Radiographic and Histologic Evaluation of a Bone Void that Formed After Recombinant Human Bone Morphogenetic Protein-2-Mediated Sinus Graft Augmentation: A Case Report.

In the present case report, the authors describe radiographic and histologic observations of a bone void that formed after a sinus augmentation using a graft material that contained recombinant human bone morphogenetic protein-2 (rhBMP-2) and discuss clinical and histologic implications of their findings. Sinus augmentation was performed using a graft material comprising 1 g of hydroxyapatite/ß-tricalcium phosphate, which contained 1 mg of rhBMP-2. Radiographic evaluation was conducted with panoramic radiographs and computed tomography images of the augmented maxillary sinus, which were analyzed using a three-dimensional image-reconstruction program. Histologic evaluation was also performed on a biopsy specimen obtained 6 months after the sinus augmentation. The total augmented volume increased from 1,582.2 mm(3) immediately after the sinus augmentation to 3,344.9 mm3 at 6 months after the augmentation because of the formation of a bone void. Twenty-six months after the sinus augmentation, the bone void remained but had reduced in volume, with the total augmented volume reduced to 2,551.7 mm(3). Histologically, new bone was observed to be in contact with the grafted particles, and a fatty marrow-like tissue was present in the area of the bone void. This case report shows that the bone void that had formed after sinus augmentation resolved over time and seemed to be partially replaced with new bone. Furthermore, none of the implants failed, and clinical adverse events were not observed during the follow-up period.

Evaluation of In Vivo Osteogenic Potential of Bone Morphogenetic Protein 2-Overexpressing Human Periodontal Ligament Stem Cells Combined with Biphasic Calcium Phosphate Block Scaffolds in a Critical-Size Bone Defect Model.

Human periodontal ligament stem cells (hPDLSCs) are considered potential cellular carriers for gene delivery in the field of tissue regeneration. This study tested the osseoregenerative potential of hPDLSCs transduced with replication-deficient recombinant adenovirus (rAd) containing the gene encoding bone morphogenetic protein-2 (BMP2; hPDLSCs/rAd-BMP2) in both in vivo and in vitro osteogenic environments. After the optimal condition for rAd-mediated transduction was determined, hPDLSCs were transduced to express BMP2. In vivo bone formation was evaluated in a critical-size rat calvarial bone defect model that more closely mimics the harsher in vivo milieu for bone regeneration than subcutaneous transplantation model. As support materials for bone regeneration, block-type biphasic calcium phosphate (BCP) scaffolds were combined with hPDLSCs and/or BMP2 and transplanted into critical-size bone defects in rats. Experimental groups were as follows: BCP scaffold control (group 1 [Gr1]), scaffold containing recombinant human BMP2 (rhBMP2; group 2 [Gr2]), scaffold loaded with normal hPDLSCs (group 3 [Gr3]), scaffold combined with both normal hPDLSCs and rhBMP2 (group 4 [Gr4]), and scaffold loaded with hPDLSCs transduced with rAd-BMP2 (hPDLSCs/rAd-BMP2; group 5 [Gr5]). Our data showed that new bone formation was highest in Gr2. Less mineralization was observed in Gr3, Gr4, and Gr5 in which hPDLSCs were transplanted. In vitro transwell assay demonstrated that hPDLSCs exert an inhibitory activity on BMP2-induced osteogenic differentiation. Our findings suggest that the in vivo bone regenerative potential of BMP2-overexpressing hPDLSCs could be compromised in a critical-size rat calvarial bone defect model. Thus, further investigations are required to elucidate the underlying mechanisms and to develop efficient techniques for improved tissue regeneration.

Three-Dimensional Bone Regeneration of Alveolar Ridge Defects Using Corticocancellous Allogeneic Block Grafts: Histologic and Immunohistochemical Analysis.

In this study, the effectiveness of a corticocancellous block allograft for restoring alveolar ridge defects in preparation for the placement of dental implants was assessed. Significant ridge defects in four partially edentulous patients were reconstructed using an irradiated corticocancellous allogeneic block soaked in platelet-rich plasma, which was also covered with a resorbable collagen membrane. After 5 or 6 months, the sites were reentered and a trephine bone core specimen was obtained from each augmented site for histologic, histomorphometric, and immunohistochemical assessment. In all four cases, histologic evaluation of the augmented site showed areas of new vital bone formation around the graft material (mean newly formed bone fraction, 23.7%; mean total mineralized tissue fraction, 40.1%), in which osteocytes were frequently observed within the lacunae. Immunohistochemical analysis showed the presence of biomarkers commonly related to active bone formation (alkaline phosphatase, osteocalcin, and bone morphogenetic protein-2), confirming that the biochemical environment was conducive to new bone formation. The findings of this study demonstrate that the use of allogeneic block grafts for restoring alveolar ridge defects prior to the placement of dental implants may be an effective and advantageous alternative to autograft procedures.

