Evaluation of the osteogenic potential of demineralized and decellularized bovine bone granules following implantation in rat calvaria critical-size defect model.

The aim of this study was to compare the ability of demineralized (DMB) and decellularized (DCC) bovine bone granules to support bone regeneration in rat calvaria critical-size defects. DMB and DCC were prepared using a previously published method. The granule size used ranged between 500 and 750 µm. A total of forty-eight Sprague-Dawley rats were divided into two groups (n = 24). A pair of 5 mm diameter defects were created on the calvaria of the rats in the right and left parietal bone in both groups. Group A animals received DMB granules and Group B received DCC granules in the right parietal defect side while the left parietal untreated defect acted as sham surgery for both groups. Four animals per group were euthanized in a CO2 chamber at day 7, 14 and 21 post-surgery and the calvaria implantation site biopsy harvested was subjected to osteogenic gene expression analysis. Another four animals per group were euthanized at days 15, 30 and 60 post surgery and the calvaria implantation site biopsy harvested was subjected to histological, immunohistochemistry, RAMAN spectroscopy and Micro-CT analysis at the mentioned time points. Statistical analysis was conducted using t-tests and ANOVA. Histomorphometry showed significantly higher new bone formation in the DCC sites (p<0.05) compared to DMB. Both DMB and DCC implantation sites showed distinct staining for osteocalcin and osteopontin proteins compared to their respective sham sites. By day 21 after implantation, DCC sites demonstrated significantly elevated mRNA levels of osteonectin (p<0.001), osteopontin (p<0.001), osteocalcin (p<0.0001), ALP (p<0.01), and BMP-2 (p<0.001) compared to DMB. However, VEGF expression showed no significant differences at this time point between the two groups. Micro-CT analysis also showed enhanced defect closure and higher bone density in DCC implanted sites while RAMAN spectra demonstrated increased abundance of collagen and bone minerals, especially, PO43- ions than DMB. In conclusion, both DMB and DCC granules demonstrated favorable osteogenic potential in critical-sized defects, with DCC exhibited superior osteoconductive, osteoinductive and osteogenesis properties.

Evaluation of decellularization process for developing osteogenic bovine cancellous bone scaffolds in-vitro.

Current immunological issues in bone grafting regarding the transfer of xenogeneic donor bone cells into the recipient are challenging the industry to produce safer acellular natural matrices for bone regeneration. The aim of this study was to investigate the efficacy of a novel decellularization technique for producing bovine cancellous bone scaffold and compare its physicochemical, mechanical, and biological characteristics with demineralized cancellous bone scaffold in an in-vitro study. Cancellous bone blocks were harvested from a bovine femoral head (18-24 months old) subjected to physical cleansing and chemical defatting, and further processed in two ways. Group I was subjected to demineralization, while Group II underwent decellularization through physical, chemical, and enzymatic treatments. Both were then freeze-dried, and gamma radiated, finally producing a demineralized bovine cancellous bone (DMB) scaffold and decellularized bovine cancellous bone (DCC) scaffold. Both DMB and DCC scaffolds were subjected to histological evaluation, scanning electron microscopy/energy-dispersive X-ray spectroscopy (SEM/EDS), fourier-transform infrared spectroscopy (FTIR), quantification of lipid, collagen, and residual nucleic acid content, and mechanical testing. The osteogenic potential was investigated through the recellularization of scaffolds with human osteoblast cell seeding and examined for cell attachment, proliferation, and mineralization by Alizarin staining and gene expression. DCC produced a complete acellular extracellular matrix (ECM) with the absence of nucleic acid content, wider pores with extensive interconnectivity and partially retaining collagen fibrils. DCC demonstrated a higher cell proliferation rate, upregulation of osteogenic differentiation markers, and substantial mineralized nodules production. Our findings suggest that the decellularization technique produced an acellular DCC scaffold with minimal damage to ECM and possesses osteogenic potential through the mechanisms of osteoconduction, osteoinduction, and osteogenesis in-vitro.

The Efficacy of Immediate Implant Placement in Extraction Sockets for Alveolar Bone Preservation: A Clinical Evaluation Using Three-Dimensional Cone Beam Computerized Tomography and Resonance Frequency Analysis Value.

The aim of this study was to determine the efficacy of immediate implant placement with alveolar bone augmentation on socket preservation following atraumatic tooth extraction and comparing it with a tooth alveolar socket that was allowed to heal in a conventional way.Twenty medically fit patients (8 males and 12 females aged between 18 and 40 years) who needed noncomplicated tooth extraction of mandibular premolar teeth were divided randomly and equally into 2 groups. In Group I, the empty extraction socket was left untreated and allowed to heal in a conventional way. In Group II, the immediate implant was placed and the gap between the implant and the inner buccal plate surface of the socket wall was filled with lyophilized bovine bone granules and the wound was covered with pericardium membrane. The patients were followed up clinically and radiologically for regular reviews at 1 week, 3 months, and 9 months postoperative. Cone beam computerized tomography images of the alveolar ridge and socket were analyzed to determine the structural changes of the alveolar ridge. Resonance frequency analysis was measured at 9 months for Group II to assess the degree of secondary stability of the implants by using Osstell machine.A significant difference of bone resorption of 1.49 mm (confidence interval, CI 95%, 0.63-2.35) was observed within the control group at 3 months, and 1.84 mm (P ≤ 0.05) at 9 months intervals. No significant changes of bone resorption were observed in Group II. Comparison between groups showed a highly significant difference at 3 months; 2.56 mm (CI 95% 4.22-0.90) and at 9 months intervals; 3.2 mm (CI 95%, 4.70-1.62) P ≤ 0.001 between Group I and II. High resonance frequency analysis values were observed at 9 months postoperative in Group II.In conclusion, the insertion of immediate implants in fresh extraction sockets together with grafting the circumferential gap between the bony socket wall and the implant surface with bovine bone granules was able to preserve a greater amount of alveolar ridge volume when compared with an extraction socket that was left to heal in a conventional way.