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    Bcl-2 inhibitors reduce steroid-insensitive airway inflammation. Tian Bao-Ping,Xia Li-Xia,Bao Zheng-Qiang,Zhang Hao,Xu Zhi-Wei,Mao Yuan-Yuan,Cao Chao,Che Luan-Qing,Liu Jin-Kai,Li Wen,Chen Zhi-Hua,Ying Songmin,Shen Hua-Hao The Journal of allergy and clinical immunology BACKGROUND:Asthmatic inflammation is dominated by accumulation of either eosinophils, neutrophils, or both in the airways. Disposal of these inflammatory cells is the key to disease control. Eosinophilic airway inflammation is responsive to corticosteroid treatment, whereas neutrophilic inflammation is resistant and increases the burden of global health care. Corticosteroid-resistant neutrophilic asthma remains mechanistically poorly understood and requires novel effective therapeutic strategies. OBJECTIVE:We sought to explore the underlying mechanisms of airway inflammation persistence, as well as corticosteroid resistance, and to investigate a new strategy of effective treatment against corticosteroid-insensitive neutrophilic asthma. METHODS:Mouse models of either eosinophil-dominated or neutrophil-dominated airway inflammation were used in this study to test corticosteroid sensitivity in vivo and in vitro. We also used vav-Bcl-2 transgenic mice to confirm the importance of granulocytes apoptosis in the clearance of airway inflammation. Finally, the Bcl-2 inhibitors ABT-737 or ABT-199 were tested for their therapeutic effects against eosinophilic or neutrophilic airway inflammation and airway hyperresponsiveness. RESULTS:Overexpression of Bcl-2 protein was found to be responsible for persistence of granulocytes in bronchoalveolar lavage fluid after allergic challenge. This was important because allergen-induced airway inflammation aggravated and persisted in vav-Bcl-2 transgenic mice, in which nucleated hematopoietic cells were overexpressed with Bcl-2 and resistant to apoptosis. The Bcl-2 inhibitors ABT-737 or ABT-199 play efficient roles in alleviation of either eosinophilic or corticosteroid-resistant neutrophilic airway inflammation by inducing apoptosis of immune cells, such as eosinophils, neutrophils, T2 cells, T17 cells, and dendritic cells. Moreover, these inhibitors were found to be more efficient than steroids to induce granulocyte apoptosis ex vivo from patients with severe asthma. CONCLUSION:Apoptosis of inflammatory cells is essential for clearance of allergen-induced airway inflammation. The Bcl-2 inhibitors ABT-737 or ABT-199 might be promising drugs for the treatment of airway inflammation, especially for corticosteroid-insensitive neutrophilic airway inflammation. 10.1016/j.jaci.2016.11.027
    Role for NLRP3 Inflammasome-mediated, IL-1β-Dependent Responses in Severe, Steroid-Resistant Asthma. Kim Richard Y,Pinkerton James W,Essilfie Ama T,Robertson Avril A B,Baines Katherine J,Brown Alexandra C,Mayall Jemma R,Ali M Khadem,Starkey Malcolm R,Hansbro Nicole G,Hirota Jeremy A,Wood Lisa G,Simpson Jodie L,Knight Darryl A,Wark Peter A,Gibson Peter G,O'Neill Luke A J,Cooper Matthew A,Horvat Jay C,Hansbro Philip M American journal of respiratory and critical care medicine RATIONALE:Severe, steroid-resistant asthma is the major unmet need in asthma therapy. Disease heterogeneity and poor understanding of pathogenic mechanisms hampers the identification of therapeutic targets. Excessive nucleotide-binding oligomerization domain-like receptor family, pyrin domain-containing 3 (NLRP3) inflammasome and concomitant IL-1β responses occur in chronic obstructive pulmonary disease, respiratory infections, and neutrophilic asthma. However, the direct contributions to pathogenesis, mechanisms involved, and potential for therapeutic targeting remain poorly understood, and are unknown in severe, steroid-resistant asthma. OBJECTIVES:To investigate the roles and therapeutic targeting of the NLRP3 inflammasome and IL-1β in severe, steroid-resistant asthma. METHODS:We developed mouse models of Chlamydia and Haemophilus respiratory infection-mediated, ovalbumin-induced severe, steroid-resistant allergic airway disease. These models share the hallmark features of human disease, including elevated airway neutrophils, and NLRP3 inflammasome and IL-1β responses. The roles and potential for targeting of NLRP3 inflammasome, caspase-1, and IL-1β responses in experimental severe, steroid-resistant asthma were examined using a highly selective NLRP3 inhibitor, MCC950; the specific caspase-1 inhibitor Ac-YVAD-cho; and neutralizing anti-IL-1β antibody. Roles for IL-1β-induced neutrophilic inflammation were examined using IL-1β and anti-Ly6G. MEASUREMENTS AND MAIN RESULTS:Chlamydia and Haemophilus infections increase NLRP3, caspase-1, IL-1β responses that drive steroid-resistant neutrophilic inflammation and airway hyperresponsiveness. Neutrophilic airway inflammation, disease severity, and steroid resistance in human asthma correlate with NLRP3 and IL-1β expression. Treatment with anti-IL-1β, Ac-YVAD-cho, and MCC950 suppressed IL-1β responses and the important steroid-resistant features of disease in mice, whereas IL-1β administration recapitulated these features. Neutrophil depletion suppressed IL-1β-induced steroid-resistant airway hyperresponsiveness. CONCLUSIONS:NLRP3 inflammasome responses drive experimental severe, steroid-resistant asthma and are potential therapeutic targets in this disease. 10.1164/rccm.201609-1830OC
    Is IL-1β inhibition the next therapeutic target in asthma? Peebles R Stokes The Journal of allergy and clinical immunology 10.1016/j.jaci.2017.03.018
    TNF is required for TLR ligand-mediated but not protease-mediated allergic airway inflammation. Whitehead Gregory S,Thomas Seddon Y,Shalaby Karim H,Nakano Keiko,Moran Timothy P,Ward James M,Flake Gordon P,Nakano Hideki,Cook Donald N The Journal of clinical investigation Asthma is associated with exposure to a wide variety of allergens and adjuvants. The extent to which overlap exists between the cellular and molecular mechanisms triggered by these various agents is poorly understood, but it might explain the differential responsiveness of patients to specific therapies. In particular, it is unclear why some, but not all, patients benefit from blockade of TNF. Here, we characterized signaling pathways triggered by distinct types of adjuvants during allergic sensitization. Mice sensitized to an innocuous protein using TLR ligands or house dust extracts as adjuvants developed mixed eosinophilic and neutrophilic airway inflammation and airway hyperresponsiveness (AHR) following allergen challenge, whereas mice sensitized using proteases as adjuvants developed predominantly eosinophilic inflammation and AHR. TLR ligands, but not proteases, induced TNF during allergic sensitization. TNF signaled through airway epithelial cells to reprogram them and promote Th2, but not Th17, development in lymph nodes. TNF was also required during the allergen challenge phase for neutrophilic and eosinophilic inflammation. In contrast, TNF was dispensable for allergic airway disease in a protease-mediated model of asthma. These findings might help to explain why TNF blockade improves lung function in only some patients with asthma. 10.1172/JCI90890
