Pathogenesis Of Severe Asthma Research Articles

Cellular senescence-Related genes define the immune microenvironment and molecular characteristics in severe asthma patients

Recent studies have shown that cellular senescence is involved in the pathogenesis of severe asthma (SA). The objective of this study was to investigate the role of cellular senescence-related genes (CSGs) in the pathogenesis of SA. Here, 54 differentially expressed CSGs were identified in SA patients compared to healthy control individuals. Among the 54 differentially expressed CSGs, 3 CSGs (ETS2, ETS1 and AURKA) were screened using the LASSO regression analysis and logistic regression analysis to establish the CSG-based prediction model to predict severe asthma. Moreover, we found that the protein expression levels of ETS2, ETS1 and AURKA were increased in the severe asthma mouse model. Then, two distinct senescence subtypes of SA with distinct immune microenvironments and molecular biological characteristics were identified. Cluster 1 was characterized by increased infiltration of immature dendritic cells, regulatory T cells, and other cells. Cluster 2 was characterized by increased infiltration levels of eosinophils, neutrophils, and other cells. The molecular biological characteristics of Cluster 1 included aerobic respiration and oxidative phosphorylation, whereas the molecular biological characteristics of Cluster 2 included activation of the immune response and immune receptor activity. Then, we established an Random Forest model to predict the senescence subtypes of SA to guide treatment. Finally, potential drugs were searched for each senescence subgroup of SA patients via the Connectivity Map database. A peroxisome proliferator-activated receptor agonist may be a potential therapeutic drug for patients in Cluster 1, whereas a tachykinin antagonist may be a potential therapeutic drug for patients in Cluster 2. In summary, CSGs are likely involved in the pathogenesis of SA, which may lead to new therapeutic options for SA patients.

Aerobic physical training reduces severe asthma phenotype involving kinins pathway.

Aerobic physical training (APT) reduces eosinophilic airway inflammation, but its effects and mechanisms in severe asthma remain unknown. An in vitro study employing key cells involved in the pathogenesis of severe asthma, such as freshly isolated human eosinophils, neutrophils, and bronchial epithelial cell lineage (BEAS-2B) and lung fibroblasts (MRC-5 cells), was conducted. Additionally, an in vivo study using male C57Bl/6 mice, including Control (Co; n = 10), Trained (Exe; n = 10), house dust mite (HDM; n = 10), and HDM + Trained (HDM + Exe; n = 10) groups, was carried out, with APT performed at moderate intensity, 5x/week, for 4 weeks. HDM and bradykinin, either alone or in combination, induced hyperactivation in human neutrophils, eosinophils, BEAS-2B, and MRC-5 cells. In contrast, IL-10, the primary anti-inflammatory molecule released during APT, inhibited these inflammatory effects, as evidenced by the suppression of numerous cytokines and reduced mRNA expression of the B1 receptor and ACE-2. The in vivo study demonstrated that APT decreased bronchoalveolar lavage levels of bradykinin, IL-1β, IL-4, IL-5, IL-17, IL-33, TNF-α, and IL-13, while increasing levels of IL-10, klotho, and IL-1RA. APT reduced the accumulation of polymorphonuclear cells, lymphocytes, and macrophages in the peribronchial space, as well as collagen fiber accumulation, epithelial thickness, and mucus accumulation. Furthermore, APT lowered the expression of the B1 receptor and ACE-2 in lung tissue and reduced bradykinin levels in the lung tissue homogenate compared to the HDM group. It also improved airway resistance, tissue resistance, and tissue damping. On a systemic level, APT reduced total leukocytes, eosinophils, neutrophils, basophils, lymphocytes, and monocytes in the blood, as well as plasma levels of IL-1β, IL-4, IL-5, IL-17, TNF-α, and IL-33, while elevating the levels of IL-10 and IL-1RA. These findings indicate that APT inhibits the severe asthma phenotype by targeting kinin signaling.

