Human mesenchymal stromal cells (hMSCs) are increasingly used in regenerative medicine for restoring worn-out or damaged tissue. Newly engineered tissues need to be properly vascularized and current candidates for in vitro tissue pre-vascularization are endothelial cells and endothelial progenitor cells. However, their use in therapy is hampered by their limited expansion capacity and lack of autologous sources. Our approach to engineering large grafts is to use hMSCs both as a source of cells for regeneration of targeted tissue and at the same time as the source of endothelial cells. Here we investigate how different stimuli influence endothelial differentiation of hMSCs. Although growth supplements together with shear force were not sufficient to differentiate hMSCs with respect to expression of endothelial markers such as CD31 and KDR, these conditions did prime the cells to differentiate into cells with an endothelial gene expression profile and morphology when seeded on Matrigel. In addition, we show that endothelial-like hMSCs are able to create a capillary network in 3D culture both in vitro and in vivo conditions. The expansion phase in the presence of growth supplements was crucial for the stability of the capillaries formed in vitro. To conclude, we established a robust protocol for endothelial differentiation of hMSCs, including an immortalized MSC line (iMSCs) which allows for reproducible in vitro analysis in further studies.

A material-based bottom-up approach is proposed towards an assembly of cells and engineered micro-objects at the macroscale. We show how shape, size and wettability of engineered micro-objects play an important role in the behavior of cells on these objects. This approach can, among other applications, be used as a tool to engineer complex 3D tissues of clinically relevant size.

Tissue engineering scaffolds having specific topographical cues offer exciting possibilities for stimulating cells differentiation and growth of new tissue. Although electrospun scaffolds have been extensively investigated in tissue engineering and regenerative medicine, little is known about the influence of introducing nanoroughness on their surface for cellular differentiation. The present study provides a method to engineer electrospun scaffolds with tailoring surface nanoroughness and investigates the effect of such topographical cues on the process of human mesenchymal stromal cells differentiation into osteoblasts and chondrocytes linages. This strategy may help the design of nanostructured scaffolds for skeletal tissue engineering.

A robust and simple method was developed for large-scale fabrication of free-standing and sub-μm PDMS through-hole membranes for biomedical applications.