    mTOR regulates metabolic adaptation of APCs in the lung and controls the outcome of allergic inflammation. Sinclair Charles,Bommakanti Gayathri,Gardinassi Luiz,Loebbermann Jens,Johnson Matthew Joseph,Hakimpour Paul,Hagan Thomas,Benitez Lydia,Todor Andrei,Machiah Deepa,Oriss Timothy,Ray Anuradha,Bosinger Steven,Ravindran Rajesh,Li Shuzhao,Pulendran Bali Science (New York, N.Y.) Antigen-presenting cells (APCs) occupy diverse anatomical tissues, but their tissue-restricted homeostasis remains poorly understood. Here, working with mouse models of inflammation, we found that mechanistic target of rapamycin (mTOR)-dependent metabolic adaptation was required at discrete locations. mTOR was dispensable for dendritic cell (DC) homeostasis in secondary lymphoid tissues but necessary to regulate cellular metabolism and accumulation of CD103 DCs and alveolar macrophages in lung. Moreover, while numbers of mTOR-deficient lung CD11b DCs were not changed, they were metabolically reprogrammed to skew allergic inflammation from eosinophilic T helper cell 2 (T2) to neutrophilic T17 polarity. The mechanism for this change was independent of translational control but dependent on inflammatory DCs, which produced interleukin-23 and increased fatty acid oxidation. mTOR therefore mediates metabolic adaptation of APCs in distinct tissues, influencing the immunological character of allergic inflammation. 10.1126/science.aaj2155
    Concomitant suppression of T2 and T17 cell responses in allergic asthma by targeting retinoic acid receptor-related orphan receptor γt. Na Hyeongjin,Lim Hoyong,Choi Garam,Kim Byung-Keun,Kim Sae-Hoon,Chang Yoon-Seok,Nurieva Roza,Dong Chen,Chang Seon Hee,Chung Yeonseok The Journal of allergy and clinical immunology BACKGROUND:Allergic asthma is a heterogeneous chronic inflammatory disease of the airways with a massive infiltration of eosinophils or neutrophils mediated by allergen-specific T2 and T17 cells, respectively. Therefore successful treatment of allergic asthma will require suppression of both T2 and T17 cells. OBJECTIVE:We sought to investigate the role of the T17 cell pathway in regulating T2 cell responses in allergic asthma. METHODS:Allergic asthma was induced by intranasal challenge with proteinase allergens in C57BL/6, Il17aIl17f, and retinoic acid receptor-related orphan receptor γt (RORγt) mice. A pharmacologic RORγt inhibitor was used to evaluate its preventive and therapeutic effects in allergic asthma. Characteristics of allergic airway inflammation were analyzed by using flow cytometry, histology, quantitative real-time PCR, and ELISA. Mixed bone marrow chimeric mice, fate mapping analysis, short hairpin RNA transduction, and in vitro T-cell differentiation were used for mechanistic studies. RESULTS:Mice deficient in IL-17A and IL-17F, as well as RORγt, exhibited a significant reduction not only in T17 cell responses but also in T2 cell responses in an animal model of allergic asthma. Similarly, mice treated with an RORγt inhibitor had significantly diminished T17 and T2 cell responses, leading to reduced neutrophil and eosinophil numbers in the airway. RORγt-deficient T cells were intrinsically defective in differentiating into T2 cells and expressed increased levels of B-cell lymphoma 6 (Bcl6). Bcl6 knockdown resulted in a remarkable restoration of T2 cell differentiation in RORγt-deficient T cells. Blockade of RORγt also significantly hampered the differentiation of human T2 and T17 cells from naive CD4 T cells. CONCLUSION:RORγt in T cells is required for optimal T2 cell differentiation by suppressing Bcl6 expression; this finding suggests that targeting RORγt might be a promising approach for the treatment of allergic asthma by concomitantly suppressing T17 and T2 cell responses in the airway. 10.1016/j.jaci.2017.07.050
    Cysteinyl leukotriene receptor 1 expression identifies a subset of neutrophils during the antiviral response that contributes to postviral atopic airway disease. Cheung Dorothy S,Sigua Jerome A,Simpson Pippa M,Yan Ke,Hussain Syed-Rehan A,Santoro Jennifer L,Buell Erika J,Hunter Desire A,Rohlfing Michelle,Patadia Deepa,Grayson Mitchell H The Journal of allergy and clinical immunology BACKGROUND:Viral respiratory tract infections increase the risk of development and exacerbation of atopic disease. Previously, we demonstrated the requirement for a neutrophil (PMN) subset expressing CD49d to drive development of postviral atopic airway disease in mice. OBJECTIVE:We sought to determine whether human CD49d PMNs are present in the nasal mucosa during acute viral respiratory tract infections and further characterize this PMN subset in human subjects and mice. METHODS:Sixty subjects (5-50 years old) were enrolled within 4 days of acute onset of upper respiratory symptoms. Nasal lavage for flow cytometry and nasal swabs for viral PCR were performed at enrollment and during convalescence. The Sendai virus mouse model was used to investigate the phenotype and functional relevance of CD49d PMNs. RESULTS:CD49d PMN frequency was significantly higher in nasal lavage fluid during acute respiratory symptoms in all subjects (2.9% vs 1.0%, n = 42, P < .001). In mice CD49d PMNs represented a "proatopic" neutrophil subset that expressed cysteinyl leukotriene receptor 1 (CysLTR1) and produced TNF, CCL2, and CCL5. Inhibition of CysLTR1 signaling in the first days of a viral respiratory tract infection was sufficient to reduce accumulation of CD49d PMNs in the lungs and development of postviral atopic airway disease. Similar to the mouse, human CD49d PMNs isolated from nasal lavage fluid during a viral respiratory tract infection expressed CysLTR1. CONCLUSION:CD49d and CysLTR1-coexpressing PMNs are present during symptoms of an acute viral respiratory tract infection in human subjects. Further study is needed to examine selective targeting of proatopic neutrophils as a potential therapeutic strategy to prevent development of postviral atopic airway disease. 10.1016/j.jaci.2017.11.026
    Saturated fatty acids, obesity, and the nucleotide oligomerization domain-like receptor protein 3 (NLRP3) inflammasome in asthmatic patients. Wood Lisa G,Li Qian,Scott Hayley A,Rutting Sandra,Berthon Bronwyn S,Gibson Peter G,Hansbro Philip M,Williams Evan,Horvat Jay,Simpson Jodie L,Young Paul,Oliver Brian G,Baines Katherine J The Journal of allergy and clinical immunology BACKGROUND:Both obesity and high dietary fat intake activate the nucleotide oligomerization domain-like receptor protein 3 (NLRP3) inflammasome. OBJECTIVE:We aimed to examine NLRP3 inflammasome activity in the airways of obese asthmatic patients after macronutrient overload and in immune cells challenged by inflammasome triggers. METHODS:Study 1 was a cross-sectional observational study of nonobese (n = 51) and obese (n = 76) asthmatic adults. Study 2 was a randomized, crossover, acute feeding study in 23 asthmatic adults (n = 12 nonobese and n = 11 obese subjects). Subjects consumed 3 isocaloric meals on 3 separate occasions (ie, saturated fatty acid, n-6 polyunsaturated fatty acid, and carbohydrate) and were assessed at 0 and 4 hours. For Studies 1 and 2, airway inflammation was measured based on sputum differential cell counts, IL-1β protein levels (ELISA), and sputum cell gene expression (Nanostring nCounter). In Study 3 peripheral blood neutrophils and monocytes were isolated by using Ficoll density gradient and magnetic bead separation and incubated with or without palmitic acid, LPS, or TNF-α for 24 hours, and IL-1β release was measured (ELISA). RESULTS:In Study 1 NLRP3 and nucleotide oligomerization domain 1 (NOD1) gene expression was upregulated, and sputum IL-1β protein levels were greater in obese versus nonobese asthmatic patients. In Study 2 the saturated fatty acid meal led to increases in sputum neutrophil percentages and sputum cell gene expression of Toll-like receptor 4 (TLR4) and NLRP3 at 4 hours in nonobese asthmatic patients. In Study 3 neutrophils and monocytes released IL-1β when challenged with a combination of palmitic acid and LPS or TNF-α. CONCLUSION:The NLRP3 inflammasome is a potential therapeutic target in asthmatic patients. Behavioral interventions that reduce fatty acid exposure, such as weight loss and dietary saturated fat restriction, warrant further exploration. 10.1016/j.jaci.2018.04.037
