Small airway fibroblasts from Chronic Obstructive Pulmonary Disease patients exhibit cellular senescence

Abstract

Rationale: Small airways disease (SAD) is a key early-stage pathology of chronic obstructive pulmonary disease (COPD). COPD is associated with cellular senescence whereby cells undergo growth arrest and express the senescence-associated secretory phenotype (SASP) leading to chronic inflammation and tissue remodelling. Parenchymal derived fibroblasts have been shown to display senescent properties in COPD, however small airway fibroblasts (SAF) have not been investigated. Therefore, this study investigated the role of these cells in COPD and their potential contribution to SAD. Objectives: To investigate the senescent and fibrotic phenotype of SAF in COPD. Methods: SAF were isolated from non-smoker, smoker and COPD lung resection tissue (n=9-17). Senescence and fibrotic marker expression were determined using iCELLigence (proliferation), qPCR, Seahorse assay and ELISAs. COPD SAF were further enriched for senescent cells using FACSAria Fusion based on cell size and autofluorescence (10% largest/autofluorescent v 10% smallest/non-autofluorescent). The phenotype of the senescence-enriched population was investigated using RNA-sequencing and pathway analysis. Main results: Markers of senescence were observed in COPD SAF, including senescence-associated β-galactosidase, SASP release and reduced proliferation. Because the pathways driving this phenotype were unclear, we used cell-sorting to enrich for senescent COPD SAF. This population displayed increased p21CIP1 and p16INK4a expression and mitochondrial dysfunction. RNA-sequencing suggested these senescent cells express genes involved in oxidative stress response, fibrosis and mitochondrial dysfunction pathways. Conclusions: These data suggest COPD SAF are senescent and may be associated with fibrotic properties and mitochondrial dysfunction. Further understanding cellular senescence in SAF may lead to potential therapies to limit SAD progression.