RESUMO
Epithelial cells orchestrate pulmonary homeostasis and pathogen defense and play a crucial role in the initiation of allergic immune responses. Maintaining the balance between homeostasis and inappropriate immune activation and associated pathology is particularly complex at mucosal sites that are exposed to billions of potentially antigenic particles daily. We demonstrated that epithelial cell-derived cytokine TGF-ß had a central role in the generation of the pulmonary immune response. Mice that specifically lacked epithelial cell-derived TGF-ß1 displayed a reduction in type 2 innate lymphoid cells (ILCs), resulting in suppression of interleukin-13 and hallmark features of the allergic response including airway hyperreactivity. ILCs in the airway lumen were primed to respond to TGF-ß by expressing the receptor TGF-ßRII and ILC chemoactivity was enhanced by TGF-ß. These data demonstrate that resident epithelial cells instruct immune cells, highlighting the central role of the local environmental niche in defining the nature and magnitude of immune reactions.
Assuntos
Células Epiteliais/imunologia , Imunidade Inata/imunologia , Pulmão/imunologia , Linfócitos/imunologia , Fator de Crescimento Transformador beta1/imunologia , Animais , Células Cultivadas , Interleucina-13/imunologia , Camundongos , Proteínas Serina-Treonina Quinases/imunologia , Receptor do Fator de Crescimento Transformador beta Tipo II , Receptores de Fatores de Crescimento Transformadores beta/imunologia , Hipersensibilidade Respiratória/imunologiaRESUMO
BACKGROUND: Early life represents a major risk window for asthma development. However, the mechanisms controlling the threshold for establishment of allergic airway inflammation in early life are incompletely understood. Airway macrophages (AMs) regulate pulmonary allergic responses and undergo TGF-ß-dependent postnatal development, but the role of AM maturation factors such as TGF-ß in controlling the threshold for pathogenic immune responses to inhaled allergens remains unclear. OBJECTIVE: Our aim was to test the hypothesis that AM-derived TGF-ß1 regulates pathogenic immunity to inhaled allergen in early life. METHODS: Conditional knockout (Tgfb1ΔCD11c) mice, with TGF-ß1 deficiency in AMs and other CD11c+ cells, were analyzed throughout early life and following neonatal house dust mite (HDM) inhalation. The roles of specific chemokine receptors were determined by using in vivo blocking antibodies. RESULTS: AM-intrinsic TGF-ß1 was redundant for initial population of the neonatal lung with AMs, but AMs from Tgfb1ΔCD11c mice failed to adopt a mature homeostatic AM phenotype in the first weeks of life. Evidence of constitutive TGF-ß1 signaling was also observed in pediatric human AMs. TGF-ß1-deficient AMs expressed enhanced levels of monocyte-attractant chemokines, and accordingly, Tgfb1ΔCD11c mice exposed to HDM throughout early life accumulated CCR2-dependent inflammatory CD11c+ mononuclear phagocytes into the airway niche that expressed the proallergic chemokine CCL8. Tgfb1ΔCD11c mice displayed augmented TH2, group 2 innate lymphoid cell, and airway remodeling responses to HDM, which were ameliorated by blockade of the CCL8 receptor CCR8. CONCLUSION: Our results highlight a causal relationship between AM maturity, chemokines, and pathogenic immunity to environmental stimuli in early life and identify TGF-ß1 as a key regulator of this.
Assuntos
Alérgenos/imunologia , Macrófagos Alveolares/imunologia , Fator de Crescimento Transformador beta1/imunologia , Animais , Quimiocinas/imunologia , Hipersensibilidade/imunologia , Pulmão/imunologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pyroglyphidae/imunologia , Fator de Crescimento Transformador beta1/genéticaRESUMO
BACKGROUND: Although originally defined as a type 2 (T2) immune-mediated condition, non-T2 cytokines, such as IFN-γ and IL-17A, have been implicated in asthma pathogenesis, particularly in patients with severe disease. IL-10 regulates TH cell phenotypes and can dampen T2 immunity to allergens, but its functions in controlling non-T2 cytokine responses in asthmatic patients are unclear. OBJECTIVE: We sought to determine how IL-10 regulates the balance of TH cell responses to inhaled allergen. METHODS: Allergic airway disease was induced in wild-type, IL-10 reporter, and conditional IL-10 or IL-10 receptor α (IL-10Rα) knockout mice by means of repeated intranasal administration of house dust mite (HDM). IL-10 and IFN-γ signaling were disrupted by using blocking antibodies. RESULTS: Repeated HDM inhalation induced a mixed IL-13/IL-17A response and accumulation of IL-10-producing forkhead box P3-negative effector CD4+ T cells in the lungs. Ablation of T cell-derived IL-10 increased the IFN-γ and IL-17A response to HDM, reducing IL-13 levels and airway eosinophilia without affecting IgE levels or airway hyperresponsiveness. The increased IFN-γ response could be recapitulated by IL-10Rα deletion in CD11c+ myeloid cells or local IL-10Rα blockade. Disruption of the T cell-myeloid IL-10 axis resulted in increased pulmonary monocyte-derived dendritic cell numbers and increased IFN-γ-dependent expression of CXCR3 ligands by airway macrophages, which is suggestive of a feedforward loop of TH1 cell recruitment. Augmented IFN-γ responses in the HDM allergic airway disease model were accompanied by increased disruption of airway epithelium, which was reversed by therapeutic blockade of IFN-γ. CONCLUSIONS: IL-10 from effector T cells signals to CD11c+ myeloid cells to suppress an atypical and pathogenic IFN-γ response to inhaled HDM.