    Asthma exacerbated by neutrophil ghosts. Bird Lucy Nature reviews. Immunology 10.1038/s41577-018-0059-6
    Neutrophils promote T-cell-mediated inflammation in allergy. Polak Dominika,Samadi Nazanin,Vizzardelli Caterina,Sánchez Acosta Gabriela,Rosskopf Sandra,Steinberger Peter,Jahn-Schmid Beatrice,Bohle Barbara The Journal of allergy and clinical immunology 10.1016/j.jaci.2018.11.035
    IL-4 receptor engagement in human neutrophils impairs their migration and extracellular trap formation. Impellizzieri Daniela,Ridder Frederike,Raeber Miro E,Egholm Cecilie,Woytschak Janine,Kolios Antonios G A,Legler Daniel F,Boyman Onur The Journal of allergy and clinical immunology BACKGROUND:Type 2 immunity serves to resist parasitic helminths, venoms, and toxins, but the role and regulation of neutrophils during type 2 immune responses are controversial. Helminth models suggested a contribution of neutrophils to type 2 immunity, whereas neutrophils are associated with increased disease severity during type 2 inflammatory disorders, such as asthma. OBJECTIVE:We sought to evaluate the effect of the prototypic type 2 cytokines IL-4 and IL-13 on human neutrophils. METHODS:Human neutrophils from peripheral blood were assessed without or with IL-4 or IL-13 for (1) expression of IL-4 receptor subunits, (2) neutrophil extracellular trap (NET) formation, (3) migration toward CXCL8 in vitro and in humanized mice, and (4) CXCR1, CXCR2, and CXCR4 expression, as well as (5) in nonallergic versus allergic subjects. RESULTS:Human neutrophils expressed both types of IL-4 receptors, and their stimulation through IL-4 or IL-13 diminished their ability to form NETs and migrate toward CXCL8 in vitro. Likewise, in vivo chemotaxis in NOD-scid-Il2rg mice was reduced in IL-4-stimulated human neutrophils compared with control values. These effects were accompanied by downregulation of the CXCL8-binding chemokine receptors CXCR1 and CXCR2 on human neutrophils on IL-4 or IL-13 stimulation in vitro. Ex vivo analysis of neutrophils from allergic patients or exposure of neutrophils from nonallergic subjects to allergic donor serum in vitro impaired their NET formation and migration toward CXCL8, thereby mirroring IL-4/IL-13-stimulated neutrophils. CONCLUSION:IL-4 receptor signaling in human neutrophils affects several neutrophil effector functions, which bears important implications for immunity in type 2 inflammatory disorders. 10.1016/j.jaci.2019.01.042
    Extracellular DNA, Neutrophil Extracellular Traps, and Inflammasome Activation in Severe Asthma. Lachowicz-Scroggins Marrah E,Dunican Eleanor M,Charbit Annabelle R,Raymond Wilfred,Looney Mark R,Peters Michael C,Gordon Erin D,Woodruff Prescott G,Lefrançais Emma,Phillips Brenda R,Mauger David T,Comhair Suzy A,Erzurum Serpil C,Johansson Mats W,Jarjour Nizar N,Coverstone Andrea M,Castro Mario,Hastie Annette T,Bleecker Eugene R,Fajt Merritt L,Wenzel Sally E,Israel Elliot,Levy Bruce D,Fahy John V American journal of respiratory and critical care medicine Extracellular DNA (eDNA) and neutrophil extracellular traps (NETs) are implicated in multiple inflammatory diseases. NETs mediate inflammasome activation and IL-1β secretion from monocytes and cause airway epithelial cell injury, but the role of eDNA, NETs, and IL-1β in asthma is uncertain. To characterize the role of activated neutrophils in severe asthma through measurement of NETs and inflammasome activation. We measured sputum eDNA in induced sputum from 399 patients with asthma in the Severe Asthma Research Program-3 and in 94 healthy control subjects. We subdivided subjects with asthma into eDNA-low and -high subgroups to compare outcomes of asthma severity and of neutrophil and inflammasome activation. We also examined if NETs cause airway epithelial cell damage that can be prevented by DNase. We found that 13% of the Severe Asthma Research Program-3 cohort is "eDNA-high," as defined by sputum eDNA concentrations above the upper 95th percentile value in health. Compared with eDNA-low patients with asthma, eDNA-high patients had lower Asthma Control Test scores, frequent history of chronic mucus hypersecretion, and frequent use of oral corticosteroids for maintenance of asthma control (all values <0.05). Sputum eDNA in asthma was associated with airway neutrophilic inflammation, increases in soluble NET components, and increases in caspase 1 activity and IL-1β (all values <0.001). In studies, NETs caused cytotoxicity in airway epithelial cells that was prevented by disruption of NETs with DNase. High extracellular DNA concentrations in sputum mark a subset of patients with more severe asthma who have NETs and markers of inflammasome activation in their airways. 10.1164/rccm.201810-1869OC
    IL-17-high asthma with features of a psoriasis immunophenotype. Östling Jörgen,van Geest Marleen,Schofield James P R,Jevnikar Zala,Wilson Susan,Ward Jonathan,Lutter Rene,Shaw Dominick E,Bakke Per S,Caruso Massimo,Dahlen Sven-Erik,Fowler Stephen J,Horváth Ildikó,Krug Norbert,Montuschi Paolo,Sanak Marek,Sandström Thomas,Sun Kai,Pandis Ioannis,Auffray Charles,Sousa Ana R,Guo Yike,Adcock Ian M,Howarth Peter,Chung Kian Fan,Bigler Jeanette,Sterk Peter J,Skipp Paul J,Djukanović Ratko,Vaarala Outi, The Journal of allergy and clinical immunology BACKGROUND:The role of IL-17 immunity is well established in patients with inflammatory diseases, such as psoriasis and inflammatory bowel disease, but not in asthmatic patients, in whom further study is required. OBJECTIVE:We sought to undertake a deep phenotyping study of asthmatic patients with upregulated IL-17 immunity. METHODS:Whole-genome transcriptomic analysis was performed by using epithelial brushings, bronchial biopsy specimens (91 asthmatic patients and 46 healthy control subjects), and whole blood samples (n = 498) from the Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes (U-BIOPRED) cohort. Gene signatures induced in vitro by IL-17 and IL-13 in bronchial epithelial cells were used to identify patients with IL-17-high and IL-13-high asthma phenotypes. RESULTS:Twenty-two of 91 patients were identified with IL-17, and 9 patients were identified with IL-13 gene signatures. The patients with IL-17-high asthma were characterized by risk of frequent exacerbations, airway (sputum and mucosal) neutrophilia, decreased lung microbiota diversity, and urinary biomarker evidence of activation of the thromboxane B2 pathway. In pathway analysis the differentially expressed genes in patients with IL-17-high asthma were shared with those reported as altered in psoriasis lesions and included genes regulating epithelial barrier function and defense mechanisms, such as IL1B, IL6, IL8, and β-defensin. CONCLUSION:The IL-17-high asthma phenotype, characterized by bronchial epithelial dysfunction and upregulated antimicrobial and inflammatory response, resembles the immunophenotype of psoriasis, including activation of the thromboxane B2 pathway, which should be considered a biomarker for this phenotype in further studies, including clinical trials targeting IL-17. 10.1016/j.jaci.2019.03.027
