Lung epithelial progenitors use a complex network of known and predicted transcriptional regulators to influence early lung development. In this study, we evaluated the function of one predicted regulator, CUX1, that we identified from transcriptional regulatory analysis of the SOX9+ distal lung progenitor network. We generated a new Cux1-floxed mouse model and created an epithelium-specific knockout of CUX1 using Shh-Cre (Cux1ShhCre-LOF). Postnatal Cux1ShhCre-LOF animals recapitulated key skin phenotypic features found in prior constitutive CUX1 knockout animals, confirming the functionality of our new floxed model. Postnatal Cux1ShhCre-LOF mice displayed subtle alveolar simplification and a transient delay in alveologenesis and alveolar type 1 cell development without persistent lung phenotypes. Cux1ShhCre-LOF mice developed failure to thrive in their second and third weeks of life because of delayed ileal maturation, which similarly resolves by Postnatal Day 35. Finally, we challenged Cux1ShhCre-LOF with influenza-mediated lung injury to demonstrate that Cux1ShhCre-LOF mice undergo productive alveolar regeneration that is indistinguishable from that in wild-type animals. Together, these findings indicate that epithelium-specific loss of CUX1 leads to transient developmental delays in the skin, lung, and intestine without defects in definitive organogenesis. We conclude that CUX1 function is required for temporal optimization of developmental maturation in multiple organs with implications for susceptibility windows in developmental disease pathogenesis.

Caspase recruitment domain family member 14 (CARD14) and its variants are associated with both atopic dermatitis (AD) and psoriasis, but their mechanistic impact on skin barrier homeostasis is largely unknown. CARD14 is known to signal via NF-κB; however, CARD14-NF-κB signaling does not fully explain the heterogeneity of CARD14-driven disease. Here, we describe a direct interaction between CARD14 and MYC and show that CARD14 signals through MYC in keratinocytes to coordinate skin barrier homeostasis. CARD14 directly binds MYC and influences barrier formation in an MYC-dependent fashion, and this mechanism is undermined by disease-associated CARD14 variants. These studies establish a paradigm that CARD14 activation regulates skin barrier function by two distinct mechanisms, including activating NF-κB to bolster the antimicrobial (chemical) barrier and stimulating MYC to bolster the physical barrier. Finally, we show that CARD14-dependent MYC signaling occurs in other epithelia, expanding the impact of our findings beyond the skin.

