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[Display omitted] The abundance and diversity of intestinal commensal bacteria influence systemic immunity with impact on disease susceptibility and severity. For example, loss of short chain fatty acid (SCFA)-fermenting bacteria in early life (humans and mice) is associated with enhanced type 2 immune responses in peripheral tissues including the lung. Our goal was to reveal the microbiome-dependent cellular and molecular mechanisms driving enhanced susceptibility to type 2 allergic lung disease. We used low-dose vancomycin to selectively deplete SCFA-fermenting bacteria in wild-type mice. We then examined the frequency and activation status of innate and adaptive immune cell lineages with and without SCFA supplementation. Finally, we used ILC2-deficient and signal transducer and activator of transcription 6 (STAT6)-deficient transgenic mouse strains to delineate the cellular and cytokine pathways leading to enhanced allergic disease susceptibility. Mice with vancomycin-induced dysbiosis exhibited a 2-fold increase in lung ILC2 primed to produce elevated levels of IL-2, -5, and -13. In addition, upon IL-33 inhalation, mouse lung ILC2 displayed a novel ability to produce high levels of IL-4. These expanded and primed ILC2s drove B1 cell expansion and IL-4–dependent production of IgE that in turn led to exacerbated allergic inflammation. Importantly, these enhanced lung inflammatory phenotypes in mice with vancomycin-induced dysbiosis were reversed by administration of dietary SCFA (specifically butyrate). SCFAs regulate an ILC2–B1 cell–IgE axis. Early-life administration of vancomycin, an antibiotic known to deplete SCFA-fermenting gut bacteria, primes and amplifies this axis and leads to lifelong enhanced susceptibility to type 2 allergic lung disease. [ABSTRACT FROM AUTHOR]

Evolution has created complex mechanisms to sense environmental danger and protect tissues, with the nervous and immune systems playing pivotal roles. These systems work together, coordinating local and systemic reflexes to restore homeostasis in response to tissue injury and infection. By sharing receptors and ligands, they influence the pathogenesis of various diseases. Recently, a less-explored aspect of neuroimmune communication has emerged: the release of neuropeptides from immune cells and cytokines/chemokines from sensory neurons. This article reviews evidence of this unique neuroimmune interplay and its impact on the development of allergy, inflammation, itch, and pain. We highlight the effects of this neuroimmune signaling on vital processes such as host defense, tissue repair, and inflammation resolution, providing avenues for exploration of the underlying mechanisms and therapeutic potential of this signaling. [ABSTRACT FROM AUTHOR]

Group 2 innate lymphoid cells (ILC2s), the innate counterpart of T H 2 cells, play a critical role in type 2 immune responses. However, the molecular regulatory mechanisms of ILC2s are still unclear. The aim of this study was to explore the importance of signal transducer and activator of transcription 3 (STAT3) to ILC2 function in allergic lung inflammation. Acute and chronic asthma models were established by intranasal administration of the protease allergen papain in Vav icre Stat3 fl/fl, Il5 tdtomato-cre Stat3 fl/fl, and Rorc cre Stat3 fl/fl mice to verify the necessity of functional STAT3 for ILC2 allergic response. The intrinsic role of STAT3 in regulating ILC2 function was examined by generation of bone marrow chimera mice. The underlying mechanism was studied through confocal imaging, metabolomics analysis, and chromatin immunoprecipitation quantitative PCR. STAT3 is essential for ILC2 effector function and promotes ILC2-driven allergic inflammation in the lung. Mechanistically, the alarmin cytokine IL-33 induces a noncanonical STAT3 phosphorylation at serine 727 in ILC2s, leading to translocation of STAT3 into the mitochondria. Mitochondrial STAT3 further facilitates adenosine triphosphate synthesis to fuel the methionine cycle and generation of S-adenosylmethionine, which supports the epigenetic reprogramming of type 2 cytokines in ILC2s. STAT3 deficiency, inhibition of STAT3 mitochondrial translocation, or blockade of methionine metabolism markedly dampened the ILC2 allergic response and ameliorated allergic lung inflammation. The mitochondrial STAT3-methionine metabolism pathway is a key regulator that shapes ILC2 effector function through epigenetic regulation, and the related proteins or metabolites represent potential therapeutic targets for allergic lung inflammation. [Display omitted] [ABSTRACT FROM AUTHOR]