    Neutrophilic inflammation in asthma and defective epithelial translational control. Ravi Abilash,Chowdhury Saheli,Dijkhuis Annemiek,Bonta Peter I,Sterk Peter J,Lutter René The European respiratory journal Neutrophilic inflammation in asthma is associated with interleukin (IL)-17A, corticosteroid-insensitivity and bronchodilator-induced forced expiratory volume in 1 s (FEV) reversibility. IL-17A synergises with tumour necrosis factor (TNF)-α in the production of the neutrophil chemokine CXCL-8 by primary bronchial epithelial cells (PBECs).We hypothesised that local neutrophilic inflammation in asthma correlates with IL-17A and TNF-α-induced CXCL-8 production by PBECs from asthma patients.PBECs from most asthma patients displayed an exaggerated CXCL-8 production in response to TNF-α and IL-17A, but not to TNF-α alone, and which was also insensitive to corticosteroids. This hyperresponsiveness of PBECs strongly correlated with CXCL-8 levels and neutrophil numbers in bronchoalveolar lavage from the corresponding patients, but not with that of eosinophils. In addition, this hyperresponsiveness also correlated with bronchodilator-induced FEV % reversibility. At the molecular level, epithelial hyperresponsiveness was associated with failure of the translational repressor T-cell internal antigen-1 related protein (TiAR) to translocate to the cytoplasm to halt CXCL-8 production, as confirmed by TiAR knockdown. This is in line with the finding that hyperresponsive PBECs also produced enhanced levels of other inflammatory mediators.Hyperresponsive PBECs in asthma patients may underlie neutrophilic and corticosteroid-insensitive inflammation and a reduced FEV, irrespective of eosinophilic inflammation. Normalising cytoplasmic translocation of TiAR is a potential therapeutic target in neutrophilic, corticosteroid-insensitive asthma. 10.1183/13993003.00547-2019
    Associations between fungal and bacterial microbiota of airways and asthma endotypes. Sharma Anukriti,Laxman Bharathi,Naureckas Edward T,Hogarth D Kyle,Sperling Anne I,Solway Julian,Ober Carole,Gilbert Jack A,White Steven R The Journal of allergy and clinical immunology BACKGROUND:The relationship between asthma, atopy, and underlying type 2 (T2) airway inflammation is complex. Although the bacterial airway microbiota is known to differ in asthmatic patients, the fungal and bacterial markers that discriminate T2-high (eosinophilic) and T2-low (neutrophilic/mixed-inflammation) asthma and atopy are still incompletely identified. OBJECTIVES:The aim of this study was to demonstrate the fungal microbiota structure of airways in asthmatic patients associated with T2 inflammation, atopy, and key clinical parameters. METHODS:We collected endobronchial brush (EB) and bronchoalveolar lavage (BAL) samples from 39 asthmatic patients and 19 healthy subjects followed by 16S gene and internal transcribed spacer-based microbiota sequencing. The microbial sequences were classified into exact sequence variants. The T2 phenotype was defined by using a blood eosinophil count with a threshold of 300 cells/μL. RESULTS:Fungal diversity was significantly lower in EB samples from patients with T2-high compared with T2-low inflammation; key fungal genera enriched in patients with T2-high inflammation included Trichoderma species, whereas Penicillium species was enriched in patients with atopy. In BAL fluid samples the dominant genera were Cladosporium, Fusarium, Aspergillus, and Alternaria. Using generalized linear models, we identified significant associations between specific fungal exact sequence variants and FEV, fraction of exhaled nitric oxide values, BAL fluid cell counts, and corticosteroid use. Investigation of interkingdom (bacterial-fungal) co-occurrence patterns revealed different topologies between asthmatic patients and healthy control subjects. Random forest models with fungal classifiers predicted asthma status with 75% accuracy for BAL fluid samples and 80% accuracy for EB samples. CONCLUSIONS:We demonstrate clear differences in bacterial and fungal microbiota in asthma-associated phenotypes. Our study provides additional support for considering microbial signatures in delineating asthma phenotypes. 10.1016/j.jaci.2019.06.025
    A Network of Sputum MicroRNAs Is Associated with Neutrophilic Airway Inflammation in Asthma. Gomez Jose L,Chen Ailu,Diaz Maria Paula,Zirn Nicholas,Gupta Amolika,Britto Clemente,Sauler Maor,Yan Xiting,Stewart Emma,Santerian Kyle,Grant Nicole,Liu Qing,Fry Rebecca,Rager Julia,Cohn Lauren,Alexis Neil,Chupp Geoffrey L American journal of respiratory and critical care medicine MicroRNAs are potent regulators of biologic systems that are critical to tissue homeostasis. Individual microRNAs have been identified in airway samples. However, a systems analysis of the microRNA-mRNA networks present in the sputum that contribute to airway inflammation in asthma has not been published. Identify microRNA and mRNA networks in the sputum of patients with asthma. We conducted a genome-wide analysis of microRNA and mRNA in the sputum from patients with asthma and correlated expression with clinical phenotypes. Weighted gene correlation network analysis was implemented to identify microRNA networks (modules) that significantly correlate with clinical features of asthma and mRNA expression networks. MicroRNA expression in peripheral blood neutrophils and lymphocytes and hybridization of the sputum were used to identify the cellular sources of microRNAs. MicroRNA expression obtained before and after ozone exposure was also used to identify changes associated with neutrophil counts in the airway. Six microRNA modules were associated with clinical features of asthma. A single module () was associated with a history of hospitalizations, lung function impairment, and numbers of neutrophils and lymphocytes in the sputum. Of the 12 microRNAs in the module, was the highest expressed microRNA in neutrophils and was associated with increased neutrophil counts in the sputum in response to ozone exposure. Multiple microRNAs in the module correlated with two mRNA modules enriched for TLR (Toll-like receptor) and T-helper cell type 17 (Th17) signaling and endoplasmic reticulum stress. was a key regulator of the TLR and Th17 pathways in the sputum of subjects with asthma. This study of sputum microRNA and mRNA expression from patients with asthma demonstrates the existence of microRNA networks and genes that are associated with features of asthma severity. Among these, a neutrophil-derived microRNA, regulates TLR/Th17 signaling and endoplasmic reticulum stress. 10.1164/rccm.201912-2360OC
    Bronchoalveolar lavage cytokine patterns in children with severe neutrophilic and paucigranulocytic asthma. Steinke John W,Lawrence Monica G,Teague W Gerald,Braciale Thomas J,Patrie James T,Borish Larry The Journal of allergy and clinical immunology BACKGROUND:Asthma is a complex heterogeneous disease occurring in adults and children that is characterized by distinct inflammatory patterns. While numerous studies have been performed in adults, little is known regarding the heterogeneity of severe asthma in children, particularly inflammatory signatures involving the air spaces. OBJECTIVE:We sought to determine the relationship of bronchoalveolar lavage (BAL) cytokine/chemokine expression patterns in children with severe therapy-resistant asthma stratified according to neutrophilic versus nonneutrophilic BAL inflammatory cell patterns. METHODS:Children with severe asthma with inadequate symptom control despite therapy underwent diagnostic bronchoscopy and BAL. Inflammatory cytokine/chemokine concentrations were determined using a multiplex protein bead assay. RESULTS:Analysis of BAL constituents with an unbiased clustering approach revealed distinct cytokine/chemokine patterns, and these aligned with pathways associated with type 2 innate lymphoid cells, monocytes, neutrophil trafficking, and T effector cells. All cytokines examined (n = 27) with 1 exception (vascular endothelial growth factor) were overexpressed with BAL neutrophilia compared with nonneutrophilic asthma, and this was confirmed in a cross-validation analysis. Cytokines specifically responsible for T17 (IL-17, IL-6, G-CSF) and T1 differentiation and expression (IL-12, TNF-α, IFN-γ) were enhanced in the neutrophilic cohorts. Neutrophilic groups were also characterized by higher prevalence of bacterial and viral pathogens; however, cytokine expression patterns manifested independently of pathogen expression. CONCLUSIONS:The results demonstrate that children with refractory asthma and neutrophilic inflammation had a BAL cytokine pattern consistent with a mixed T17/T1/T2 response. In contrast, nonneutrophilic asthma presented independently of cytokine overexpression. 10.1016/j.jaci.2020.05.039