Atopic dermatitis (AD) affects between 15% and 24% of children in the United States1 and up to 20% of children worldwide.2 Adults with AD have increased cardiovascular risk including thromboembolic events compared to adults without AD.3 Indeed, associations between altered cutaneous and systemic clotting and atopy in adults have been observed,4-6 including a recent study which showed that adults with AD have a significant increase in cardiovascular/atherosclerosis-related proteins compared with controls.7 However, whether children with AD have altered hemostatic function remains unclear. Moreover, despite these associations in adults, the causal relationship and directionality of the association between AD and clotting has not been investigated. Herein, we investigate clotting's role in AD using the Mechanisms of Progression of Atopic Dermatitis to Asthma in Children (MPAACH) cohort8 and an AD mouse model. Briefly, the MPAACH cohort is a longitudinal cohort of children with AD. Children with no history of AD or other allergic disease are recruited in parallel. The MPAACH cohort biorepository includes extensive demographic, clinical, and biomarker data including skin AD severity and transepidermal water loss (TEWL) measurements from lesional (active or historical) and never-lesional (skin never affected by AD and ≥ 10 cm away from a site of active or historical lesions), along with extensive banked longitudinal biospecimens. A full, detailed description of MPAACH and the methods used in this study8 are found within the supplementary materials. We analyzed plasma samples that had not undergone multiple freeze–thaw cycles from 99 MPAACH children. Except for age, the subset and full MPAACH cohort were similar (Table S1). Control participants without AD were recruited in parallel with the MPAACH cohort and were also included in this substudy (Table S2A). The controls were similar to the cases in terms of sex and race, but they were younger than the cases. Due to the noted age difference between the cases and controls, we restricted the analysis to children under 1 year of age including additional demographic comparisons in the <1 year of age group (Table S2B). In this age-controlled analysis, we found that children with AD had no difference in time to clot initiation (Figure 1A p = .98), but children with AD had increased peak thrombin (Figure 1B p = 8.5e-07), decreased time to peak thrombin (Figure 1C p = .04), increased rate of thrombin generation/velocity index (Figure 1D p = 8e-04), and increased total thrombin generated (Figure 1E p = .002) compared to children without AD. Insufficient children without AD that are over the age of 1 as part of MPAACH, therefore a similar analysis is not presented for this subset. Moreover, compared with controls, in the entire cohort, children with AD had shorter clot initiation phase time (p < .001, Figure S1A), higher peak thrombin (p = .0015, Figure S1B), shorter peak thrombin time (p < .001, Figure S1C), and faster thrombin generation rate (p = .003, Figure S1D). Total thrombin generation did not differ between groups (Figure S1E). Children with moderate-to-severe AD had increased total thrombin generation compared to those with mild disease (p = .043, Figure S1A). Lastly, in all children with AD, increased total plasma thrombin was associated with lesional TEWL (β = .30, p = .001, Figure S1F) and never-lesional TEWL (β = .17, p = .02, Figure S1G).These differences reflect a significant increase in the thrombin amplification phase of thrombin generation in children with AD under 1 year of age. Thrombin cleaves fibrinogen to initiate fibrin clot formation.9 Total plasma fibrinogen was inversely associated with lesional TEWL (β = −.1, p = .04). Plasma fibrinogen was not associated with other AD biomarkers (Table S3). Platelet count was not associated with lesional or never-lesional TEWL (Table S3). Lastly, two common prothrombotic variants, factor V Leiden (rs6025) and prothrombin G20210A (rs1799963) frequencies in the children with AD were similar to the general population (Table S4). To determine if thrombin is mechanistically coupled to AD pathogenesis, mice were given chow with dabigatran (7.5 mg/kg of chow), a direct thrombin inhibitor,10 or control diet. Mice (n = 40) underwent three repeated cutaneous allergen patches to induce an AD-like phenotype.11 After three patches, mice receiving dabigatran had decreased TEWL (p = .03) compared with control (Figure 2A) and a substantial decrease in symptom score (p = .07, Figure 2B). To determine whether fibrinogen impacted AD development we performed immunohistochemistry and ELISA-based quantification of fibrinogen on mouse skin. Dabigatran administration resulted in decreased fibrinogen staining (Figure 2C) and significantly lower fibrinogen (p = .0025, Figure 2D) in the Asp group. To determine fibrinogen's direct impact on AD pathogenesis, we used mice with complete (Fib−/−) and partial (Fib+/−) fibrinogen deficiency and wildtype controls (FibWT) in our AD model (n = 30). After three patches, Fib−/− and Fib+/− mice had decreased disease development evidenced by decreased TEWL (p = .0079 and p = .016, respectively; Figure 2E) and median disease severity (p = .0092 and p = .014, respectively; Figure 2F) compared with WT. Notably, Fib+/− mice carry fibrinogen levels that are half of normal, which does not impair hemostasis, but resulted in significant attenuation in AD severity. Our studies are the first to establish associations of pediatric AD with thrombin and contributions of thrombin and fibrinogen to AD, demonstrating that hemostatic system regulation in childhood AD is pathologically altered. Based on our findings we suggest a mechanistic model whereby increased thrombin generation leads to increased conversion of fibrinogen to fibrin, which drives AD pathogenesis and is associated with skin barrier dysfunction and disease severity in pediatric AD. In addition thrombin may act in a synergic role with other skin barrier damage mediators, thus blocking thrombin contributes to decreased barrier damage. Our findings are novel and provide insights into the link between clotting and AD in adults.3 Previously, directionality of associations between clotting and AD remained speculative, our findings provide initial support that thrombin and fibrinogen may contribute to AD pathogenesis. Our results suggest that thrombin and fibrinogen could be used as novel targets for therapy in in patients with AD. Future investigations should elucidate the role thrombin mediated conversion of fibrinogen to fibrin as well as clotting mechanisms impact AD pathogenesis. This work is funded by NIH U19AI070235 (GKKH, JBM, LJM) and 5K12HD28827–27 (MGS) and the American Academy of Allergy, Asthma, and Immunology Foundation (Faculty Development Award, MGS). Alyssa Filuta: Investigation; Peter Amezcua: Investigation; Brandy Ruff: Investigation; Jocelyn M. Biagini, PhD: Resources, Writing–Review and Editing; John Kroner: Resources, Data Curation; Hua He: Data curation, Writing–Original Draft, Visualization; Eric Brandt: Investigation; Lisa J. Martin: Validation; Data Curation, Visualization, Writing–Review and Editing; Joseph S. Palumbo: Conceptualization, Resources, Writing–Review and Editing; Gurjit K. Khurana Hershey: Conceptualization, Resources, Writing–Review and Editing, Funding Acquisition; Michael G. Sherenian: Conceptualization, Supervision, Project administration; Writing–Original Draft, Review, and Editing, Resources, Methodology, Funding Acquisition. We would like to acknowledge and thank the participants and their families who volunteered to be a part of the MPAACH study. No authors have competing interests to disclose. All data may be made available upon request, there are no restrictions on materials or data for the experiments conducted in this study. Data S1: Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

