The Primary Research Of Constructing Tracheal Cartilage Tissue Via Tissue Engineering Approaches Combined With 3D-printed Supports

Background:The incidence of tracheomalacia/tracheal stenosis is not high in general population,the major cause of them in children is congenital tracheal anomalies which is usually companied with multiple congenital anomalies.Particularly,the incidence of tracheal anomalies is much higher in children suffering congenital heart diseases.The present surgical methods of tracheal end-to-end anastomosis and slide tracheoplasty can be used to correct tracheal anomalies with short segment but not long segment.While,fortunately,the transplantation based on tissue engineered trachea may be a promising method.Purpose: To identify the feasibility of constructing tissue engineered tracheal cartilage tissue,which is aimed to treat tracheal anomalies in children by using chondrocyte sheets or electrospun gelatin/polycaprolactone nanofibrous membranes combined with 3D-printed supports.Methods: According to the published research results of cartilage tissue engineering and our previous work on engineering tracheal cartilage,we selected the cell sheet-and electrospun nanofibrous membrane-based tissue engineering methods to construct tracheal cartilage tissue;Furthermore,the application of 3D-printed supports was also involved as the difference between children at different ages and between individuals is concerned.The research was performed in three parts: A.Engineering tracheal cartilage through using chondrocyte sheets and 3D-printed polyamide supports: Prepare chondrocyte sheets through culturing chondrocytes of secondary passage at high density;Construct engineered tracheal cartilage tissue through rolling chondrocyte sheets on 3D-printed polyamide(PA)supports;After in-vitro/-vivo culture,the tissue was estimated with histological staining method.B.Engineer tracheal cartilage through using electrospun gelatin/polycaprolactone nanofibrous membranes and 3D-printed biodegradable supports: Fabricate extracellular matrix-mimic scaffolds with electrospinning technology;Construct engineered tracheal cartilage tissue through rolling electrospun membranes reseeded with chondrocytes on biodegradable 3D-printed support fabricated with polycaprolactone(PCL);After in-vitro/-vivo culture,the tissue was estimated with histological staining method.C.The establishment of an animal model of tracheomalacia/ tracheal stenosis: establish the rabbit model of tracheomalacia/tracheal stenosis through breaking up and striping tracheal cartilage rings.Results: After the in-vitro/-vivo culture of constructed tracheal cartilage tissue through utilizing chondrocyte sheets combined with 3D-printed polyamide support,the histological results displayed the formation of cartilage tissue rich with cartilage-specific matrix;After the in-vivo culture,the C-shaped tracheal cartilage tissue was engineered with electrospun gelatin/polycaprolactone nanofibrous membranes and biodegradable 3D-printed support.And the histological results indicated better tracheal-like cartilage tissue formation with using 3D-printed support than conventional silicon tubes,and the better mimicking of native tracheal cartilage ring in morphology and bio-mechanism was also demonstrated with gross observation;Through breaking up and striping the tracheal cartilage rings,the rabbit tracheomalacia model and the tracheal stenosis model were established at early and late stages,respectively.Conclusion: It is feasible to construct tracheal cartilage tissue using tissue engineering methods combined with 3D-printed supports.The expected animal model of tracheomalacia/ tracheal stenosis could be established with surgical method of breaking up and striping cartilage rings,which will be applied to and benefit for the further estimation of the bio-function of engineered tracheal cartilage.