Hyun-Chang Lim

Regeneration of rabbit calvarial defects using cells-implanted nano-hydroxyapatite coated silk scaffolds

BackgroundThe aim of this study was to characterize the efficacy of nano-hydroxyapatite-coated silk fibroin constructs as a scaffold for bone tissue engineering and to determine the osteogenic effect of human dental pulp and periodontal ligament derived cells at an early stage of healing in rabbits. 3D silk fibroin constructs were developed and coated using nano-hydroxyapatite crystals. Dental pulp and periodontal ligament cells from extracted human third molars were cultured and seeded onto the silk scaffolds prior to in vivo implantation into 8 male New Zealand White rabbits. Four circular windows 8 mm in diameter were created in the calvarium of each animal. The defects were randomly allocated to the groups; (1) silk scaffold with dental pulp cells (DPSS), (2) silk scaffold with PDL cells (PDLSS), (3) normal saline-soaked silk scaffold (SS), and (4) empty control. The animals were sacrificed 2 (n = 4) or 4 weeks (n = 4) postoperatively. The characteristics of the silk scaffolds before and after cell seeding were analyzed using SEM. Samples were collected for histologic and histomorphometic analysis. ANOVA was used for statistical analysis.ResultHistologic view of the experimental sites showed well-maintained structure of the silk scaffolds mostly unresorbed at 4 weeks. The SEM observations after cell-seeding revealed attachment of the cells onto silk fibroin with production of extracellular matrix. New bone formation was observed in the 4 week groups occurring from the periphery of the defects and the silk fibers were closely integrated with the new bone. There was no significant difference in the amount of bone formation between the SS group and the DPSS and PDLSS groups.ConclusionWithin the limitations of this study, silk scaffold is a biocompatible material with potential expediency as an osteoconductive scaffold in bone tissue engineering. However, there was no evidence to suggest that the addition of hDPCs and hPDLCs to the current rabbit calvarial defect model can produce an early effect in augmenting osteogenesis.

Osteoconductive effects of calcium phosphate glass cement grafts in rabbit calvarial defects.

Calcium phosphate glass (CPG) is well-documented alloplastic bone graft material. The objective of this study was to evaluate the osteoconductive effect of newly developed calcium phosphate glass cement (CPGC) in rabbit calvarial defects. Three circular defects (8 mm diameter) were created on the rabbit calvarium. One defect was filled with biphasic calcium phosphate (BCP group) and one defect was filled with CPGC (CPGC group). The remaining defect was not filled as the control. Histologic and histometric analysis were performed at four and eight weeks following the implantation of materials. One-way ANOVA method was used to evaluated the significance between three groups (p < 0.05). The CPGC group did not show a statistical difference in new bone area compared with the control at all healing periods, but the bone formation rate of CPGC seemed to increase between four and eight weeks. This suggests the bone formation rate of CPGC is initially slow, but increases at a specific time, showing the possibility of greater bone formation with time. The resorption rate of CPGC was greater than BCP. Within the limits of this study, CPGC demonstrated good space maintaining capacity and had an osteoconductive effect, suggesting it could be successfully used to improve bone formation capacity.

Effect of different hydroxyapatite:β-tricalcium phosphate ratios on the osteoconductivity of biphasic calcium phosphate in the rabbit sinus model

