Airway soluble CSF1R predicts progression in patients with idiopathic pulmonary fibrosis.

This study provides the first evidence for a role of airway sCSF1R in IPF https://bit.ly/3KTBrCA.

Immuno-proteomic profiling reveals aberrant immune cell regulation in the airways of individuals with ongoing post-COVID-19 respiratory disease.

Some patients hospitalized with acute COVID-19 suffer respiratory symptoms that persist for many months. We delineated the immune-proteomic landscape in the airways and peripheral blood of healthy controls and post-COVID-19 patients 3 to 6 months after hospital discharge. Post-COVID-19 patients showed abnormal airway (but not plasma) proteomes, with an elevated concentration of proteins associated with apoptosis, tissue repair, and epithelial injury versus healthy individuals. Increased numbers of cytotoxic lymphocytes were observed in individuals with greater airway dysfunction, while increased B cell numbers and altered monocyte subsets were associated with more widespread lung abnormalities. A one-year follow-up of some post-COVID-19 patients indicated that these abnormalities resolved over time. In summary, COVID-19 causes a prolonged change to the airway immune landscape in those with persistent lung disease, with evidence of cell death and tissue repair linked to the ongoing activation of cytotoxic T cells.

Autoantibodies are present in the bronchoalveolar lavage but not circulation in patients with fibrotic interstitial lung disease.

BACKGROUND: Fibrotic interstitial lung disease (fILD) has previously been associated with the presence of autoantibody. While studies have focused on systemic autoimmunity, the role of local autoantibodies in the airways remains unknown. We therefore extensively characterised the airway and peripheral autoantibody profiles in patients with fILD, and assessed association with disease severity and outcome. METHODS: Bronchoalveolar lavage (BAL) fluid was collected from a cohort of fILD patients and total BAL antibody concentrations were quantified. An autoantigen microarray was used to measure IgG and IgA autoantibodies against 122 autoantigens in BAL from 40 idiopathic pulmonary fibrosis (IPF), 20 chronic hypersensitivity pneumonitis (CHP), 20 connective tissue disease-associated ILD (CTD-ILD) patients and 20 controls. RESULTS: A subset of patients with fILD but not healthy controls had a local autoimmune signature in their BAL that was not present systemically, regardless of disease. The proportion of patients with IPF with a local autoantibody signature was comparable to that of CTD-ILD, which has a known autoimmune pathology, identifying a potentially novel subset of patients. The presence of an airway autoimmune signature was not associated with reduced survival probability or changes in lung function in the cohort as a whole. Patients with IPF had increased BAL total IgA and IgG1 while subjects with CHP had increased BAL IgA, IgG1 and IgG4. In patients with CHP, increased BAL total IgA was associated with reduced survival probability. CONCLUSION: Airway autoantibodies that are not present systemically identify a group of patients with fILD and the mechanisms by which these autoantibodies contribute to disease requires further investigation.

DNA Methylome Alterations Are Associated with Airway Macrophage Differentiation and Phenotype during Lung Fibrosis.

Rationale: Airway macrophages (AMs) are key regulators of the lung environment and are implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF), a fatal respiratory disease with no cure. However, knowledge about the epigenetics of AMs in IPF is limited. Objectives: To assess the role of epigenetic regulation of AMs during lung fibrosis. Methods: We undertook DNA methylation (DNAm) profiling by using Illumina EPIC (850k) arrays in sorted AMs from healthy donors (n = 14) and donors with IPF (n = 30). Cell-type deconvolution was performed by using reference myeloid-cell DNA methylomes. Measurements and Main Results: Our analysis revealed that epigenetic heterogeneity was a key characteristic of IPF AMs. DNAm "clock" analysis indicated that epigenetic alterations in IPF AMs were not associated with accelerated aging. In differential DNAm analysis, we identified numerous differentially methylated positions (n = 11) and differentially methylated regions (n = 49) between healthy and IPF AMs, respectively. Differentially methylated positions and differentially methylated regions encompassed genes involved in lipid (LPCAT1 [lysophosphatidylcholine acyltransferase 1]) and glucose (PFKFB3 [6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3]) metabolism, and importantly, the DNAm status was associated with disease severity in IPF. Conclusions: Collectively, our data identify that changes in the epigenome are associated with the development and function of AMs in the IPF lung.

The Respiratory Microbiome in Chronic Hypersensitivity Pneumonitis Is Distinct from That of Idiopathic Pulmonary Fibrosis.

