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The extracellular matrix (ECM) is not just a three-dimensional scaffold that provides stable support for all cells in the lungs, but also an important component of chronic fibrotic airway, vascular, and interstitial diseases. It is a bioactive entity that is dynamically modulated during tissue homeostasis and disease, that controls structural and immune cell functions and drug responses, and that can release fragments that have biological activity and that can be used to monitor disease activity. There is a growing recognition of the importance of considering ECM changes in chronic airway, vascular, and interstitial diseases, including 1) compositional changes, 2) structural and organizational changes, and 3) mechanical changes and how these affect disease pathogenesis. As altered ECM biology is an important component of many lung diseases, disease models must incorporate this factor to fully recapitulate disease-driver pathways and to study potential novel therapeutic interventions. Although novel models are evolving that capture some or all of the elements of the altered ECM microenvironment in lung diseases, opportunities exist to more fully understand cell–ECM interactions that will help devise future therapeutic targets to restore function in chronic lung diseases. In this perspective article, we review evolving knowledge about the ECM's role in homeostasis and disease in the lung. [ABSTRACT FROM AUTHOR]

We estimated the particle number distributions (PNDs), particle number concentrations (PNCs), physicochemical characteristics, meteorological effects, and respiratory deposition doses (RDD) in the human respiratory tract for three different particle modes: nucleation (N6–30), accumulation (N30–300), and coarse (N300–10,000) modes. This study was conducted in three different microenvironments (MEs) in London (indoor, IN; traffic intersection, TI; park, PK) measuring particles in the range of 6 nm–10,000 nm using an electrical low-pressure impactor (ELPI+). Mean PNCs were 1.68 ± 1.03 × 104 #cm−3, 7.00 ± 18.96 × 104 #cm−3, and 0.76 ± 0.95 × 104 #cm−3 at IN, TI, and PK, respectively. The PNDs were high for nucleation-mode particles at the TI site, especially during peak traffic hours. Wind speeds ranging from 0 to 6 ms−1 exhibit higher PNCs for nucleation- and accumulation-mode particles at TI and PK sites. Physicochemical characterisation shows trace metals, including Fe, O, and inorganic elements, that were embedded in a matrix of organic material in some samples. Alveolar RDD was higher for the nucleation and accumulation modes than the coarse-mode particles. The chemical signatures from the physicochemical characterisation indicate the varied sources at different MEs. These findings enhance our understanding of the different particle profiles at each ME and should help devise ways of reducing personal exposure at each ME. [ABSTRACT FROM AUTHOR]

COVID-19, caused by SARS-CoV-2, requires new approaches to control the disease. Programmed cell death protein (PD-1) and cytotoxic T-lymphocyte–associated protein 4 (CTLA4) play important roles in T-cell exhaustion in severe COVID-19. This study evaluated the frequency of whole blood lymphocytes expressing PD-1 and CTLA-4 in COVID-19 patients upon admission to the intensive care unit (ICU) (i.e., severe) or infection ward (i.e., moderate) and after 7 days of antiviral therapy. COVID-19 patients were treated with either favipiravir or Kaletra (FK group, 11 severe and 11 moderate) or dexamethasone plus remdesivir (DR group, 7 severe and 10 moderate) for 7 days in a pilot study. Eight healthy control subjects were also enrolled. The frequency of PD-1 + and CTLA4 + lymphocytes in whole blood was evaluated by flow cytometry. Patients on DR therapy had shorter hospital stays than those on FK therapy. The frequency of PD-1 + lymphocytes in the FK group at baseline differed between COVID-19 patients and healthy controls, while the frequency of both PD-1 + and CTLA-4 + cells increased significantly 7 days of FK therapy. The response was similar in both moderate and severe patients. In contrast, the frequency of PD-1 + and CTLA-4 + lymphocytes varied significantly between patients and healthy controls before DR treatment. DR therapy enhanced PD-1 + but not the CTLA-4 + frequency of these cells after 7 days. We show that the frequency of PD-1 and CTAL-4-bearing lymphocytes during hospitalization was increased in Iranian ICU COVID-19 patients who received FK treatment, but that the frequency of CTLA-4 + cells was higher at baseline and did not increase in patients who received DR. The effectiveness of DR treatment may reflect differences in T-cell activation or exhaustion status, particularly in CTLA-4-expressing cells. [ABSTRACT FROM AUTHOR]

