3D Protein Gradients on Scaffolds for Tissue Engineering: A Polymer Brush-Assisted Fabrication

In order to mimic the compositional diversity of natural extra-cellular matrices (ECMs) we propose here a new and versatile method to obtain synthetic ECMs, which are potentially directly applicable to clinical practices. Fabrication of three-dimensional (3D) gradients of proteins within microporous, biodegradable tissue engineering scaffolds making use of solution wetting is described. 3D regularly layered starting supports are manufactured by rapid prototyping of poly-ε-caprolactone (PCL). Uniform coating of the scaffold surfaces with “grafted-from”, poly[(oligoethylene gycol)methacrylate] (POEGMA) brushes is employed for the covalent immobilization of proteins. The 3D gradient formation processes make use of surface energy and capillary forces, which administer proteins from their solutions inside the pores. Following these approaches multidirectional gradients of different protein species can be produced with precise control over the protein coverage morphology. In addition, 3D gradients of brush-supported fibronectin permit the controlled immobilization of human mesenchymal stem cells (hMSCs) in spatially determined cultures. PCL-POEGMA scaffolds with 3D protein gradients demonstrate to be easily reproduced also in clinics since they do not require any inert environment or complicated chemistry. Brush-coated scaffolds could be simply incubated with the required proteins and directly applied to patients in or without the presence of cell preparations. This technique supports the combination of multiple cues which could eventually trigger cell adhesion, migration or differentiation, in a single 3D porous structure. All the above-mentioned features make the method presented a practical and affordable strategy to synthetically mimic natural ECMs and their 3D multidirectional diversity.