    Cross-reactive antibodies against dust mite-derived enolase induce neutrophilic airway inflammation. The European respiratory journal BACKGROUND AND AIMS:Neutrophilic inflammation is a hallmark of some specific asthma phenotypes; its aetiology is not yet fully understood. House dust mite (HDM) is the most common factor in the pathogenesis of airway inflammation. This study aims to elucidate the role of cross-antibodies against HDM-derived factors in the development of neutrophilic inflammation in the airway. METHODS:Blood samples were collected from asthma patients with chronic neutrophilic asthma for analysis of HDM-specific cross-reactive antibodies. The role of an antibody against HDM-derived enolase (EnoAb) in the impairment of airway epithelial barrier function and induction of airway inflammation was assessed in a cell culture model and an animal model. RESULTS:High similarity (72%) of the enolase gene sequences was identified between HDM and human. Serum EnoAb was detected in patients with chronic neutrophilic asthma. The EnoAb bound to airway epithelial cells to form complexes with enolase, which activated complement, impaired airway epithelial barrier functions and induced neutrophilic inflammation in the airway tissues. CONCLUSIONS:HDM-derived enolase can induce specific cross-antibodies in humans, which induce neutrophilic inflammation in the airway. 10.1183/13993003.02375-2019
    Single-cell transcriptomic analysis reveals key immune cell phenotypes in the lungs of patients with asthma exacerbation. Li Hui,Wang Huaqi,Sokulsky Leon,Liu Shaoxia,Yang Rui,Liu Xiaojie,Zhou Lujia,Li Juan,Huang Chun,Li Fangfang,Lei Xu,Jia Hongxia,Cheng Jiuling,Li Fuguang,Yang Ming,Zhang Guojun The Journal of allergy and clinical immunology BACKGROUND:Asthma exacerbations are associated with heightened asthma symptoms, which can result in hospitalization in severe cases. However, the molecular immunologic processes that determine the course of an exacerbation remain poorly understood, impeding the progression of development of effective therapies. OBJECTIVE:Our aim was to identify candidate genes that are strongly associated with asthma exacerbation at a cellular level. METHODS:Subjects with asthma exacerbation and healthy control subjects were recruited, and bronchoalveolar lavage fluid was isolated from these subjects via bronchoscopy. Cells were isolated through fluorescence-activated cell sorting, and single-cell RNA sequencing was performed on enriched cell populations. RESULTS:We showed that the levels of monocytes, CD8 T cells, and macrophages are significantly elevated in the bronchoalveolar lavage fluid of patients. A set of cytokines and intracellular transduction regulators are associated with asthma exacerbations and are shared across multiple cell clusters, forming a complicated molecular framework. An additional group of core exacerbation-associated modules is activated, including eukaryotic initiation factor 2 signaling, ephrin receptor signaling, and C-X-C chemokine receptor type 4 signaling in the subpopulations of CD8 T cells (C1-a) and monocyte clusters (C7 clusters), which are associated with infection. CONCLUSION:Our study identified a significant number of severe asthma-associated genes that are differentially expressed by multiple cell clusters. 10.1016/j.jaci.2020.09.032
    Ceramide in apoptosis and oxidative stress in allergic inflammation and asthma. The Journal of allergy and clinical immunology BACKGROUND:Nothing is known about the mechanisms by which increased ceramide levels in the lung contribute to allergic responses and asthma severity. OBJECTIVE:We sought to investigate the functional role of ceramide in mouse models of allergic airway disease that recapitulate the cardinal clinical features of human allergic asthma. METHODS:Allergic airway disease was induced in mice by repeated intranasal administration of house dust mite or the fungal allergen Alternaria alternata. Processes that can be regulated by ceramide and are important for severity of allergic asthma were correlated with ceramide levels measured by mass spectrometry. RESULTS:Both allergens induced massive pulmonary apoptosis and also significantly increased reactive oxygen species in the lung. Prevention of increases in lung ceramide levels mitigated allergen-induced apoptosis, reactive oxygen species, and neutrophil infiltration. In contrast, dietary supplementation of the antioxidant α-tocopherol decreased reactive oxygen species but had no significant effects on elevation of ceramide level or apoptosis, indicating that the increases in lung ceramide levels in allergen-challenged mice are not mediated by oxidative stress. Moreover, specific ceramide species were altered in bronchoalveolar lavage fluid from patients with severe asthma compared with in bronchoalveolar lavage fluid from individuals without asthma. CONCLUSION:Our data suggest that elevation of ceramide level after allergen challenge contributes to the apoptosis, reactive oxygen species generation, and neutrophilic infiltrate that characterize the severe asthmatic phenotype. Ceramide might be the trigger of formation of Creola bodies found in the sputum of patients with severe asthma and could be a biomarker to optimize diagnosis and to monitor and improve clinical outcomes in this disease. 10.1016/j.jaci.2020.10.024
    CC16 Binding to αβ Integrin Protects against Infection. Johnson Michael D L,Younis Usir S,Menghani Sanjay V,Addison Kenneth J,Whalen Michael,Pilon Aprile L,Cress Anne E,Polverino Francesca,Romanoski Casey E,Kraft Monica,Martinez Fernando D,Guerra Stefano,Ledford Julie G American journal of respiratory and critical care medicine CC16 (club cell secretory protein) is a pneumoprotein produced predominantly by pulmonary club cells. Circulating CC16 is associated with protection from the inception and progression of the two most common obstructive lung diseases (asthma and chronic obstructive pulmonary disease). Although exact mechanisms remain elusive, studies consistently suggest a causal role of CC16 in mediating antiinflammatory and antioxidant functions in the lung. We sought to determine any novel receptor systems that could participate in CC16's role in obstructive lung diseases. Protein alignment of CC16 across species led to the discovery of a highly conserved sequence of amino acids, leucine-valine-aspartic acid (LVD), a known integrin-binding motif. Recombinant CC16 was generated with and without the putative integrin-binding site. A mouse model and a fluorescent cellular adhesion assay were used to determine the impact of the LVD site regarding CC16 function during live infection and on cellular adhesion during inflammatory conditions. CC16 bound to integrin αβ), also known as the adhesion molecule VLA-4 (very late antigen 4), dependent on the presence of the LVD integrin-binding motif. During infection, recombinant CC16 rescued lung function parameters both when administered to the lung and intravenously but only when the LVD integrin-binding site was intact; likewise, neutrophil recruitment during infection and leukocyte adhesion were both impacted by the loss of the LVD site. We discovered a novel receptor for CC16, VLA-4, which has important mechanistic implications for the role of CC16 in circulation as well as in the lung compartment. 10.1164/rccm.202006-2576OC