Activated non-neuronal cholinergic system correlates with non-type 2 inflammation and exacerbations in severe asthma

BackgroundNon-neuronal cholinergic system (NNCS) contributes to various inflammatory airway diseases. However, the role of NNCS in severe asthma (SA) remains largely unexplored. ObjectiveTo explore airway NNCS in SA. MethodsIn this prospective cohort study based on the Australasian Severe Asthma Network in a real-world setting, patients with SA (n = 52) and non-SA (n = 104) underwent clinical assessment and sputum induction. The messenger RNA (mRNA) levels of NNCS components and proinflammatory cytokines in the sputum were detected using real-time quantitative polymerase chain reaction, and the concentrations of acetylcholine (Ach)-related metabolites were evaluated using liquid chromatography coupled with tandem mass spectrometry. Asthma exacerbations were prospectively investigated during the next 12 months. The association between NNCS and future asthma exacerbations was also analyzed. ResultsPatients with SA were less controlled and had worse airway obstruction, a lower bronchodilator response, higher doses of inhaled corticosteroids, and more add-on treatments. The sputum mRNA levels of NNCS components, such as muscarinic receptors M1R-M5R, OCT3, VACHT, and ACHE; proinflammatory cytokines; and Ach concentration in the SA group were significantly higher than those in the non-SA group. Furthermore, most NNCS components positively correlated with non-type (T) 2 inflammatory profiles, such as sputum neutrophils, IL8, and IL1B. In addition, the mRNA levels of sputum M2R, M3R, M4R, M5R, and VACHT were independently associated with an increased risk of moderate-to-severe asthma exacerbations. ConclusionThis study indicated that the NNCS was significantly activated in SA, leading to elevated Ach and was associated with clinical features, non-T2 inflammation, and future exacerbations of asthma, highlighting the potential role of the NNCS in the pathogenesis of SA. Clinical Trial RegistrationChiCTR-OOC-16009529 (http://www.chictr.org.cn).

The role of mitochondria in eosinophil function: implications for severe asthma pathogenesis.

Mitochondria are key metabolic hubs involved in cellular energy production and biosynthesis. ATP is generated primarily by glucose and fatty acid oxidation through the tricarboxylic acid (TCA) cycle and oxidative phosphorylation (OXPHOS) in the mitochondria. During OXPHOS there is also production of reactive oxygen species (ROS), which are involved in the regulation of cellular function. Mitochondria are also central in the regulating cell survival and death, particularly in the intrinsic apoptosis pathway. Severe asthma is a heterogeneous disease driven by various immune mechanisms. Severe eosinophilic asthma entails a type 2 inflammatory response and peripheral and lung eosinophilia, associated with severe airflow obstruction, frequent exacerbations and poor response to treatment. Mitochondrial dysfunction and altered metabolism have been observed in airway epithelial and smooth muscle cells from patients with asthma. However, the role of mitochondria in the development of eosinophilia and eosinophil-mediated inflammation in severe asthma is unknown. In this review, we discuss the currently limited literature on the role of mitochondria in eosinophil function and how it is regulated by asthma-relevant cytokines, including interleukin (IL)-5 and granulocyte-macrophage colony-stimulating factor (GM-CSF), as well as by corticosteroid drugs. Moreover, we summarise the evidence on the role of mitochondria in the regulation of eosinophils apoptosis and eosinophil extracellular trap formation. Finally, we discuss the possible role of altered mitochondrial function in eosinophil dysfunction in severe asthma and suggest possible research avenues in order to better understand their role in disease pathogenesis, and identify novel therapeutic targets.

