Functionality of endothelial cells on silk fibroin nets: comparative study of micro- and nanometric fibre size

Biomimetic material design, such as mimicking nanostructured components of the extracellular matrix, is an actual challenge for biomaterial research with a high impact on tissue engineering and regenerative medicine. Thus, understanding the cellular response at the cell biological and molecular level and the consequences of various chemically or physically modified biomaterials is highly important. In the present study we assessed the response of human umbilical vein endothelial cells (HUVEC) and outgrowth endothelial cells (OEC) from endothelial progenitor cells to different variants of nanofibrous silk fibroin nets in comparison to microfibrous silk fibroin scaffolds with regard to cellular morphology, proliferation, formation of intercellular contacts as well as integrin-dependent adhesion. Endothelial cells (ECs) grown on nanometric nets formed a differentiated and interconnected endothelial monolayer with no significant changes in the expression of intercellular contact molecules or proliferation rates compared with cells grown on micrometric nets. Nevertheless, quantitative real-time PCR revealed a higher expression level of integrin-beta1 in ECs grown on nanofibrous fibroin nets compared to the microfibrous samples. In addition, single nano-fibres were recognised by the integrin-receptor mechanism supporting the formation of focal adhesion at the interface of ECs and nanometric nets. These findings indicate that the nanometric silk fibroin scaffolds did not interfere with the formation of a differentiated and interconnected EC layer. On the contrary, nanofibre variation of the fibroin net architecture induced changes in ECs at the molecular level in terms of the increased expression of adhesion molecules such as integrin-beta1.