Aspirin-exacerbated respiratory disease (AERD) is associated with high levels of cysteinyl leukotrienes, prostaglandin D 2 , and low levels of prostaglandin E 2. Further, 15-hydroxyeicosatetraenoic acid (15-HETE) levels may have predictive value in therapeutic outcomes of aspirin desensitization. Accumulation of nasal group 2 innate lymphoid cells (ILC2s) has been demonstrated during COX-1 inhibition in AERD, although the relationships between tissue ILC2 accumulation, reaction symptom severity, and novel lipid biomarkers are unknown. We sought to determine whether novel lipid mediators are predictive of nasal ILC2 accumulation and symptom scores during COX-1 inhibitor challenge in patients with AERD. Blood and nasal scraping samples from patients with AERD were collected at baseline and COX-1 inhibitor reaction and then processed for flow cytometry for nasal ILC2s and serum for lipidomic analysis. Eight patients with AERD who were undergoing aspirin desensitization were recruited. Of the 161 eicosanoids tested, 42 serum mediators were detected. Baseline levels of 15-HETE were negatively correlated with the change in numbers of airway ILC2s (r = –0.6667; P =.0428). Docosahexaenoic acid epoxygenase metabolite 19,20-dihydroxy-4Z,7Z,10Z,13Z,16Z-docosapentaenoic acid (19,20-diHDPA) was positively correlated with both changes in airway ILC2s (r = 0.7143; P =.0305) and clinical symptom scores (r = 0.5000; P =.0081). Low levels of baseline 15-HETE predicted a greater accumulation of airway ILC2s in patients with AERD who were receiving COX-1 inhibition. Further, increases in the cytochrome P pathway metabolite 19,20-dihydroxy-4Z,7Z,10Z,13Z,16Z-docosapentaenoic acid (19,20-diHDPA) were associated with increased symptoms and nasal ILC2 accumulation. Future studies to assess how these mediators might control ILC2s may improve the understanding of AERD pathogenesis. [ABSTRACT FROM AUTHOR]

Cannabinoids modulate the activation of immune cells and physiologic processes in the lungs. Group 2 innate lymphoid cells (ILC2s) are central players in type 2 asthma, but how cannabinoids modulate ILC2 activation remains to be elucidated. Our goal was to investigate the effects of cannabinoids on ILC2s and their role in asthma. A combination of cannabinoid receptor (CB) 2 knockout (KO) mice, CB 2 antagonist and agonist were used in the mouse models of IL-33, IL-25, and Alternaria alternata ILC2-dependent airway inflammation. RNA sequencing was performed to assess transcriptomic changes in ILC2s, and humanized mice were used to assess the role of CB 2 signaling in human ILC2s. We provide evidence that CB 2 signaling in ILC2s is important for the development of ILC2-driven airway inflammation in both mice and human. We showed that both naive and activated murine pulmonary ILC2s express CB 2. CB 2 signaling did not affect ILC2 homeostasis at steady state, but strikingly it stimulated ILC2 proliferation and function upon activation. As a result, ILC2s lacking CB 2 induced lower lung inflammation, as we made similar observations using a CB 2 antagonist. Conversely, CB 2 agonism remarkably exacerbated ILC2-driven airway hyperreactivity and lung inflammation. Mechanistically, transcriptomic and protein analysis revealed that CB 2 signaling induced cyclic adenosine monophosphate–response element binding protein (CREB) phosphorylation in ILC2s. Human ILC2s expressed CB 2 , as CB 2 antagonism and agonism showed opposing effects on ILC2 effector function and development of airway hyperreactivity in humanized mice. Collectively, our results define CB 2 signaling in ILC2s as an important modulator of airway inflammation. [Display omitted] [ABSTRACT FROM AUTHOR]