Background: Interleukin (IL)‐33 is an upstream regulator of type 2 (T2) eosinophilic inflammation and has been proposed as a key driver of some asthma phenotypes. Objective: To derive gene signatures from in vitro studies of IL‐33‐stimulated cells and use these to determine IL‐33‐associated enrichment patterns in asthma. Methods: Signatures downstream of IL‐33 stimulation were derived from our in vitro study of human mast cells and from public datasets of in vitro stimulated human basophils, type 2 innate lymphoid cells (ILC2), regulatory T cells (Treg) and endothelial cells. Gene Set Variation Analysis (GSVA) was used to probe U‐BIOPRED and ADEPT sputum transcriptomics to determine enrichment scores (ES) for each signature according to asthma severity, sputum granulocyte status and previously defined molecular phenotypes. Results: IL‐33‐activated gene signatures were cell‐specific with little gene overlap. Individual signatures, however, were associated with similar signalling pathways (TNF, NF‐κB, IL‐17 and JAK/STAT signalling) and immune cell differentiation pathways (Th17, Th1 and Th2 differentiation). ES for IL‐33‐activated gene signatures were significantly enriched in asthmatic sputum, particularly in patients with neutrophilic and mixed granulocytic phenotypes. IL‐33 mRNA expression was not elevated in asthma whereas the expression of mRNA for IL1RL1, the IL‐33 receptor, was up‐regulated in the sputum of severe eosinophilic asthma. The mRNA expression for IL1RAP, the IL1RL1 co‐receptor, was greatest in severe neutrophilic and mixed granulocytic asthma. Conclusions: IL‐33‐activated gene signatures are elevated in neutrophilic and mixed granulocytic asthma corresponding with IL1RAP co‐receptor expression. This suggests incorporating T2‐low asthma in anti‐IL‐33 trials. [ABSTRACT FROM AUTHOR]

Indoor, airborne, transmission of SARS-CoV-2 is a key infection route. We monitored fourteen different indoor spaces in order to assess the risk of SARS-CoV-2 transmission. PM2.5 and CO2 concentrations were simultaneously monitored in order to understand aerosol exposure and ventilation conditions. Average PM2.5 concentrations were highest in the underground station (261 ± 62.8 μgm−3), followed by outpatient and emergency rooms in hospitals located near major arterial roads (38.6 ± 20.4 μgm−3), the respiratory wards, medical day units and intensive care units recorded concentrations in the range of 5.9 to 1.1 μgm−3. Mean CO2 levels across all sites did not exceed 1000 ppm, the respiratory ward (788 ± 61 ppm) and the pub (bar) (744 ± 136 ppm) due to high occupancy. The estimated air change rates implied that there is sufficient ventilation in these spaces to manage increased levels of occupancy. The infection probability in the medical day unit of hospital 3, was 1.6-times and 2.2-times higher than the emergency and outpatient waiting rooms in hospitals 4 and 5, respectively. The temperature and relative humidity recorded at most sites was below 27 °C, and 40% and, in sites with high footfall and limited air exchange, such as the hospital medical day unit, indicate a high risk of airborne SARS-CoV-2 transmission. [ABSTRACT FROM AUTHOR]

The airway epithelium is remodelled in patients with SA and plays a pivotal role in orchestrating airway inflammation.1 However, airway epithelial cell-directed therapies have not been explored due to the lack of specific targets. CEACAM5 is an IL-13-regulated epithelial gene that mediates transcription in type-2 (T2) high severe asthma Severe asthma (SA) is a heterogeneous disease characterized by airflow restriction, frequent exacerbations and lack of effective treatments that improve quality of life and prevent exacerbations. [Extracted from the article]

Rationale: Mast cells (MCs) play a role in inflammation and both innate and adaptive immunity, but their involvement in severe asthma (SA) remains undefined. Objectives: 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. Methods: 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. Measurements and Main Results: 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. Conclusions: 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. [ABSTRACT FROM AUTHOR]

This letter to the editor has not an abstract. [ABSTRACT FROM AUTHOR]