Assuntos
Asma/imunologia , Interferon gama/imunologia , Interleucina-10/imunologia , Células Mieloides/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Alérgenos/imunologia , Animais , Modelos Animais de Doenças , Feminino , Hipersensibilidade/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pyroglyphidae/imunologiaRESUMO
Rationale: Idiopathic pulmonary fibrosis (IPF) is a devastating progressive disease with limited therapeutic options. Airway macrophages (AMs) are key components of the defense of the airways and are implicated in the pathogenesis of IPF. Alterations in iron metabolism have been described during fibrotic lung disease and in murine models of lung fibrosis. However, the role of transferrin receptor 1 (CD71)-expressing AMs in IPF is not known. Objectives: To assess the role of CD71-expressing AMs in the IPF lung. Methods: We used multiparametric flow cytometry, gene expression analysis, and phagocytosis/transferrin uptake assays to delineate the role of AMs expressing or lacking CD71 in the BAL of patients with IPF and of healthy control subjects. Measurements and Main Results: There was a distinct increase in proportions of AMs lacking CD71 in patients with IPF compared with healthy control subjects. Concentrations of BAL transferrin were enhanced in IPF-BAL, and furthermore, CD71- AMs had an impaired ability to sequester transferrin. CD71+ and CD71- AMs were phenotypically, functionally, and transcriptionally distinct, with CD71- AMs characterized by reduced expression of markers of macrophage maturity, impaired phagocytosis, and enhanced expression of profibrotic genes. Importantly, proportions of AMs lacking CD71 were independently associated with worse survival, underlining the importance of this population in IPF and as a potential therapeutic target. Conclusions: Taken together, these data highlight how CD71 delineates AM subsets that play distinct roles in IPF and furthermore show that CD71- AMs may be an important pathogenic component of fibrotic lung disease.
Assuntos
Antígenos CD/metabolismo , Fibrose Pulmonar Idiopática/metabolismo , Macrófagos Alveolares/metabolismo , Fagocitose , Receptores da Transferrina/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/citologia , Estudos de Casos e Controles , Feminino , Citometria de Fluxo , Perfilação da Expressão Gênica , Humanos , Ferro/metabolismo , Masculino , Pessoa de Meia-Idade , Prognóstico , Reação em Cadeia da Polimerase em Tempo Real , Taxa de Sobrevida , Transferrina/metabolismo , Adulto JovemRESUMO
BACKGROUND: Allergic asthma is characterized by chronic inflammation and remodelling of the airways, associated with dysregulated type 2 immune responses and allergen-specific IgE. T follicular helper cells (TFH ) are crucial in T-dependent B-cell responses and have been implicated in allergic airway disease (AAD). TFH , unlike other CD4+ T cells, are uniquely reliant on continuous ICOS signalling to maintain their phenotype after T-cell priming; therefore, disrupting this signal can impair TFH responses. However, the contribution of TFH to disease during chronic aero-allergen exposure and the therapeutic potential of targeting these cells have not been evaluated. METHODS: To establish AAD, female BALB/c mice were repeatedly exposed to house dust mite or Alternaria alternata three times a week for up to 5 weeks. To examine the impact of TFH on AAD, mice were allergen exposed for 5 weeks and co-administered anti-ICOS Ligand-targeted antibodies, three times a week for the last 2 weeks. RESULTS: TFH were first observed in the lung-draining lymph nodes and with further exposure were also found locally within the lungs. TFH accumulated with sustained allergen exposure, alongside germinal centre (GC) B cells. Blockade of ICOS signalling after AAD establishment successfully depleted TFH but did not affect the differentiation of other CD4+ T-cell subsets. This reduced GC responses, allergen-specific IgE, inflammation, pulmonary IL-13 and airway hyper-responsiveness. CONCLUSIONS: TFH are crucial in the regulation of AAD and the ICOS/ICOS-L pathway could represent a novel therapeutic target in allergic asthma.
