In Vivo Behavior of a Custom-Made 3D Synthetic Bone Substitute in Sinus Augmentation Procedures in Sheep
In this study, the in vivo behavior of a custom-made three-dimensional (3D) synthetic bone substitute was evaluated when used as scaffold for sinus augmentation procedures in an animal model. The scaffold was a calcium phosphate ceramic fabricated by the direct rapid prototyping technique, dispense-plotting. The geometrical and chemical properties of the scaffold were first analyzed through light and electron scanning microscopes, helium picnometer, and semi-quantitative X-ray diffraction measurements. Then, 6 sheep underwent monolateral sinus augmentation with the fabricated scaffolds. The animals were euthanized after healing periods of 45 and 90 days, and block sections including the grafted area were obtained. Bone samples were subjected to micro computerized tomography, morphological and histomorphometric analyses. A complete integration of the scaffold was reported, with abundant deposition of newly formed bone tissue within the biomaterial pores. Moreover, initial foci of bone remodeling were mainly localized at the periphery of the implanted area after 45 days, while continuous bridges of mature lamellar bone were recorded in 90-day specimens. This evidence supports the hypothesis that bone regeneration proceeds from the periphery to the center of the sinus cavity. These results showed how a technique allowing control of porosity, pore design, and external shape of a ceramic bone substitute may be valuable for producing synthetic bone grafts with good clinical performances.
Figure 1. Schematic representation of ovine monolateral sinus augmentation procedure performed with the insertion of two blocks of the scaffold (1.4 cm3 each). A macroscopic transverse section of the sheep head has been carried out, as indicated by the blue boxes, between the oral and caudal edges of the oval osteotomy hole. Figure 2. The haematoxylin-eosin stained sections (a) were digitally converted in colored images to quantify the extension of the tissues of interest. (b) In detail, black, green, and red have been used to identify scaffold, newly formed bone, and fibrous tissue, respectively.
Scanning electron microscopy images of surface and fracture face of dispense-plotted calcium phosphate ceramic scaffold in different magnifications. (a) Fracture through ceramic scaffold. (b) Surface (white star) and fracture face (white arrow) of material rod. (c) Surface of material rod. (d) Fracture face through material rod.
An example of micro-CT 2D analysis of an explant at 90 days postgraft showing, with an arbitrary colored scale, the complete integration of the two scaffold blocks within the maxillary sinus. The dark blue color indicates the preexistent or the newly deposited bone, the white blue the softer connective/fibrous tissues and the green/yellow the biomaterial. The asterisk (*) indicates the infraorbital canal.
An example of micro-CT 3D analysis of an explant at 90 days postgraft showing, with an arbitrary grey color scale, different points of bone integration between the scaffold and the maxillary preexistent bone (arrows). The asterisk (*) indicates the infraorbital canal.
Two examples of histological cryosections stained with haematoxylin and eosin (HE) of sinus explants isolated at 45 (top) and 90 days after augmentation with the scaffolds. The lower magnification images show the whole implantation area (a and d) demonstrating the excellent integration of the scaffold (dark structure) within the newly formed tissues that completely filled the interscaffold spaces. The higher magnifications indicate the presence of a rich vascularized fibrous tissue in the central portion of the grafted area (b and e), and the abundant newly deposited bone at the periphery (c and f), where the scaffold contacted the woven bone (white arrows). Active fronts of bone deposition with osteoblastic active cells are frequently identified in 45-day explants (black arrows). The asterisk (*) indicates the infraorbital canal.
(a) An example of double immunostaining for vWF (green fluorescence, left image) and α-SMA (red fluorescence, central image) carried out on cryosectioned maxillary sinus explants. The merged image (right picture) displays a rich vascularization with blood vessels of variable diameters and with different degrees of maturity as indicated by the presence of absence of α-SMA positive mural cells. (b) Topographic quantitative analysis of VA demonstrated that blood vessel extension progressively increases in central (C) and peripheral (P) fields, while the higher VA is recorded at the periphery of the grafted area. The histograms represent the data as mean ± SD. Double asterisks (**) indicates data significantly different for P < 0.01 and P < 0.05, respectively, within each time group (C vs P at 45 or 90 days); aindicates C or P data significantly different for P < 0.01 statistically compared between the two time groups (45 vs 90 days).
The quantitative topographic analysis confirms a significant greater peripheral (P) extension of newly generated bone either 45 or 90 days after graft, and a significant bone deposition in the central portion (C) of the grafted area, exclusively at day 90. The histograms represent the data as mean ± SD. The asterisk (*) indicates data significantly different for P < 0.01 within each time group (C vs P); aindicates C or P data significantly different for P < 0.01 between the two time groups (45 vs 90 days).
Contributor Notes