Exploring the Autoimmune Pathogenesis in Severe Asthma

Introduction: Severe asthma has a poor response to hormone therapy and a poor level of control, so the discovery of new pathogenetic mechanisms is important for diagnosing and treating severe asthma. IL-35 may play a protective role in autoimmune diseases by directly or indirectly inhibiting the secretion of IL-17, which is an important proinflammatory factor involved in the occurrence and development of autoimmune diseases. The autologous serum skin test (ASST) is a good sensitivity and specificity screening test for autoimmune functional autoantibodies. We compared the levels of IL-35 and IL-17 in serum samples, the positive rate of ASST, the level of exhaled nitric oxide (FeNO), and the atopic constitution in patients with severe asthma to those with mild-to-moderate asthma so as to explore the possible autoimmune pathogenesis of severe asthma. Methods: Patients with mild-to-moderate and severe asthma were enrolled. Their age, gender, smoking history, family history of asthma, history of allergic rhinitis, positive allergen results, serum total IgE (TlgE), allergen-specific IgE (slgE), routine blood, ASST results, and FeNO test results were compared and analyzed. The IL-35 and IL-17 levels in serum samples from both groups were measured by enzyme-linked immunosorbent assay for comparison and analysis. The SPSS 22.0 software package was used for statistical analysis. Results: A total of 50 patients with mild-to-moderate asthma and 31 patients with severe asthma were included in this study. The proportion of patients with a history of smoking and a family history of asthma was significantly higher in the severe asthma group compared to the mild-to-moderate asthma group (all p < 0.05); the number of positive allergen tests was significantly lower in patients with severe asthma compared to those with mild-to-moderate asthma (p < 0.001). The rate of positive ASST was significantly higher in patients with severe asthma than in patients with mild-to-moderate asthma (p < 0.05). Serum IL-17 levels were significantly higher in patients with severe asthma than in patients with mild-to-moderate asthma (p < 0.05), but serum IL-35 level between the two group was not significantly different (p = 0.113). ASST-positive patients had a statistically significant increase in the risk of developing severe asthma, while patients with allergen positive were less likely to develop severe asthma (positive ASST: OR = 5.277, p = 0.024; allergen positivity: OR = 0.123, p = 0.001). Conclusions: IL-35 has a weaker inhibitory effect on high IL-17 expression in patients with severe asthma, and the rate of positive ASST was significantly higher in patients with severe asthma, which all suggested the possibility of autoimmune pathogenesis in patients with severe asthma.

Construction of lncRNA-miRNA-mRNA regulatory network in severe asthmatic bronchial epithelial cells: A bioinformatics study.

Asthma is a chronic respiratory disease caused by environment-host interactions. Bronchial epithelial cells (BECs) are the first line of defense against environmental toxins. However, the mechanisms underlying the role of BECs in severe asthma (SA) are not yet fully understood. Long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) have been shown to play important roles in the regulation of gene expression in the pathogenesis of SA. In this study, bioinformatics was used for the first time to reveal the lncRNA-miRNA-mRNA regulatory network of BECs in SA. Five mRNA datasets of bronchial brushing samples from patients with SA and healthy controls (HC) were downloaded from the Gene Expression Omnibus (GEO) database. A combination of the Venn diagram and robust rank aggregation (RRA) method was used to identify core differentially expressed genes (DEGs). Protein-protein interaction (PPI) analysis of core DEGs was performed to screen hub genes. The miRDB, miRWalk, and ENCORI databases were used to predict the miRNA-mRNA relationships, and the ENCORI and starBase v2.0 databases were used to predict the upstream lncRNAs of the miRNA-mRNA relationships. Four core DEGs were identified: carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM5), interleukin-1 receptor type 2 (IL1R2), trefoil factor 3 (TFF3), and vascular endothelial growth factor A (VEGFA). These 4 core DEGs indicated that SA was not significantly associated with sex. Enrichment analysis showed that the MAPK, Rap1, Ras, PI3K-Akt and Calcium signaling pathways may serve as the principal pathways of BECs in SA. A lncRNA-miRNA-mRNA regulatory network of the severe asthmatic bronchial epithelium was constructed. The top 10 competing endogenous RNAs (ceRNAs) were FGD5 antisense RNA 1 (FGD5-AS1), metastasis associated lung adenocarcinoma transcript 1 (MALAT1), X inactive specific transcript (XIST), HLA complex group 18 (HCG18), small nucleolar RNA host gene 16 (SNHG16), has-miR-20b-5p, has-miR-106a-5p, hsa-miR-106b-5p, has-miR-519d-3p and Fms related receptor tyrosine kinase 1 (FLT1). Our study revealed a potential mechanism for the lncRNA-miRNA-mRNA regulatory network in BECs in SA.

Serum Zonulin Is a Biomarker for Severe Asthma.

