Rapid Preparation Method for Preparing Tracheal Decellularized Scaffolds: Vacuum Assistance and Optimization of DNase I

[Image: see text] Decellularized scaffolds are an effective way for tracheal tissue engineering to perform alternative treatments. However, clinically used decellularized tracheal scaffolds have a long preparation cycle. The purpose of this study is to improve the efficiency of decellularization by vacuum assistance and optimizing the concentration of DNase I in the decellularization process and to quickly obtain tracheal decellularized scaffolds. The trachea of New Zealand white rabbits was decellularized with 2, 4, 6, and 8 KU/mL DNase I under vacuum. The performance of the decellularized tracheal scaffold was evaluated through histological analysis, immunohistochemical staining, DNA residue, extracellular matrix composition, scanning electron microscopy, mechanical properties, cell compatibility, and in vivo experiments. Histological analysis and immunohistochemical staining showed that compared with the native trachea, the hierarchical structure of the decellularized trachea remained unchanged after decellularization, nonchondrocytes were effectively removed, and the antigenicity of the scaffold was significantly weakened. Deoxyribonucleic acid (DNA) quantitative analysis showed that the amount of residual DNA in the 6-KU group was significantly decreased. Scanning electron microscopy and mechanical tests showed that small gaps appeared in the basement membrane of the 6-KU group, and the mechanical properties decreased. The CCK-8 test results and in vivo experiments showed that the 6-KU group’s acellular scaffold had good cell compatibility and new blood vessels were visible on the surface. Taken together, the 6-KU group could quickly prepare rabbit tracheal scaffolds with good decellularization effects in only 2 days, which significantly shortened the preparation cycle reducing the required cost.