PURPOSE This study compared the volume stability and bone-forming capacity of biphasic calcium phosphate (BCP) with a high versus a low ratio of (β-tricalcium phosphate (β-TCP) relative to hydroxyapatite (HA), i.e., 70:30 vs 30:70, in the rabbit sinus model. MATERIALS AND METHODS Bilateral sinus windows were created in eight adult New Zealand white rabbits (2.5 to 3.5 kg); each sinus in each rabbit was assigned to one of two experimental BCP groups according to the HA: β-TCP ratio. One sinus was grafted with BCP with a high ratio of β-TCP (30:70; TCP70), and the contralateral sinus was grafted with BCP with a low ratio of β-TCP (70:30; TCP30). The animals were sacrificed after 2 weeks (n = 4) or 8 weeks (n = 4) of healing. Biopsy specimens were harvested and evaluated histologically, histomorphometrically, and with microcomputed tomography. RESULTS The bone volume did not differ significantly between the two groups at each healing point, or between 2 and 8 weeks of healing in both groups. The amount of new bone increased significantly between 2 and 8 weeks of healing in both groups, and it did not differ significantly between the TCP30 and TCP70 groups. The residual material was significantly more resorbed in the TCP70 group than in the TCP30 group at both 2 and 8 weeks. In the TCP70 group, a greater number of multinucleated giant cells were observed at both weeks. The bone-to-residual material contact ratio did not differ significantly between the two groups. CONCLUSION The volume stability and osteoconductive capacity of BCP with an HA: β-TCP ratio of 30:70 was comparable to that with an HA: β-TCP ratio of 70:30. Thus, within the limitations of this study, it can be argued that BCP with an HA: β-TCP ratio of 30:70 can be successfully used for sinus augmentation.

Delayed intentional replantation of periodontally hopeless teeth: a retrospective study.

Purpose The purpose of this study was to retrospectively evaluate the survival of periodontally hopeless teeth that were intentionally extracted and replanted after a delay and to compare the radiographic characteristics of the survival group with those of the failure group. Methods The clinical and radiographic data from patients who underwent delayed intentional replantation between March 2000 and July 2010 were reviewed. Twenty-seven periodontally hopeless teeth were extracted and preserved in medium supplemented with antibiotics for 10-14 days. The teeth were then repositioned in the partially healed extraction socket and followed for 3 to 21 months. The radiographic parameters were analyzed using a paired t test and the cumulative survival rate was analyzed using Kaplan-Meier analysis. Results Seven replanted teeth failed and the overall cumulative survival rate was 66.4%. In the survival group, the amount of bone loss was reduced from 68.45% to 34.66% three months after replantation. There was radiologic and clinical evidence of ankylosis with 5 teeth. However, no root resorption was found throughout the follow-up period. In the failure group, bone formation occurred from the bottom of the socket. However, a remarkable radiolucent line along the root of a replanted tooth existed. The line lengthened and thickened as time passed. Finally, in each case of failure, the tooth was extracted due to signs of inflammation and increased mobility. Conclusions Delayed intentional replantation has many advantages compared to immediate intentional replantation and could serve as an alternative treatment for periodontally involved hopeless teeth. However, techniques for maintaining the vitality of periodontal structures on the tooth surface should be developed for improved and predictable results. Graphical Abstract

Sulfuretin promotes osteoblastic differentiation in primary cultured osteoblasts and in vivo bone healing

Although sulfuretin, the major flavonoid of Rhus verniciflua Stokes, has a variety of biological actions, its in vitro and in vivo effects on osteogenic potential remain poorly understood. The objective of the present study was to investigate the effects of sulfuretin on in vitro osteoblastic differentiation and the underlying signal pathway mechanisms in primary cultured osteoblasts and on in vivo bone formation using critical-sized calvarial defects in mice. Sulfuretin promoted osteogenic differentiation of primary osteoblasts, with increased ALP activity and mineralization, and upregulated differentiation markers, including ALP, osteocalcin, and osteopontin, in a concentration-dependent manner. The expression levels of Runx2, BMP-2, and phospho-Smad1/5/8 were upregulated by sulfuretin. Moreover, sulfuretin increased phosphorylation of Akt, mTOR, ERK, and JNK. Furthermore, sulfuretin treatment increased mRNA expression of Wnt ligands, phosphorylation of GSK3, and nuclear β-catenin protein expression. In vivo studies with calvarial bone defects revealed that sulfuretin significantly enhanced new bone formation by micro-computed tomography and histologic analysis. Collectively, these data suggest that sulfuretin acts through the activation of BMP, mTOR, Wnt/β-catenin, and Runx2 signaling to promote in vitro osteoblast differentiation and facilitate in vivo bone regeneration, and might be have therapeutic benefits in bone disease and regeneration.