Zonulin is a regulator of epithelial and endothelial barrier function. It regulates intestinal permeability through disrupting tight junctions. Defective epithelial barrier function is a hallmark of airway inflammation in asthma. This study aimed to investigate the role of zonulin in the pathogenesis of severe asthma. We enrolled 56 adult patients with asthma (29 severe asthma and 27 mild-to-moderate asthma) and 33 normal controls. The clinical data, sera, and lung tissues of the patients were provided by the Cohort for Reality and Evolution of adult Asthma in Korea (COREA) and the Biobank of Soonchunhyang University Bucheon Hospital, South Korea. Serum zonulin levels were estimated using an enzyme-linked immunosorbent assay, and zonulin expression in the bronchial tissue was evaluated by immunohistochemical staining. The serum zonulin levels were significantly higher in patients with severe asthma (51.98 ± 19.66 ng/mL) than in those with mild-to-moderate asthma and normal controls (26.35 ± 13.70 vs. 17.26 ± 10.29 ng/mL, P < 0.001). They significantly correlated with percent predicted forced expiratory volume in one second (%FEV1) (r = -0.35, P = 0.009). The zonulin expression in the bronchial epithelium was greater in patients with severe asthma. A serum zonulin cutoff value to distinguish between severe and mild-to-moderate asthmatics was 38.83 ng/mL. Zonulin may play an important role in the pathogenesis of severe asthma, and serum zonulin could be a potential biomarker for severe asthma.

Tissue Inhibitor of Metalloproteinase-1 Enhances Eosinophilic Airway Inflammation in Severe Asthma.

Severe asthma (SA) is characterized by persistent airway inflammation and remodeling, followed by lung function decline. The present study aimed to evaluate the role of tissue inhibitor of metalloproteinase-1 (TIMP-1) in the pathogenesis of SA. We enrolled 250 adult asthmatics (54 with SA and 196 with non-SA) and 140 healthy controls (HCs). Serum TIMP-1 levels were determined by enzyme-linked immunosorbent assay. The release of TIMP-1 from airway epithelial cells (AECs) in response to stimuli as well as the effects of TIMP-1 on the activations of eosinophils and macrophages were evaluated in vitro and in vivo. Significantly higher levels of serum TIMP-1 were noted in asthmatics than in HCs, in the SA group than in non-SA group, and in the type 2 SA group than in non-type 2 SA group (P < 0.01 for all). A negative correlation between serum TIMP-1 and FEV1% values (r = -0.400, P = 0.003) was noted in the SA group. In vitro study demonstrated that TIMP-1 was released from AECs in response to poly I:C, IL-13, eosinophil extracellular traps (EETs) and in coculture with eosinophils. TIMP-1-stimulated mice showed eosinophilic airway inflammation, which was not completely suppressed by steroid treatment. In vitro and in vivo functional studies showed that TIMP-1 directly activated eosinophils and macrophages, and induced the release of EETs and macrophages to polarize toward M2 subset, which was suppressed by anti-TIMP-1 antibody. These findings suggest that TIMP-1 enhances eosinophilic airway inflammation and that serum TIMP-1 may be a potential biomarker and/or therapeutic target for type 2 SA.

MiR-365-3p is a negative regulator in IL-17-mediated asthmatic inflammation.

BackgroundInterleukin-17, the major proinflammatory cytokine secreted by Th17 cells, makes essential contribution to pathogenesis of severe asthma, while the detailed mechanisms, especially the involvement of microRNAs which are also important participants in asthma progression, remains largely unclear.MethodsIn this study, we established a house dust mite (HDM) extract-induced murine asthmatic models and the miRNA expression in the lung tissues of mice were profiled by miRNA microarray assay. The effect of miR-365-3p on IL-17-mediated inflammation was examined by qRT-PCR and immunoblotting analysis. The involvement of ARRB2 as target gene of miR-365-3p was verified by overexpression or RNA interference.ResultsHDM extract-induced asthmatic inflammation was proved to be IL17-mediated and miR-365-3p was screened out to be the only miRNA exclusively responsive to IL-17. miR-365-3p, whose expression was significantly downregulated upon IL-17 stimulation, was demonstrated to exert remarkable anti-inflammatory effect to decrease IL-17-provoked inflammatory cytokines (KC/IL-8 and IL-6) in both airway epithelial cells and macrophages of murine and human origins, verifying its universal antagonizing activity against IL-17-initiated inflammation across the two species. ARRB2 was characterized as the key target of miR-365-3p to negate IL-17-induced inflammatory cytokines.ConclusionTaken together, our data supported the notion that miR-365-3p, which was diminished by IL-17 in murine and human asthmatic pathogenesis, functioned as an essential negative mediator in IL-17-stimuated inflammatory response by targeting ARRB2, which would shed new light to the understanding and therapeutics thereof of asthmatic inflammation.

