Expansion of the sagittal suture induces proliferation of skeletal stem cells and sustains endogenous calvarial bone regeneration.

In newborn humans, and up to approximately 2 y of age, calvarial bone defects can naturally regenerate. This remarkable regeneration potential is also found in newborn mice and is absent in adult mice. Since previous studies showed that the mouse calvarial sutures are reservoirs of calvarial skeletal stem cells (cSSCs), which are the cells responsible for calvarial bone regeneration, here we hypothesized that the regenerative potential of the newborn mouse calvaria is due to a significant amount of cSSCs present in the newborn expanding sutures. Thus, we tested whether such regenerative potential can be reverse engineered in adult mice by artificially inducing an increase of the cSSCs resident within the adult calvarial sutures. First, we analyzed the cellular composition of the calvarial sutures in newborn and in older mice, up to 14-mo-old mice, showing that the sutures of the younger mice are enriched in cSSCs. Then, we demonstrated that a controlled mechanical expansion of the functionally closed sagittal sutures of adult mice induces a significant increase of the cSSCs. Finally, we showed that if a calvarial critical size bone defect is created simultaneously to the mechanical expansion of the sagittal suture, it fully regenerates without the need for additional therapeutic aids. Using a genetic blockade system, we further demonstrate that this endogenous regeneration is mediated by the canonical Wnt signaling. This study shows that controlled mechanical forces can harness the cSSCs and induce calvarial bone regeneration. Similar harnessing strategies may be used to develop novel and more effective bone regeneration autotherapies.

Biocompatibility and antibiofilm activity of graphene-oxide functionalized titanium discs and collagen membranes.

The aims of the study were: 1) to evaluate the effect on biofilm formation of barrier membranes and titanium surfaces coated with graphene-oxide (GO); 2) to analyze the connection between the superficial topography of the tested materials and the amount of bacterial accumulation on them and 3) to analyze the biocompatibility of GO functionalized discs using the zebrafish model. METHODS: Single species bacterial biofilms (Streptococcus oralis, Veilonella parvula, Fusobacterium nucleatum, Porphyomonas gingivalis) were grown on GO-free membranes, membranes coated with 2 and 10 µg/ml of GO, GO-free and GO-coated titanium discs. The biofilms were analyzed by determining the CFU count and by Scanning Electron Microscopy (SEM) and the materials' topography by Atomic Force Microscopy (AFM). Zebrafish model was used to determine the materials' toxicity and inflammatory effects. RESULTS: AFM showed similar roughness of control and GO-coated materials. CFU counts on GO-coated discs were significantly lower than on control discs for all species. CFU counts of S. oralis, V. parvula and P. gingivalis were lower on biofilms grown on both types of GO-coated membranes than on GO-free membrane. SEM analysis showed different formation of single species biofilm of S. oralis on control and GO-coated materials. GO-functionalized titanium discs do not induce toxic or inflammatory effects. SIGNIFICANCE: Titanium implant surfaces functionalized with GO have shown to be biocompatible and less susceptible to biofilm formation. These results encourage further in vivo investigation of the tested materials on infection prevention, specifically in prevention and reduction of peri-implant mucositis and periimplantitis incidence.

Osteoblastic Differentiation on Graphene Oxide-Functionalized Titanium Surfaces: An In Vitro Study.

BACKGROUND: Titanium implant surfaces are continuously modified to improve biocompatibility and to promote osteointegration. Graphene oxide (GO) has been successfully used to ameliorate biomaterial performances, in terms of implant integration with host tissue. The aim of this study is to evaluate the Dental Pulp Stem Cells (DPSCs) viability, cytotoxic response, and osteogenic differentiation capability in the presence of GO-coated titanium surfaces. METHODS: Two titanium discs types, machined (control, Crtl) and sandblasted and acid-etched (test, Test) discs, were covalently functionalized with GO. The ability of the GO-functionalized substrates to allow the proliferation and differentiation of DPSCs, as well as their cytotoxic potential, were assessed. RESULTS: The functionalization procedures provide a homogeneous coating with GO of the titanium surface in both control and test substrates, with unchanged surface roughness with respect to the untreated surfaces. All samples show the deposition of extracellular matrix, more pronounced in the test and GO-functionalized test discs. GO-functionalized test samples evidenced a significant viability, with no cytotoxic response and a remarkable early stage proliferation of DPSCs cells, followed by their successful differentiation into osteoblasts. CONCLUSIONS: The described protocol of GO-functionalization provides a novel not cytotoxic biomaterial that is able to stimulate cell viability and that better and more quickly induces osteogenic differentiation with respect to simple titanium discs. Our findings pave the way to exploit this GO-functionalization protocol for the production of novel dental implant materials that display improved integration with the host tissue.

Covalent Decoration of Cortical Membranes with Graphene Oxide as a Substrate for Dental Pulp Stem Cells.

(1) Background: The aim of this study was to optimize, through a cheap and facile protocol, the covalent functionalization of graphene oxide (GO)-decorated cortical membrane (Lamina®) in order to promote the adhesion, the growth and the osteogenic differentiation of DPSCs (Dental Pulp Stem Cells); (2) Methods: GO-coated Laminas were fully characterized by Scannsion Electron Microscopy (SEM) and Atomic Force Microscopy (AFM) analyses. In vitro analyses of viability, membrane integrity and calcium phosphate deposition were performed; (3) Results: The GO-decorated Laminas demonstrated an increase in the roughness of Laminas, a reduction in toxicity and did not affect membrane integrity of DPSCs; and (4) Conclusions: The GO covalent functionalization of Laminas was effective and relatively easy to obtain. The homogeneous GO coating obtained favored the proliferation rate of DPSCs and the deposition of calcium phosphate.