Purpose: The aim of this review is to introduce a novel bone-graft material for hard-tissue regeneration based on the calcium phosphate glass(CPG). Materials and Methods: CPG was synthesized by melting and subsequent quenching process in the system of CaO-$CaF_2-P_2O_5$-MgO-ZnO having a much lower Ca/P ratio than that of conventional calcium phosphates such as HA or TCP. The biodegradability and bioactivity were performed. Effects on the proliferation, calcification and mineralization of osteoblast-like cells were examined in vitro. Influence in new bone and cementum formations was investigated in vivo using calvarial defects of Sprague-Dawley rats as well as 1-wall intrabony defect of beagle dogs. The application to the tissue-engineered macroporous scaffold and in vitro and in vivo tests was explored. Results: The extent of dissolution decreased with increasing Ca/P ratio. Exposure to either simulated body fluid or fetal bovine serum caused precipitation on the surface. The calcification and mineralization of osteoblast-like cells were enhanced by CPG. CPG promoted new bone and cementum formation in the calvarial defect of Sprague-Dawley rats after 8 weeks. The macroporous scaffolds can be fabricated with $500{\sim}800{\mu}m$ of pore size and a three-dimensionally interconnected open pore system. The stem cells were seeded continuously proliferated in CPG scaffold. Extracellular matrix and the osteocalcin were observed at the $2^{nd}$ days and $4^{th}$ week. A significant difference in new bone and cementum formations was observed in vivo (p<0.05). Conclusion: The novel calcium phosphate glass may play an integral role as potential biomaterial for regeneration of new bone and cementum.

Purpose: The aim of this study was to evaluate adhesion and gene expression of the MC3T3-E1 cells cultured on machined titanium surface (MS) and anodized titanium surface (AS) using MTT test, Scanning electron micrograph and cDNA microarray. Materials and Methods: The MTT test assay was used for examining the proliferation of MC3T3-E1 cells, osteoblast like cells from Rat calvaria, on MS and AS for 24 hours and 48 hours. Cell cultures were incubated for 24 hours to evaluate the influence of the substrate geometry on both surfaces using a Scanning Electron Micrograph (SEM). The cDNA microarray Agilent Rat 22K chip was used to monitor expressions of genes. Results: After 24 hours of adhesion, the cell density on AS was higher than MS (p < 0.05). After 48 hours the cell density on both titanium surfaces were similar (p > 0.05). AS had the irregular, rough and porous surface texture. After 48 hours incubation of the MC3T3-E1 cells, connective tissue growth factor (CTGF) was up-regulated on AS than MS (more than 2 fold) and the insulin-like growth factor 1 receptor was down-regulated (more than 2 fold) on AS than MS. Conclusion: Microarray assay at 48 hours after culturing the cells on both surfaces revealed that osteoinductive molecules appeared more prominent on AS, whereas the adhesion molecules on the biomaterial were higher on MS than AS, which will affect the phenotype of the plated cells depending on the surface morphology.

Purpose: The aim of this study was to determine the survival rates of implants placed in grafted maxillary sinuses and compare the results obtained with graft materials, implant surfaces and timing of implant placement. Materials and Methods: Between January 1996 and December 2005, 391 implants were placed in 161 patients who underwent sinus grafting treatment simultaneously or separately at Ewha Womans University Hospital. According to inclusion criteria, 272 implants were placed in 102 patients with 112 sinus grafts (30 females, 72 males), aged 26 to 88 years (mean age $49.0{\pm}9.7$). The follow-up period ranged from 12 to 134 months (mean F/U $47{\pm}32$). Survival rates were evaluated according to graft material, implant surface and timing of implant placement. The Kaplan-Meier procedure and the log rank (Mantel-Cox) test were used to estimate survival rates and test for equality of survival rates between different groups of patients. Results: Ten-year cumulative survival rate for implants placed in the grafted sinuses was 90.1%. The survival rates for autogenous bone, combination and bone substitutes were 94.6%, 85.9% and 100%, respectively (p > 0.05). According to implant surface, survival rates were 84.8% in machined group and 97.5% in rough group (p < 0.05). The survival rates were 92.9% in delayed group and 86.0% in simultaneous group (p > 0.05). Conclusion: Ten-year cumulative survival rate for implants placed in the grafted sinuses was 90.1%. Rough-surfaced implants have a higher survival rate than machined-surfaced implants when placed in grafted sinuses (p < 0.05).