Spray-Dried and Spray-Freeze-Dried Powder Formulations of an Anti-Interleukin-4Rα Antibody for Pulmonary Delivery.

The therapeutic options for severe asthma are limited, and the biological therapies are all parenterally administered. The purpose of this study was to formulate a monoclonal antibody that targets the receptor for IL-4, an interleukin implicated in the pathogenesis of severe asthma, into a dry powder intended for delivery via inhalation. Dehydration was achieved using either spray drying or spray freeze drying, which exposes the thermolabile biomacromolecules to stresses such as shear and adverse temperatures. 2-hydroxypropyl-beta-cyclodextrin was incorporated into the formulation as protein stabiliser and aerosol performance enhancer. The powder formulations were characterised in terms of physical and aerodynamic properties, while the antibody was assessed with regard to its structural stability, antigen-binding ability, and in vitro biological activity after drying. The spray-freeze-dried formulations exhibited satisfactory aerosol performance, with emitted fraction exceeding 80% and fine particle fraction of around 50%. The aerosolisation of the spray-dried powders was hindered possibly by high residual moisture. Nevertheless, the antigen-binding ability and inhibitory potency were unaffected for the antibody in the selected spray-dried and spray-freeze-dried formulations, and the antibody was physically stable even after one-year storage at ambient conditions. The findings of this study establish the feasibility of developing an inhaled dry powder formulation of an anti-IL-4R antibody using spray drying and spray freeze drying techniques with potential for the treatment of severe asthma.

Pathways linked to unresolved inflammation and airway remodelling characterize the transcriptome in two independent severe asthma cohorts.

Background and objectiveSevere asthma (SA) is a heterogeneous disease. Transcriptomic analysis contributes to the understanding of pathogenesis necessary for developing new therapies. We sought to identify and validate mechanistic pathways of SA across two independent cohorts.MethodsTranscriptomic profiles from U‐BIOPRED and Australian NOVocastrian Asthma cohorts were examined and grouped into SA, mild/moderate asthma (MMA) and healthy controls (HCs). Differentially expressed genes (DEGs), canonical pathways and gene sets were identified as central to SA mechanisms if they were significant across both cohorts in either endobronchial biopsies or induced sputum.ResultsThirty‐six DEGs and four pathways were shared across cohorts linking to tissue remodelling/repair in biopsies of SA patients, including SUMOylation, NRF2 pathway and oxidative stress pathways. MMA presented a similar profile to HCs. Induced sputum demonstrated IL18R1 as a shared DEG in SA compared with healthy subjects. We identified enrichment of gene sets related to corticosteroid treatment; immune‐related mechanisms; activation of CD4+ T cells, mast cells and IL18R1; and airway remodelling in SA.ConclusionOur results identified differentially expressed pathways that highlight the role of CD4+ T cells, mast cells and pathways linked to ongoing airway remodelling, such as IL18R1, SUMOylation and NRF2 pathways, as likely active mechanisms in the pathogenesis of SA.

Establishment method and experimental application of the mouse model of severe asthma

Severe asthma remains the bottleneck of clinical treatment. In order to deeply analyze the pathogenesis of severe asthma, so as to develop the new targeted drugs and evaluate the curative effects, it is urgent to establish an ideal animal model of severe asthma, which is of great significance to simulate the pathophysiological and clinical characteristics of patients with severe asthma and study the potential therapeutic targets of severe asthma. Here we summarized the establishment method and experimental application of the mouse model of severe asthma.

Cross-talk of four types of RNA modification writers defines the immune microenvironment in severe asthma.