Characterization of the enhanced bone regenerative capacity of human periodontal ligament stem cells engineered to express the gene encoding bone morphogenetic protein 2.

Human periodontal ligament stem cells (hPDLSCs) are considered an appropriate cell source for therapeutic strategies. The aims of this study were to investigate the sustainability of bone morphogenetic protein 2 (BMP2) secretion and the bone regenerative capacity of hPDLSCs that had been genetically modified to express the gene encoding BMP2 (BMP2). hPDLSCs isolated from healthy third molars were transduced using replication-deficient recombinant adenovirus (rAd) encoding BMP2 (hPDLSCs/rAd-BMP2), and the cellular characteristics and osteogenic potentials of hPDLSCs/rAd-BMP2 were analyzed both in vitro and in vivo. hPDLSCs/rAd-BMP2 successfully secreted BMP2, formed colonies, and expressed immunophenotypes similar to their nontransduced counterparts. As to their osteogenic potential, hPDLSCs/rAd-BMP2 formed greater mineralized nodules and exhibited significantly higher levels of expression of BMP2 and the gene encoding alkaline phosphatase, and formed more and better quality bone than other hPDLSC-containing or recombinant human BMP2-treated groups, being localized at the initial site until 8 weeks. The findings of the present study demonstrate that hPDLSCs/rAd-BMP2 effectively promote osteogenesis not only in vitro but also in vivo. The findings also suggest that hPDLSCs can efficiently carry and deliver BMP2, and that hPDLSCs/rAd-BMP2 could be used in an attractive novel therapeutic approach for the regeneration of deteriorated bony defects.

Optimal medium formulation for the long-term expansion and maintenance of human periodontal ligament stem cells.

BACKGROUND: Human periodontal ligament stem cells (hPDLSCs) are promising mesenchymal stem cells that are readily accessible. However, there is as yet no consensus as to the optimal culture medium for hPDLSCs. Thus, the purpose of the present study is to determine the optimal culture medium for long-term expansion of hPDLSCs. METHODS: hPDLSCs were isolated from healthy third molars, and the most widely used medium formulations in previous studies were used: 1) an α minimum essential medium-based medium formulation (MBM); and 2) a Dulbecco's minimum essential medium-based medium formulation. Passage 5 (P5) and P8 were evaluated with the two media for cell proliferation, differentiation, and immunophenotype. RESULTS: hPDLSCs that were primarily cultured in MBM were far more proliferated than those grown in DBM. In general, application of the MBM for longer periods produced greater cell growth and osteogenic differentiation. Furthermore, MBM-precultured hPDLSCs exhibited a greater degree of cell proliferation and a greater production of mineralized tissue and alkaline phosphatase (ALP) activity in vitro, although the levels of both were dependent on the culture medium used. With respect to long-term expansion, the P5 hPDLSCs grew and produced the largest amount of mineralized nodules faster than the P8 hPDLSCs, but both passages exhibited a similar phenotype for stemness and ALP activity. CONCLUSION: The present study indicates that the inherent capacity of hPDLSCs could be maintained until a later passage, P8 in MBM, and MBM appears to be an optimal choice for manipulating the finest and most stable hPDLSCs.

Secondary closure of an extraction socket using the double-membrane guided bone regeneration technique with immediate implant placement.

PURPOSE: Immediate implantation presents challenges regarding site healing, osseointegration, and obtaining complete soft-tissue coverage of the extraction socket, especially in the posterior area. This last issue is addressed herein using the double-membrane (collagen membrane+high-density polytetrafluoroethylene [dPTFE] membrane) technique in two clinical cases of posterior immediate implant placement. METHODS: An implant was placed immediately after atraumatically extracting the maxillary posterior tooth. The gap between the coronal portion of the fixture and the adjacent bony walls was filled with allograft material. In addition, a collagen membrane (lower) and dPTFE membrane (upper) were placed in a layer-by-layer manner to enable the closure of the extraction socket without a primary flap closure, thus facilitating the preservation of keratinized mucosa. The upper dPTFE membrane was left exposed for 4 weeks, after which the membrane was gently removed using forceps without flap elevation. RESULTS: There was considerable plaque deposition on the outer surface of the dPTFE membrane but not on the inner surface. Moreover, scanning electron microscopy of the removed membrane revealed only a small amount of bacteria on the inner surface of the membrane. The peri-implant tissue was favorable both clinically and radiographically after a conventional dental-implant healing period. CONCLUSIONS: Secondary closure of the extraction socket and immediate guided bone regeneration using the double-membrane technique may produce a good clinical outcome after immediate placement of a dental implant in the posterior area.