Keywords: asthma; bioinformatics; endotypes; precision medicine; systems biology EN asthma bioinformatics endotypes precision medicine systems biology 380 383 4 01/09/21 20210101 NES 210101 I To the Editor, i Asthma is a complex heterogeneous disease that presents with varying degrees of severity. We identified four optimal clusters (TAC*1, TAC*2, TAC*3a and TAC*3b) (Figure S3), in agreement with our previous clustering1 where TAC*3a and TAC*3b were combined. Asthma, bioinformatics, endotypes, precision medicine, systems biology. [Extracted from the article]

Epidemiological and challenge studies in healthy subjects and in individuals with asthma highlight the health impact of environmental ozone even at levels considered safe. Acute ozone exposure in man results in sputum neutrophilia in 30% of subjects particularly young children, females, and those with ongoing cardiopulmonary disease. This may be associated with systemic inflammation although not in all cases. Chronic exposure amplifies these effects and can result in the formation of asthma-like symptoms and immunopathology. Asthmatic patients who respond to ozone (responders) induce a greater number of genes in bronchoalveolar (BAL) macrophages than healthy responders with up-regulation of inflammatory and immune pathways under the control of cytokines and chemokines and the enhanced expression of remodeling and repair programmes including those associated with protease imbalances and cell-cell adhesion. These pathways are under the control of several key transcription regulatory factors including nuclear factor (NF)-κB, anti-oxidant factors such as nuclear factor (erythroid-derived 2)-like 2 NRF2, the p38 mitogen activated protein kinase (MAPK), and priming of the immune system by up-regulating toll-like receptor (TLR) expression. Murine and cellular models of acute and chronic ozone exposure recapitulate the inflammatory effects seen in humans and enable the elucidation of key transcriptional pathways. These studies emphasize the importance of distinct transcriptional networks in driving the detrimental effects of ozone. Studies indicate the critical role of mediators including IL-1, IL-17, and IL-33 in driving ozone effects on airway inflammation, remodeling and hyperresponsiveness. Transcription analysis and proof of mechanisms studies will enable the development of drugs to ameliorate the effects of ozone exposure in susceptible individuals. [ABSTRACT FROM AUTHOR]

Background and Objective. Progressive pulmonary fibrosis is the main cause of death in patients with systemic sclerosis (SSc) with interstitial lung disease (ILD) and in those with idiopathic pulmonary fibrosis (IPF). Transforming growth factor-β (TGF-β) and NADPH oxidase- (NOX-) derived reactive oxygen species (ROS) are drivers of lung fibrosis. We aimed to determine the role of the epigenetic readers, bromodomain and extraterminal (BET) proteins in the regulation of redox balance in activated myofibroblasts. Methods. In TGF-β-stimulated fibroblasts, we investigated the effect of the BET inhibitor JQ1 on the mRNA expression of the prooxidant gene NOX4 and the antioxidant gene superoxide dismutase (SOD2) by quantitative RT-PCR, the antioxidant transcription factor NF-E2-related factor 2 (Nrf2) activity by a reporter assay, and intracellular ROS levels by dichlorofluorescein staining. Myofibroblast activation was determined by α-smooth muscle actin immunocytochemistry. The role of specific BET protein isoforms in NOX4 gene regulation was studied by siRNA silencing and chromatin-immunoprecipitation. Results and Conclusions. Affymetrix gene array analysis revealed increased NOX4 and reduced SOD2 expression in SSc and IPF fibroblasts. SOD2 silencing in non-ILD control fibroblasts induced a profibrotic phenotype. TGF-β increased NOX4 and inhibited SOD2 expression, while increasing ROS production and myofibroblast differentiation. JQ1 reversed the TGF-β-mediated NOX4/SOD2 imbalance and Nrf2 inactivation and attenuated ROS production and myofibroblast differentiation. The BET proteins Brd3 and Brd4 were shown to bind to the NOX4 promoter and drive TGF-β-induced NOX4 expression. Our data indicate a critical role of BET proteins in promoting redox imbalance and pulmonary myofibroblast activation and support BET bromodomain inhibitors as a potential therapy for fibrotic lung disease. [ABSTRACT FROM AUTHOR]