Adenine modifications, including m6 A, m1 A, APA, and A-to-I modifications, are the most impactful RNA modifications. These modifications are primarily produced by enzymes called writers. The main purpose of this study was to explore the cross-talk and potential roles of these writers in severe asthma. We found 13 RNA writers potentially related to severe asthma and three RNA modification patterns. Cluster 3 showed predominant neutrophil infiltration and C-type lectin receptor signaling; cluster 1 showed predominant innate immune cell infiltration and ubiquitin-proteasome system activation; and cluster 2 did not show obvious immune infiltration characteristics. We found that RNA modification writers modified immune cell-related genes and led to both accumulation of different immune cells in the airways and activation of a series of biological processes, which ultimately leads to severe asthma. TRMT6, WTAP, and TRMT6A were included in a random forest model as predictors. Cromoglicic acid, thioperamide, and fluvastatin were potential drugs for clusters 1, 2, and 3, respectively. We found that cross-talk of RNA modifications is significant in severe asthma, which provides insight into severe asthma pathogenesis and possible treatment avenues.

Cytotoxic CD4+ tissue-resident memory T cells are associated with asthma severity.

Abstract Patients with severe uncontrolled asthma represent a distinct endotype with persistent airway inflammation and remodeling that is refractory to corticosteroid treatment. To determine T cell subsets and effector molecules that drive pathogenesis of severe asthma, we performed single-cell transcriptome analysis of &amp;gt;50,000 airway CD4+ T cells isolated from bronchoalveolar lavage (BAL) samples from 30 patients with mild and severe asthma. We observed striking heterogeneity in the nature of CD4+ T cells present in asthmatics' airways with tissue-resident memory (TRM) cells making a dominant contribution. Notably, in severe asthmatics a subset of CD4+ TRM cells (CD103-expressing) was significantly increased, comprising nearly 65% of all CD4+ T cells in the airways of male patients with severe asthma when compared to mild asthma (13%). This subset was enriched for transcripts linked to T cell receptor (TCR) activation (HLA-DRB1, HLA-DPA1, CD40LG) and cytotoxicity (GZMB, GZMH), and following stimulation expressed high levels of transcripts encoding for pro-inflammatory non-TH2 cytokines (CCL3, CCL4, CCL5, TNF, LIGHT) that could fuel persistent airway inflammation and remodeling. Our findings indicate the need to look beyond the traditional T2 model of severe asthma to better understand the heterogeneity of this disease. This work was supported by (i) NIH research grant R01HL114093 (PV) and equipment grants (S10RR027366 - BD FACSAria Fusion, and S10OD025052 - Illumina Novaseq 6000); the William K. Bowes Jr. Foundation (P.V.). The WATCH study is supported by the Southampton NIHR Biomedical Research Centre and the Southampton NIHR Clinical Research Facility which are funded by the NIHR and are a partnership between the University of Southampton and University Hospital Southampton NHS Foundation Trust.

Decreased expression of airway epithelial Axl is associated with eosinophilic inflammation in severe asthma

BackgroundAirway epithelium-derived cytokines are critical to provoke and perpetuate type 2 inflammation in asthma. Yet it is poorly understood how this epithelial cell-driven inflammatory response is negatively regulated. We previously reported that Axl receptor tyrosine kinase was expressed by basal cells in the airway epithelium and had a role in defining their stem cell identity. However, whether and how Axl regulates airway type 2 inflammation remains unknown. MethodsWe performed immunofluorescence staining to compare Axl expression in airway epithelium between non-asthmatic subjects, mild-moderate asthma and severe asthma. We confirmed this result by interrogating public databases of global gene expression in endobronchial biopsies. We then quantified eosinophil numbers infiltrating into the trachea of wild-type or Axl-knockout mice that were intranasally treated with house dust mite extracts (HDM). Cell-based assays using siRNA targeting Axl were further performed to identify molecules involved in Axl-mediated regulation of inflammation. ResultsHistological assessments and transcriptome analyses revealed decreases in protein and mRNA of Axl in airway basal cells of severe asthmatics. This reduction of Axl expression was correlated with infiltration of eosinophils and mast cells in severe asthmatics. Eosinophil infiltration was more evident in the trachea of Axl-knockout mice in response to repetitive HDM administration. siRNA-mediated knockdown of Axl increased mRNA and protein expression of granulocyte macrophage-colony stimulating factor (GM-CSF) in human bronchial epithelial cells. ConclusionsAxl kinase expressed by basal cells may suppress excessive eosinophilic inflammation via inhibition of GM-CSF in the airway. Axl reduction has clinical implications for the pathogenesis of severe asthma.