Background: A major route of sensitization to food allergen is through an impaired skin barrier. IL-33 and thymic stromal lymphopoietin (TSLP) have both been implicated in epicutaneous sensitization and food allergy, albeit in different murine models. Objective: We assessed the respective contributions of TSLP and IL-33 to the development of atopic dermatitis (AD) and subsequent food allergy in TSLP and IL-33 receptor (ST2)-deficient mice using an AD model that does not require tape stripping.Method: TSLP receptor (TSLPR)-/-, ST2-/-, and BALB/cJ control mice were exposed to 3 weekly epicutaneous skin patches of one of saline, ovalbumin (OVA), or a combination of OVA and Aspergillus fumigatus (ASP), followed by repeated intragastric OVA challenges and development of food allergy.Results: ASP and/or OVA patched, but not OVA-alone patched, BALB/cJ mice developed an AD-like skin phenotype. However, epicutaneous OVA sensitization occurred in OVA patched mice and was decreased in ST2-/-mice, resulting in lower intestinal mast cell degranulation and accumulation, as well as OVA -induced diarrhea occurrences on intragastric OVA challenges. In TSLPR-/-mice, intestinal mast cell accumulation was abrogated, and no diarrhea was observed. AD was significantly milder in OVA + ASP patched TSLPR-/-mice compared to wild type and ST2-/-mice. Accordingly, intestinal mast cell accumulation and degranulation were impaired in OVA + ASP patched TSLPR-/-mice compared to wild type and ST2-/-mice, protecting TSLPR-/-mice from developing allergic diarrhea.Conclusion: Epicutaneous sensitization to food allergen and development of food allergy can occur without skin inflammation and is partly mediated by TSLP, suggesting that prophylactic targeting of TSLP may be useful in mitigating the development of AD and food allergy early in life in at-risk infants.

