American Journal of Respiratory Cell and Molecular Biology最新文献

Endurance exercise is broadly beneficial to cardiopulmonary function, with these benefits thought to be driven by extrapulmonary factors rather than direct structural changes in the lungs. Thus, to address how endurance exercise training and sex impact molecular responses in the lungs, we used a multi-omics approach to study 6-month-old Fischer 344 rats that undertook a progressive endurance treadmill training protocol for 1 to 8 weeks. Specifically, we reannotated publicly accessible transcriptomics, metabolomics, proteomics and phosphoproteomics data from the Molecular Transducers of Physical Activity Consortium (MoTrPAC) and integrated newly analyzed acetylproteomics data to assess multi-omic sex differences in sedentary and treadmill trained rats. Female rats displayed enrichment in immune-related features and pathways at the transcriptome and proteome level that were largely maintained with training. However, both sexes exhibited decreases in immune pathway activity following 8 weeks of training, although the effect was more pronounced in males. Shared responses to training included increased enrichment in transcriptomic pathways related to type I alveoli, proteomic pathways related to cilia, and decreased acetylation of pathways linked to mitochondrial function. Furthermore, features known to be enriched in lung diseases were attenuated with training in both sexes. Together, our findings provide novel insight into responses to endurance exercise training in the healthy rat lung and may offer translational insight into sex-specific differences in lung disease pathogenesis and treatment.

Pulmonary blood volumes (PBV), currently not assessed by computed tomography pulmonary angiography (CTPA), could provide additional information to routine investigations performed for chronic thromboembolic pulmonary hypertension (CTEPH). We investigated CTPA-based PBV in evaluating hemodynamic outcome from pulmonary endarterectomy (PEA) surgery. A deep learning-based CTPA vascular segmentation model, differentiating arteries and veins, was applied for automated PBV measurements in CTEPH patients who underwent PEA at UK's national CTEPH service. Pulmonary arteries were compartmentalised into "central" (main pulmonary and proximal lobar) and "intrapulmonary". Mean pulmonary arterial pressure >30 mmHg post-PEA defined "clinically relevant" residual PH. Logistic regression models applying CTPA-based PBV to identify residual PH were trained and tested on the discovery and validation cohorts respectively. Paired pre- and postoperative CTPA, in the discovery (n=71) and validation (n=102) cohorts showed that central pulmonary artery volume and total artery to vein volume ratio (A-VR) decreased and pulmonary vein volume increased with hemodynamic improvement post-PEA. Preoperative central pulmonary artery volume and A-VR helped identify patients at risk for clinically relevant residual PH post-PEA (AUROC 0.88 and 0.82 in the discovery and validation cohorts). Postoperative central pulmonary artery volume, A-VR and pulmonary vein volume helped to non-invasively identify patients without clinically relevant residual PH (AUROC 0.91 and 0.88 in the discovery and validation cohorts). Automated quantification of CTPA-based PBV at diagnosis can help stratify risk for residual PH in patients managed with PEA. Utilizing CTPA-derived PBV post-PEA to identify patients without residual PH can potentially reduce the need for routine postoperative right heart catheterization.

Extracellular vesicles (EVs) have emerged as versatile carriers of therapeutic cargo, including nucleic acids, proteins, and small molecules. However, their potential to deliver bioactive lipid mediators remains largely unexplored. Here, we present a novel synthetic biology-based strategy to selectively load EVs with pro-resolving lipid mediators of the Resolvin D and E series by co-expressing the resolvin biosynthetic enzymes cyclooxygenase 2 (COX-2), 5-lipoxygenase (5-LOX), and 15-lipoxygenase (15-LOX) using a custom-designed multigene expression vector. Human embryonic kidney 293T (HEK293T) cells transfected with the multigene expression vector and cultured in the presence of fatty acid free bovine serum albumen (BSA)-complexed docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and aspirin produced multiple members of the Resolvin D, aspirin-triggered Resolvin D series, and Resolvin E1 and E2, along with their biosynthetic precursors, which were subsequently packaged into EVs (referred to as Resolvin-EVs). Resolvin-EVs attenuated neutrophil adhesion to endothelial cells both under static and flow conditions and preserved endothelial barrier integrity by upregulating VE-cadherin. In macrophages, Resolvin-EVs suppressed nuclear factor kappa B (NF-κB) reporter activity, release of IL6 and TNFα. Effects of Resolvin-EVs on endothelial permeability and macrophage activation were abrogated by pharmacological inhibition of EV uptake using nystatin and cytochalasin D. Furthermore, Resolvin-EVs enhanced efferocytosis in THP-1 derived macrophages compared to Control-EVs. Notably, post-injury administration of Resolvin-EVs attenuated pulmonary inflammation in LPS-treated mice without inducing systemic or pulmonary toxicity. Together, these findings establish a novel, scalable platform for generating Resolvin-loaded EVs and highlight their therapeutic potential for acute lung injury and other chronic inflammatory disorders.

