G‐CSFR antagonism reduces mucosal injury and airways fibrosis in a virus‐dependent model of severe asthma

Abstract

Background and Purpose: Asthma is a chronic disease that displays heterogeneous clinical and molecular features. A phenotypic subset of late-onset severe asthmatics has debilitating fixed airflow obstruction, increased neutrophilic inflammation and a history of pneumonia. Influenza A virus (IAV) is an important viral cause of pneumonia and asthmatics are frequently hospitalised during IAV epidemics. This study aims to determine whether antagonising granulocyte colony stimulating factor receptor (G-CSFR) prevents pneumonia-associated severe asthma. Experimental Approach: Mice were sensitised to house dust mite (HDM) to establish allergic airway inflammation and subsequently infected with IAV (HKx31/H3N2 subtype). A neutralising monoclonal antibody against G-CSFR was therapeutically administered. Key Results: In IAV-infected mice with prior HDM sensitisation, a significant increase in airway fibrotic remodelling and airways hyper-reactivity was observed. A mixed granulocytic inflammatory profile consisting of neutrophils, macrophages and eosinophils was prominent and at a molecular level, G-CSF expression was significantly increased in HDMIAV-treated mice. Blockage of G-CSFR reduced neutrophilic inflammation in the bronchoalveolar and lungs by over 80% in HDMIAV-treated mice without altering viral clearance. Markers of NETosis (dsDNA and myeloperoxidase in bronchoalveolar), tissue injury (LDH activity in bronchoalveolar) and oedema (total bronchoalveolar-fluid protein) were also significantly reduced with anti-G-CSFR treatment. In addition, anti-G-CSFR antagonism significantly reduced bronchoalveolar gelatinase activity, active TFGβ lung levels, collagen lung expression, airways fibrosis and airways hyper-reactivity in HDMIAV-treated mice. Conclusions and Implications: We have shown that antagonising G-CSFR-dependent neutrophilic inflammation reduced pathological disruption of the mucosal barrier and airways fibrosis in an IAV-induced severe asthma model.

Blocking the human common beta subunit of the GM-CSF, IL-5 and IL-3 receptors markedly reduces hyperinflammation in ARDS models

“…The rest of the lungs were snap-frozen in liquid nitrogen prior to −80 °C storage. Myeloperoxidase (MPO) activity in lung tissue homogenates was performed as previously described [ 23 ]. Total protein levels in the BAL fluid were quantified using the BCA protein assay (Thermofisher Scientific, US) and dsDNA levels in the BAL fluid were measured using the Quant-iT™ PicoGreen™ dsDNA Assay Kit (Thermofisher Scientific, US).…”

“…Lung cells were pelleted, and red blood cells were lysed with ACK lysis buffer. After blocking with CD16/CD32 antibody, cells were then stained with a myeloid cell antibody cocktail containing FITC-CD45, PE-Siglec F, APC-F4/80, eFluor 450-CD11b, PE/Cy7-CD11c, PerCp/eFluor710-Ly6G and LIVE/DEAD™ Fixable Yellow Dead Cell Stain, or a lymphoid cell antibody cocktail containing APC/eFluor 780-CD45, PerCP/Cyanine5.5-CD3e, PE/Cy7-NK-1.1 (BD, US), PE-CD8a (BioLegend, US), PE/eFluor 610-CD4, Alexa Fluor 488-FOXP3 (BioLegend, US) and LIVE/DEAD™ Fixable Violet Dead Cell Stain as previously described [ 23 , 26 , 27 ]. A Foxp3 / Transcription Factor Staining Buffer Set was used for cell permeabilization before Foxp3 staining.…”

“…To determine lung viral load, qPCR on IAV polymerase A subunit ( PA ) gene was performed using customed TaqMan probes and normalized to Gapdh . Lung viral load is expressed as fold change vs saline controls [ 23 ].…”

The dual effect of MBD2 regulating Th2 cell differentiation

“…Asthma is one of the most common chronic immunological diseases in humans, affecting people from childhood to old age. Reversible airflow obstruction, airway hyperresponsiveness (AHR) and airway inflammation are distinguishing feature of asthma，and a phenotypic subset of late-onset severe asthmatics has debilitating fixed airflow obstruction, increased neutrophilic inflammation and a history of pneumonia [1]. Eosinophilic airway inflammation and the imbalance between Th1/Th2 cells [2] are generally recognized as pivotal factors in the pathogenesis of asthma.…”

Correction

“…The uncorrected version of the paper “G‐CSFR antagonism reduces mucosal injury and airways fibrosis in a virus‐dependent model of severe asthma” by Wang et al (2021) was inadvertently published in the issue.…”