The most common endotype of asthma is type 2–high asthma, which is sometimes driven by adaptive allergen-specific T H 2 lymphocytes that react to allergens presented by dendritic cells (DCs), or sometimes by an innate immune response dominated by type 2 innate lymphocytes (ILC2s). Understanding the underlying pathophysiology of asthma is essential to improve patient-tailored therapy. The STE20 kinase thousand-and-one kinase 3 (TAOK3) controls key features in the biology of DCs and lymphocytes, but to our knowledge, its potential usefulness as a target for asthma therapy has not yet been addressed. We examined if and how loss of Taok3 affects the development of house dust mite (HDM)-driven allergic asthma in an in vivo mouse model. Wild-type Taok3 +/+ and gene-deficient Taok3 −/− mice were sensitized and challenged with HDM, and bronchoalveolar lavage fluid composition, mediastinal lymph node cytokine production, lung histology, and bronchial hyperreactivity measured. Conditional Taok3 fl/fl mice were crossed to tissue- and cell-specific specific deletor Cre mice to understand how Taok3 acted on asthma susceptibility. Kinase-dead (KD) Taok3 KD mice were generated to probe for the druggability of this pathway. Activation of HDM-specific T cells was measured in adoptively transferred HDM-specific T-cell receptor–transgenic CD4+ T cells. ILC2 biology was assessed by in vivo and in vitro IL-33 stimulation assays in Taok3 −/− and Taok3 +/+ , Taok3 KD, and Red5-Cre Taok3 fl/fl mice. Taok3 −/− mice failed to mount salient features of asthma, including airway eosinophilia, T H 2 cytokine production, IgE secretion, airway goblet cell metaplasia, and bronchial hyperreactivity compared to controls. This was due to intrinsic loss of Taok3 in hematopoietic and not epithelial cells. Loss of Taok3 resulted in hampered HDM-induced lung DC migration to the draining lymph nodes and defective priming of HDM-specific T H 2 cells. Strikingly, HDM and IL-33–induced ILC2 proliferation and function were also severely affected in Taok3-deficient and Taok3 KD mice. Absence of Taok3 or loss of its kinase activity protects from HDM-driven allergic asthma as a result of defects in both adaptive DC-mediated T H 2 activation and innate ILC2 function. This identifies Taok3 as an interesting drug target, justifying further testing as a new treatment for type 2–high asthma. [Display omitted] [ABSTRACT FROM AUTHOR]

Type 2 innate lymphoid cells (ILC2s) are relevant players in type 2 asthma. They initiate eosinophil infiltration and airway hyperreactivity (AHR) through cytokine secretion. Leukocyte-associated immunoglobulin-like receptor 1 (LAIR-1) is an inhibitory receptor considered to be an immune checkpoint in different inflammatory diseases. Our aim here was to investigate the expression of LAIR-1 and assess its role in human and murine ILC2s. Wild-type and LAIR-1 knockout mice were intranasally challenged with IL-33, and pulmonary ILC2s were sorted to perform an ex vivo comparative study based on RNA sequencing and flow cytometry. We next studied the impact of LAIR-1 deficiency on AHR and lung inflammation by using knockout mice and adoptive transfer experiments in Rag2 −/− Il2rg −/− mice. Knockdown antisense strategies and humanized mice were used to assess the role of LAIR-1 in human ILC2s. We have demonstrated that LAIR-1 is inducible on activated ILC2s and downregulates cytokine secretion and effector function. LAIR-1 signaling in ILC2s was mediated via inhibitory pathways, including SHP1/PI3K/AKT, and LAIR-1 deficiency led to exacerbated ILC2-dependent AHR in IL-33 and Alternaria alternata models. In adoptive transfer experiments, we confirmed the LAIR-1–mediated regulation of ILC2s in vivo. Interestingly, LAIR-1 was expressed and inducible in human ILC2s, and knockdown approaches of Lair1 resulted in higher cytokine production. Finally, engagement of LAIR-1 by physiologic ligand C1q significantly reduced ILC2-dependent AHR in a humanized ILC2 murine model. Our results unravel a novel regulatory axis in ILC2s with the capacity to reduce allergic AHR and lung inflammation. [Display omitted] [ABSTRACT FROM AUTHOR]