Purpose: Aggregatibacter actinomycetemcomitans was associated with localized aggressive periodontitis, endocarditis, meningitis, and osteomyelitis. The cytolethal distending toxin (CDT) of A. actinomycetemcomitans was considered as a key factor of these diseases is composed of five open reading frames (ORFs). Among of them, An enzymatic subunit of the CDT, CdtB has been known to be internalized into the host cell in order to induce its genotoxic effect. However, CdtB can not be localized in host cytoplasm without the help of a heterodimeric complex consisting of CdtA and CdtC. So, some studies suggested that CdtC functions as a ligand to interact with GM3 ganglioside of host cell surface. The precise role of the CdtC protein in the mechanism of action of the holotoxin is unknown at the present time. The aim of this study was to generate recombinant CdtC proteins expression from A. actinomycetemcomitans, through gene cloning and protein used to investigate the function of Cdt C protein in the bacterial pathogenesis. Materials and Methods: The genomic DNA of A. actinomycetemcomitans Y4 (ATCC29522) was isolated using the genomic DNA extraction kit and used as template to yield cdtC genes by PCR. The amplifed cdtC genes were cloned into T-vector and cloned cdt C gene was then subcloned to pET28a expression vector. The pET28a-cdtC plasmid expressed in BL21 (DE3) Escherichia coli system. Diverse conditons were tested to opitimize the expression and purification of functional CdtC protein in E. coli. Results: In this study we reconstructed CdtC subunit of A. actinomycetemcomitans Y4 and comfirmed the recombinant CdtC expression by SDS-PAGE and Western Blotting. The expression level of the recombinant CdtC was about 2% of total bacterial proteins. Conclusion: The lab condition of procedure for the purification of functionally active recombinant CdtC protein is established. The active recombinant CdtC protein will serve to examine the role of CdtC proteins in the host recognition and enzyme activity of CDT and investigate the pathological process of A. actinomycetemcomitans in periodontal disease.

Purpose: The carrier used as delivery agent for bone morphogenetic proteins(BMPs) should also act as a scaffold for new bone formation. Moreover, bone formation should be predictable in terms of the volume and shape. This study examined the osteogenic effect of macroporous biphasic calcium phosphate (MBCP) block combined with ePTFE membrane as a carrier for recombinant human bone morphogenetic proteins (rhBMP-2). In addition, the additive effect of ePTFE membrane on bone formation was evaluated. Materials and Methods: Eight-millimeter critical sized calvarial defects were created surgically in 28 male Sprague-Dawley rats. The animals were divided into 2 groups containing 14 animals each. The defects were treated with either rhBMP-2/MBCP block (rhBMP-2/MBCP group) or rhBMP-2/MBCP block/ePTFE membrane (rhBMP-2/MBCP/ePTFE group). A disc-shaped MBCP block (3 mm height and 8 mm diameter) was used as the carrier for the rhBMP-2 and ePTFE membrane was used to cover the rhBMP-2/MBCP block. The histologic and histometric parameters were used to evaluate the defects after 2- or 8-week healing period (7 animals/group/healing interval). Results: The level of bone formation in the defects of both groups was significantly higher at 8 weeks than that at 2 weeks (P < 0.05). The ePTFE membrane has no additional effect compared with the rhBMP-2/MBCP block only. However, at 8 weeks, rhBMP-2/MBCP/ePTFE group showed more even bone formation on the top of the MBCP block than the rhBMP-2/MBCP group. Conclusion: These results suggest that the ePTFE membrane has no additive effect on bone formation when a MBCP block is used as a carrier for rhBMP-2.

Purpose: Cytolethal distending toxin (CDT) considered as a key factor of localized aggressive periodontitis, endocarditis, meningitis, and osteomyelitis is composed of five open reading frames (ORFs). Among of them, the individual role of CdtA and CdtC is not clear; several reports presents that CDT is an AB2 toxin and they enters the host cell via clathrin-coated pits or through the interaction with GM3 ganglioside. So, CdtA, CdtC, or both seem to be required for the delivery of the CdtB protein into the host cell. Moreover, recombinant CDT was suggested as good vaccine material and antibody against CDT can be used for neutralization or for a detection kit. Materials and Methods: We constructed the pET28a-cdtC plasmid from Aggregatibacter actinomycetemcomitans Y4 by genomic DNA PCR and expressed in BL21 (DE3) Escherichia coli system. We obtained the antibody against the recombinant CdtC in mice system. Using the anti-CdtC antibody, we test the native CdtC detection by ELISA and Western Blotting and confirm the expression time of native CdtC protein during the growth phase of A. actinomycetemcomitans. Results: In this study we reconstructed CdtC subunit of A. actinomycetemcomitans Y4 and generated the anti CdtC antibody against recombinant CdtC subunit expressed in E. coli system. Our anti CdtC antibody can be interacting with recombinant CdtC and native CDT in ELISA and Western system. Also, CDT holotoxin existed at 24h but not at 48h meaning that CDT holotoxin was assembled at specific time during the bacterial growth. Conclusion: In conclusion, we thought that our anti CdtC antibody could be used mucosal adjuvant or detection kit development, because it could interact with native CDT holotoxin.