Assuntos
Asma/tratamento farmacológico , Ligante Coestimulador de Linfócitos T Induzíveis/antagonistas & inibidores , Proteína Coestimuladora de Linfócitos T Induzíveis/antagonistas & inibidores , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Asma/patologia , Linfócitos B/imunologia , Feminino , Centro Germinativo/imunologia , Centro Germinativo/patologia , Ligante Coestimulador de Linfócitos T Induzíveis/metabolismo , Proteína Coestimuladora de Linfócitos T Induzíveis/metabolismo , Pulmão/patologia , Camundongos , Camundongos Endogâmicos BALB C , Pyroglyphidae/imunologiaRESUMO
IL-8-dependent inflammation is a hallmark of host lung innate immunity to bacterial pathogens, yet in many human lung diseases, including chronic obstructive pulmonary disease, bronchiectasis, and pulmonary fibrosis, there are progressive, irreversible, pathological changes associated with elevated levels of IL-8 in the lung. To better understand the duality of IL-8-dependent host immunity to bacterial infection and lung pathology, we expressed human IL-8 transgenically in murine bronchial epithelium, and investigated the impact of overexpression on lung bacterial clearance, host immunity, and lung pathology and function. Persistent IL-8 expression in bronchial epithelium resulted in neutrophilia, neutrophil maturation and activation, and chemotaxis. There was enhanced protection against challenge with Pseudomonas aeruginosa, and significant changes in baseline expression of innate and adaptive immunity transcripts for Ccl5, Tlr6, IL-2, and Tlr1. There was increased expression of Tbet and Foxp3 in response to the Pseudomonas antigen OprF, indicating a regulatory T-cell phenotype. However, this enhanced bacterial immunity came at a high price of progressive lung remodeling, with increased inflammation, mucus hypersecretion, and fibrosis. There was increased expression of Ccl3 and reduced expression of Claudin 18 and F11r, with damage to epithelial organization leading to leaky tight junctions, all of which resulted in impaired lung function with reduced compliance, increased resistance, and bronchial hyperreactivity as measured by whole-body plethysmography. These results show that IL-8 overexpression in the bronchial epithelium benefits lung immunity to bacterial infection, but specifically drives lung damage through persistent inflammation, lung remodeling, and damaged tight junctions, leading to impaired lung function.
Assuntos
Imunidade Inata/imunologia , Interleucina-8/metabolismo , Pulmão/imunologia , Pneumonia/patologia , Infecções por Pseudomonas/imunologia , Pseudomonas aeruginosa/imunologia , Fibrose Pulmonar/patologia , Animais , Doença Crônica , Humanos , Interleucina-8/genética , Pulmão/metabolismo , Pulmão/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Pneumonia/etiologia , Pneumonia/metabolismo , Infecções por Pseudomonas/metabolismo , Infecções por Pseudomonas/microbiologia , Fibrose Pulmonar/etiologia , Fibrose Pulmonar/metabolismoRESUMO
BACKGROUND: Genome-wide association studies have identified the ORM (yeast)-like protein isoform 3 (ORMDL3) gene locus on human chromosome 17q to be a highly significant risk factor for childhood-onset asthma. OBJECTIVE: We sought to investigate in vivo the functional role of ORMDL3 in disease inception. METHODS: An Ormdl3-deficient mouse was generated and the role of ORMDL3 in the generation of allergic airways disease to the fungal aeroallergen Alternaria alternata was determined. An adeno-associated viral vector was also used to reconstitute ORMDL3 expression in airway epithelial cells of Ormdl3 knockout mice. RESULTS: Ormdl3 knockout mice were found to be protected from developing allergic airways disease and showed a marked decrease in pathophysiology, including lung function and airway eosinophilia induced by Alternaria. Alternaria is a potent inducer of cellular stress and the unfolded protein response, and ORMDL3 was found to play a critical role in driving the activating transcription factor 6-mediated arm of this response through Xbp1 and downstream activation of the endoplasmic reticulum-associated degradation pathway. In addition, ORMDL3 mediated uric acid release, another marker of cellular stress. In the knockout mice, reconstitution of Ormdl3 transcript levels specifically in the bronchial epithelium resulted in reinstatement of susceptibility to fungal allergen-induced allergic airways disease. CONCLUSIONS: This study demonstrates that ORMDL3, an asthma susceptibility gene identified by genome-wide association studies, contributes to key pathways that promote changes in airway physiology during allergic immune responses.
