ECM Mechano-Sensing Regulates Cytoskeleton Assembly and Receptor-Mediated Endocytosis of Nanoparticles.

It is possible to create sophisticated and target-specific devices for nanomedicine thanks to technological advances in the engineering of nanomaterials. When on target, these nanocarriers often have to be internalized by cells in order to accomplish their diagnostic or therapeutic function. Therefore, the control of such uptake mechanism by active targeting strategy has today become the new challenge in nanoparticle designing. It is also well-known that cells are able to sense and respond to the local physical environment and that the substrate stiffness, and not only the nanoparticle design, influences the cellular internalization mechanisms. In this frame, our work reports on the cyclic relationship among substrate stiffness, cell cytoskeleton assembly and internalization mechanism. Nanoparticles uptake has been investigated in terms of the mechanics of cell environment, the resulting cytoskeleton activity and the opportunity of activate molecular specific molecular pathways during the internalization process. To this aim, the surface of 100 nm polystyrene nanoparticles was decorated with a tripeptide (RGD and a scrambled version as a control), which was able to activate an internalization pathway directly correlated to the dynamics of the cell cytoskeleton, in turn, directly correlated to the elastic modulus of the substrates. We found that the substrate stiffness modulates the uptake of nanoparticles by regulating structural parameters of bEnd.3 cells as spreading, volume, focal adhesion, and mechanics. In fact, the nanoparticles were internalized in larger amounts both when decorated with RGD, which activated an internalization pathway directly correlated to the cell cytoskeleton, and when cells resided on stiffer material that, in turn, promoted the formation of a more structured cytoskeleton. This evidence indicates the directive role of the mechanical environment on cellular uptake of nanoparticles, contributing new insights to the rational design and development of novel nanocarrier systems.