Plastic Surgery Research Council
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Presenter: Ning Zeng, MD
Co-Authors: Zeng N; Wang L; Zang M; Johnson J; Yu P; Zhang Q
the University of Texas MD Anderson Cancer Center

Introduction: Combination of tissue engineering techniques with allograft provides a promising approach for large size tracheal reconstruction. This study aims to engineer a neo-trachea by using cryopreserved tracheal allograft as a scaffold.

Methods: Tracheal segments harvested from Brown Norway Rat (BN) were programmed frozen for 30 days. The tracheal epithelial cells were removed by washing with PBS for 4 hours at 40C. Resultant tracheal scaffold was re-epithelized by luminal seeded with epithelial cells isolated from Lewis rat (LEW). These engineered neo-tracheae were subcutaneously implanted to LEW for 30 days. Fresh tracheae from BN and LEW, cryopreserved tracheae and de-epithelized cryopreserved tracheae from BN were transplanted to LEW as controls.

Results: The epithelial cells were removed while the chondrocytes well remained within washed tracheal scaffold as shown by H&E and DAPI stain. The tracheal scaffold maintained lumen rigidity, strong mechanical properties and natural ECM in cartilage (collagen II+, GAG+). The chondrocytes preserved viability confirmed by cell culture and MTT test. The alloantigen was significantly decreased (MHCI-, MHCII-). The epithelial cells (Cytokeratin peptide 17+) isolated from LEW contained all three type cells of native tracheal epithelium (cilia cells (Beta Tubulin+), goblet cells (Mucin5AC+), and basal cells (P63+)). The viability assay indicated the de-epithelized tracheal scaffold provided a niche for epithelial cells integration and proliferation. In vivo study showed that neo-trachea achieved complete epithelial coverage and preserved luminal patency with histoarchitecture integrity (Masson Trechrome stain). There was no immunorejection in neo-tracheae and tracheal isograft groups (few CD4+, CD8+ and CD68+ cells). Tracheal luminal obstruction and severe histoarchitecture damage with high level of CD4+, CD8+, and CD68+ cells infiltration were observed in other three groups.

Conclusions: Engineered neo-trachea from de-epithelized cryopreserved tracheal allograft scaffold showed promise to achieve suppressants free tracheal reconstruction.

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