In Silico Bioinformatics Followed by Molecular Validation Using Archival FFPE Tissue Biopsies Identifies a Panel of Transcripts Associated with Severe Asthma and Lung Cancer.

Simple SummaryThe present study identified a panel of transcripts involved in the pathogenesis of both severe asthma and lung cancer. The genes identified using publicly available transcriptomics data were validated on cell lines, plasma samples, and archival tissue biopsies from asthmatic and lung cancer patients. The functional roles of the identified markers in both the diseases were ascertained from the literature. These molecular markers might be useful for diagnosing lung cancer at early stages.Severe asthma and lung cancer are both heterogeneous pathological diseases affecting the lung tissue. Whilst there are a few studies that suggest an association between asthma and lung cancer, to the best of our knowledge, this is the first study to identify common genes involved in both severe asthma and lung cancer. Publicly available transcriptomic data for 23 epithelial brushings from severe asthmatics and 55 samples of formalin-fixed paraffin-embedded (FFPE) lung cancer tissue at relatively early stages were analyzed by absolute gene set enrichment analysis (GSEA) in comparison to 37 healthy bronchial tissue samples. The key pathways enriched in asthmatic patients included adhesion, extracellular matrix, and epithelial cell proliferation, which contribute to tissue remodeling. In the lung cancer dataset, the main pathways identified were receptor tyrosine kinase signaling, wound healing, and growth factor response, representing the early cancer pathways. Analysis of the enriched genes derived from the pathway analysis identified seven genes expressed in both the asthma and lung cancer sets: BCL3, POSTN, PPARD, STAT1, MYC, CD44, and FOSB. The differential expression of these genes was validated in vitro in the cell lines retrieved from different lung cancer and severe asthma patients using real-time PCR. The effect of the expression of the seven genes identified in the study on the overall survival of lung cancer patients (n = 1925) was assessed using a Kaplan–Meier plot. In vivo validation performed in the archival biopsies obtained from patients diagnosed with both the disease conditions provided interesting insights into the pathogenesis of severe asthma and lung cancer, as indicated by the differential expression pattern of the seven transcripts in the mixed group as compared to the asthmatics and lung cancer samples alone.

Nrf2 regulates downstream genes by targeting miR-29b in severe asthma and the role of grape seed proanthocyanidin extract in a murine model of steroid-insensitive asthma

Context Grape seed proanthocyanidin extract (GSPE) is effective in treating severe asthma (SA). Objective To examine the relationship between Nrf2-miR-29b axis and SA, and to detect whether preventive use of GSPE relieves SA via it. Materials and methods We recruited 10 healthy controls, 10 patients with non-severe asthma (nSA), and 9 patients with SA from February 2017 to December 2017. Peripheral blood mononuclear cells from these volunteers were extracted. A murine model of steroid-insensitive asthma was established in six-week-old female BALB/c mice that were sensitised and challenged with OVA, Al(OH)3 and LPS for 31 days. Mice in the treated groups were injected with DXM (5 mg/kg/d), with or without GSPE (100 mg/kg/d). Control group received PBS. We performed quantitative real-time PCR, western blot and luciferase reporter assay in animal and cell models. Results SA group demonstrated significantly lower concentrations of Nrf2 protein, Nrf2 mRNA, and miR-29b than nSA group and control group. Conversely, higher levels of platelet derived growth factor C (PDGFC), phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1), and collagen type III alpha 1 (COL3A1) were measured in SA than in the other two groups. PDGFC, PIK3R1, and COL3A1 were the target genes of miR-29b. GSPE + DXM significantly elevated the expression of Nrf2 (+188%), Nrf2 mRNA (+506%), and miR-29b (+201%), and significantly reduced the expression of PDGFC (−72%), PIK3R1 (−40%), and COL3A1 (−65%) compared with OVA + LPS. Conclusions Nrf2-miR-29b axis is involved in the pathogenesis of SA. GSPE, as an adjuvant drug, maybe a potential therapeutic agent for SA.

