Diminished γδ T Cells during Murine Allergic Skin Inflammation Is Mediated by IL-4 Signaling in Keratinocytes.

Atopic dermatitis results in diminished barrier function and altered production of antimicrobial peptides. Dendritic epidermal T cells (DETCs) play an important role in the wound repair and inflammation process. Our previous work identified an IL-4-dependent loss of DETCs in Stat6VT mice and in the MC903-induced skin inflammation mouse model. However, the mechanisms through which IL-4 mediates the loss of DETCs are unclear. In this study, we show that IL-4Rα germline knockout mice (Il4ra-/-) have increased DETCs, faster wound healing, and increased epidermal differentiation complex gene and fibronectin expression. The absence of IL-4Rα minimized the MC903-induced loss of DETCs, and reciprocal bone marrow chimera experiments in Il4ra-/- and wild-type mice demonstrated structural nonhematopoietic IL-4-responsive cell-mediated DETC homeostasis. Skin keratinocyte-derived IL-15 decreased dramatically in the MC903 model, while injection of IL-15 rescued DETC loss by promoting DETC proliferation and limiting apoptosis. Conditional deletion of IL-4Rα from keratinocytes using Il4rafl/fl K14-Cre mice showed an increase of DETCs, increased IL-15 production, and diminished skin inflammation following wounding. These results suggest that IL-4-dependent effects on DETCs in allergic skin inflammation are mediated by the IL-4Rα receptor of keratinocytes.

TL1A priming induces a multi-cytokine Th9 cell phenotype that promotes robust allergic inflammation in murine models of asthma.

Multi-cytokine-producing Th9 cells secrete IL-9 and type 2 cytokines and mediate mouse and human allergic inflammation. However, the cytokines that promote a multi-cytokine secreting phenotype have not been defined. Tumor necrosis factor superfamily member TL1A signals through its receptor DR3 to increase IL-9. Here we demonstrate that TL1A increases expression of IL-9 and IL-13 co-expressing cells in murine Th9 cell cultures, inducing a multi-cytokine phenotype. Mechanistically, this is linked to histone modifications allowing for increased accessibility at the Il9 and Il13 loci. We further show that TL1A alters the transcription factor network underlying expression of IL-9 and IL-13 in Th9 cells and increases binding of transcription factors to Il9 and Il13 loci. TL1A-priming enhances the pathogenicity of Th9 cells in murine models of allergic airway disease through the increased expression of IL-9 and IL-13. Lastly, in both chronic and memory-recall models of allergic airway disease, blockade of TL1A signaling decreases the multi-cytokine Th9 cell population and attenuates the allergic phenotype. Taken together, these data demonstrate that TL1A promotes the development of multi-cytokine Th9 cells that drive allergic airway diseases and that targeting pathogenic T helper cell-promoting cytokines could be an effective approach for modifying disease.

The Dichotomy of Interleukin-9 Function in the Tumor Microenvironment.

Interleukin 9 (IL-9) is a cytokine with potent proinflammatory properties that plays a central role in pathologies such as allergic asthma, immunity to parasitic infection, and autoimmunity. More recently, IL-9 has garnered considerable attention in tumor immunity. Historically, IL-9 has been associated with a protumor function in hematological malignancies and an antitumor function in solid malignancies. However, recent discoveries of the dynamic role of IL-9 in cancer progression suggest that IL-9 can act as both a pro- or antitumor factor in various hematological and solid malignancies. This review summarizes IL-9-dependent control of tumor growth, regulation, and therapeutic applicability of IL-9 blockade and IL-9-producing cells in cancer.

The World according to IL-9.

Among the cytokines regulating immune cells, IL-9 has gained considerable attention for its ability to act on multiple cell types as a regulator of beneficial and pathologic immune responses. Yet, it is still not clearly defined how IL-9 impacts immune responses. IL-9 demonstrates a remarkable degree of tissue-specific functionality and has cellular sources that vary by tissue site and the context of the inflammatory milieu. Here, we provide perspective to summarize the biological activities of IL-9 and highlight cell type-specific roles in the immune pathogenesis of diseases. This perspective will be important in defining the diseases where targeting IL-9 as a therapeutic strategy would be beneficial and where it has the potential to complicate clinical outcomes.