Assuntos
Asma/imunologia , Proteínas de Membrana/imunologia , Alérgenos/imunologia , Alternaria/imunologia , Animais , Asma/fisiopatologia , Modelos Animais de Doenças , Pulmão/imunologia , Masculino , Proteínas de Membrana/genética , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mucosa Respiratória/imunologia , Ácido Úrico/metabolismoRESUMO
BACKGROUND: The mechanism underlying severe asthma with fungal sensitization (SAFS) is unknown. IL-33 is important in fungus-induced asthma exacerbations, but its role in fungal sensitization is unexplored. OBJECTIVE: We sought to determine whether fungal sensitization in children with severe therapy-resistant asthma is mediated by IL-33. METHODS: Eighty-two children (median age, 11.7 years; 63% male) with severe therapy-resistant asthma were included. SAFS (n = 38) was defined as specific IgE or skin prick test response positivity to Aspergillus fumigatus, Alternaria alternata, or Cladosporium herbarum. Clinical features and airway immunopathology were assessed. Chronic exposure to house dust mite and A alternata were compared in a neonatal mouse model. RESULTS: Children with SAFS had earlier symptom onset (0.5 vs 1.5 years, P = .006), higher total IgE levels (637 vs 177 IU/mL, P = .002), and nonfungal inhalant allergen-specific IgE. Significantly more children with SAFS were prescribed maintenance oral steroids (42% vs 14%, P = .02). SAFS was associated with higher airway IL-33 levels. In neonatal mice A alternata exposure induced higher serum IgE levels, pulmonary IL-33 levels, and IL-13(+) innate lymphoid cell (ILC) and TH2 cell numbers but similar airway hyperresponsiveness (AHR) compared with those after house dust mite exposure. Lung IL-33 levels, IL-13(+) ILC numbers, TH2 cell numbers, IL-13 levels, and AHR remained increased with inhaled budesonide during A alternata exposure, but all features were significantly reduced in ST2(-/-) mice lacking a functional receptor for IL-33. CONCLUSION: Pediatric SAFS was associated with more oral steroid therapy and higher IL-33 levels. A alternata exposure resulted in increased IL-33-mediated ILC2 numbers, TH2 cell numbers, and steroid-resistant AHR. IL-33 might be a novel therapeutic target for SAFS.
Assuntos
Antiasmáticos/uso terapêutico , Asma/tratamento farmacológico , Asma/imunologia , Budesonida/uso terapêutico , Interleucinas/imunologia , Micoses/tratamento farmacológico , Micoses/imunologia , Adolescente , Alternaria/imunologia , Animais , Animais Recém-Nascidos , Anticorpos Anti-Idiotípicos/uso terapêutico , Anticorpos Monoclonais Humanizados/uso terapêutico , Aspergillus fumigatus/imunologia , Asma/complicações , Asma/patologia , Criança , Cladosporium/imunologia , Modelos Animais de Doenças , Feminino , Humanos , Imunoglobulina E/genética , Imunoglobulina E/imunologia , Proteína 1 Semelhante a Receptor de Interleucina-1 , Interleucina-13/genética , Interleucina-13/imunologia , Interleucina-33 , Interleucinas/genética , Masculino , Camundongos , Micoses/complicações , Micoses/patologia , Omalizumab , Pyroglyphidae/química , Pyroglyphidae/imunologia , Receptores de Interleucina/deficiência , Receptores de Interleucina/genética , Receptores de Interleucina/imunologia , Índice de Gravidade de Doença , Testes Cutâneos , Células Th2/imunologia , Células Th2/patologiaRESUMO
Myofibroblast accumulation, subepithelial fibrosis, and vascular remodeling are complicating features of chronic asthma, but the mechanisms are not clear. Platelet-derived growth factors (PDGFs) regulate the fate and function of various mesenchymal cells and have been implicated as mediators of lung fibrosis. However, it is not known whether PDGF-BB signaling via PDGFRß, which is critical for the recruitment of pericytes to blood vessels, plays a role in airway remodeling in chronic asthma. In the present study, we used a selective PDGFRß inhibitor (CP-673451) to investigate the role of PDGFRß signaling in the development of airway remodeling and lung dysfunction in an established mouse model of house dust mite-induced chronic allergic asthma. Unexpectedly, we found that pharmacological inhibition of PDGFRß signaling in the context of chronic aeroallergen exposure led to exacerbated lung dysfunction and airway smooth muscle thickening. Further studies revealed that the inflammatory response to aeroallergen challenge in mice was associated with decreased PDGF-BB expression and the loss of pericytes from the airway microvasculature. In parallel, cells positive for pericyte markers accumulated in the subepithelial region of chronically inflamed airways. This process was exacerbated in animals treated with CP-673451. The results indicate that perturbed PDGF-BB/PDGFRß signaling and pericyte accumulation in the airway wall may contribute to airway remodeling in chronic allergic asthma.