    GM-CSF instigates a dendritic cell-T-cell inflammatory circuit that drives chronic asthma development. Nobs Samuel Philip,Pohlmeier Lea,Li Fengqi,Kayhan Merve,Becher Burkhard,Kopf Manfred The Journal of allergy and clinical immunology BACKGROUND:Steroid-resistant asthma is often characterized by high levels of neutrophils and mixed T2/T17 immune profiles. Indeed, neutrophils are key drivers of chronic lung inflammation in multiple respiratory diseases. Their numbers correlate strongly with disease severity, and their presence is often associated with exacerbation of chronic lung inflammation. OBJECTIVE:What factors drive development of neutrophil-mediated chronic lung disease remains largely unknown, and we sought to study the role of GM-CSF as a potential regulator in chronic asthma. METHODS:Different experimental animal models of chronic asthma were used in combination with alveolar macrophage-reconstitution of global GM-CSF receptor knockout mice as well as cell-type-specific knockout animals to elucidate the role of GM-CSF signaling in chronic airway inflammation. RESULTS:We identify GM-CSF signaling as a critical factor regulating pulmonary accumulation of neutrophils. We show that although being not required for intrinsically regulating neutrophil migration, GM-CSF controls lung dendritic cell function, which in turn promotes T-cell-dependent recruitment of neutrophils to the airways. We demonstrate that GM-CSF regulates lung dendritic cell antigen uptake, transport, and T2/T17 cell priming in an intrinsic fashion, which in turn drives pulmonary granulocyte recruitment and contributes to development of airway hyperresponsiveness in chronic disease. CONCLUSIONS:We identify GM-CSF as a potentially novel therapeutic target in chronic lung inflammation, describing a GM-CSF-dependent lung conventional dendritic cell-T-cell-neutrophil axis that drives chronic lung disease. 10.1016/j.jaci.2020.12.638
    Neutrophilic asthma: misconception or misnomer? Nair Parameswaran,Surette Michael G,Virchow J Christian The Lancet. Respiratory medicine 10.1016/S2213-2600(21)00023-0
    Mapping atopic dermatitis and anti-IL-22 response signatures to type 2-low severe neutrophilic asthma. Badi Yusef Eamon,Pavel Ana B,Pavlidis Stelios,Riley John H,Bates Stewart,Kermani Nazanin Zounemat,Knowles Richard,Kolmert Johan,Wheelock Craig E,Worsley Sally,Uddin Mohib,Alving Kjell,Bakke Per S,Behndig Annelie,Caruso Massimo,Chanez Pascal,Fleming Louise J,Fowler Stephen J,Frey Urs,Howarth Peter,Horváth Ildikó,Krug Norbert,Maitland-van der Zee Anke H,Montuschi Paolo,Roberts Graham,Sanak Marek,Shaw Dominick E,Singer Florian,Sterk Peter J,Djukanovic Ratko,Dahlen Sven-Eric,Guo Yi-Ke,Chung Kian Fan,Guttman-Yassky Emma,Adcock Ian M, The Journal of allergy and clinical immunology BACKGROUND:Transcriptomic changes in patients who respond clinically to biological therapies may identify responses in other tissues or diseases. OBJECTIVE:We sought to determine whether a disease signature identified in atopic dermatitis (AD) is seen in adults with severe asthma and whether a transcriptomic signature for patients with AD who respond clinically to anti-IL-22 (fezakinumab [FZ]) is enriched in severe asthma. METHODS:An AD disease signature was obtained from analysis of differentially expressed genes between AD lesional and nonlesional skin biopsies. Differentially expressed genes from lesional skin from therapeutic superresponders before and after 12 weeks of FZ treatment defined the FZ-response signature. Gene set variation analysis was used to produce enrichment scores of AD and FZ-response signatures in the Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes asthma cohort. RESULTS:The AD disease signature (112 upregulated genes) encompassing inflammatory, T-cell, T2, and T17/T22 pathways was enriched in the blood and sputum of patients with asthma with increasing severity. Patients with asthma with sputum neutrophilia and mixed granulocyte phenotypes were the most enriched (P < .05). The FZ-response signature (296 downregulated genes) was enriched in asthmatic blood (P < .05) and particularly in neutrophilic and mixed granulocytic sputum (P < .05). These data were confirmed in sputum of the Airway Disease Endotyping for Personalized Therapeutics cohort. IL-22 mRNA across tissues did not correlate with FZ-response enrichment scores, but this response signature correlated with T22/IL-22 pathways. CONCLUSIONS:The FZ-response signature in AD identifies severe neutrophilic asthmatic patients as potential responders to FZ therapy. This approach will help identify patients for future asthma clinical trials of drugs used successfully in other chronic diseases. 10.1016/j.jaci.2021.04.010
    Role of the NLRP3 inflammasome in asthma: Relationship with neutrophilic inflammation, obesity, and therapeutic options. Williams Evan J,Negewo Netsanet A,Baines Katherine J The Journal of allergy and clinical immunology 10.1016/j.jaci.2021.04.022
    Plasma proteins elevated in severe asthma despite oral steroid use and unrelated to Type-2 inflammation. The European respiratory journal RATIONALE:Asthma phenotyping requires novel biomarker discovery. OBJECTIVES:To identify plasma biomarkers associated with asthma phenotypes by application of a new proteomic panel to samples from two well-characterised cohorts of severe (SA) and mild-to-moderate (MMA) asthmatics, COPD subjects and healthy controls (HCs). METHODS:An antibody-based array targeting 177 proteins predominantly involved in pathways relevant to inflammation, lipid metabolism, signal transduction and extracellular matrix was applied to plasma from 525 asthmatics and HCs in the U-BIOPRED cohort, and 142 subjects with asthma and COPD from the validation cohort BIOAIR. Effects of oral corticosteroids (OCS) were determined by a 2-week, placebo-controlled OCS trial in BIOAIR, and confirmed by relation to objective OCS measures in U-BIOPRED. RESULTS:In U-BIOPRED, 110 proteins were significantly different, mostly elevated, in SA compared to MMA and HCs. 10 proteins were elevated in SA MMA in both U-BIOPRED and BIOAIR (alpha-1-antichymotrypsin, apolipoprotein-E, complement component 9, complement factor I, macrophage inflammatory protein-3, interleukin-6, sphingomyelin phosphodiesterase 3, TNF receptor superfamily member 11a, transforming growth factor-β and glutathione S-transferase). OCS treatment decreased most proteins, yet differences between SA and MMA remained following correction for OCS use. Consensus clustering of U-BIOPRED protein data yielded six clusters associated with asthma control, quality of life, blood neutrophils, high-sensitivity C-reactive protein and body mass index, but not Type-2 inflammatory biomarkers. The mast cell specific enzyme carboxypeptidase A3 was one major contributor to cluster differentiation. CONCLUSIONS:The plasma proteomic panel revealed previously unexplored yet potentially useful Type-2-independent biomarkers and validated several proteins with established involvement in the pathophysiology of SA. 10.1183/13993003.00142-2021