Interleukin-9 promotes mast cell progenitor proliferation and CCR2-dependent mast cell migration in allergic airway inflammation.

Allergic asthma is a chronic lung disease characterized by airway hyperresponsiveness and cellular infiltration that is exacerbated by immunoglobulin E-dependent mast cell (MC) activation. Interleukin-9 (IL-9) promotes MC expansion during allergic inflammation but precisely how IL-9 expands tissue MCs and promotes MC function is unclear. In this report, using multiple models of allergic airway inflammation, we show that both mature MCs (mMCs) and MC progenitors (MCp) express IL-9R and respond to IL-9 during allergic inflammation. IL-9 acts on MCp in the bone marrow and lungs to enhance proliferative capacity. Furthermore, IL-9 in the lung stimulates the mobilization of CCR2+ mMC from the bone marrow and recruitment to the allergic lung. Mixed bone marrow chimeras demonstrate that these are intrinsic effects in the MCp and mMC populations. IL-9-producing T cells are both necessary and sufficient to increase MC numbers in the lung in the context of allergic inflammation. Importantly, T cell IL-9-mediated MC expansion is required for the development of antigen-induced and MC-dependent airway hyperreactivity. Collectively, these data demonstrate that T cell IL-9 induces lung MC expansion and migration by direct effects on the proliferation of MCp and the migration of mMC to mediate airway hyperreactivity.

Chronic developmental hypoxia alters rat lung immune cell transcriptomes during allergic airway inflammation.

Populations that are born and raised at high altitude develop under conditions of chronic developmental hypoxia (CDH), which results in pulmonary adaptations of increased lung volume and diffusion capacity to increase gas exchange. It is not clear how CDH may alter allergic inflammation in the lung. In this study, we sought to characterize the impact of CDH on immune cell populations in the rat lung during a murine model of asthma. Rats were bred and raised in either hypoxic (15% oxygen, CDH) or normobaric room air (20% oxygen). At 3-weeks of age, animals were sensitized to ovalbumin (OVA) or physiologic saline (phosphate-buffered saline [PBS]) as a control, followed by three consecutive days of intra-nasal OVA or PBS at 6-weeks of age. We then assessed airway reactivity and allergic-associated cytokine levels. This was followed by single-cell transcriptomic profiling of lung cell populations. In scRNA-seq analysis, we assessed differentially expressed genes, differentially enriched functional pathways, immune cell exhaustion/activation markers, and immune cell secretory products. Our results show that while OVA heightened airway reactivity, CDH suppressed airway reactivity in OVA-challenged and control animals. Through scRNA-seq analysis, we further demonstrate that CDH alters the transcriptional landscape in the lung and alters transcriptional programs in immune cells. These data define CDH-dependent changes in the lung that impact airway reactivity.

Mouse pulmonary interstitial macrophages mediate the pro-tumorigenic effects of IL-9.

Although IL-9 has potent anti-tumor activity in adoptive cell transfer therapy, some models suggest that it can promote tumor growth. Here, we show that IL-9 signaling is associated with poor outcomes in patients with various forms of lung cancer, and is required for lung tumor growth in multiple mouse models. CD4+ T cell-derived IL-9 promotes the expansion of both CD11c+ and CD11c- interstitial macrophage populations in lung tumor models. Mechanistically, the IL-9/macrophage axis requires arginase 1 (Arg1) to mediate tumor growth. Indeed, adoptive transfer of Arg1+ but not Arg1- lung macrophages to Il9r-/- mice promotes tumor growth. Moreover, targeting IL-9 signaling using macrophage-specific nanoparticles restricts lung tumor growth in mice. Lastly, elevated expression of IL-9R and Arg1 in tumor lesions is associated with poor prognosis in lung cancer patients. Thus, our study suggests the IL-9/macrophage/Arg1 axis is a potential therapeutic target for lung cancer therapy.

γδ T Cell‒Mediated Wound Healing Is Diminished by Allergic Skin Inflammation.