Assuntos
Remodelação das Vias Aéreas , Asma/patologia , Pericitos/fisiologia , Resistência das Vias Respiratórias , Animais , Asma/fisiopatologia , Becaplermina , Benzimidazóis/farmacologia , Brônquios/imunologia , Brônquios/metabolismo , Brônquios/patologia , Doença Crônica , Modelos Animais de Doenças , Elasticidade , Feminino , Camundongos Endogâmicos C57BL , Músculo Liso/patologia , Proteínas Proto-Oncogênicas c-sis/metabolismo , Quinolinas/farmacologia , Receptor beta de Fator de Crescimento Derivado de Plaquetas/antagonistas & inibidores , Receptor beta de Fator de Crescimento Derivado de Plaquetas/metabolismoRESUMO
BACKGROUND: Current data concerning maternal paracetamol intake during pregnancy, or intake during infancy and risk of wheezing or asthma in childhood is inconclusive based on epidemiological studies. We have investigated whether there is a causal link between maternal paracetamol intake during pregnancy and lactation and the development of house dust mite (HDM) induced allergic airways disease (AAD) in offspring using a neonatal mouse model. METHODS: Pregnant mice were administered paracetamol or saline by oral gavage from the day of mating throughout pregnancy and/or lactation. Subsequently, their pups were exposed to intranasal HDM or saline from day 3 of life for up to 6 weeks. Assessments of airway hyper-responsiveness, inflammation and remodelling were made at weaning (3 weeks) and 6 weeks of age. RESULTS: Maternal paracetamol exposure either during pregnancy and/or lactation did not affect development of AAD in offspring at weaning or at 6 weeks. There were no effects of maternal paracetamol at any time point on airway remodelling or IgE levels. CONCLUSIONS: Maternal paracetamol did not enhance HDM induced AAD in offspring. Our mechanistic data do not support the hypothesis that prenatal paracetamol exposure increases the risk of childhood asthma.
Assuntos
Acetaminofen/farmacologia , Analgésicos não Narcóticos/farmacologia , Hiper-Reatividade Brônquica , Exposição Materna , Animais , Animais Recém-Nascidos , Hiper-Reatividade Brônquica/imunologia , Hiper-Reatividade Brônquica/fisiopatologia , Modelos Animais de Doenças , Feminino , Camundongos , GravidezRESUMO
BACKGROUND: The fungal allergen Alternaria alternata is implicated in severe asthma and rapid onset life-threatening exacerbations of disease. However, the mechanisms that underlie this severe pathogenicity remain unclear. OBJECTIVE: We sought to investigate the mechanism whereby Alternaria was capable of initiating severe, rapid onset allergic inflammation. METHODS: IL-33 levels were quantified in wild-type and ST2(-/-) mice that lacked the IL-33 receptor given inhaled house dust mite, cat dander, or Alternaria, and the effect of inhibiting allergen-specific protease activities on IL-33 levels was assessed. An exacerbation model of allergic airway disease was established whereby mice were sensitized with house dust mite before subsequently being challenged with Alternaria (with or without serine protease activity), and inflammation, remodeling, and lung function assessed 24 hours later. RESULTS: Alternaria, but not other common aeroallergens, possessed intrinsic serine protease activity that elicited the rapid release of IL-33 into the airways of mice through a mechanism that was dependent upon the activation of protease activated receptor-2 and adenosine triphosphate signaling. The unique capacity of Alternaria to drive this early IL-33 release resulted in a greater pulmonary inflammation by 24 hours after challenge relative to the common aeroallergen house dust mite. Furthermore, this Alternaria serine protease-IL-33 axis triggered a rapid, augmented inflammation, mucus release, and loss of lung function in our exacerbation model. CONCLUSION: Alternaria-specific serine protease activity causes rapid IL-33 release, which underlies the development of a robust TH2 inflammation and exacerbation of allergic airway disease.
Assuntos
Alternaria/imunologia , Alternariose/microbiologia , Proteínas Fúngicas/imunologia , Hipersensibilidade/microbiologia , Serina Proteases/imunologia , Trifosfato de Adenosina/metabolismo , Alternariose/imunologia , Animais , Antígenos de Dermatophagoides/imunologia , Modelos Animais de Doenças , Progressão da Doença , Feminino , Humanos , Hipersensibilidade/imunologia , Proteína 1 Semelhante a Receptor de Interleucina-1 , Interleucina-33 , Interleucinas/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Pyroglyphidae , Receptor PAR-2/metabolismo , Receptores de Interleucina/genética , Transdução de SinaisRESUMO
BACKGROUND: TH2 cytokines are not responsible for the ongoing symptoms and pathology in children with severe therapy-resistant asthma (STRA). IL-33 induces airway hyperresponsiveness, but its role in airway remodeling and steroid resistance is unknown. OBJECTIVE: We sought to investigate the relationship between IL-33 and airway remodeling in pediatric patients with STRA. METHODS: IL-33 levels were quantified in neonatal mice given inhaled house dust mite (HDM), and the effect of blocking IL-13 on remodeling and IL-33 levels was assessed. HDM-induced allergic airways disease (AAD) in neonatal ST2(-/-) mice lacking the IL-33 receptor was assessed, together with collagen production after IL-33 administration. The effect of steroid therapy on IL-33 levels in patients with neonatal AAD was explored. IL-33 expression was quantified in endobronchial biopsy (EB) specimens from children with STRA and related to remodeling, and collagen production by airway fibroblasts from pediatric patients stimulated with IL-33 and budesonide was quantified. RESULTS: Blocking IL-13 after AAD was established in neonatal mice and did not reduce remodeling or IL-33 levels; airway hyperresponsiveness was only partially reduced. IL-33 promoted collagen synthesis both from asthmatic fibroblasts from pediatric patients and after intranasal administration in mice. Increased cellular expression of IL-33, but not IL-13, was associated with increased reticular basement membrane thickness in EB specimens from children with STRA, whereas remodeling was absent in HDM-exposed ST2(-/-) mice. IL-33 levels were maintained, whereas IL-13 levels were abrogated by steroid treatment in neonatal HDM-exposed mice and in EB specimens from children with STRA. CONCLUSION: IL-33 is a relatively steroid-resistant mediator that promotes airway remodeling in patients with STRA and is an important therapeutic target.