    Association of Differential Mast Cell Activation with Granulocytic Inflammation in Severe Asthma. Tiotiu Angelica,Badi Yusef,Kermani Nazanin Zounemat,Sanak Marek,Kolmert Johan,Wheelock Craig E,Hansbro Philip M,Dahlén Sven-Erik,Sterk Peter J,Djukanovic Ratko,Guo Yike,Mumby Sharon,Adcock Ian M,Chung Kian Fan, American journal of respiratory and critical care medicine Mast cells (MCs) play a role in inflammation and both innate and adaptive immunity, but their involvement in severe asthma (SA) remains undefined. We investigated the phenotypic characteristics of the U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Diseases Outcomes) asthma cohort by applying published MC activation signatures to the sputum cell transcriptome. Eighty-four participants with SA, 20 with mild/moderate asthma (MMA), and 16 healthy participants without asthma were studied. We calculated enrichment scores (ESs) for nine MC activation signatures by asthma severity, sputum granulocyte status, and three previously defined sputum molecular phenotypes or transcriptome-associated clusters (TACs) 1, 2, and 3 using gene set variation analysis. MC signatures except unstimulated, repeated FcεR1-stimulated and IFN-γ-stimulated signatures were enriched in SA. A FcεR1-IgE-stimulated and a single-cell signature from asthmatic bronchial biopsies were highly enriched in eosinophilic asthma and in the TAC1 molecular phenotype. Subjects with a high ES for these signatures had elevated sputum amounts of similar genes and pathways. IL-33- and LPS-stimulated MC signatures had greater ES in neutrophilic and mixed granulocytic asthma and in the TAC2 molecular phenotype. These subjects exhibited neutrophil, NF-κB (nuclear factor-κB), and IL-1β/TNF-α (tumor necrosis factor-α) pathway activation. The IFN-γ-stimulated signature had the greatest ES in TAC2 and TAC3 that was associated with responses to viral infection. Similar results were obtained in an independent ADEPT (Airway Disease Endotyping for Personalized Therapeutics) asthma cohort. Gene signatures of MC activation allow the detection of SA phenotypes and indicate that MCs can be induced to take on distinct transcriptional phenotypes associated with specific clinical phenotypes. IL-33-stimulated MC signature was associated with severe neutrophilic asthma, whereas IgE-activated MC was associated with an eosinophilic phenotype. 10.1164/rccm.202102-0355OC
    T17 cells and corticosteroid insensitivity in severe asthma. Xie Yan,Abel Peter W,Casale Thomas B,Tu Yaping The Journal of allergy and clinical immunology Asthma is classically described as having either a type 2 (T2) eosinophilic phenotype or a non-T2 neutrophilic phenotype. T2 asthma usually responds to classical bronchodilation therapy and corticosteroid treatment. Non-T2 neutrophilic asthma is often more severe. Patients with non-T2 asthma or late-onset T2 asthma show poor response to the currently available anti-inflammatory therapies. These therapeutic failures result in increased morbidity and cost associated with asthma and pose a major health care problem. Recent evidence suggests that some non-T2 asthma is associated with elevated T17 cell immune responses. T17 cells producing Il-17A and IL-17F are involved in the neutrophilic inflammation and airway remodeling processes in severe asthma and have been suggested to contribute to the development of subsets of corticosteroid-insensitive asthma. This review explores the pathologic role of T17 cells in corticosteroid insensitivity of severe asthma and potential targets to treat this endotype of asthma. 10.1016/j.jaci.2021.12.769
    Steroid-induced fibroblast growth factors drive an epithelial-mesenchymal inflammatory axis in severe asthma. Science translational medicine Asthma and inflammatory airway diseases restrict airflow in the lung, compromising gas exchange and lung function. Inhaled corticosteroids (ICSs) can reduce inflammation, control symptoms, and improve lung function; however, a growing number of patients with severe asthma do not benefit from ICS. Using bronchial airway epithelial brushings from patients with severe asthma or primary human cells, we delineated a corticosteroid-driven fibroblast growth factor (FGF)-dependent inflammatory axis, with FGF-responsive fibroblasts promoting downstream granulocyte colony-stimulating factor (G-CSF) production, hyaluronan secretion, and neutrophilic inflammation. Allergen challenge studies in mice demonstrate that the ICS, fluticasone propionate, inhibited type 2-driven eosinophilia but induced a concomitant increase in FGFs, G-CSF, hyaluronan, and neutrophil infiltration. We developed a model of steroid-induced neutrophilic inflammation mediated, in part, by induction of an FGF-dependent epithelial-mesenchymal axis, which may explain why some individuals do not benefit from ICS. In further proof-of-concept experiments, we found that combination therapy with pan-FGF receptor inhibitors and corticosteroids prevented both eosinophilic and steroid-induced neutrophilic inflammation. Together, these results establish FGFs as therapeutic targets for severe asthma patients who do not benefit from ICS. 10.1126/scitranslmed.abl8146
    Deficiency of leukocyte-specific protein 1 (LSP1) alleviates asthmatic inflammation in a mouse model. Respiratory research BACKGROUND:Asthma is a major cause of morbidity and mortality in humans. The mechanisms of asthma are still not fully understood. Leukocyte-specific protein-1 (LSP-1) regulates neutrophil migration during acute lung inflammation. However, its role in asthma remains unknown. METHODS:An OVA-induced mouse asthma model in LSP1-deficient (Lsp1) and wild-type (WT) 129/SvJ mice were used to test the hypothesis that the absence of LSP1 would inhibit airway hyperresponsiveness and lung inflammation. RESULTS:Light and electron microscopic immunocytochemistry and Western blotting showed that, compared with normal healthy lungs, the levels of LSP1 were increased in lungs of OVA-asthmatic mice. Compared to Lsp1 OVA mice, WT OVA mice had higher levels of leukocytes in broncho-alveolar lavage fluid and in the lung tissues (P < 0.05). The levels of OVA-specific IgE but not IgA and IgG1 in the serum of WT OVA mice was higher than that of Lsp1 OVA mice (P < 0.05). Deficiency of LSP1 significantly reduced the levels of IL-4, IL-5, IL-6, IL-13, and CXCL1 (P < 0.05) but not total proteins in broncho-alveolar lavage fluid in asthmatic mice. The airway hyper-responsiveness to methacholine in Lsp1 OVA mice was improved compared to WT OVA mice (P < 0.05). Histology revealed more inflammation (inflammatory cells, and airway and blood vessel wall thickening) in the lungs of WT OVA mice than in those of Lsp1 OVA mice. Finally, immunohistology showed localization of LSP1 protein in normal and asthmatic human lungs especially associated with the vascular endothelium and neutrophils. CONCLUSION:These data show that LSP1 deficiency reduces airway hyper-responsiveness and lung inflammation, including leukocyte recruitment and cytokine expression, in a mouse model of asthma. 10.1186/s12931-022-02078-7