Layered approach with autogenous bone and bone substitute for ridge augmentation on implant dehiscence defects in dogs

OBJECTIVES This study compared the efficacies of different layered approaches using autogenous bone and synthetic bone substitute for ridge augmentation on implant dehiscence defects in dogs. MATERIALS AND METHODS Right mandibular second, third, and fourth premolars and the first molar were extracted, followed by standardized one-wall defect preparation in five dogs. After a healing period of 12 weeks, three implants (Implantium(®) ) were installed. Each of the three implant dehiscence defects was grafted with a different material as follows: (i) synthetic bone substitute combined with collagen (SBC; SBC group), (ii) inner autogenous bone layer and outer SBC layer (IAB group), and (iii) inner SBC layer and outer autogenous bone layer (OAB group). The grafted sites were covered with a resorbable collagen membrane. Quantitative and qualitative analyses of the subsequent bone regeneration were performed at 12 weeks postoperatively. RESULTS The dome-like augmented shape was relatively well maintained in the IAB and OAB groups, while the graft particles in the SBC group were dispersed. The bone-to-implant contact values tended to be significantly higher in the OAB group (49.51%) than in the SBC (36.58%) group. The amounts of newly formed bone within an area designated as 1 × 3 mm (width × height) from the implant platform in the IAB, OAB, and SBC groups were 35.59%, 28.10%, and 16.71%, respectively. CONCLUSION Application of the layered approach using autogenous bone and synthetic biomaterial resulted in substantial new bone formation and volume maintenance on implant dehiscence defects, irrespective of the position of the autogenous bone layer.

Assessment of dehydrothermally cross‐linked collagen membrane for guided bone regeneration around peri-implant dehiscence defects: a randomized single-blinded clinical trial

Purpose The aim of this study was to determine the clinical feasibility of using dehydrothermally cross‐linked collagen membrane (DCM) for bone regeneration around peri-implant dehiscence defects, and compare it with non-cross-linked native collagen membrane (NCM). Methods Dehiscence defects were investigated in twenty-eight patients. Defect width and height were measured by periodontal probe immediately following implant placement (baseline) and 16 weeks afterward. Membrane manipulation and maintenance were clinically assessed by means of the visual analogue scale score at baseline. Changes in horizontal thickness at 1 mm, 2 mm, and 3 mm below the top of the implant platform and the average bone density were assessed by cone-beam computed tomography at 16 weeks. Degradation of membrane was histologically observed in the soft tissue around the implant prior to re-entry surgery. Results Five defect sites (two sites in the NCM group and three sites in the DCM group) showed soft-tissue dehiscence defects and membrane exposure during the early healing period, but there were no symptoms or signs of severe complications during the experimental postoperative period. Significant clinical and radiological improvements were found in all parameters with both types of collagen membrane. Partially resorbed membrane leaflets were only observed histologically in the DCM group. Conclusions These findings suggest that, compared with NCM, DCM has a similar clinical expediency and possesses more stable maintenance properties. Therefore, it could be used effectively in guided bone regeneration around dehiscence-type defects.

Bone formation around rhBMP-2-coated implants in rabbit sinuses with or without absorbable collagen sponge grafting

