Short-term effects of microstructured surfaces: role in cell differentiation toward a contractile phenotype

Cell adhesion plays a key role in cell behavior, in terms of migration, proliferation, differentiation and apoptosis. All of these events concur with tissue regeneration and remodeling mechanisms, integrating a complex network of intracellular signaling modules. Morphogenetic responses, which involve changes in cell shape, proliferation and differentiation, are thought to be controlled by both biochemical and biophysical cues. Indeed, the extracellular matrix not only displays adhesive ligands necessary for cell adhesion but also plays an essential biomechanical role - responsible, for instance, for the acquisition of the contractile phenotype. The substrate topography around the forming tissues and the associated mechanical stresses that are generated regulate cellular morphology, proliferation and differentiation. Thus, the ability to tailor topographical features around cells can be a crucial design parameter in tissue engineering applications, inducing cells to exhibit the required performances.In this work, we designed micropillared substrates using highly spaced arrays (interspacing equal to 25 µm) to evaluate the effects of topography on C2C12 myoblasts' adhesion and differentiation. Optical and fluorescence microscopy images were used to observe cell adhesion, together with Western blot analysis on vinculin and focal adhesion kinase (FAK) expression, a protein highly involved in adhesive processes. Differentiation marker (Myf5, myogenin and myosin heavy chain [MHC]) expression was also studied, in relation to the effect of different substrate topographies on the enhancement of a contractile phenotype. Our results demonstrated that microstructured surfaces may play a key role in the regeneration of functional tissues.