Atopic dermatitis (AD) is a chronic inflammatory skin disease, characterized by clinical manifestations of dry skin, chronic eczema and severe itching, often combined with asthma and allergic rhinitis. AD can affect as many as 20% of children and 1–3% of adults. Skin barrier dysfunction, inflammatory and immunological abnormalities have long been noted in patients with AD. However, conflicting evidence suggests that enhanced innervation of sensory neurons may contribute to disease pathology in AD. To better understand this phenomenon, we investigated the role of sensory neurons and their relationship with mast cells in itch sensations at different ages of AD initiation. AD was induced by epicutaneous exposure of extracts from Aspergillus fumigatus (ASP) in a sensory neuron reporter animal (Pirt;tdTomato) initiated either at postnatal day14 (P14) or P35. Mice were exposed to ASP extract (or saline) for 3 weeks via patches on the back skin. We performed immunohistochemistry on back skin for mast cell protease 6 (MCPT-6), assessed trans-epidermal water loss (TEWL), and determined scratching behaviors using an artificial intelligence system. We demonstrated an increased total innervation in the skin of ASP treated mice started at P35 but not at P14. There was significant increase in TEWL and thickness of epidermal layers in both AD groups. However, increased thickening at P35 was 2x greater than that observed at P14. This resulted in decreased sensory innervation density specifically in the epidermal layer at P35. We also found increased MCs at the epidermal/dermal border in both ASP treated groups. Surprisingly, ASP treated mice at P14 did not develop robust scratching behaviors as observed at P35. Results suggest that distinctions in innervation density in the epidermis vs dermis in association with MCs may underlie scratching behaviors that contribute to AD progression and could reveal novel treatment strategies for AD. Grant support from 5U19 AI70235-15 R01NS105715 R01NS113965. Atopic dermatitis (AD) is a chronic inflammatory skin disease, characterized by clinical manifestations of dry skin, chronic eczema and severe itching, often combined with asthma and allergic rhinitis. AD can affect as many as 20% of children and 1–3% of adults. Skin barrier dysfunction, inflammatory and immunological abnormalities have long been noted in patients with AD. However, conflicting evidence suggests that enhanced innervation of sensory neurons may contribute to disease pathology in AD. To better understand this phenomenon, we investigated the role of sensory neurons and their relationship with mast cells in itch sensations at different ages of AD initiation. AD was induced by epicutaneous exposure of extracts from Aspergillus fumigatus (ASP) in a sensory neuron reporter animal (Pirt;tdTomato) initiated either at postnatal day14 (P14) or P35. Mice were exposed to ASP extract (or saline) for 3 weeks via patches on the back skin. We performed immunohistochemistry on back skin for mast cell protease 6 (MCPT-6), assessed trans-epidermal water loss (TEWL), and determined scratching behaviors using an artificial intelligence system. We demonstrated an increased total innervation in the skin of ASP treated mice started at P35 but not at P14. There was significant increase in TEWL and thickness of epidermal layers in both AD groups. However, increased thickening at P35 was 2x greater than that observed at P14. This resulted in decreased sensory innervation density specifically in the epidermal layer at P35. We also found increased MCs at the epidermal/dermal border in both ASP treated groups. Surprisingly, ASP treated mice at P14 did not develop robust scratching behaviors as observed at P35. Results suggest that distinctions in innervation density in the epidermis vs dermis in association with MCs may underlie scratching behaviors that contribute to AD progression and could reveal novel treatment strategies for AD. Grant support from 5U19 AI70235-15 R01NS105715 R01NS113965.

Adults with atopic dermatitis (AD) have an increased risk for thromboembolic events. However, the interplay between pediatric AD and clotting and the underlying mechanisms remain under investigated. To assess thrombin in pediatric AD we performed a plasma thrombin generation assay on samples from the Mechanisms of Progression of Atopic Dermatitis to Asthma in Children (MPAACH) cohort. We used disease severity and transepidermal water loss (TEWL) as AD measures. To determine whether thrombin contributes mechanistically to AD pathogenesis we used dabigatran, a direct thrombin inhibitor, in our established murine model of AD. To determine whether fibrinogen, a major thrombin substrate, impacts AD pathogenesis we used mice with fibrinogen deficiency. All mice underwent objective disease severity assessments and TEWL evaluations. Analyses were performed using GraphPad Prism, version 9.0.0. In children, we found that increased total thrombin generation was associated with moderate-to-severe AD (P-value=0.02) and increased TEWL at lesional (P-value=0.001) and never-lesional (P-value=0.02) sites; moreover, increased peak thrombin generation was associated with moderate-to-severe AD (P-value=0.05) and increased lesional TEWL (P-value=0.01). Further, thrombin inhibition attenuated disease development as evidenced by decreased disease severity (P-value<0.001) and TEWL (P-value<0.001) in a murine AD model. Last, we found that complete and partial fibrinogen deficiency markedly attenuated disease development as evidenced by decreased disease severity (P-value<0.0001) and TEWL (P-value<0.0001) in mice. TEWL in complete and partial fibrinogen deficient mice was comparable to unchallenged controls. Our combined human and murine findings highly suggest a key mechanistic role for thrombin and fibrinogen and their contribution to AD in children.