Allergic asthma is more severe and frequent in women than in men. In male mice, androgens negatively control group 2 innate lymphoid cell (ILC2) development and function by yet unknown mechanisms. We sought to investigate the impact of androgen on ILC2 homeostasis and IL-33–mediated inflammation in female lungs. We evaluated the role of androgen receptor (AR) signaling and the contribution of the putative inhibitory receptor killer cell lectin-like receptor G1 (KLRG1). Subcutaneous pellets mimicking physiological levels of androgen were used to treat female mice together with mice expressing a reporter enzyme under the control of androgen response elements and mixed bone marrow chimeras to assess the cell-intrinsic role of AR activation within ILC2s. We generated KLRG1-deficient mice. We established that lung ILC2s express a functionally active AR that can be in vivo targeted with exogenous androgens to negatively control ILC2 homeostasis, proliferation, and function. Androgen signaling upregulated KLRG1 on ILC2s, which inhibited their proliferation on E-cadherin interaction. Despite evidence that KLRG1 impaired the competitive fitness of lung ILC2s during inflammation, KLRG1 deficiency neither alters in vivo ILC2 numbers and functions, nor did it lead to hyperactive ILC2s in either sexes. AR agonists can be used in vivo to inhibit ILC2 homeostatic numbers and ILC2-dependent lung inflammation through cell-intrinsic AR activation. Although androgen signals in ILC2s to upregulate KLRG1, we demonstrate that KLRG1 is dispensable for androgen-mediated inhibition of pulmonary ILC2s. [ABSTRACT FROM AUTHOR]

Systemic mastocytosis is a hematological disease in which aberrant mast cells accumulate because of gain-of-function mutations in the KIT receptor. Group 2 innate lymphoid cells (ILC2s) are effector cells of type 2 immune responses that also express KIT and colocalize with mast cells at barrier tissue sites. In mouse models, mast cell-ILC2 crosstalk can drive local inflammation. However, a possible role for ILC2s in the pathophysiology of mastocytosis remains unexplored. We sought to characterize circulating ILC2s in a clinically diverse cohort of patients with mastocytosis. We included 21 adults with systemic mastocytosis and 18 healthy controls. Peripheral blood ILC2 abundance and phenotype were analyzed by flow cytometry and correlated to clinical characteristics, including the presence of the D816V KIT mutation. ILC2 levels were significantly higher in D816V+ patients with mastocytosis compared with D816V− patients or healthy controls. We observed increased proportions of KIT+ ILC2s among patients with mastocytosis, regardless of D816V status. Patients with skin involvement and itch showed the highest levels of ILC2s, which was independent from atopy or serum tryptase levels. Allele-specific quantitative PCR showed that the vast majority of ILC2s did not carry the D816V mutation. Our findings suggest a role for ILC2s and pathogenic ILC2-mast cell crosstalk in mastocytosis. We hypothesize that a high cutaneous D816V+ mast cell burden alters the skin microenvironment to induce chronic local ILC2 activation and their dissemination into the circulation. Activated ILC2s could contribute to skin symptoms by producing inflammatory mediators and by further augmenting mast cell mediator release. [Display omitted] [ABSTRACT FROM AUTHOR]

Group 2 innate lymphoid cells (ILC2s) are the dominant innate lymphoid cell population in the lungs at steady state, and their release of type 2 cytokines is a central driver in responding eosinophil infiltration and increased airway hyperreactivity. Our laboratory has identified a unique subset of ILC2s in the lungs that actively produce IL-10 (ILC2 10 s). Our aim was to characterize the effector functions of ILC2 10 s in the development and pathology of allergic asthma. IL-4–stimulated ILC2 10 s were isolated to evaluate cytokine secretion, transcription factor signaling, metabolic dependence, and effector functions in vitro. ILC2 10 s were also adoptively transferred into Rag2–/–γc–/– mice, which were then challenged with IL-33 and assessed for airway hyperreactivity and lung inflammation. We have determined that the transcription factors cMaf and Blimp-1 regulate IL-10 expression in ILC2 10 s. Strikingly, our results demonstrate that ILC2 10 s can utilize both autocrine and paracrine signaling to suppress proinflammatory ILC2 effector functions in vitro. Further, this subset dampens airway hyperreactivity and significantly reduces lung inflammation in vivo. Interestingly, ILC2 10 s demonstrated a metabolic dependency on the glycolytic pathway for IL-10 production, shifting from the fatty acid oxidation pathway conventionally utilized for proinflammatory effector functions. These findings provide an important and previously unrecognized role of ILC2 10 s in diseases associated with ILC2s such as allergic lung inflammation and asthma. They also provide new insights into the metabolism dependency of proinflammatory and anti-inflammatory ILC2 phenotypes. [ABSTRACT FROM AUTHOR]