The multifunctional role of mast cells (MCs) in the immune system is complex and has not fully been explored. MCs reside in tissues and mucous membranes such as the lung, digestive tract, and skin which are strategically located at interfaces with the external environment. These cells, therefore, will encounter external stimuli and pathogens. MCs modulate both the innate and the adaptive immune response in inflammatory disorders including transplantation. MCs can have pro- and anti-inflammatory functions, thereby regulating the outcome of lung transplantation through secretion of mediators that allow interaction with other cell types, particularly innate lymphoid cells (ILC2). ILC2 cells are a unique population of hematopoietic cells that coordinate the innate immune response against a variety of threats including infection, tissue damage, and homeostatic disruption. In addition, MCs can modulate alloreactive T cell responses or assist in T regulatory (Treg) cell activity. This paper outlines the current understanding of the role of MCs in lung transplantation, with a specific focus on their interaction with ILC2 cells within the engrafted organ. [ABSTRACT FROM AUTHOR]

Exosomes are nanosized vesicles and have recently been recognized as important players in cell-to-cell communication. Exosomes contain different mediators such as proteins, nucleic acids (DNA, mRNA, miRNAs, and other ncRNAs), and lipid mediators and can shuttle their exosomal content to both neighboring and distal cells. Exosomes are very effective in orchestrating immune responses in the airways and all cell types can contribute to the systemic exosome pool. Intracellular communication between the broad range of cell types within the lung is crucial in disease emphasizing the importance of exosomes. In asthma, exosomes affect the inflammatory microenvironment which ultimately determines the development or alleviation of the pathological symptoms. Recent studies in this area have provided insight into the underlying mechanisms of disease and led to interest in using exosomes as potential novel therapeutic agents. [ABSTRACT FROM AUTHOR]

Chronic cough is associated with airway inflammation and remodelling. Abnormal airway smooth muscle cell (ASMC) function may underlie mechanisms of chronic cough. Our objective was to examine the transcriptome and focused secretome of ASMCs from chronic cough patients and healthy non-cough volunteers. ASMC gene expression profiling was performed at baseline and/or after stimulation with polyinosinic:polycytidylic acid (poly(I:C)) to mimic viral infection. Supernatants were collected formultiplex analysis. Our results showed no significant differentially expressed genes (DEGs, false discovery rate (FDR) <0.05) between chronic cough and healthy non-cough ASMCs at baseline. Poly(I:C) stimulation resulted in 212 DEGs (>1.5 fold-change, FDR <0.05) in ASMCs from chronic cough patients compared with 1674 DEGs in healthy non-cough volunteers. The top up-regulated genes included chemokine (C–X–Cmotif) ligand (CXCL) 11 (CXCL11), CXCL10, chemokine (C–Cmotif) ligand (CCL) 5 (CCL5) and interferon-induced protein 44 like (IFI44L) corresponding with inflammation and innate immune response pathways. ASMCs from cough subjects had enhanced activation of viral response pathways in response to poly(I:C) compared with healthy non-cough subjects, reduced activation of pathways involved in chronic inflammation and equivalent activation of neuroregulatory genes. The poly(I:C)-induced release of inflammatory mediators, including CXCL8, interleukin (IL)-6 and CXCL1, from ASMCs from cough patients was significantly impaired compared with healthy non-cough subjects. Addition of fluticasone propionate (FP) to poly(I:C)-treated ASMCs resulted in greater gene expression changes in healthy non-cough ASMCs. FP had a differential effect on poly(I:C)-induced mediator release between chronic cough and healthy non-cough volunteers. In conclusion, altered innate immune and inflammatory gene profiles within ASMCs, rather than infiltrating cells or nerves, may drive the cough response following respiratory viral infection. [ABSTRACT FROM AUTHOR]

Rationale: Nitric oxide synthase (NOS) is a biomarker/target in sepsis. NOS activity is driven by amino acids, which cycle to regulate the substrate L-arginine in parallel with cycles which regulate the endogenous inhibitors ADMA and L-NMMA. The relationship between amines and the consequence of plasma changes on iNOS activity in early sepsis is not known. Objective: Our objective was to apply a metabolomics approach to determine the influence of sepsis on a full array of amines and what consequence these changes may have on predicted iNOS activity. Methods and measurements: 34 amino acids were measured using ultra purification mass spectrometry in the plasma of septic patients (n = 38) taken at the time of diagnosis and 24–72 hours post diagnosis and of healthy volunteers (n = 21). L-arginine and methylarginines were measured using liquid-chromatography mass spectrometry and ELISA. A top down approach was also taken to examine the most changed metabolic pathways by Ingenuity Pathway Analysis. The iNOS supporting capacity of plasma was determined using a mouse macrophage cell-based bioassay. Main results: Of all the amines measured 22, including L-arginine and ADMA, displayed significant differences in samples from patients with sepsis. The functional consequence of increased ADMA and decreased L-arginine in context of all cumulative metabolic changes in plasma resulted in reduced iNOS supporting activity associated with sepsis. Conclusions: In early sepsis profound changes in amine levels were defined by dominant changes in the iNOS canonical pathway resulting in functionally meaningful changes in the ability of plasma to regulate iNOS activity ex vivo. [ABSTRACT FROM AUTHOR]

