The osteocyte-osteoblast interactions affect bone repair and remodeling, which are relevant to theosseointegration of titanium (Ti) implants. We have demonstrated that Ti with nanotopography (Ti-Nano) generated by treatment with H2SO4 and H2O2 favors osteoblast differentiation by modulating several cell signaling pathways and attenuates the inhibitory effect of osteoclasts on osteoblast differentiation. In this context, the aim of this study is to evaluate the effect of osteocytes on osteoblast differentiation induced by Ti-Nano. The experimental design consists of co-cultures where mesenchymal stem cells (MSCs) will be grown on Ti-Control and Ti-Nano discs and cells from osteocyte lineage Ocy454 will be cultured into inserts positioned on MSCs cultures. Cells will be cultured under osteogenic conditions for up to 10 days and osteoblast differentiation of MSCs will be evaluated by: gene expression of Runx2, osteocalcin (Oc), osteopontin (Opn) and bone sialoprotein (Bsp) by real-time PCR, protein expression of RUNX2 and alkaline phosphatase (ALP) by western blot, and ALP activity by fast red detection. The numerical data will be submitted to the normal distribution test to select the adequate statistical analysis and the level of significance wil be set at 5%. The development of this project could contribute to the understanding of the cellular events that regulate osseointegration, especially that ones related to the osteocyte-osteoblast interactions, and thedevelopment of new therapies in the implantology field.
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