Thesis Defence: Developing mechanodynamic alveolar epithelial-fibroblast in vitro models via the Flexcell bioreactor

Safiya Al Yazeedi, supervised by Dr. Emmanuel T. Osei, will defend their thesis titled “Developing mechanodynamic alveolar epithelial-fibroblast in vitro models via the Flexcell bioreactor” in partial fulfillment of the requirements for the degree of Master of Science in Biology.

An abstract for Safiya Al Yazeedi’s thesis is included below.

Defences are open to all members of the campus community as well as the general public. Registration is not required for in-person defences.

---

ABSTRACT

Mechanical strain plays a significant role in lung physiology and pathophysiology, influencing cellular behavior, tissue homeostasis, and disease progression. Current in-vitro models, particularly 2D monolayer cultures and animal models, fail to capture the complexity of the lung’s mechanodynamic environment. This study addresses the gap by developing advanced mechanodynamic 2D and 3D alveolar epithelial-fibroblast co-culture and organoid models to simulate the lung’s mechanical environment, using the unique mechanical Flexcell bioreactor that enables the application of strain to mimic the mechanical lung environment. Human lung fibroblasts (MRC-5) and alveolar epithelial cells (A549) were utilised to establish 2D and 3D alveolar co-cultures as well as 3D alveolar organoids in BioFlex plates and Tissue Train plates. The models were then subjected to equibiaxial strain of 18% amplitude at 0.4Hz using the Flexcell tension system. The impact of mechanical strain on cell proliferation, morphology, cytoskeletal & tight junctional, IL-6 & IL-8 inflammatory cytokine release, and viability was assessed via cell counts, immunocytochemistry & confocal imaging, ELISAs, and lactate dehydrogenase (LDH) assays. Mechanical strain of 18%, 0.4Hz, significantly caused predominantly increased cell proliferation rates in 3D co-culture models but not in 2D cultures. Morphological analysis revealed a marked transition of fibroblasts into broadened-shaped cells under strain, indicating myofibroblast differentiation in the 3D co-cultures. This was in line with increased F-actin intensity in 3D co-cultures compared to decreased F-actin in in fibroblast and epithelial cells in 2D models after pathological strain. The expression of the tight junctional protein ZO-1 was decreased under after strain across all 2D and 3D models. Further, there was increased release of pro-inflammatory cytokines, IL-6 and IL-8, in response to strain, and increased strain-induced cell death all models, which was higher in 3D cultures. The study demonstrates that multicellular alveolar models mimic the mechanical lungs. Specifically, 3D alveolar models in the Flexcell provide the 3D configuration and multicellular environment necessary to replicate the in vivo mechanical environment of lung tissue, highlighting their importance in studying strain-induced cellular responses. The developed models provide a valuable platform for exploring the mechanisms underlying lung disease progression and for testing therapeutic interventions aimed at restoring normal cellular responses to fatal injury from aberrant mechanical forces.