Early-life wheezing-associated respiratory tract infection by rhinovirus (RV) is a risk factor for asthma development. Infants are infected with many different RV strains per year. We previously showed that RV infection of 6-day-old BALB/c mice induces a mucous metaplasia phenotype that is dependent on type 2 innate lymphoid cells (ILC2s). We hypothesized that early-life RV infection alters the response to subsequent heterologous infection, inducing an exaggerated asthma-like phenotype. Wild-type BALB/c mice and Rora fl/fl Il7r cre mice lacking ILC2s were treated as follows: (1) sham on day 6 of life plus sham on day 13 of life, (2) RV-A1B on day 6 plus sham on day 13, (3) sham on day 6 plus RV-A2 on day 13, and (4) RV-A1B on day 6 plus RV-A2 on day 13. Mice infected with RV-A1B at day 6 and sham at day 13 showed an increased number of bronchoalveolar lavage eosinophils and increased expression of IL-13 mRNA but not expression of IFN-γ mRNA (which is indicative of a type 2 immune response), whereas mice infected with sham on day 6 and RV-A2 on day 13 of life demonstrated increased IFN-γ expression (which is a mature antiviral response). In contrast, mice infected with RV-A1B on day 6 before RV-A2 infection on day 13 showed increased expression of IL-13, IL-5, Gob5, Muc5b, and Muc5ac mRNA; increased numbers of eosinophils and IL-13–producing ILC2s; and exaggerated mucus metaplasia and airway hyperresponsiveness. Compared with Rora fl/fl mice, Rora fl/fl Il7r cre mice showed complete suppression of bronchoalveolar lavage eosinophils and mucous metaplasia. Early-life RV infection alters the response to subsequent heterologous infection, inducing an intensified asthma-like phenotype that is dependent on ILC2s. [ABSTRACT FROM AUTHOR]

Atopic dermatitis skin lesions demonstrate increased expression of IL-25 by keratinocytes and increased numbers of type 2 innate lymphoid cells (ILC2s) that express high levels of IL-25 receptor (IL-25R). IL-13 is expressed in atopic dermatitis skin lesions and plays an important role in pathogenesis of the disease. Our aim was to determine the role of IL-25 and ILC2s in a mouse model of antigen-driven allergic skin inflammation. Wild-type mice; mice that express an Il13- driven enhanced green fluorescent protein; and mice that lack IL-25R, IL-25 in keratinocytes, or IL-13 or IL-25R in ILC2s were subjected to acute or chronic epicutaneous sensitization with ovalbumin. Sensitized skin was examined by histology for epidermal thickening. Cellular infiltrates were analyzed for surface markers and intracellular expression of enhanced green fluorescent protein by flow cytometry. Gene expression was quantitated by RT quantitative PCR. In both acute and chronic antigen-driven allergic skin inflammation, signaling by keratinocyte-derived IL-25 in ILC2s is important for epidermal hyperplasia, dermal infiltration by CD4 + T cells, and cutaneous expression of Il13 and the IL-13–dependent T H 2-cell–attracting chemokines Cc17 and Ccl22. ILCs are the major source of IL-13 in acutely sensitized mouse skin, whereas T cells are its major source in chronically sensitized mouse skin. ILC2 activation by IL-25 is essential for IL-13 expression at sites of allergic skin inflammation. [ABSTRACT FROM AUTHOR]