Adults with atopic dermatitis (AD) have an increased risk for thromboembolic events. While coagulation is critical for almost all healing responses, dysregulated clotting and fibrin(ogen) deposition lead to inflammation that exacerbates tissue damage and impairs tissue repair. While plasma fibrin(ogen) has been associated with established asthma, fibrin(ogen)'s mechanistic role in AD pathogenesis and allergic sensitization has not been investigated. Here we show that fibrinogen plays a critical role in a murine model atopic dermatitis pathogenesis by markedly attenuating disease severity, barrier dysfunction, and allergic sensitization.

Abstract Background: A major route of sensitization to food allergen is through an impaired skin barrier. IL33 and TSLP have both been implicated in epicutaneous sensitization and development of food allergy. Method: We assessed the respective contributions of TSLP and IL33 to the development of food allergy in TSLP and IL33 receptor deficient mice following epicutaneous food allergen sensitization, where mice were exposed to thrice weekly skin patches of either saline, OVA or a combination of OVA and Aspergillus fumigatus extract (ASP). Results: ASP+OVA-patched but not OVA-patched mice developed an atopic dermatitis (AD)-like skin phenotype, while sensitization to OVA occurred in both OVA and OVA+ASP patched mice. OVA-specific IgE levels were significantly lower in OVA(±ASP)-patched TSLPR−/− mice compared to wild type mice and ST2−/− mice. Repeated intragastric challenges with 50mg of OVA, induced intestinal accumulation of mast cells but not ILC2s in OVA(±ASP)-patched WT and ST2−/− mice, and to a much lesser degree in TSLPR−/− mice. While OVA-induced mast cell degranulation assessed by measuring MCPT1 blood levels and the development of food allergy (diarrhea occurrences) was observed in almost all WT mice and two thirds of OVA+ASP patched ST2−/− mice, all TSLPR−/− mice were protected from developing food allergy and had significantly lower MCPT1 levels. Conclusion: Epicutaneous sensitization to food allergen and subsequent development of food allergy does not require atopic dermatitis skin lesions and is dependent on TSLP, suggesting that prophylactic targeting of TSLP may be useful in mitigating the development of food allergy early in life.

Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

Single nucleotide polymorphisms (SNPs) in the gene encoding kinesin family member 3A, KIF3A, have been associated with atopic dermatitis (AD), a chronic inflammatory skin disorder. We find that KIF3A SNP rs11740584 and rs2299007 risk alleles create cytosine-phosphate-guanine sites, which are highly methylated and result in lower KIF3A expression, and this methylation is associated with increased transepidermal water loss (TEWL) in risk allele carriers. Kif3aK14∆/∆ mice have increased TEWL, disrupted junctional proteins, and increased susceptibility to develop AD. Thus, KIF3A is required for skin barrier homeostasis whereby decreased KIF3A skin expression causes disrupted skin barrier function and promotes development of AD.

