METHODS: The porous PCL scaffold with cylindrical structure was generated by a negative mold. The chondrocytes were seeded onto PCL scaffolds and in vitro-cultured in chondrogenic defined medium for four weeks. The construct was surrounded by fibroblast-based tissue-engineered tracheal adventitia, and was cultivated for 1 week. The PCL neo-trachea was implanted subcutaneously in nude mice for four weeks, and the chondrogenesis of construct were evaluated by histological analysis.
RESULTS: The cylindrical structure of TETA-surrounded PCL neo-trachea remained after four-week in vivo cultivation. The immunofluorescence staining revealed that the TETA layer stayed intact without invasion from peripheral tissue to the adventitia or the neo-trachea. In comparison to the neo-trachea without adventitia, the chondrocytes density increased in the construct with TETA surrounding. The cartilaginous matrix, including proteoglycans and type II collagen, also showed prominent increasing in the TETA group. The terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining showed that the apoptosis of seeded chondrocytes was attenuated in the presence of TETA.
CONCLUSION: We demonstrated that TETA protected the chondrocyte-seeded construct from deformation after in vivo cultivation, increased the chondrocytes density and cartilaginous matrix and decreased apoptotic cell death in the construct. Our work provides a novel approach to replace muscle flap and simultaneously accelerate chondrogenesis which can be applied in tracheal tissue engineering for the treatment of tracheal resection patients.
- Tsao C-K, Ko C-Y, Yang S-R, et al. An ectopic approach for engineering a vascularized tracheal substitute. Biomaterials. 2014;35(4):1163-1175.