A Key Role of Autophagy in Osteoblast Differentiation on Titanium-Based Dental Implants

Abstract

Autophagy plays an important role in embryogenesis, for the maintenance of tissue homeostasis and the elimination of damaged subcellular structures. Furthermore, autophagy could be a mode of physiological cell death and also be implicated in cell differentiation. Thus, we hypothesized that autophagy may have an impact on the differentiation of osteoblast cells influenced by various titanium-based surfaces. Interactions between smooth, commercially available pure titanium (Ti cp), rough Ticer, acid-etched Ti cp (SS) and M1-M3 (comprised of the monoclinic phase of sodium-titanium oxides and rutile; M2 contains amorphous calcium phosphates) and human osteoblast cells were investigated. Immunofluorescent staining was used for detecting autophagy, cell cluster formation and collagen type I (Col-1) expression. Flow cytometry was employed to identify autophagy, the production of endogenous nitric oxide (NO) and the size and granularity of the cells. Rough surfaces caused osteoblast differentiation via the autophagic-dependent PI3/Akt signalling pathway. These surfaces induced the formation of discrete populations of large, granular cells, i.e. mature osteoblasts. In addition, M1-M3 provoked the development of a third population of small, granular cells, responsible for cell cluster formation, which are important for the formation of bone noduli and mineralisation. The same surfaces induced faster osteoblast maturation and enhanced NO production, a hallmark of the already mentioned processes. Neither the mature osteoblasts nor the small cells appeared after the inhibition of autophagy. Inhibition of autophagy also prevented cell cluster formation. We demonstrate that autophagy plays an essential role in the osteoblast differentiation on titanium-based surfaces with rough topography.

Ti-SLActive and TiZr-SLActive Dental Implant Surfaces Promote Fast Osteoblast Differentiation

“…Contrary to that, on Ti-SLActive and TiZr-SLActive surfaces on day 2, migrations of cells and cell clustering were observed. Those clusters were exclusively formed from the small polygonal cells, highly differentiated ones, presenting the next step in the process of osteoblast maturation, as reported previously [12]. On Ticer and Ti-SLA, cell clusters were formed on day 5 of culture.…”

“…As previously shown using Ti cp [12], cell cluster formation was not observed throughout the present study when smooth surface materials such as Cercon and Ti cp were applied. Also, no variation in cell size in either morphology was observed.…”

“…Recently, it was elucidated that autophagy plays an important role in osteoblast differentiation when cultivated on anodical titanium-based topographies [12]. In addition to three white novel titanium-based surfaces, Ticer, a clinically used surface, was employed in the previously reported study.…”

“…In order to find out whether chemical composition of dental implant surfaces affects osteoblast differentiation, Ti-SLA, Ti-SLActive and TiZr-SLActive (all rough; R a = 1.5, 1.6 and 1.6 µm, respectively) [68,69] materials, as well as Cercon (smooth; R a < 0.2 µm), zirconia material, were selected for the present study. Ti cp (smooth; R a < 0.2 µm) and Ticer (rough; R a = 1.4 µm) [22] titanium-based surfaces (investigated previously) [12], served as controls. The results of this work will confirm and extend the hypothesis that surface topography, as well as the composition of dental implants influence not only cell proliferation, but also morphology and expression of BSP, OC and Col-1 of osteoblasts.…”

“…Recently, we reported a pioneering work on the role of autophagy in the osteoblast differentiation process [12]. It has been found that rough topographies of titanium-based materials obtained by anodic oxidation under spark discharge (Ticer and 3 analogue white surfaces) and acid etched (SS) induced osteoblast differentiation via autophagic PI3/Akt dependent signalling pathway.…”