BACKGROUND:Exposure to traffic pollution, notably diesel exhaust particles (DEP), increases risk for asthma and asthma exacerbations. The contribution of cytokines generated by stressed lung epithelial cells (IL25, IL33, TSLP) to DEP-induced asthma severity remains poorly understood.METHODS:BALB/c mice were exposed intratracheally once to DEP or 9 times over 3-weeks to either saline, DEP, and/or house dust mite extract (HDM). Airway hyper-responsiveness (AHR), pulmonary inflammation, and T-cell subsets were assessed 24 hours after the last exposure in mice sufficient and deficient for the IL33 receptor ST2.RESULTS:DEP exposure induces oxidative stress, IL6, neutrophils and pulmonary accumulation of IL33, but not IL25 or TSLP or other features of allergic disease. When mice are co-exposed to DEP and low doses of HDM, DEP increases IL33 lung levels and Th2 responses. ST2 deficiency partially protected mice from HDM + DEP induced AHR in association with decreased type 2 inflammation and lung levels of IL5+ IL17A+ co-producing T-cells. Upon in vitro HDM challenge of lung cells from HDM ± DEP exposed ST2-/- mice, secretion of IL5, IL13, IL6 and IL17A was abrogated by a mechanism involving IL33 signaling in both dendritic cells and T-cells. HDM + DEP exposed bone marrow derived dendritic cells and IL33 pulsed BMDC promote a mixed Th2/Th17 response that was dependent on ST2 expression by CD4+ T-cells.CONCLUSION:IL33 contributes to DEP mediated increase in allergen-induced Th2 inflammation and AHR in a mouse model of severe steroid resistant asthma, potentially through the accumulation of pathogenic IL5+ IL17A+ CD4+ effector T-cells.

The authors declare no conflict of interest.

Previous studies have suggested a role for Tet1 in the pathogenesis of childhood asthma. However, how Tet1 contributes to asthma remains unknown. Here we used mice deficient for Tet1 in a well-established model of allergic airway inflammation and demonstrated that loss of Tet1 increased disease severity including airway hyperresponsiveness and lung eosinophilia. Increased expression of Muc5ac , Il1 3 , Il33 , Il 1 7a , Egfr , and Tff 2 were observed in HDM-challenged Tet1-deficient mice compared to Tet1 +/+ littermates. Further, transcriptomic analysis of lung RNA followed by pathway and protein network analysis showed that the IFN signaling pathway was significantly upregulated and the aryl hydrocarbon receptor (AhR) pathway was significantly downregulated in HDM-challenged Tet1 −/− mice. This transcriptional regulation of the IFN and AhR pathways by Tet1 was also present in human bronchial epithelial cells at base line and following HDM challenges. Genes in these pathways were further associated with changes in DNA methylation, predicted binding of transcriptional factors with relevant functions in their promoters, and the presence of histone marks generated by histone enzymes that are known to interact with Tet1. Collectively, our data suggest that Tet1 inhibits HDM-induced allergic airway inflammation by direct regulation of the IFN and AhR pathways.

BACKGROUND:GTPase of immunity-associated protein 5 (GIMAP5) is essential for lymphocyte homeostasis and survival. Recently, human GIMAP5 single nucleotide polymorphisms have been linked to an increased risk for asthma, whereas loss of Gimap5 in mice has been associated with severe CD4+ T cell-driven immune pathology. OBJECTIVE:We sought to identify the molecular and cellular mechanisms by which Gimap5 deficiency predisposes to allergic airway disease. METHODS:CD4+ T-cell polarization and development of pathogenic CD4+ T cells were assessed in Gimap5-deficient mice and a human patient with a GIMAP5 loss-of-function (LOF) mutation. House dust mite-induced airway inflammation was assessed by using a complete Gimap5 LOF (Gimap5sph/sph) and conditional Gimap5fl/flCd4Cre/ert2 mice. RESULTS:GIMAP5 LOF mutations in both mice and human subjects are associated with spontaneous polarization toward pathogenic TH17 and TH2 cells in vivo. Mechanistic studies in vitro reveal that impairment of Gimap5-deficient TH cell differentiation is associated with increased DNA damage, particularly during TH1-polarizing conditions. DNA damage in Gimap5-deficient CD4+ T cells could be controlled by TGF-β, thereby promoting TH17 polarization. When challenged with house dust mite in vivo, Gimap5-deficient mice displayed an exacerbated asthma phenotype (inflammation and airway hyperresponsiveness), with increased development of TH2, TH17, and pathogenic TH17/TH2 cells. CONCLUSION:Activation of Gimap5-deficient CD4+ T cells is associated with increased DNA damage and reduced survival that can be overcome by TGF-β. This leads to selective survival of pathogenic TH17 cells but also TH2 cells in human subjects and mice, ultimately promoting allergic airway disease.