RNA-Binding Protein HuR Promotes Airway Inflammation in a House Dust Mite-Induced Allergic Asthma Model.

Mounting evidence indicates that interleukin 17 (IL-17) is critically involved in the pathogenesis of severe asthma. We have previously reported that upon IL-17 stimulation, Act1, an IL-17-receptor-complex adaptor, directly binds to its target mRNAs and utilizes other proteins, such as HuR, to upregulate mRNA stability and translation. HuR mRNA targets include multiple asthma-related genes. In this study, we have used house dust mite (HDM), a natural allergen, to test the role of HuR in the pathogenesis of allergic asthma. We found that HuR deletion in airway epithelium diminished HDM-induced lung inflammation, including neutrophil and eosinophil infiltration. While Th2 cytokines were not altered, the production of CXCL1, CXCL5 and CCL11 chemokines was significantly diminished. Airway smooth muscle (ASM) cells contribute to the pathogenesis of allergic asthma by orchestrating inflammatory and remodeling responses.We found that IL-17 treatment of ASM cells induced translocation of HuR from nucleus to cytoplasm, where it bound directly to Cxcl1 and Ccl11 mRNA. Deletion of HuR in ASM cells decreased their proliferation as well as CXCL1 and CCL11 production in response to IL-17. Taken together, our findings demonstrate the importance of HuR-mediated regulation of gene expression to the pathogenesis of allergic asthma, in both airway epithelial and ASM cells.

Epithelial Autoantigen-Specific IgG Antibody Enhances Eosinophil Extracellular Trap Formation in Severe Asthma.

PurposeThere have been autoimmune mechanisms for the pathogenesis of severe asthma (SA) involving epithelial autoantigen-specific antibodies. This study aimed to find the function of these antibodies in the formation of eosinophil extracellular traps (EETs), contributing to the development of SA.MethodsPatients with SA (n = 11), those with patients with nonsevere asthma (NSA, n = 41), and healthy controls (HCs, n = 26) were recruited to evaluate levels of epithelial antigens and autoantigen-specific antibodies. Moreover, the significance of epithelial autoantigen-specific antibodies in association with EET production was investigated ex vivo and in vivo.ResultsSignificantly higher levels of serum cytokeratin (CK) 18 and CK18-specific IgG were observed in patients with SA than in those with NSA (P = 0.001 and P = 0.031, respectively), while no differences were found in serum CK19 or CK19-specific immunoglobulin G (IgG). Moreover, levels of serum CK18 were positively correlated with total eosinophil counts (r = 0.276, P = 0.048) in asthmatics, while a negative correlation was noted between levels of serum CK18 and forced expiratory volume in 1 second (FEV1) %. In the presence of CK18-specific IgG, peripheral eosinophils from asthmatics released EETs, which further increased CK18 production from airway epithelial cells. In severe asthmatic mice, CK18 expression and CK18-specific IgG production were enhanced in the lungs, where EET treatment enhanced CK18 expression and CK18-specific IgG production, either of which was not suppressed by dexamethasone.ConclusionsThese suggest that EETs could enhance epithelial autoantigen (CK18)-induced autoimmune responses, further stimulating EET production and type 2 airway responses, which is a new therapeutic target for SA.

Investigation of Janus Kinase (JAK) Inhibitors for Lung Delivery and the Importance of Aldehyde Oxidase Metabolism

The Janus family of tyrosine kinases (JAK1, JAK2, JAK3, and TYK2) play an essential role in the receptor signaling of cytokines that have been implicated in the pathogenesis of severe asthma, and there is emerging interest in the development of small-molecule-inhaled JAK inhibitors as treatments. Here, we describe the optimization of a quinazoline series of JAK inhibitors and the results of mouse lung pharmacokinetic (PK) studies where only low concentrations of parent compound were observed. Subsequent investigations revealed that the low exposure was due to metabolism by aldehyde oxidase (AO), so we sought to identify quinazolines that were not metabolized by AO. We found that specific substituents at the quinazoline 2-position prevented AO metabolism and this was rationalized through computational docking studies in the AO binding site, but they compromised kinome selectivity. Results presented here highlight that AO metabolism is a potential issue in the lung.