Assuntos
Remodelação das Vias Aéreas , Asma/patologia , Hiper-Reatividade Brônquica/imunologia , Interleucinas/imunologia , Adolescente , Remodelação das Vias Aéreas/fisiologia , Animais , Animais Recém-Nascidos , Anti-Inflamatórios/uso terapêutico , Asma/tratamento farmacológico , Asma/imunologia , Hiper-Reatividade Brônquica/patologia , Budesonida/uso terapêutico , Criança , Colágeno/imunologia , Resistência a Medicamentos , Feminino , Glucocorticoides/uso terapêutico , Humanos , Interleucina-13/imunologia , Interleucina-33 , Interleucinas/administração & dosagem , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Pyroglyphidae/imunologiaRESUMO
Biomaterials that can improve the healing of articular cartilage lesions are needed. To address this unmet need, we developed novel 3D printed silica/poly(tetrahydrofuran)/poly(ε-caprolactone) (SiO2/PTHF/PCL-diCOOH) hybrid scaffolds. Our aim was to carry out essential studies to advance this medical device towards functional validation in pre-clinical trials. First, we show that the chemical composition, microarchitecture and mechanical properties of these scaffolds were not affected by sterilisation with gamma irradiation. To evaluate the systemic and local immunogenic reactivity of the sterilised 3D printed hybrid scaffolds, they were implanted subcutaneously into Balb/c mice. The scaffolds did not trigger a systemic inflammatory response over one week of implantation. The interaction between the host immune system and the implanted scaffold elicited a local physiological reaction with infiltration of mononuclear cells without any signs of a chronic inflammatory response. Then, we investigated how these 3D printed hybrid scaffolds direct chondrogenesis in vitro. Human bone marrow-derived mesenchymal stem/stromal cells (hBM-MSCs) seeded within the 3D printed hybrid scaffolds were cultured under normoxic or hypoxic conditions, with or without chondrogenic supplements. Chondrogenic differentiation assessed by both gene expression and protein production analyses showed that 3D printed hybrid scaffolds support hBM-MSC chondrogenesis. Articular cartilage-specific extracellular matrix deposition within these scaffolds was enhanced under hypoxic conditions (1.7 or 3.7 fold increase in the median of aggrecan production in basal or chondrogenic differentiation media). Our findings show that 3D printed SiO2/PTHF/PCL-diCOOH hybrid scaffolds have the potential to support the regeneration of cartilage tissue.
RESUMO
Allergic asthma generally starts during early life and is linked to substantial tissue remodeling and lung dysfunction. Although angiogenesis is a feature of the disrupted airway, the impact of allergic asthma on the pulmonary microcirculation during early life is unknown. Here, using quantitative imaging in precision-cut lung slices (PCLSs), we report that exposure of neonatal mice to house dust mite (HDM) extract disrupts endothelial cell/pericyte interactions in adventitial areas. Central to the blood vessel structure, the loss of pericyte coverage was driven by mast cell (MC) proteases, such as tryptase, that can induce pericyte retraction and loss of the critical adhesion molecule N-cadherin. Furthermore, spatial transcriptomics of pediatric asthmatic endobronchial biopsies suggests intense vascular stress and remodeling linked with increased expression of MC activation pathways in regions enriched in blood vessels. These data provide previously unappreciated insights into the pathophysiology of allergic asthma with potential long-term vascular defects.