Abstract Rationale Exposure to traffic pollution, notably diesel exhaust particles (DEP), promote asthma exacerbations and a mixed Th2/Th17 response. The contribution of TSLP and IL33, generated by sressed lung epithelial cells, to DEP-induced asthma exacerbations remains poorly understood. Method We used TSLP receptor deficient mice, mice lacking the IL33 receptor (ST2) and Balb/c control mice and exposed them 9 times over a 3-week period to saline, house dust mite extract (HDM; 10ug) and/or DEP (100ug). Seven weeks later some mice received a single HDM challenge to assess memory responses. Airway hyper-responsiveness (AHR), BALF inflammation and lung T-cells and ILC subsets were assessed after primary and recall responses. Results DEP co-exposure with HDM exacerbates HDM-induced AHR and Th2 responses. AHR was similar between TSLPR deficient mice and control mice exposed to HDM+DEP. In contrast, AHR was significantly decreased in ST2-deficient mice compared to wild type mice. The impact of ST2 deficiency on lung accumulation of ILC2 and Th2 cells was modest. Following HDM+DEP co-exposures, decreases in ILC3, γδ T-cells and Th17 cells were observed in the lungs of ST2 deficient mice compared to control mice. The in vivo HDM recall response recapitulated the impaired AHR associated with decreases in IL17A secreting cells observed after the primary response in HDM+DEP exposed ST2 deficient mice. Finally, a decrease in lung levels of pathogenic IL5/IL13/IL17A producing CD4+ T-cells was observed after the primary and recall responses. Conclusions IL33 but not TSLP contributes to DEP-induced asthma exacerbations by affecting the accumulation of innate and adaptive type 2 and type 17 cells, notably pathogenic Th2/Th17 cells.

Mucosal barriers are densely colonized by pathobiont microbes such as Candida albicans, capable of invasive disseminated infection. However, systemic infections occur infrequently in healthy individuals, suggesting that pathobiont commensalism may elicit host benefits. We show that intestinal colonization with C. albicans drives systemic expansion of fungal-specific Th17 CD4(+) T cells and IL-17 responsiveness by circulating neutrophils, which synergistically protect against C. albicans invasive infection. Protection conferred by commensal C. albicans requires persistent fungal colonization and extends to other extracellular invasive pathogens such as Staphylococcus aureus. However, commensal C. albicans does not protect against intracellular influenza virus infection and exacerbates allergic airway inflammation susceptibility, indicating that positively calibrating systemic Th17 responses is not uniformly beneficial. Thus, systemic Th17 inflammation driven by CD4(+) T cells responsive to tonic stimulation by commensal C. albicans improves host defense against extracellular pathogens, but with potentially harmful immunological consequences.

Background: Recent literature suggests that children who are vitamin D deficient are uniquely susceptible to the effects of traffic-related air pollution (TRAP) exposure. This is highly significant because large segments of the population reside in zones of high TRAP exposure. Objective: We sought to determine whether vitamin D supplementation mitigates the effect of TRAP exposure on asthma development, asthma exacerbation, and/or airway inflammation and to determine the timing of vitamin D supplementation that confers maximal health benefit. Methods: Using established mouse models of asthma, we examined the effect of prenatal and postnatal vitamin D supplementation on asthma development, as well as the utility of vitamin D as a treatment for established asthma in the context of diesel exhaust particle (DEP) exposure. Results: DEP and allergen coexposure resulted in increased airway hyperresponsiveness (AHR) and accumulation of pathogenic T(H)2/T(H)17 cells in the lungs of vitamin D-deficient mice compared with control mice. Prenatal and postnatal vitamin D supplementation significantly attenuated the development of AHR and decreased pulmonary accumulation of T(H)2/T(H)17 cells after coexposure to TRAP and allergen but not to allergen alone. Restoration of normal vitamin D status had no effect on AHR once asthma was already established. Conclusions: Our data establish that vitamin D confers protection against asthma development specifically in the context of TRAP exposure. Although vitamin D replacement did not reverse established asthma, restoration of normal vitamin D status in early life significantly attenuated the development of AHR in the setting of DEP-exacerbated allergic asthma and reduced numbers of lung T(H)2/T(H)17 cells, which portend the development of severe asthma.

