Sharma Dhavan, Ferguson Morgan, Zhao Feng
Department of Biomedical Engineering, Michigan Technological University, Houghton, MI, United States.
Department of Biomedical Engineering, Michigan Technological University, Houghton, MI, United States.
Methods Cell Biol. 2020;156:3-13. doi: 10.1016/bs.mcb.2019.10.010. Epub 2019 Dec 5.
Native extracellular matrix (ECM) based scaffolds are far more superior in structural and compositional complexity than other engineered scaffolding materials such as hydrogels, electrospun fibers, and three-dimensional (3D) printed substrates. Due to the presence of native structural proteins and other macromolecules, native ECM can better restore the crucial cell-ECM crosstalk and provide a highly biomimetic microenvironment to cells. Allogenic or xenogeneic tissues have been derived by decellularization to obtain native ECM scaffolds. However, their applicability is limited by batch to batch variation, risk of pathogen transfer, undesirable immune response and scarcity of donors. Human dermal fibroblasts (hDFs) can be prescreened and maintained in a pathogen-free condition. Herein, we have described a step-by-step protocol to generate a completely biological ECM scaffold by decellularization of hDF cell sheets. Decellularization was achieved by using an anionic surfactant sodium dodecyl sulfate (SDS) and ethylene diamine tetraacetate (EDTA). The resulting ECM sheet was organized into a nanofibrous scaffold, containing major ECM structural proteins as well as other macromolecules including collagens, fibronectin, laminin and elastin. This cell-derived nanofibrous ECM is a promising scaffold material for constructing highly biomimetic functional tissues.
基于天然细胞外基质（ECM）的支架在结构和成分复杂性方面远比其他工程支架材料优越，如水凝胶、电纺纤维和三维（3D）打印基质。由于存在天然结构蛋白和其他大分子，天然ECM能够更好地恢复关键的细胞-ECM相互作用，并为细胞提供高度仿生的微环境。同种异体或异种组织已通过去细胞化处理来获得天然ECM支架。然而，它们的适用性受到批次间差异、病原体传播风险、不良免疫反应和供体稀缺性的限制。人真皮成纤维细胞（hDFs）可以预先筛选并在无病原体条件下保存。在此，我们描述了一种通过hDF细胞片去细胞化生成完全生物性ECM支架的分步方案。使用阴离子表面活性剂十二烷基硫酸钠（SDS）和乙二胺四乙酸（EDTA）实现去细胞化。所得的ECM片被组装成纳米纤维支架，包含主要的ECM结构蛋白以及其他大分子，包括胶原蛋白、纤连蛋白、层粘连蛋白和弹性蛋白。这种细胞衍生的纳米纤维ECM是一种用于构建高度仿生功能组织的有前景的支架材料。