Purpose The purpose of this study was to evaluate bone formation around recombinant human bone morphogenetic protein (rhBMP-2)-coated implants placed with or without absorbable collagen sponge (ACS) in rabbit maxillary sinuses. Methods The Schneiderian membrane was elevated and an implant was placed in 24 sinuses in 12 rabbits. The space created beneath the elevated membrane was filled with either blood (n=6) or ACS (n=6). In the rabbits in which this space was filled with blood, rhBMP-2-coated and non-coated implants were alternately placed on different sides. The resulting groups were referred to as the BC and BN groups, respectively. The AC and AN groups were produced in ACS-grafted rabbits in the same manner. Radiographic and histomorphometric analyses were performed after eight weeks of healing. Results In micro-computed tomography analysis, the total augmented volume and new bone volume were significantly greater in the ACS-grafted sinuses than in the blood-filled sinuses (P<0.05). The histometric analysis showed that the areas of new bone and bone-to-implant contact were significantly larger in the AC group than in the AN group (P<0.05). In contrast, none of the parameters differed significantly between the BC and BN groups. Conclusions The results of this pilot study indicate that the insertion of ACS after elevating the Schneiderian membrane, simultaneously with implant placement, can significantly increase the volume of the augmentation. However, in the present study, the rhBMP-2 coating exhibited limited effectiveness in enhancing the quantity and quality of regenerated bone.

Paracrine effect of the bone morphogeneticprotein-2 at the experimental site on healing of the adjacent control site: a study in the rabbit calvarial defect model

Purpose The aim of this study was to assess the possible paracrine effect of bone morphogeneticprotein-2 (BMP-2) at the experimental site on the adjacent control site for validating a rabbit calvarial defect model as a means of verifying the effect of BMP-2. Methods Sixteen rabbits were divided into two groups (n=8 in each) according to whether or not BMP-2 would be used. Two circular defects (8 mm in diameter) were created side by side, 2 mm apart, in the calvarium of all of the rabbits. In each animal, one of the defects was grafted with either BMP-2-loaded carrier or carrier material alone. The control defects adjacent to these grafted defects, designated CB (the nongrafted defect adjacent BMP-2-loaded carrier-grafted defect) and CC (the nongrafted defect adjacent to carrier only-grafted defect), respectively, were the focus of this study, and were filled only with a blood clot in all of the animals. Histologic observation and histomorphometric analysis were performed at 2 and 8 weeks (n=4 animals per point in time) after surgery. Results There was no noteworthy difference in the healing pattern, and no statistically significant differences in histomorphometric parameters such as the defect closure, new bone area, or total augmented area between the CC and CB groups. Conclusions The results of this study suggest that rabbit calvarial defects separated by a distance of 2 mm are suitable for evaluating the effects of BMP-2 and the control defect can be regarded not to be affected by BMP-2 applied defect. Graphical Abstract

The hidden X suture: a technical note on a novel suture technique for alveolar ridge preservation

Purpose The present study investigated the impact of 2 different suture techniques, the conventional crossed mattress suture (X suture) and the novel hidden X suture, for alveolar ridge preservation (ARP) with an open healing approach. Methods This study was a prospective randomized controlled clinical trial. Fourteen patients requiring extraction of the maxillary or mandibular posterior teeth were enrolled and allocated into 2 groups. After extraction, demineralized bovine bone matrix mixed with 10% collagen (DBBM-C) was grafted and the socket was covered by porcine collagen membrane in a double-layer fashion. No attempt to obtain primary closure was made. The hidden X suture and conventional X suture techniques were performed in the test and control groups, respectively. Cone-beam computed tomographic (CBCT) images were taken immediately after the graft procedure and before implant surgery 4 months later. Additionally, the change in the mucogingival junction (MGJ) position was measured and was compared after extraction, after suturing, and 4 months after the operation. Results All sites healed without any complications. Clinical evaluations showed that the MGJ line shifted to the lingual side immediately after the application of the X suture by 1.56±0.90 mm in the control group, while the application of the hidden X suture rather pushed the MGJ line slightly to the buccal side by 0.25±0.66 mm. It was demonstrated that the amount of keratinized tissue (KT) preserved on the buccal side was significantly greater in the hidden X suture group 4 months after the procedure (P<0.05). Radiographic analysis showed that the hidden X suture had a significant effect in preserving horizontal width and minimizing vertical reduction in comparison to X suture (P<0.05). Conclusions Our study provided clinical and radiographic verification of the efficacy of the hidden X suture in preserving the width of KT and the dimensions of the alveolar ridge after ARP.