Assuntos
Asma , Mastócitos , Humanos , Criança , Animais , Camundongos , Mastócitos/patologia , Pericitos/metabolismo , Células Endoteliais/metabolismo , Asma/patologia , Pulmão/patologia , Alérgenos , Pyroglyphidae , Modelos Animais de DoençasRESUMO
BACKGROUND: Overexpression of the transforming growth factor ß family signalling molecule smad2 in the airway epithelium provokes enhanced allergen-induced airway remodelling in mice, concomitant with elevated levels of interleukin (IL)-25. OBJECTIVE: We investigated whether IL-25 plays an active role in driving this airway remodelling. METHODS: Anti-IL-25 antibody was given to mice exposed to either inhaled house dust mite (HDM) alone, or in conjunction with an adenoviral smad2 vector which promotes an enhanced remodelling phenotype. RESULTS: Blocking IL-25 in allergen-exposed mice resulted in a moderate reduction in pulmonary eosinophilia and levels of T helper type 2 associated cytokines, IL-5 and IL-13. In addition, IL-25 neutralisation abrogated peribronchial collagen deposition, airway smooth muscle hyperplasia and airway hyperreactivity in control mice exposed to HDM and smad2-overexpressing mice. IL-25 was shown to act directly on human fibroblasts to induce collagen secretion. Recruitment of endothelial progenitor cells to the lung and subsequent neovascularisation was also IL-25 dependent, demonstrating a direct role for IL-25 during angiogenesis in vivo. Moreover, the secretion of innate epithelial derived cytokines IL-33 and thymic stromal lymphopoietin (TSLP) was completely ablated. CONCLUSIONS: In addition to modulating acute inflammation, we now demonstrate a role for IL-25 in orchestrating airway remodelling. IL-25 also drives IL-33 and TSLP production in the lung. These data delineate a wider role for IL-25 in mediating structural changes to the lung following allergen exposure and implicate IL-25 as a novel therapeutic target for the treatment of airway remodelling in asthma.
Assuntos
Remodelação das Vias Aéreas/imunologia , Hiper-Reatividade Brônquica/imunologia , Interleucinas/imunologia , Pyroglyphidae/imunologia , Remodelação das Vias Aéreas/genética , Animais , Asma/imunologia , Asma/metabolismo , Biópsia por Agulha , Hiper-Reatividade Brônquica/metabolismo , Líquido da Lavagem Broncoalveolar/imunologia , Células Cultivadas , Modelos Animais de Doenças , Feminino , Fibroblastos/imunologia , Fibroblastos/metabolismo , Regulação da Expressão Gênica/imunologia , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos BALB C , Distribuição Aleatória , Sensibilidade e Especificidade , Proteína Smad2/imunologiaRESUMO
Aberrant expansion of KRT5+ basal cells in the distal lung accompanies progressive alveolar epithelial cell loss and tissue remodelling during fibrogenesis in idiopathic pulmonary fibrosis (IPF). The mechanisms determining activity of KRT5+ cells in IPF have not been delineated. Here, we reveal a potential mechanism by which KRT5+ cells migrate within the fibrotic lung, navigating regional differences in collagen topography. In vitro, KRT5+ cell migratory characteristics and expression of remodelling genes are modulated by extracellular matrix (ECM) composition and organisation. Mass spectrometry- based proteomics revealed compositional differences in ECM components secreted by primary human lung fibroblasts (HLF) from IPF patients compared to controls. Over-expression of ECM glycoprotein, Secreted Protein Acidic and Cysteine Rich (SPARC) in the IPF HLF matrix restricts KRT5+ cell migration in vitro. Together, our findings demonstrate how changes to the ECM in IPF directly influence KRT5+ cell behaviour and function contributing to remodelling events in the fibrotic niche.
Assuntos
Fibrose Pulmonar Idiopática , Humanos , Matriz Extracelular , Células Epiteliais Alveolares , Transporte Biológico , Movimento Celular , Queratina-5RESUMO
RATIONALE: Airway hyperreactivity and remodeling are characteristic features of asthma. Interactions between the airway epithelium and environmental allergens are believed to be important in driving development of pathology, particularly because altered epithelial gene expression is common in individuals with asthma. OBJECTIVES: To investigate the interactions between a modified airway epithelium and a common aeroallergen in vivo. METHODS: We used an adenoviral vector to generate mice overexpressing the transforming growth factor-beta signaling molecule, Smad2, in the airway epithelium and exposed them to house dust mite (HDM) extract intranasally. MEASUREMENTS AND MAIN RESULTS: Smad2 overexpression resulted in enhanced airway hyperreactivity after allergen challenge concomitant with changes in airway remodeling. Subepithelial collagen deposition was increased and smooth muscle hyperplasia was evident resulting in thickening of the airway smooth muscle layer. However, there was no increase in airway inflammation in mice given the Smad2 vector compared with the control vector. Enhanced airway hyperreactivity and remodeling did not correlate with elevated levels of Th2 cytokines, such as IL-13 or IL-4. However, mice overexpressing Smad2 in the airway epithelium showed significantly enhanced levels of IL-25 and activin A after HDM exposure. Blocking activin A with a neutralizing antibody prevented the increase in lung IL-25 and inhibited subsequent collagen deposition and also the enhanced airway hyperreactivity observed in the Smad2 overexpressing HDM-exposed mice. CONCLUSIONS: Epithelial overexpression of Smad2 can specifically alter airway hyperreactivity and remodeling in response to an aeroallergen. Moreover, we have identified novel roles for IL-25 and activin A in driving airway hyperreactivity and remodeling.