Background Group 2 innate lymphoid cells (ILC2s) are involved in the initial phase of type 2 inflammation and can amplify allergic immune responses by orchestrating other type 2 immune cells. Prostaglandin (PG) E 2 is a bioactive lipid that plays protective roles in the lung, particularly during allergic inflammation. Objective We set out to investigate how PGE 2 regulates human ILC2 function. Methods The effects of PGE 2 on human ILC2 proliferation and intracellular cytokine and transcription factor expression were assessed by means of flow cytometry. Cytokine production was measured by using ELISA, and real-time quantitative PCR was performed to detect PGE 2 receptor expression. Results PGE 2 inhibited GATA-3 expression, as well as production of the type 2 cytokines IL-5 and IL-13, from human tonsillar and blood ILC2s in response to stimulation with a combination of IL-25, IL-33, thymic stromal lymphopoietin, and IL-2. Furthermore, PGE 2 downregulated the expression of IL-2 receptor α (CD25). In line with this observation, PGE 2 decreased ILC2 proliferation. These effects were mediated by the combined action of E-type prostanoid receptor (EP) 2 and EP4 receptors, which were specifically expressed on ILC2s. Conclusion Our findings reveal that PGE 2 limits ILC2 activation and propose that selective EP2 and EP4 receptor agonists might serve as a promising therapeutic approach in treating allergic diseases by suppressing ILC2 function. [ABSTRACT FROM AUTHOR]

Background Allergic airway inflammation is triggered by allergen exposure through several steps including release of IL-33, which promotes cytokine (IL-5, IL-13) production by type 2 innate lymphoid cells (ILC2s). MicroRNA (miR)-155 has recently been described to regulate adaptive responses in allergic inflammation. However, the role of miR-155 in the regulation of ILC2s remains unexplored. Objective We sought to elucidate the contribution of miR-155 in ILC2 expansion using experimental murine models of allergic airway inflammation. Methods To determine the role of miR-155 in the regulation of ILC2s in allergic airway inflammation, miR-155 deficient (miR-155 −/− ) and wild-type (WT) mice were subjected to acute or chronic allergen-induced inflammation or treated with recombinant IL-33. Results miR-155 was 10-fold upregulated in WT-derived ILC2s in response to IL-33. Furthermore, miR-155 −/− mice demonstrated impaired lung IL-33 levels in response to allergen challenge and the number of ILC2s was significantly reduced in allergen-challenged miR-155 −/− mice compared with WT mice. Exogenous IL-33 treatment revealed that miR-155 is needed for IL-33–induced ILC2 expansion and eosinophilic airway inflammation. Indeed, ILC2s from IL-33–challenged miR-155 −/− lungs exhibited impaired proliferation, GATA-3 expression, and IL-13 production as compared with IL-33–challenged WT ILC2s. Conclusions Our findings for the first time demonstrate that ILC2s and IL-33 signaling are regulated by miR-155 in allergic airway inflammation. [ABSTRACT FROM AUTHOR]

Background In mice, group 2 innate lymphoid cells (ILC2s) likely mediate helminth immunity, inflammation, and tissue repair and remodeling. However, the involvement of ILC2s in human diseases, such as asthma, is not well understood. Objectives The goals of this study were to investigate whether peripheral blood specimens can be used to monitor innate type 2 immunity in human subjects and to examine whether ILC2s are involved in human asthma. Methods PBMCs from subjects with allergic asthma (AA), subjects with allergic rhinitis (AR), or healthy control (HC) subjects were cultured in vitro with IL-25 or IL-33. Flow cytometry and cell sorting were used to identify, isolate, and quantitate ILC2s in PBMCs. Results Human PBMCs produced IL-5 and IL-13 when stimulated with IL-33 or IL-25 in the presence of IL-2 without antigens. In addition, IL-7 or thymic stromal lymphopoietin were able to replace IL-2. The cell population with phenotypic ILC2 characteristics, lineage − CD127 + CRTH2 + cells, responded to IL-33 and produced large quantities of IL-5 and IL-13 but undetectable levels of IL-4. PBMCs from subjects with AA produced significantly larger amounts of IL-5 and IL-13 in response to IL-25 or IL-33 than from subjects with AR or HC. The prevalence of ILC2s in blood was greater in the AA group than in the AR group or the HC group. Conclusions Innate type 2 immune responses are increased in asthma but not in AR, suggesting potential differences in the immunopathogenesis of these diseases. Peripheral blood is useful for evaluating innate type 2 immunity in humans. [ABSTRACT FROM AUTHOR]