Use of a decellularised dermis scaffold and human bronchial epithelial cells to tissue engineer airway mucosa suitable for tracheal transplantation

Abstract

BackgroundTracheal reconstruction relies on the use of a split skin graft to re-epithelialise the mucosal layer. Since split skin grafts are made up of a keratinising stratified epithelial layer, sloughing occurs within the airway with mucus retention and subsequent airway obstruction. The delivery of a graft with the same mucociliary function as the native airway would overcome these limitations and greatly improve the safety and effectiveness of this type of surgery. We aimed to generate a transplantable tissue-engineered respiratory epithelial graft with mucociliary function. MethodsCadaveric human skin was decellularised and the epidermal layer removed. Human bronchial epithelial cells were seeded with human respiratory fibroblasts onto the dermis at densities of 1 × 106 per cm2 and 1 × 104 per cm2, respectively, and cultured at air–liquid interface in a transwell system. At 3 weeks, the constructs were transplanted onto a decellularised trachea that had been prevascularised within a muscle wrap in an immunosuppressed New Zealand White rabbit. FindingsAfter 3 weeks of air–liquid interface culture, high-speed video microscopy showed beating cilia on the surface of the dermis, and the epithelial layer stained positively for the ciliated cell marker acetylated α-tubulin, the secretory cell marker MUC5AC, and the epithelial cell marker pan-cytokeratin on top-down whole-mount confocal microscopy. Staining with haematoxylin and eosin (H&E) demonstrated a pseudostratified mucociliary layer along the length of the dermis. 24 h after transplantation, a pseudostratified, ciliated layer could be observed on H&E staining of sections of trachea. At 5 days, the respiratory epithelial layer consisted of a single layer of cytokeratin 5-positive epithelial cells. InterpretationThis study is the first, to our knowledge, to report the delivery of a transplantable tissue-engineered respiratory epithelial graft with mucociliary function. 24 h after transplantation the mucociliary layer was preserved although only a basal layer was demonstrated by 5 days, possibly due to the loss of the air–liquid interface within the muscle wrap. FundingMedical Research Council.