    Mechanisms of non-type 2 asthma. Hudey Stephanie N,Ledford Dennis K,Cardet Juan Carlos Current opinion in immunology Non-type 2 inflammation (Non-T2)-mediated asthma is difficult to define due to lack of signature biomarkers. It exists in the absence of T2-high or eosinophilic inflammation and includes neutrophilic and paucigranulocytic subtypes. Several cell types and cytokines, including Th1, Th17, IL-6, and IL-17, contribute to mechanisms of non-T2 asthma. Neutrophil extracellular traps (NETs) and inflammasome activation likely play a role in severe neutrophilic asthma. Several mechanisms lead to uncoupling of airway hyperresponsiveness and remodeling from airway inflammation in paucigranulocytic asthma. Recent research on transcriptomics and proteomics in non-T2 asthma is discussed in this review. Investigations of specific drug therapies for non-T2 asthma have been disappointing, and remain an important area for future clinical studies. 10.1016/j.coi.2020.10.002
    Molecular mechanisms of oxidative stress in asthma. Molecular aspects of medicine The lungs are exposed to reactive oxygen species oxygen (ROS) produced as a result of inhalation of oxygen, as well as smoke and other air pollutants. Cell metabolism and the NADPH oxidases (Nox) generate low levels of intracellular ROS that act as signal transduction mediators by inducing oxidative modifications of histones, enzymes and transcription factors. Redox signalling is also regulated by localised production and sensing of ROS in mitochondria, the endoplasmic reticulum (ER) and inside the nucleus. Intracellular ROS are maintained at low levels through the action of a battery of enzymatic and non-enzymatic antioxidants. Asthma is a heterogeneous airway inflammatory disease with different immune endotypes; these include atopic or non-atopic Th2 type immune response associated with eosinophilia, or a non-Th2 response associated with neutrophilia. Airway remodelling and hyperresponsiveness accompany the inflammatory response in asthma. Over-production of ROS resulting from infiltrating immune cells, particularly eosinophils and neutrophils, and a concomitant impairment of antioxidant responses lead to development of oxidative stress in asthma. Oxidative stress is augmented in severe asthma and during exacerbations, as well as by air pollution and obesity, and causes oxidative damage of tissues promoting airway inflammation and hyperresponsiveness. Furthermore, deregulated Nox activity, mitochondrial dysfunction, ER stress and/or oxidative DNA damage, resulting from exposure to irritants, inflammatory mediators or obesity, may lead to redox-dependent changes in cell signalling. ROS play a central role in airway epithelium-mediated sensing, development of innate and adaptive immune responses, and airway remodelling and hyperresponsiveness. Nonetheless, antioxidant compounds have proven clinically ineffective as therapeutic agents for asthma, partly due to issues with stability and in vivo metabolism of these compounds. The compartmentalised nature of ROS production and sensing, and the role of ROS in homeostatic responses and in the action of corticosteroids and β2-adrenergic receptor agonists, adds another layer of complexity to antioxidant therapy development. Nox inhibitors and mitochondrial-targeted antioxidants are in clinical development for a number of diseases but they have not yet been investigated in asthma. A better understanding of the complex role of ROS in the pathogenesis of asthma will highlight new opportunities for more targeted and effective redox therapies. 10.1016/j.mam.2021.101026
    Role of Cellular Metabolism in the Formation of Neutrophil Extracellular Traps in Airway Diseases. Frontiers in immunology Neutrophil extracellular traps (NETs) are a recently described mechanism of neutrophils that play an important role in health and disease. NETs are an innate defense mechanism that participate in clearance of pathogens, but they may also cause collateral damage in unrelated host tissues. Neutrophil dysregulation and NETosis occur in multiple lung diseases, such as pathogen-induced acute lung injury, pneumonia, chronic obstructive pulmonary disease (COPD), severe asthma, cystic fibrosis, and recently, the novel coronavirus SARS-CoV-2. More recently, research into immunometabolism has surged due to the possibility of reprogramming metabolism in order to modulate immune functions. The present review analyzes the different metabolic pathways associated with NETs formation, and how these impact on pathologies of the airways. 10.3389/fimmu.2022.850416
    Neutrophil extracellular traps in chronic lung disease: implications for pathogenesis and therapy. Keir Holly R,Chalmers James D European respiratory review : an official journal of the European Respiratory Society Neutrophilic inflammation has a key role in the pathophysiology of multiple chronic lung diseases. The formation of neutrophil extracellular traps (NETs) has emerged as a key mechanism of disease in neutrophilic lung diseases including asthma, COPD, cystic fibrosis and, most recently, bronchiectasis. NETs are large, web-like structures composed of DNA and anti-microbial proteins that are able to bind pathogens, prevent microbial dissemination and degrade bacterial virulence factors. The release of excess concentrations of proteases, antimicrobial proteins, DNA and histones, however, also leads to tissue damage, impaired mucociliary clearance, impaired bacterial killing and increased inflammation. A number of studies have linked airway NET formation with greater disease severity, increased exacerbations and overall worse disease outcomes across the spectrum of airway diseases. Treating neutrophilic inflammation has been challenging in chronic lung disease because of the delicate balance between reducing inflammation and increasing the risk of infections through immunosuppression. Novel approaches to suppressing NET formation or the associated inflammation are in development and represent an important therapeutic target. This review will discuss the relationship between NETs and the pathophysiology of cystic fibrosis, asthma, COPD and bronchiectasis, and explore the current and future development of NET-targeting therapies. 10.1183/16000617.0241-2021
    The enigmatic role of the neutrophil in asthma: Friend, foe or indifferent? Snelgrove R J,Patel D F,Patel T,Lloyd C M Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology Whilst severe asthma has classically been categorized as a predominantly Th2-driven pathology, there has in recent years been a paradigm shift with the realization that it is a heterogeneous disease that may manifest with quite disparate underlying inflammatory and remodelling profiles. A subset of asthmatics, particularly those with a severe, corticosteroid refractory disease, present with a prominent neutrophilic component. Given the potential of neutrophils to impart extensive tissue damage and promote inflammation, it has been anticipated that these cells are closely implicated in the underlying pathophysiology of severe asthma. However, uncertainty persists as to why the neutrophil is present in the asthmatic lung and what precisely it is doing there, with evidence supporting its role as a protagonist of pathology being primarily circumstantial. Furthermore, our view of the neutrophil as a primitive, indiscriminate killer has evolved with the realization that neutrophils can exhibit a marked anti-inflammatory, pro-resolving and wound healing capacity. We suggest that the neutrophil likely exhibits pleiotropic and potentially conflicting roles in defining asthma pathophysiology-some almost certainly detrimental and some potentially beneficial-with context, timing and location all critical confounders. Accordingly, indiscriminate blockade of neutrophils with a broad sword approach is unlikely to be the answer, but rather we should first seek to understand their complex and multifaceted roles in the disease state and then target them with the same subtleties and specificity that they themselves exhibit. 10.1111/cea.13191