Atopic dermatitis results in profound changes in the function of the skin that include diminished barrier function and altered production of antimicrobial peptides. Our previous work in a model of allergic skin inflammation identified a defect in the wound healing process that was dependent on IL-4. In this report, we show that allergic skin inflammation results in a dramatic decrease in the presence of the Vγ3+ dendritic epidermal T-cell (DETC) population of γδ T cells in the skin. In mice that express an active signal transducer and activator of transcription 6 in T cells, DETCs are lost early in life. The loss of DETCs is entirely dependent on IL-4 and is recovered with a genetic deficiency of IL-4. Moreover, injection of IL-4 into wild-type mice results in acute loss of the DETC population. A similar loss of DETCs was observed in mice treated topically with MC903. Wounding of skin from Stat6VT-transgenic or MC903-treated mice resulted in decreased production of DETC-dependent cytokines in the skin, coincident with diminished wound closure. Importantly, intradermal injection of the DETC-produced cytokine fibroblast GF 7 rescued the rate of wound closure in mice with allergic skin inflammation. Together, these results suggest that the atopic environment diminishes prohealing T-cell populations in the skin, resulting in attenuated wound healing responses.

Allergic airway recall responses require IL-9 from resident memory CD4+ T cells.

Asthma is a chronic inflammatory lung disease with intermittent flares predominately mediated through memory T cells. Yet, the identity of long-term memory cells that mediate allergic recall responses is not well defined. In this report, using a mouse model of chronic allergen exposure followed by an allergen-free rest period, we characterized a subpopulation of CD4+ T cells that secreted IL-9 as an obligate effector cytokine. IL-9-secreting cells had a resident memory T cell phenotype, and blocking IL-9 during a recall challenge or deleting IL-9 from T cells significantly diminished airway inflammation and airway hyperreactivity. T cells secreted IL-9 in an allergen recall-specific manner, and secretion was amplified by IL-33. Using scRNA-seq and scATAC-seq, we defined the cellular identity of a distinct population of T cells with a proallergic cytokine pattern. Thus, in a recall model of allergic airway inflammation, IL-9 secretion from a multicytokine-producing CD4+ T cell population was required for an allergen recall response.

An IL-9-pulmonary macrophage axis defines the allergic lung inflammatory environment.

Despite IL-9 functioning as a pleiotropic cytokine in mucosal environments, the IL-9-responsive cell repertoire is still not well defined. Here, we found that IL-9 mediates proallergic activities in the lungs by targeting lung macrophages. IL-9 inhibits alveolar macrophage expansion and promotes recruitment of monocytes that develop into CD11c+ and CD11c- interstitial macrophage populations. Interstitial macrophages were required for IL-9-dependent allergic responses. Mechanistically, IL-9 affected the function of lung macrophages by inducing Arg1 activity. Compared with Arg1-deficient lung macrophages, Arg1-expressing macrophages expressed greater amounts of CCL5. Adoptive transfer of Arg1+ lung macrophages but not Arg1- lung macrophages promoted allergic inflammation that Il9r-/- mice were protected against. In parallel, the elevated expression of IL-9, IL-9R, Arg1, and CCL5 was correlated with disease in patients with asthma. Thus, our study uncovers an IL-9/macrophage/Arg1 axis as a potential therapeutic target for allergic airway inflammation.

γδ T Cells in Skin Inflammation.

Gamma delta (γδ) T cells are a subset of T lymphocytes that express T cell receptor γ and 5 chains and display structural and functional heterogeneity. γδ T cells are typically of low abundance in the body and account for 1-5% of the blood lymphocytes and peripheral lymphoid tissues. As a bridge between innate and adaptive immunity, γδ T cells are uniquely poised to rapidly respond to stimulation and can regulate immune responses in peripheral tissues. The dendritic epidermal T cells in the skin epidermis can secrete growth factors to regulate skin homeostasis and re-epithelization and release inflammatory factors to mediate wound healing during skin inflammatory responses. Dermal γδ T cells can regulate the inflammatory process by producing interleukin-17 and other cytokines or chemokines. Here, we offer a review of the immune functions of γδ T cells, intending to understand their role in regulating skin barrier integrity and skin wound healing, which may be crucial for the development of novel therapeutics in skin diseases like atopic dermatitis and psoriasis.