Rationale: Chronic hypersensitivity pneumonitis (CHP) is a condition that arises after repeated exposure and sensitization to inhaled antigens. The lung microbiome is increasingly implicated in respiratory disease, but, to date, no study has investigated the composition of microbial communities in the lower airways in CHP.Objectives: To characterize and compare the airway microbiome in subjects with CHP, subjects with idiopathic pulmonary fibrosis (IPF), and control subjects.Methods: We prospectively recruited individuals with a CHP diagnosis (n = 110), individuals with an IPF diagnosis (n = 45), and control subjects (n = 28). Subjects underwent BAL and bacterial DNA was isolated, quantified by quantitative PCR and the 16S ribosomal RNA gene was sequenced to characterize the bacterial communities in the lower airways.Measurements and Main Results: Distinct differences in the microbial profiles were evident in the lower airways of subjects with CHP and IPF. At the phylum level, the prevailing microbiota of both subjects with IPF and subjects with CHP included Firmicutes, Bacteroidetes, Proteobacteria, and Actinobacteria. However, in IPF, Firmicutes dominated, whereas the percentage of reads assigned to Proteobacteria in the same group was significantly lower than the percentage found in subjects with CHP. At the genus level, the Staphylococcus burden was increased in CHP, and Actinomyces and Veillonella burdens were increased in IPF. The lower airway bacterial burden in subjects with CHP was higher than that in control subjects but lower than that of those with IPF. In contrast to IPF, there was no association between bacterial burden and survival in CHP.Conclusions: The microbial profile of the lower airways in subjects with CHP is distinct from that of IPF, and, notably, the bacterial burden in individuals with CHP fails to predict survival.

Itaconate controls the severity of pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease in which airway macrophages (AMs) play a key role. Itaconate has emerged as a mediator of macrophage function, but its role during fibrosis is unknown. Here, we reveal that itaconate is an endogenous antifibrotic factor in the lung. Itaconate levels are reduced in bronchoalveolar lavage, and itaconate-synthesizing cis-aconitate decarboxylase expression (ACOD1) is reduced in AMs from patients with IPF compared with controls. In the murine bleomycin model of pulmonary fibrosis, Acod1−/− mice develop persistent fibrosis, unlike wild-type (WT) littermates. Profibrotic gene expression is increased in Acod1−/− tissue-resident AMs compared with WT, and adoptive transfer of WT monocyte-recruited AMs rescued mice from disease phenotype. Culture of lung fibroblasts with itaconate decreased proliferation and wound healing capacity, and inhaled itaconate was protective in mice in vivo. Collectively, these data identify itaconate as critical for controlling the severity of lung fibrosis, and targeting this pathway may be a viable therapeutic strategy.

Global expression and CpG methylation analysis of primary endothelial cells before and after TNFa stimulation reveals gene modules enriched in inflammatory and infectious diseases and associated DMRs.

Endothelial cells are a primary site of leukocyte recruitment during inflammation. An increase in tumor necrosis factor-alpha (TNFa) levels as a result of infection or some autoimmune diseases can trigger this process. Several autoimmune diseases are now treated with TNFa inhibitors. However, genomic alterations that occur as a result of TNF-mediated inflammation are not well understood. To investigate molecular targets and networks resulting from increased TNFa, we measured DNA methylation and gene expression in 40 human umbilical vein endothelial cell primary cell lines before and 24 hours after stimulation with TNFa via microarray. Weighted gene co-expression network analysis identified 15 gene groups (modules) with similar expression correlation patterns; four modules showed a strong association with TNFa treatment. Genes in the top TNFa-associated module were all up-regulated, had the highest proportion of hypomethylated regions, and were associated with 136 Disease Ontology terms, including autoimmune/inflammatory, infectious and cardiovascular diseases, and cancers. They included chemokines CXCL1, CXCL10 and CXCL8, and genes associated with autoimmune diseases including HLA-C, DDX58, IL4, NFKBIA and TNFAIP3. Cardiovascular and metabolic disease genes, including APOC1, ACLY, ELOVL6, FASN and SCD, were overrepresented in a module that was not associated with TNFa treatment. Of 223 hypomethylated regions identified, several were in promoters of autoimmune disease GWAS loci (ARID5B, CD69, HDAC9, IL7R, TNIP1 and TRAF1). Results reveal specific gene groups acting in concert in endothelial cells, delineate those driven by TNFa, and establish their relationship to DNA methylation changes, which has strong implications for understanding disease etiology and precision medicine approaches.

