Severe bone injuries can result in incapacities and thus affect a person's quality of life. Mesenchymal stem cells (MSCs) can be an alternative for bone healing by growing them on scaffolds that provide mechanical signals for differentiation. Such scaffolds can give the appropriate ques to the cells in order to induce their differentiation into mature osteoblasts and later on to be transplanted into the body. Until now lots of attention has been given to create appropriate nano and micro patterns that can work as signal inducers limiting the research in only 2.5 dimensions. On the other hand, our work introduces true, well- defined 3D environments to the research of MSCs differentiation.
In our approach, we fabricated hierarchical auxetic mechanical metamaterials and ultra-light ultra-stiff scaffolds via two photon polymerization and used them as scaffolds to investigate the differentiation of MSCs into osteoblasts. Those scaffolds consist of unit cells comparable to the diameter of MSCs which is approximately 50 to 100μm, so only a couple of cells can fit inside thus ensuring the optimal mechanical environment for each cell. In the case of auxetic scaffolds, the unit cells are able to bend without breaking such that the cells can adapt their environment to their needs, whereas the kelvin foam is stiff non elastic scaffold that shows no deformation in response to the forces exceeded by the cells. We investigated the localization of YAP protein, a key protein transcription factor that acts as a mechanotransduction mediator and compared it to common osteogenic markers in both protein and gene levels by using confocal microscopy and qPCR .
Interestingly, YAP protein is translocated to the nucleus even after 21 days of culture and RUNX2 gene shows a 10-fold increase in auxetic scaffold in comparison with the control only after 7 days. Long term cultures up to 28 days shows high mineralization of the extracellular matrix after Alizarin red staining. Moreover, SEM pictures revealed different cell morphology in those different scaffolds because of the different geometries used. Auxetics pushes the cells into more elongated phenotypes whereas kelvin foam in more broad cells bodies.
Auxetic scaffolds are ideal for osteogenic differentiation as they can maintain and promote the osteogenesis efficiently even after 28 days of culture. Our work paves the way for the use of more complicated metamaterials into the tissue engineering field.
This work was funded by In2Sight: Horizon 2020 GA: 964481