Ontrafeling van de bijdrage van de zieke micro-omgeving aan chronische longziekten

Chronic lung diseases such as cystic fibrosis (CF) and idiopathic pulmonary fibrosis (IPF) share features such as epithelial dysfunction, immune dysregulation, and extracellular matrix remodelling. This thesis investigates CFTR's role in neutrophil dysfunction in CF and establishes a framework to study epithelial-mesenchymal interactions in IPF by studying fibroblast activation in different in vitro models and developing tools to investigate epithelial-mesenchymal transition (EMT) in basal cells. CF is characterised by ongoing bacterial infections and unresolved inflammation despite excessive neutrophil infiltration. In Chapter 1, I looked into the role of CFTR in neutrophils using inhibitor-172 (inh-172) and compared results to CF neutrophils with severe mutations. Neutrophils treated with inh-172 exhibited low phagosome acidification while retaining bacterial uptake, with reduced reactive oxygen species (ROS) and bacterial killing. Comparing CF and healthy neutrophils, phagosome acidification and ROS production varied, while neutrophil elastase and myeloperoxidase secretion were uniformly reduced upon stimulation. To mimic the CF airway, neutrophils were pre-exposed to CF sputum supernatant in monoculture or co-culture with an epithelial monolayer. Treating same-donor neutrophils with inh-172 reduced donor variability and provided a model for CF and other lung diseases involving CFTR inhibition. IPF is a chronic interstitial lung disease driven by fibroblast activation, leading to extracellular matrix remodelling and tissue stiffening. Despite the focus on the alveolar epithelium, little is known about basal cells in fibrosis. Chapter 2 is divided into two main parts. In part 1, I observed that TGF-β treatment induced a myofibroblast phenotype with different α-SMA distributions, highlighting how important culture conditions are to activate fibroblasts for pulmonary fibrosis. In part 2, I examined whether bronchial progenitor cells acquire a mesenchymal phenotype upon profibrotic exposure. Upregulation of vimentin suggested basal cells may contribute to fibroblast activation. I also evaluated a fluorescent reporter driven by a human vimentin promoter region, which detected mesenchymal vimentin but not EMT-induced vimentin changes in basal cells. While currently suboptimal for capturing partial EMT, this system can be used in mesenchymal cells and lays a foundation for tracking EMT in vitro.