Assuntos
Ativinas/metabolismo , Remodelação das Vias Aéreas/imunologia , Hiper-Reatividade Brônquica/imunologia , Interleucina-17/metabolismo , Pyroglyphidae/imunologia , Proteína Smad2/genética , Ativinas/antagonistas & inibidores , Adenoviridae , Alérgenos/efeitos adversos , Animais , Anticorpos/farmacologia , Colágeno/biossíntese , Dermatophagoides pteronyssinus , Modelos Animais de Doenças , Feminino , Expressão Gênica , Células Caliciformes/metabolismo , Hiperplasia , Imuno-Histoquímica , Pulmão/metabolismo , Pulmão/patologia , Camundongos , Camundongos Endogâmicos BALB C , Músculo Liso/patologia , Mucosa Respiratória/metabolismoRESUMO
Asthma is a highly prevalent lung disease, characterized by airway dysfunction and chronic inflammation. Asthma occurs in both children and adults, but frequently originates in early life. Heterogeneous asthma phenotypes exist, but Th2 cells are key players in a large proportion of cases, while other CD4+ T cell subsets are also implicated in driving and limiting pathology. In this chapter, we describe methods for establishing allergic airway disease to model asthma in adult and neonatal mice, along with protocols for measuring key disease parameters and quantifying and phenotyping CD4+ T cell subtypes.
Assuntos
Asma/imunologia , Linfócitos T CD4-Positivos/imunologia , Separação Celular , Citometria de Fluxo , Pulmão/imunologia , Fatores Etários , Animais , Asma/metabolismo , Asma/patologia , Linfócitos T CD4-Positivos/metabolismo , Células Cultivadas , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Imuno-Histoquímica , Pulmão/metabolismo , Pulmão/patologia , Masculino , Camundongos Endogâmicos BALB C , Fenótipo , Projetos de Pesquisa , Fluxo de TrabalhoRESUMO
Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease in which airway macrophages (AMs) play a key role. Itaconate has emerged as a mediator of macrophage function, but its role during fibrosis is unknown. Here, we reveal that itaconate is an endogenous antifibrotic factor in the lung. Itaconate levels are reduced in bronchoalveolar lavage, and itaconate-synthesizing cis-aconitate decarboxylase expression (ACOD1) is reduced in AMs from patients with IPF compared with controls. In the murine bleomycin model of pulmonary fibrosis, Acod1−/− mice develop persistent fibrosis, unlike wild-type (WT) littermates. Profibrotic gene expression is increased in Acod1−/− tissue-resident AMs compared with WT, and adoptive transfer of WT monocyte-recruited AMs rescued mice from disease phenotype. Culture of lung fibroblasts with itaconate decreased proliferation and wound healing capacity, and inhaled itaconate was protective in mice in vivo. Collectively, these data identify itaconate as critical for controlling the severity of lung fibrosis, and targeting this pathway may be a viable therapeutic strategy.
Assuntos
Carboxiliases/metabolismo , Fibrose Pulmonar Idiopática/imunologia , Macrófagos Alveolares/imunologia , Succinatos/metabolismo , Administração por Inalação , Transferência Adotiva/métodos , Adulto , Idoso , Animais , Bleomicina/administração & dosagem , Bleomicina/toxicidade , Líquido da Lavagem Broncoalveolar/imunologia , Broncoscopia , Estudos de Casos e Controles , Células Cultivadas , Modelos Animais de Doenças , Feminino , Fibroblastos , Voluntários Saudáveis , Humanos , Hidroliases/genética , Hidroliases/metabolismo , Fibrose Pulmonar Idiopática/induzido quimicamente , Fibrose Pulmonar Idiopática/diagnóstico , Fibrose Pulmonar Idiopática/terapia , Pulmão/citologia , Pulmão/imunologia , Pulmão/patologia , Macrófagos Alveolares/metabolismo , Macrófagos Alveolares/transplante , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Cultura Primária de Células , Índice de Gravidade de Doença , Succinatos/administração & dosagem , Succinatos/imunologiaRESUMO
The ontogeny of airway macrophages (AMs) in human lung and their contribution to disease are poorly mapped out. In mice, aging is associated with an increasing proportion of peripherally, as opposed to perinatally derived AMs. We sought to understand AM ontogeny in human lung during healthy aging and after transplant. We characterized monocyte/macrophage populations from the peripheral blood and airways of healthy volunteers across infancy/childhood (2-12 yr), maturity (20-50 yr), and older adulthood (>50 yr). Single-cell RNA sequencing (scRNA-seq) was performed on airway inflammatory cells isolated from sex-mismatched lung transplant recipients. During healthy aging, the proportions of blood bronchoalveolar lavage (BAL) classical monocytes peak in adulthood and decline in older adults. scRNA-seq of BAL cells from lung transplant recipients indicates that after transplant, the majority of AMs are recipient derived. These data show that during aging, the peripheral monocyte phenotype is consistent with that found in the airways and, furthermore, that the majority of human AMs after transplant are derived from circulating monocytes.