Bacterial burden in the lower airways predicts disease progression in idiopathic pulmonary fibrosis and is independent of radiological disease extent.

Increasing bacterial burden in the lower airways of patients with idiopathic pulmonary fibrosis confers an increased risk of disease progression and mortality. However, it remains unclear whether this increased bacterial burden directly influences progression of fibrosis or simply reflects the magnitude of the underlying disease extent or severity.We prospectively recruited 193 patients who underwent bronchoscopy and received a multidisciplinary diagnosis of idiopathic pulmonary fibrosis. Quantification of the total bacterial burden in bronchoalveolar lavage fluid was performed by 16S rRNA gene qPCR. Imaging was independently evaluated by two readers assigning quantitative scores for extent, severity and topography of radiographic changes and relationship of these features with bacterial burden was assessed.Increased bacterial burden significantly associated with disease progression (HR 2.1; 95% CI 1.287-3.474; p=0.0028). Multivariate stepwise regression demonstrated no relationship between bacterial burden and radiological features or extent of disease. When specifically considering patients with definite or probable usual interstitial pneumonia there was no difference in bacterial burden between these two groups. Despite a postulated association between pleuroparenchymal fibroelastosis and clinical infection, there was no relationship between either the presence or extent of pleuroparenchymal fibroelastosis and bacterial burden.We demonstrate that bacterial burden in the lower airways is not simply secondary to the extent of the underlying architectural destruction of the lung parenchyma seen in idiopathic pulmonary fibrosis. The independent nature of this association supports a relationship with the underlying pathogenic mechanisms and highlights the urgent need for functional studies.

Dynamics of human monocytes and airway macrophages during healthy aging and after transplant.

The ontogeny of airway macrophages (AMs) in human lung and their contribution to disease are poorly mapped out. In mice, aging is associated with an increasing proportion of peripherally, as opposed to perinatally derived AMs. We sought to understand AM ontogeny in human lung during healthy aging and after transplant. We characterized monocyte/macrophage populations from the peripheral blood and airways of healthy volunteers across infancy/childhood (2-12 yr), maturity (20-50 yr), and older adulthood (>50 yr). Single-cell RNA sequencing (scRNA-seq) was performed on airway inflammatory cells isolated from sex-mismatched lung transplant recipients. During healthy aging, the proportions of blood bronchoalveolar lavage (BAL) classical monocytes peak in adulthood and decline in older adults. scRNA-seq of BAL cells from lung transplant recipients indicates that after transplant, the majority of AMs are recipient derived. These data show that during aging, the peripheral monocyte phenotype is consistent with that found in the airways and, furthermore, that the majority of human AMs after transplant are derived from circulating monocytes.

The Transferrin Receptor CD71 Delineates Functionally Distinct Airway Macrophage Subsets during Idiopathic Pulmonary Fibrosis.

Rationale: Idiopathic pulmonary fibrosis (IPF) is a devastating progressive disease with limited therapeutic options. Airway macrophages (AMs) are key components of the defense of the airways and are implicated in the pathogenesis of IPF. Alterations in iron metabolism have been described during fibrotic lung disease and in murine models of lung fibrosis. However, the role of transferrin receptor 1 (CD71)-expressing AMs in IPF is not known. Objectives: To assess the role of CD71-expressing AMs in the IPF lung. Methods: We used multiparametric flow cytometry, gene expression analysis, and phagocytosis/transferrin uptake assays to delineate the role of AMs expressing or lacking CD71 in the BAL of patients with IPF and of healthy control subjects. Measurements and Main Results: There was a distinct increase in proportions of AMs lacking CD71 in patients with IPF compared with healthy control subjects. Concentrations of BAL transferrin were enhanced in IPF-BAL, and furthermore, CD71- AMs had an impaired ability to sequester transferrin. CD71+ and CD71- AMs were phenotypically, functionally, and transcriptionally distinct, with CD71- AMs characterized by reduced expression of markers of macrophage maturity, impaired phagocytosis, and enhanced expression of profibrotic genes. Importantly, proportions of AMs lacking CD71 were independently associated with worse survival, underlining the importance of this population in IPF and as a potential therapeutic target. Conclusions: Taken together, these data highlight how CD71 delineates AM subsets that play distinct roles in IPF and furthermore show that CD71- AMs may be an important pathogenic component of fibrotic lung disease.