Does FAK Mediate Implant Surface Microtopography Effects on Osteoblast Differentiation?

Our previous reports indicate implant microtopography modulates the ability of preosteoblasts to differentiate and form a mineralized matrix. Activation of FAK is considered a critical step in adhesion-mediated osteoblast differentiation. Objectives: The hypothesis of this study was variation in implant surface microtopographies will cause changes in levels of FAK gene and protein expression, FAK tyrosine phosphorylation, and influence osteoblast differentiation through FAK-RUNX2 mediated pathways. Methods: Human palatal mesenchymal preosteoblast cells (ATCC HEPM 1486) were grown in micromass cultures, in triplicate, on either 600 grit polished (machined) or 50um Al2O3 grit blasted titanium surfaces, or on tissue culture plastic for a period of 24 hours to 3 weeks. At specific time points RNA and protein lysates were collected, normalized, and then analyzed using real time PCR for FAK/RUNX2 gene expression; or Western blotting for FAK, or FAK-phosphotyrosine (PY). Results: The results (N=3) on both the 600g and SB implant microtopographies demonstrated that from 24 hours to 1 week on both implant surfaces the level of FAK protein expression increased. However, from 1 week to 2 weeks the level of FAK protein expression decreased while the level of FAK-PY signal increased (p<0.056). Real time PCR analysis demonstrated that FAK gene expression was highest at 1 week, and began to decrease from 1 to 2 weeks, similar to that of the protein levels (p<0.001). Interestingly, the level of RUNX2 gene expression began to increase at 1 week and peaked in expression at 2 weeks when the levels of FAK-PY signal were highest (p<0.001) and FAK protein levels the lowest on these implant surfaces. Conclusions: This data suggests that FAK protein levels increase, followed by FAK-PY signaling. This in turn may lead to downstream regulation of RUNX2 expression in an implant microtopography dependent fashion. This work was supported by NIH/NIDCR R21-DE016677-01.