Camacho Paula, Fainor Matthew, Seims Kelly B, Tolbert John W, Chow Lesley W
Department of Bioengineering, Lehigh University, Bethlehem, PA 18015, USA.
Integrated Degree in Engineering, Arts and Sciences Program, Lehigh University, Bethlehem, PA 18015, USA.
J Biol Methods. 2021 Mar 22;8(1):e146. doi: 10.14440/jbm.2021.353. eCollection 2021.
Three-dimensional (3D) printing of biodegradable polymers has rapidly become a popular approach to create scaffolds for tissue engineering. This technique enables fabrication of complex architectures and layer-by-layer spatial control of multiple components with high resolution. The resulting scaffolds can also present distinct chemical groups or bioactive cues on the surface to guide cell behavior. However, surface functionalization often includes one or more post-fabrication processing steps, which typically produce biomaterials with homogeneously distributed chemistries that fail to mimic the biochemical organization found in native tissues. As an alternative, our laboratory developed a novel method that combines solvent-cast 3D printing with peptide-polymer conjugates to spatially present multiple biochemical cues in a single scaffold without requiring post-fabrication modification. Here, we describe a detailed, stepwise protocol to fabricate peptide-functionalized scaffolds and characterize their physical architecture and biochemical spatial organization. We used these 3D-printed scaffolds to direct human mesenchymal stem cell differentiation and osteochondral tissue formation by controlling the spatial presentation of cartilage-promoting and bone-promoting peptides. This protocol also describes how to seed scaffolds and evaluate matrix deposition driven by peptide organization.
可生物降解聚合物的三维（3D）打印已迅速成为一种用于制造组织工程支架的流行方法。该技术能够制造复杂的结构，并以高分辨率对多种成分进行逐层空间控制。所得支架还可以在表面呈现不同的化学基团或生物活性信号，以引导细胞行为。然而，表面功能化通常包括一个或多个制造后的处理步骤，这些步骤通常会产生化学组成均匀分布的生物材料，无法模拟天然组织中发现的生化组织。作为一种替代方法，我们实验室开发了一种新方法，将溶剂浇铸3D打印与肽-聚合物共轭物相结合，在单个支架中空间呈现多种生化信号，而无需制造后修饰。在这里，我们描述了一个详细的、逐步的方案，用于制造肽功能化支架，并表征其物理结构和生化空间组织。我们使用这些3D打印支架，通过控制促进软骨和促进骨的肽的空间呈现，来指导人间充质干细胞分化和骨软骨组织形成。该方案还描述了如何接种支架以及评估由肽组织驱动的基质沉积。
