Abstract
Objective The present study aimed to investigate the participation of focal adhesion kinases (FAK) in interactions between osteoblastic cells and titanium (Ti) surfaces with three different topographies, namely, untreated (US), microstructured (MS), and nanostructured (NS).Methodology Osteoblasts harvested from the calvarial bones of 3-day-old rats were cultured on US, MS and NS discs in the presence of PF-573228 (FAK inhibitor) to evaluate osteoblastic differentiation. After 24 h, we evaluated osteoblast morphology and vinculin expression, and on day 10, the following parameters: gene expression of osteoblastic markers and integrin signaling components, FAK protein expression and alkaline phosphatase (ALP) activity. A smooth surface, porosities at the microscale level, and nanocavities were observed in US, MS, and NS, respectively.ResultsFAK inhibition decreased the number of filopodia in cells grown on US and MS compared with that in NS. FAK inhibition decreased the gene expression of Alp, bone sialoprotein, osteocalcin, and ALP activity in cells grown on all evaluated surfaces. FAK inhibition did not affect the gene expression of Fak, integrin alpha 1 ( Itga1 ) and integrin beta 1 ( Itgb1 ) in cells grown on MS, increased the gene expression of Fak in cells grown on NS, and increased the gene expression of Itga1 and Itgb1 in cells grown on US and NS. Moreover, FAK protein expression decreased in cells cultured on US but increased in cells cultured on MS and NS after FAK inhibition; no difference in the expression of vinculin was observed among cells grown on all surfaces.Conclusions Our data demonstrate the relevance of FAK in the interactions between osteoblastic cells and Ti surfaces regardless of surface topography. Nanotopography positively regulated FAK expression and integrin signaling pathway components during osteoblast differentiation. In this context, the development of Ti surfaces with the ability to upregulate FAK activity could positively impact the process of implant osseointegration.
Highlights
Excellent mechanical and biological properties render titanium (Ti) to be the most frequently used biomaterial for manufacturing dental implants.1 The bone−Ti contact is influenced by several parameters, including topography and surface chemistry of the dental implants
focal adhesion kinases (FAK) inhibition did not affect the gene expression of Fak, integrin alpha 1 (Itga1) and integrin beta 1 (Itgb1) in cells grown on MS, increased the gene expression of Fak in cells grown on NS, and increased the gene expression of Itga1 and Itgb1 in cells grown on untreated surface (US) and NS
The results indicated that FAK is relevant to osteoblastic differentiation of cells grown on Ti surfaces regardless of topographic characteristics since the inhibition of FAK reduced the differentiation of cells grown on all three surfaces
Summary
Excellent mechanical and biological properties render titanium (Ti) to be the most frequently used biomaterial for manufacturing dental implants. The bone−Ti contact is influenced by several parameters, including topography and surface chemistry of the dental implants. Osseointegration of implants is strongly associated with the responses of osteoblasts to the surface of the biomaterial, and signaling pathways involved in osteoblastic differentiation, such as integrin signaling, are known to play an important role in this process. Integrins are heterodimeric transmembrane proteins composed of α and β subunits forming a family of membrane receptors whose primary function is adhesion of cells to extracellular matrix proteins, such as collagen and fibronectin; some of these receptors involved in osteoblastic differentiation.. Focal adhesion kinases (FAK) or Src family kinases are the main integrin-activated protein tyrosine kinases that play a key role in this signaling pathway.. The associations of extracellular matrix ligands, integrins, and cytoskeletal components form a focal adhesion complex, where FAK is recruited and interacts directly or indirectly with these complexes causing their activation by autophosphorylation and consequent binding to Src kinase. Src kinases phosphorylate several components of focal adhesion sites participating in FAK signaling to generate the signal transduction mechanism.
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