Participation of focal adhesion kinases in the osteoblast differentiation of mesenchymal stem cells induced by titanium surfaces with micro and nanotopography

Abstract

Integrins are transmembrane proteins composed by two subunits (± and ²) with primary function in cell adhesion and some of them are involved in osteoblast differentiation. These proteins are capable of signaling in both directions of the cell membrane, activating multiple protein tyrosine kinases and two signaling pathways involving FAK (focal adhesion kinases). Several studies have shown that FAK are involved in the mechanisms of signal transduction that promote the maturation of osteoblasts in osteogenic-committed cells. Moreover, the FAK act at the early stages of osteoblast differentiation of human mesenchymal stem cells (MSCs). Thus, the development of biomaterials that may modulate the interaction between integrins and extracellular matrix via FAK represents an important strategy for therapies based on osseointegrated implants. Considering the importance of these kinases in the integrin signaling pathway, the aim of the present study is to investigate the role of FAK in osteoblast differentiation of MSCs induced by titanium (Ti) with nanotopography compared with a commercial available surface with microtopography and the untreated machined surface (control). We will use the PF-573228 compound, a specific inhibitor of FAK and after selecting the optimal concentration, the cells will be grown on the three Ti surfaces with or without PF-573228, and the following parameters will be evaluated: (1) gene expression of FAK and the osteoblastic markers, RUNX2, osterix, alkaline phosphatase, osteocalcin, osteopontin and bone sialoprotein by real-time PCR at 3, 7 and 10 days, (2) alkaline phosphatase activity at 10 days and (3) the formation of mineralized extracellular matrix by alizarin red staining, at 21 days.