A Tissue-Engineered Tracheobronchial In Vitro Co-Culture Model for Determining Epithelial Toxicological and Inflammatory Responses

Abstract

Translation of novel inhalable therapies for respiratory diseases is hampered due to the lack of in vitro cell models that reflect the complexity of native tissue, resulting in many novel drugs and formulations failing to progress beyond preclinical assessments. The development of physiologically-representative tracheobronchial tissue analogues has the potential to improve the translation of new treatments by more accurately reflecting in vivo respiratory pharmacological and toxicological responses. Herein, advanced tissue-engineered collagen hyaluronic acid bilayered scaffolds (CHyA-B) previously developed within our group were used to evaluate bacterial and drug-induced toxicity and inflammation for the first time. Calu-3 bronchial epithelial cells and Wi38 lung fibroblasts were grown on either CHyA-B scaffolds (3D) or Transwell® inserts (2D) under air liquid interface (ALI) conditions. Toxicological and inflammatory responses from epithelial monocultures and co-cultures grown in 2D or 3D were compared, using lipopolysaccharide (LPS) and bleomycin challenges to induce bacterial and drug responses in vitro. The 3D in vitro model exhibited significant epithelial barrier formation that was maintained upon introduction of co-culture conditions. Barrier integrity showed differential recovery in CHyA-B and Transwell® epithelial cultures. Basolateral secretion of pro-inflammatory cytokines to bacterial challenge was found to be higher from cells grown in 3D compared to 2D. In addition, higher cytotoxicity and increased basolateral levels of cytokines were detected when epithelial cultures grown in 3D were challenged with bleomycin. CHyA-B scaffolds support the growth and differentiation of bronchial epithelial cells in a 3D co-culture model with different transepithelial resistance in comparison to the same co-cultures grown on Transwell® inserts. Epithelial cultures in an extracellular matrix like environment show distinct responses in cytokine release and metabolic activity compared to 2D polarised models, which better mimic in vivo response to toxic and inflammatory stimuli offering an innovative in vitro platform for respiratory drug development.

Interaction of Neisseria meningitidis carrier and disease isolates of MenB cc32 and MenW cc22 with epithelial cells of the nasopharyngeal barrier

“…The inclusion of 3D in vitro models for infection studies could improve our knowledge of the interaction of bacteria with cells of the respiratory tract by providing a more complex 3D environment that better mimics the extracellular matrix surrounding airway tissues, along with the incorporation of co-culture strategies (van der Vaart and Clevers, 2021). 3D in vitro models for the respiratory tract are widely engineered for several sorts of different applications from basic research to drug discovery (Soriano et al, 2021) and could be adapted for studies with N. meningitidis. As an example, a study by Marrazzo et al was published recently, who developed an in vitro 3D system that reconstructs the human tracheal and bronchial mucosa with the pseudostratified epithelium and underlying stromal tissue (Marrazzo et al, 2016).…”

A miniaturized multicellular platform to mimic the 3D structure of the alveolar-capillary barrier

“…Over the past decade, only about 3% of new drugs achieved clinical approval due to the poor understanding of disease pathogenesis and the lack of effective preclinical models that rapidly predict the efficacy of new therapeutic approaches in humans (Ioannidis et al, 2018;Soriano et al, 2021). Indeed, in addition to ethical concerns, the use of preclinical animal models has revealed a lack in the ability to replicate human disease and correctly predict the effect and the response to therapy (Greek and Kramer, 2019).…”