title: The development of a tissue-engineered tracheobronchial epithelial model using a bilayered collagen-hyaluronate scaffold.
abstract: Today, chronic respiratory disease is one of the leading causes of mortality globally. Epithelial dysfunction can play a central role in its pathophysiology. The development of physiologically-representative in vitro model systems using tissue-engineered constructs might improve our understanding of epithelial tissue and disease. This study sought to engineer a bilayered collagen-hyaluronate (CHyA-B) scaffold for the development of a physiologically-representative 3D in vitro tracheobronchial epithelial co-culture model. CHyA-B scaffolds were fabricated by integrating a thin film top-layer into a porous sub-layer with lyophilisation. The film layer firmly connected to the sub-layer with delamination occurring at stresses of 12-15 kPa. Crosslinked scaffolds had a compressive modulus of 1.9 kPa and mean pore diameters of 70 μm and 80 μm, depending on the freezing temperature. Histological analysis showed that the Calu-3 bronchial epithelial cell line attached and grew on CHyA-B with adoption of an epithelial monolayer on the film layer. Immunofluorescence and qRT-PCR studies demonstrated that the CHyA-B scaffolds facilitated Calu-3 cell differentiation, with enhanced mucin expression, increased ciliation and the formation of intercellular tight junctions. Co-culture of Calu-3 cells with Wi38 lung fibroblasts was achieved on the scaffold to create a submucosal tissue analogue of the upper respiratory tract, validating CHyA-B as a platform to support co-culture and cellular organisation reminiscent of in vivo tissue architecture. In summary, this study has demonstrated that CHyA-B is a promising tool for the development of novel 3D tracheobronchial co-culture in vitro models with the potential to unravel new pathways in drug discovery and drug delivery.
authors.affiliation: Tissue Engineering Research Group, Dept. of Anatomy, Royal College of Surgeons in Ireland (RCSI), Dublin, Ireland; School of Pharmacy, RCSI, Dublin, Ireland; Centre for Research in Medical Devices (CURAM), RCSI, Dublin, Ireland; Trinity Centre for Bioengineering, Trinity College Dublin (TCD), Dublin, Ireland; Advanced Materials and Bioengineering Research (AMBER) Centre, RCSI and TCD, Dublin, Ireland. Electronic address: cianoleary@rcsi.ie.
authors.forename: Cian
authors.initials: C
authors.lastname: O'Leary
authors.affiliation: Cellular and Molecular Imaging Core, RCSI, Dublin, Ireland. Electronic address: brentoncavanagh@rcsi.ie.
authors.forename: Brenton
authors.initials: B
authors.lastname: Cavanagh
authors.affiliation: REPAIR-Lab, Institute of Pathology, University Medical Centre, Johannes Gutenberg University, Mainz, Germany. Electronic address: runger@uni-mainz.de.
authors.forename: Ronald E
authors.initials: RE
authors.lastname: Unger
authors.affiliation: REPAIR-Lab, Institute of Pathology, University Medical Centre, Johannes Gutenberg University, Mainz, Germany. Electronic address: kirkpatrick@uni-mainz.de.
authors.forename: C James
authors.initials: CJ
authors.lastname: Kirkpatrick
authors.affiliation: Biology Department, Maynooth University, Co Kildare, Ireland. Electronic address: shirley.odea@nuim.ie.
authors.forename: Shirley
authors.initials: S
authors.lastname: O'Dea
authors.affiliation: Tissue Engineering Research Group, Dept. of Anatomy, Royal College of Surgeons in Ireland (RCSI), Dublin, Ireland; Centre for Research in Medical Devices (CURAM), RCSI, Dublin, Ireland; Trinity Centre for Bioengineering, Trinity College Dublin (TCD), Dublin, Ireland; Advanced Materials and Bioengineering Research (AMBER) Centre, RCSI and TCD, Dublin, Ireland. Electronic address: fjobrien@rcsi.ie.
authors.forename: Fergal J
authors.initials: FJ
authors.lastname: O'Brien
authors.affiliation: Tissue Engineering Research Group, Dept. of Anatomy, Royal College of Surgeons in Ireland (RCSI), Dublin, Ireland; School of Pharmacy, RCSI, Dublin, Ireland; Centre for Research in Medical Devices (CURAM), RCSI, Dublin, Ireland; Trinity Centre for Bioengineering, Trinity College Dublin (TCD), Dublin, Ireland; Advanced Materials and Bioengineering Research (AMBER) Centre, RCSI and TCD, Dublin, Ireland. Electronic address: SCryan@rcsi.ie.
authors.forename: Sally-Ann
authors.initials: SA
authors.lastname: Cryan
mesh_terms: D001980:Bronchi / Q000166:cytology; D002454:Cell Differentiation; D002460:Cell Line; D018920:Coculture Techniques; D003094:Collagen / Q000737:chemistry*; D004847:Epithelial Cells / Q000166:cytology; D004848:Epithelium / Q000254:growth & development*; D005240:Feasibility Studies; D005347:Fibroblasts / Q000166:cytology; D006801:Humans; D006820:Hyaluronic Acid / Q000737:chemistry*; D023822:Tissue Engineering; D054457:Tissue Scaffolds / Q000737:chemistry*; D014132:Trachea / Q000166:cytology
chemical_list: D006820:Hyaluronic Acid; D003094:Collagen
keywords: Bilayered; Co-culture; Collagen; Epithelium; Hyaluronate; Respiratory