serum IgE, as well as the expression of Smo, Il4, Il13, and the mucin gene Muc5ac in lung tissue. Smoothened inhibitor treatment reduced cellular infiltration of eosinophils, mast cells, basophils, and CD4+ T-cells to the lung, and eosinophils and CD4+ T-cells in the bronchoalveolar lavage. In the mediastinal lymph nodes, smoothened inhibitor treatment reduced the number of CD4+ T-cells, and the cell surface expression of Th2 markers ST2 and IL-4rα and expression of Th2 cytokines. Thus, overall pharmacological smoothened inhibition attenuated T-cell infiltration to the lung and Th2 function and reduced disease severity and inflammation in the airway.

Sonic Hedgehog signaling limits atopic dermatitis via Gli2-driven immune regulation.

Hedgehog (Hh) proteins regulate development and tissue homeostasis, but their role in atopic dermatitis (AD) remains unknown. We found that on induction of mouse AD, Sonic Hedgehog (Shh) expression in skin, and Hh pathway action in skin T cells were increased. Shh signaling reduced AD pathology and the levels of Shh expression determined disease severity. Hh-mediated transcription in skin T cells in AD-induced mice increased Treg populations and their suppressive function through increased active transforming growth factor-ß (TGF-ß) in Tregs signaling to skin T effector populations to reduce disease progression and pathology. RNA sequencing of skin CD4+ T cells from AD-induced mice demonstrated that Hh signaling increased expression of immunoregulatory genes and reduced expression of inflammatory and chemokine genes. Addition of recombinant Shh to cultures of naive human CD4+ T cells in iTreg culture conditions increased FOXP3 expression. Our findings establish an important role for Shh upregulation in preventing AD, by increased Gli-driven Treg cell-mediated immune suppression, paving the way for a potential new therapeutic strategy.

IFITM proteins drive type 2 T helper cell differentiation and exacerbate allergic airway inflammation.

The interferon-inducible transmembrane (Ifitm/Fragilis) genes encode homologous proteins that are induced by IFNs. Here, we show that IFITM proteins regulate murine CD4+ Th cell differentiation. Ifitm2 and Ifitm3 are expressed in wild-type (WT) CD4+ T cells. On activation, Ifitm3 was downregulated and Ifitm2 was upregulated. Resting Ifitm-family-deficient CD4+ T cells had higher expression of Th1-associated genes than WT and purified naive Ifitm-family-deficient CD4+ T cells differentiated more efficiently to Th1, whereas Th2 differentiation was inhibited. Ifitm-family-deficient mice, but not Ifitm3-deficient mice, were less susceptible than WT to induction of allergic airways disease, with a weaker Th2 response and less severe disease and lower Il4 but higher Ifng expression and IL-27 secretion. Thus, the Ifitm family is important in adaptive immunity, influencing Th1/Th2 polarization, and Th2 immunopathology.

Foxa1 and Foxa2 in thymic epithelial cells (TEC) regulate medullary TEC and regulatory T-cell maturation.

The Foxa1 and Foxa2 transcription factors are essential for mouse development. Here we show that they are expressed in thymic epithelial cells (TEC) where they regulate TEC development and function, with important consequences for T-cell development. TEC are essential for T-cell differentiation, lineage decisions and repertoire selection. Conditional deletion of Foxa1 and Foxa2 from murine TEC led to a smaller thymus with a greater proportion of TEC and a greater ratio of medullary to cortical TEC. Cell-surface MHCI expression was increased on cortical TEC in the conditional Foxa1Foxa2 knockout thymus, and MHCII expression was reduced on both cortical and medullary TEC populations. These changes in TEC differentiation and MHC expression led to a significant reduction in thymocyte numbers, reduced positive selection of CD4+CD8+ cells to the CD4 lineage, and increased CD8 cell differentiation. Conditional deletion of Foxa1 and Foxa2 from TEC also caused an increase in the medullary TEC population, and increased expression of Aire, but lower cell surface MHCII expression on Aire-expressing mTEC, and increased production of regulatory T-cells. Thus, Foxa1 and Foxa2 in TEC promote positive selection of CD4SP T-cells and modulate regulatory T-cell production and activity, of importance to autoimmunity.

Gli3 in fetal thymic epithelial cells promotes thymocyte positive selection and differentiation by repression of Shh.

Gli3 is a Hedgehog (Hh)-responsive transcription factor that can function as a transcriptional repressor or activator. We show that Gli3 activity in mouse thymic epithelial cells (TECs) promotes positive selection and differentiation from CD4+ CD8+ to CD4+ CD8- single-positive (SP4) cells in the fetal thymus and that Gli3 represses Shh Constitutive deletion of Gli3, and conditional deletion of Gli3 from TECs, reduced differentiation to SP4, whereas conditional deletion of Gli3 from thymocytes did not. Conditional deletion of Shh from TECs increased differentiation to SP4, and expression of Shh was upregulated in the Gli3-deficient thymus. Use of a transgenic Hh reporter showed that the Hh pathway was active in thymocytes, and increased in the Gli3-deficient fetal thymus. Neutralisation of endogenous Hh proteins in the Gli3-/- thymus restored SP4 differentiation, indicating that Gli3 in TECs promotes SP4 differentiation by repression of Shh Transcriptome analysis showed that Hh-mediated transcription was increased whereas TCR-mediated transcription was decreased in Gli3-/- thymocytes compared with wild type.

IL-12 and IL-4 activate a CD39-dependent intrinsic peripheral tolerance mechanism in CD8(+) T cells.

Immune responses to protein antigens involve CD4(+) and CD8(+) T cells, which follow distinct programs of differentiation. Naïve CD8 T cells rapidly develop cytotoxic T-cell (CTL) activity after T-cell receptor stimulation, and we have previously shown that this is accompanied by suppressive activity in the presence of specific cytokines, i.e. IL-12 and IL-4. Cytokine-induced CD8(+) regulatory T (Treg) cells are one of several Treg-cell phenotypes and are Foxp3(-) IL-10(+) with contact-dependent suppressive capacity. Here, we show they also express high level CD39, an ecto-nucleotidase that degrades extracellular ATP, and this contributes to their suppressive activity. CD39 expression was found to be upregulated on CD8(+) T cells during peripheral tolerance induction in vivo, accompanied by release of IL-12 and IL-10. CD39 was also upregulated during respiratory tolerance induction to inhaled allergen and on tumor-infiltrating CD8(+) T cells. Production of IL-10 and expression of CD39 by CD8(+) T cells was independently regulated, being respectively blocked by extracellular ATP and enhanced by an A2A adenosine receptor agonist. Our results suggest that any CTL can develop suppressive activity when exposed to specific cytokines in the absence of alarmins. Thus negative feedback controls CTL expansion under regulation from both nucleotide and cytokine environment within tissues.