Department of Otorhinolaryngology - Head and Neck Surgery, College of Medicine, Ewha Womans University, Anyangcheon-ro 1071, Yang Cheon-Gu, Seoul, 07985, Korea.
Department of Molecular Medicine, College of Medicine, Ewha Womans University, Seoul, 07985, Korea.
Tissue Eng Regen Med. 2020 Aug;17(4):433-443. doi: 10.1007/s13770-020-00260-w. Epub 2020 May 10.
Respiratory mucosa defects result in airway obstruction and infection, requiring subsequent functional recovery of the respiratory epithelium. Because site-specific extracellular matrix (ECM) facilitates restoration of organ function by promoting cellular migration and engraftment, previous studies considered decellularized trachea an ideal ECM; however, incomplete cell removal from cartilage and mucosal-architecture destruction are frequently reported. Here, we developed a decellularization protocol and applied it to the respiratory mucosa of separated porcine tracheas.
The trachea was divided into groups according to decellularization protocol: native mucosa, freezing-thawing (FT), FT followed by the use of Perasafe-based chemical agents before mucosal separation (wFTP), after mucosal separation (mFTP), and followed by DNase decellularization (mFTD). Decellularization efficacy was evaluated by DNA quantification and hematoxylin and eosin staining, and ECM content of the scaffold was evaluated by histologic analysis and glycosaminoglycan and collagen assays. Biocompatibility was assessed by cell-viability assay and in vivo transplantation.
The mFTP mucosa showed low antigenicity and maintained the ECM to form a proper microstructure. Additionally, tonsil-derived stem cells remained viable when cultured with or seeded onto mFTP mucosa, and the in vivo host response showed a constructive pattern following implantation of the mFTP scaffolds.
These results demonstrated that xenogenic acellular respiratory mucosa matrix displayed suitable biocompatibility as a scaffold material for respiratory mucosa engineering.
呼吸道黏膜缺损会导致气道阻塞和感染,需要随后恢复呼吸道上皮的功能。由于特定部位的细胞外基质(ECM)通过促进细胞迁移和植入,有助于恢复器官功能,因此先前的研究认为脱细胞气管是一种理想的 ECM;然而,软骨和黏膜结构的不完全细胞去除经常被报道。在这里,我们开发了一种脱细胞方案,并将其应用于分离的猪气管的呼吸道黏膜。
根据脱细胞方案将气管分为几组:天然黏膜、冻融(FT)、FT 后在黏膜分离前使用基于 Perasafe 的化学试剂(wFTP)、黏膜分离后(mFTP)和随后进行 DNAse 脱细胞(mFTD)。通过 DNA 定量和苏木精和伊红染色评估脱细胞效果,通过组织学分析和糖胺聚糖和胶原测定评估支架的 ECM 含量。通过细胞活力测定和体内移植评估生物相容性。
mFTP 黏膜显示出低抗原性并保持 ECM 形成适当的微观结构。此外,扁桃体衍生的干细胞在与 mFTP 黏膜共培养或接种到 mFTP 黏膜上时仍然存活,并且植入 mFTP 支架后体内宿主反应显示出建设性模式。
这些结果表明,异种脱细胞呼吸道黏膜基质作为呼吸道黏膜工程的支架材料具有合适的生物相容性。