Objective: Endothelial dysfunction associated with systemic inflammation can contribute to organ injury/failure following cardiac surgery requiring cardiopulmonary bypass (CPB). Roundabout protein 4 (Robo4), an endothelial-expressed transmembrane receptor and regulator of cell activation, is an important inhibitor of endothelial hyper-permeability. We investigated the hypothesis that plasma levels of Robo4 are indicative of organ injury, in particular acute kidney injury (AKI), after cardiac surgery. Methods: Patients (n = 32) undergoing elective cardiac surgery with CPB were enrolled, prospectively. Plasma Robo4 concentrations were measured pre-, 2 and 24 h post-operatively, using a commercially available ELISA. Plasma and endothelial markers of inflammation [interleukin (IL) -6, -8, -10: von Willibrand factor (vWF) and angiopoeitin-2 (Ang-2)] and the AKI marker, neutrophil gelatinase-associated lipocalin (NGAL), were also measured by ELISA. Results: Plasma Robo4 increased significantly (p<0.001) from pre-operative levels of 2515±904 pg/ml to 4473±1915 pg/ml, 2 h after surgery; and returned to basal levels (2682±979 pg/ml) by 24 h. Plasma cytokines, vWF and NGAL also increased 2 h post-operatively and remained elevated at 24 h. Ang-2 increased 24 h post-operatively, only. There was a positive, significant correlation (r = 0.385, p = 0.0298) between Robo-4 and IL-10, but not other cytokines, 2 h post-operatively. Whilst raised Robo4 did not correlate with indices of lung dysfunction or other biomarkers of endothelial activation; there was a positive, significant correlation between raised (2 h) plasma NGAL and Robo4 (r = 0.4322, p = 0.0135). When patients were classed as AKI or non-AKI either using NGAL cut-off of 150 ng/ml, or the AKI Network (AKIN) clinical classification; plasma Robo4 was significantly higher (p = 0.0073 and 0.003, respectively) in AKI vs. non-AKI patients (NGAL cut-off: 5350±2191 ng/ml, n = 16 vs. 3595±1068 pg/ml, n = 16; AKIN: 6546 pg/ml, IQR 5025–8079, n = 6; vs. 3727 pg/ml, IQR 1962–3727, n = 26) subjects. Conclusion: Plasma Robo4 levels are increased, transiently, following cardiac surgery requiring CPB; and higher levels in patients with AKI suggest a link between endothelial dysregulation and onset of AKI. [ABSTRACT FROM AUTHOR]

Diffuse alveolar hemorrhage is characterized by the presence of red blood cells and free hemoglobin in the alveoli and complicates a number of serious medical and surgical lung conditions including the pulmonary vasculitides and acute respiratory distress syndrome. In this study we investigated the hypothesis that exposure of human alveolar epithelial cells to hemoglobin and its breakdown products regulates chemokine release via iron- and oxidant-mediated activation of the transcription factor NF-κB. Methemoglobin alone stimulated the release of IL-8 and MCP-1 from A549 cells via activation of the NF-κB pathway; additionally, IL-8 required ERK activation and MCP-1 required JNK activation. Neither antioxidants nor iron chelators and knockdown of ferritin heavy and light chains affected these responses, indicating that iron and reactive oxygen species are not involved in the response of alveolar epithelial cells to methemoglobin. Incubation of primary cultures of human alveolar type 2 cells with methemoglobin resulted in a similar pattern of chemokine release and signaling pathway activation. In summary, we have shown for the first time that methemoglobin induced chemokine release from human lung epithelial cells independent of iron- and redox-mediated signaling involving the activation of the NF-κB and MAPK pathways. Decompartmentalization of hemoglobin may be a significant proinflammatory stimulus in a variety of lung diseases. [ABSTRACT FROM AUTHOR]