Perfusion‐Decellularization of Porcine Lung and Trachea for Respiratory Bioengineering

Decellularization of native organs may provide an acellular tissue platform for organ regeneration. However, decellularization involves a trade‐off between removal of immunogenic cellular elements and preservation of biomechanical integrity. We sought to develop a bioartificial scaffold for respiratory tissue engineering by decellularization of porcine lungs and trachea while preserving organ architecture and vasculature. Lung–trachea preparations from 25 German Landrace pigs were perfused in a modified Langendorff circuit and decellularized by an SDC (sodium deoxycholate)‐based perfusion protocol. Decellularization was evaluated by histology and fluorescence microscopy, and residual DNA quantified spectrophotometrically and compared with controls. Airway compliance was evaluated by endotracheal intubation and mechanical ventilation to simulate physiological breathing‐induced stretch. Structural integrity was evaluated by bronchoscopy and biomechanical stress/strain analysis by measuring passive tensile strength, all compared with controls. Decellularized lungs and trachea lacked intracellular components but retained specific collagen fibers and elastin. Quantitative DNA analysis demonstrated a significant reduction of DNA compared with controls (32.8 ± 12.4 μg DNA/mg tissue vs. 179.7 ± 35.8 μg DNA/mg tissue, P < 0.05). Lungs and trachea decellularized by our perfusion protocol demonstrated increased airway compliance but preserved biomechanical integrity as compared with native tissue. Whole porcine lungs–tracheae can be successfully decellularized to create an acellular scaffold that preserves extracellular matrix and retains structral integrity and three‐dimensional architecture to provide a bioartifical platform for respiratory tissue engineering.     

新型脱细胞半月板细胞外基质的制备及其生物相容性的研究

目的探索新型脱细胞半月板细胞外基质(d MECM)的制备方法,并对其细胞相容性进行研究。方法无菌条件下收集新鲜的猪半月板组织,切碎,采用湿法粉碎、差速离心的方法制备dM ECM生物材料。通过天狼猩红、甲苯胺蓝染色方法分别比较天然半月板与dM ECM中胶原以及糖胺聚糖含量的区别;采用Hoechst 33258荧光染色法观察脱细胞后dM ECM中DNA残留情况。将P3代的兔内侧半月板细胞种植在铺有dM ECM的盖玻片上,体外培养7 d后行扫描电镜检测其细胞相容性。结果本实验制备的dM ECM天狼猩红、甲苯胺蓝染色均呈阳性,并且与天然半月板染色类似;dM ECM行Hoechst33258染色呈阴性;扫描电镜结果显示种植于d MECM表面上的半月板细胞黏附紧密,可见大量细胞ECM的分泌。结论本实验制备的dM ECM可以很好地保留天然半月板ECM成分,有效地去除DNA物质,并且具有良好的生物相容性,是未来半月板组织工程领域非常有前景的支架材料。