Summary Background Asthma is an allergic airway inflammation‐driven disease that affects more than 300 million people world‐wide. Targeted therapies for asthma are largely lacking. Although asthma symptoms can be prevented from worsening, asthma development cannot be prevented. Cdc42 GTPase has been shown to regulate actin cytoskeleton, cell proliferation and survival. Objectives To investigate the role and targeting of Cdc42 in Th2 cell differentiation and Th2‐mediated allergic airway inflammation. Methods Post‐thymic Cdc42‐deficient mice were generated by crossing Cdc42 flox/flox mice with dLck icre transgenic mice in which Cre expression is driven by distal Lck promoter. Effects of post‐thymic Cdc42 deletion and pharmacological targeting Cdc42 on Th2 cell differentiation were evaluated in vitro under Th2‐polarized culture conditions. Effects of post‐thymic Cdc42 deletion and pharmacological targeting Cdc42 on allergic airway inflammation were evaluated in ovalbumin‐ and/or house dust mite‐induced mouse models of asthma. Results Post‐thymic deletion of Cdc42 led to reduced peripheral CD8 + T cells and attenuated Th2 cell differentiation, with no effect on closely related Th1, Th17 and induced regulatory T (iTreg) cells. Post‐thymic Cdc42 deficiency ameliorated allergic airway inflammation. The selective inhibition of Th2 cell differentiation by post‐thymic deletion of Cdc42 was recapitulated by pharmacological targeting of Cdc42 with CASIN, a Cdc42 activity‐specific chemical inhibitor. CASIN also alleviated allergic airway inflammation. CASIN‐treated Cdc42‐deficient mice showed comparable allergic airway inflammation to vehicle‐treated Cdc42‐deficient mice, indicative of negligible off‐target effect of CASIN. CASIN had no effect on established allergic airway inflammation. Conclusion and Clinical Relevance Cdc42 is required for Th2 cell differentiation and allergic airway inflammation, and rational targeting Cdc42 may serve as a preventive but not therapeutic approach for asthma control.

BACKGROUND:Allergic sensitization to fungi has been associated with asthma severity. As a result, it has been largely assumed that the contribution of fungi to allergic disease is mediated through their potent antigenicity.OBJECTIVE:We sought to determine the mechanism by which fungi affect asthma development and severity.METHODS:We integrated epidemiologic and experimental asthma models to explore the effect of fungal exposure on asthma development and severity.RESULTS:We report that fungal exposure enhances allergen-driven TH2 responses, promoting severe allergic asthma. This effect is independent of fungal sensitization and can be reconstituted with β-glucan and abrogated by neutralization of IL-17A. Furthermore, this severe asthma is resistant to steroids and characterized by mixed TH2 and TH17 responses, including IL-13+IL-17+CD4+ double-producing effector T cells. Steroid resistance is dependent on fungus-induced TH17 responses because steroid sensitivity was restored in IL-17rc-/- mice. Similarly, in children with asthma, fungal exposure was associated with increased serum IL-17A levels and asthma severity.CONCLUSION:Our data demonstrate that fungi are potent immunomodulators and have powerful effects on asthma independent of their potential to act as antigens. Furthermore, our results provide a strong rationale for combination treatment strategies targeting IL-17A for this subgroup of fungus-exposed patients with difficult-to-treat asthma.