Graphene-Induced Osteogenic Differentiation Is Mediated by the Integrin/FAK Axis

Graphene is capable of promoting osteogenesis without chemical induction. Nevertheless, the underlying mechanism(s) remain largely unknown. The objectives here were: (i) to assess whether graphene scaffolds are capable of supporting osteogenesis in vivo and; (ii) to ascertain the participation of the integrin/FAK mechanotransduction axis during the osteogenic differentiation induced by graphene. MSC-impregnated graphene scaffolds (n = 6) were implanted into immunocompromised mice (28 days). Alternatively, MSCs were seeded onto PDMS substrates (modulus of elasticity = 130, 830 and 1300 kPa) coated with a single monomolecular layer of graphene and cultured in basal medium (10 days). The ensuing expressions of FAK-p397, integrin, ROCK1, F-actin, Smad p1/5, RUNX2, OCN and OPN were evaluated by Western blot (n = 3). As controls, MSCs were plated onto uncoated PDMS in the presence of mechanotransduction inhibitors (echistatin, Y27632 and DMH1). MSC-impregnated graphene scaffolds exhibited positive immunoexpression of bone-related markers (RUNX2 and OPN) without the assistance of osteogenic inducers. In vitro, regardless of the stiffness of the underlying PDMS substrate, MSCs seeded onto graphene-coated PDMS substrates demonstrated higher expressions of all tested osteogenic and integrin/FAK proteins tested compared to MSCs seeded onto PDMS alone. Hence, graphene promotes osteogenesis via the activation of the mechanosensitive integrin/FAK axis.