Cystic fibrosis (CF) is an autosomal recessive disease caused by variants in the gene encoding the CF transmembrane conductance regulator (CFTR) protein. Delivery of a functional CFTR transgene to airway epithelial cells (AEC) offers the potential to provide durable restoration of normal CFTR function. Adeno-associated virus (AAV) vectors are the leading platform for the delivery of in vivo gene therapy; however, wild-type AAV vectors exhibit a limited capacity to transduce airway cells and evade pre-existing human neutralizing antibodies (NAb). We therefore employed a directed evolution platform to invent a novel AAV capsid (A101) with the capacity to efficiently transduce AECs, including in the presence of NAbs, following aerosolized administration to nonhuman primates (NHP). We then engineered 4D-710, a gene therapy comprising the A101 vector and a CFTR transgene with a partial deletion in the regulatory domain (CFTRΔR) to facilitate vector packaging. 4D-710 exhibited efficient transduction of human bronchial epithelial (HBE) cell air-liquid interface (ALI) cultures in vitro and robust functional activity in CF HBE ALI cultures. Aerosolized administration of 4D-710 to NHPs was well tolerated and resulted in dose-dependent transgene expression and increased CFTR protein in diverse AEC types compared to vehicle controls. No significant differences in CFTRΔR mRNA levels were observed in lung samples from NHPs with pre-existing serum anti-capsid NAbs compared to NAb-negative NHPs. These findings demonstrate the tolerability and feasibility of A101-mediated transgene delivery and expression in primate airways. A clinical trial evaluating aerosol delivery of 4D-710 in adults with CF (NCT05248230) is underway.

Osteopontin, also known as secreted phosphoprotein 1 (Spp1), is a key molecule involved in lung fibrosis; however, the mechanism underlying the exacerbation caused by Spp1-producing cells remains unclear. In the present study, we investigated the detailed functions of Spp1-producing macrophages in lung fibrosis. Analysis of published single-cell RNA sequencing (scRNA-seq) datasets revealed the fibrogenic role of the interaction between SPP1-expressing macrophages and fibroblasts in patients with idiopathic pulmonary fibrosis. In addition, interstitial macrophages (IMs) were identified as the primary Spp1 source in the bleomycin-treated lungs of Spp1-enhanced green fluorescent protein (EGFP) knock-in reporter mice; their IMs promote lung fibrosis by enhancing fibroblast activation. Spp1-EGFP⁺ IMs expanded, peaking 7 days post-bleomycin administration and engrafting as inflammatory resident macrophages. Multi-omics analysis revealed that Spp1-EGFP⁺ IMs produced glycoprotein non-metastatic melanoma protein b (Gpnmb)-a fibrogenic, pro-inflammatory protein. Furthermore, Spp1-producing macrophages expressed the interleukin (IL)7 receptor on their surface in the fibrotic lungs of humans and mice. In the bleomycin-induced lung fibrosis model of Il7r^fl/fl Csf1r-iCre mice, macrophage expression of Spp1 and Gpnmb was reduced, and lung fibrosis was attenuated, compared with those of Il7r^fl/fl mice. These profibrotic Spp1-producing macrophages and the IL-7/macrophage/Spp1 axis may represent therapeutic targets for lung fibrosis.

We used spatial transcriptomics from idiopathic pulmonary fibrosis (IPF) and unaffected control lung tissue to further understand the pathogenesis of MUC5B-driven lung fibrosis. We captured 43 fields of view in 15 IPF and 13 controls with and without the MUC5B promoter variant using the CosMx^® platform and identified 19 cell types via semi-supervised clustering. MUC5B was ectopically expressed in AT2 cells in controls with the risk variant. We observed a decreased proportion of AT2 cells in controls and an increased proportion of aberrant basaloid cells in IPF associated with the MUC5B risk variant. We identified co-localized expression of MUC5B in respiratory bronchioles with 13 genes including the endoplasmic reticulum (ER) stress marker XBP1 and distal secretory markers SCGB3A1 and SCGB1A1. Experimentally, we demonstrated a direct relationship between MUC5B expression and ER stress in bronchiolar epithelia in vitro and validated the co-expression of MUC5B and XBP1 in the IPF lung. Based on our results, we conclude that MUC5B injures alveolar and bronchiolar epithelia that results in loss of AT2 cells and an increase in aberrant basaloid cells which initiates ER stress and a secretory phenotype in the terminal respiratory bronchiole, establishing a persistently injured distal airspace.