Stem-Cell Clinging by a Thread: AFM Measure of Polymer-Brush Lateral Deformation

If the binding strength of adhesive cues to the extracellular matrix (ECM) and the mechanisms involved in cell adhesion are synergistically correlated via a mechanical feedback, engineering of cue presentation at the ECM by designer macromolecules can enable control over cell-matrix interaction. Here, polymer brushes supporting fibronectin (FN) and presenting different grafted-chain length to modulate cell interaction at ECM cell-binding sites are exploited. Application of friction force microscopy allows us to estimate the lateral deformability and friction of oligoethylene glycol-containing brushes. These parameters are demonstrated to regulate the adhesion of human mesenchymal stem cells (hMSCs), which adopt their morphology and form focal adhesions (FAs) responding to FN brush-tether characteristics. Across a brush-thickness gradient presenting uniform FN exposure, thin brushes stimulate cell spreading and the development of FAs. Conversely, thick and more laterally deformable polymer grafts induce a decrease in cell spreading and FA formation. A correlation between frictional forces experienced at the (macro)molecular scale and the behavior of stem cells has been found. This interaction can be clarified by exploring novel aspects of FFM, which demonstrates a powerful tool to dynamically probe the ECM environment and indirectly suggest a way to structure ECM in order to trigger specific cell responses

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-\u3b5-caprolactone (PCL). Uniform coating of the scaffold surfaces with \u201cgrafted-from\u201d, 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