Purpose: Aggregatibacter actinomycetemcomitans is associated with localized aggressive periodontitis. It produces cytolethal distending toxin (CDT), which induces cell cycle arrest in the G2/M phase. The CDT holotoxin is composed of CdtA, CdtB, and CdtC. CdtB has structural homology to human DNase I and is an active component of the CDT complex acting as a DNase. In particular, the pattern homology seen in the CdtB subunit has been associated with specific DNase I residues involved in enzyme catalysis, DNA binding, and metal ion binding. So, to study the functions and regulation of recombinant CdtB, we made up a quantity of functional recombinant CdtB and tested it in relation to the metal ion effect. Materials and Methods: We constructed the pET28a-cdtB plasmid from A. actinomycetemcomitans Y4 by genomic DNA PCR and expressed it in the BL21 (DE3) Escherichia coli system. We obtained the functional recombinant CdtB by the refolding system using the dialysis method and then analyzed the DNase activity and investigated the metal ion effect from plasmid digestion. Results: The recombinant CdtB subunit was expressed as the inclusion bodies. We were able to obtain functional recombinant CdtB subunit using refolding system. We confirmed that our refolded recombinant CdtB had DNase activity and was influenced by the metal ions $Mg^{2+}$ and $Ca^{2+}$. Conclusion: We suggest that the factors influencing recombinant CdtB may contribute to CDT associated diseases, such as periodontitis, endocarditic, meningitis, and osteomyelitis.

Purpose: The carrier for the delivery of bone morphogenetic proteins(BMPs) should also serve as a scaffold for new bone growth. In addition, predictable bone formation in terms of the volume and shape should be guaranteed. This study evaluated the ectopic bone formation of recombinant human BMP-2(rhBMP-2) using a micro macroporous biphasic calcium phosphate (MBCP: mixture of ${\beta}TCP$ and HA) block as a carrier in a rat subcutaneous assay model. Materials and Methods: Subcutaneous pockets were created on the back of 40 male Sprague-Dawley rats. In the pockets, rhBMP-2/MBCP and MBCP alone were implanted. The blocks were evaluated by histological and histometric parameters after a healing interval of 2 weeks (each 10 rats; MBCP and rhBMP-2/MBCP) or 8 weeks (each 10 rats; MBCP and rhBMP-2/MBCP). Results: The shape and volume of the block was maintained stable over the healing period. No histological bone forming activity was observed in the MBCP alone sites after 2 weeks and there was minimal new bone formation at 8 weeks. In the rhBMP-2/MBCP sites, new bone formation was evident in the macropores of the block. The new bone area at 8 weeks was greater than at 2 weeks. There was a further increase in the quantity of new bone with the more advanced stage of remodeling. Conclusions: A MBCP block could serve as a carrier system for predictable bone tissue engineering using rhBMPs.

Purpose: This study compared the effects of coating implants with hydroxyapatite (HA) using an ion beam-assisted deposition (IBAD) method prepared with machined, anodized, sandblasted and large-grit acid etched (SLA) surfaces in minipigs, and verified the excellency of coating method with HA using IBAD. Material and Methods: 4 male Minipigs(Prestige World Genetics, Korea), 18 to 24 months old and weighing approximately 35 to 40 kg, were chosen. All premolars and first molars of the maxilla were carefully extracted on each side. The implants were placed on the right side after an 8 week healing period. The implant stability was assessed by resonance frequency analysis (RFA) at the time of placement. 40 implants were divided into 5 groups; machined, anodized, anodized plus IBAD, SLA, and SLA plus IBAD surface implants. 4 weeks after implantation on the right side, the same surface implants were placed on the left side. After 4 weeks of healing, the minipigs were sacrificed and the implants were analyzed by RFA, histology and histometric. Results: RFA showed a mean implant stability quotient (ISQ) of $75.625{\pm}5.021$, $76.125{\pm}3.739$ ISQ and $77.941{\pm}2.947$ at placement, after 4 weeks healing and after 8 weeks, respectively. Histological analysis of the implants demonstrated newly formed, compact, mature cortical bone with a nearby marrow spaces. HA coating was not separated from the HA coated implant surfaces using IBAD. In particular, the SLA implants coated with HA using IBAD showed better contact osteogenesis. Statistical and histometric analysis showed no significant differences in the bone to implant contact and bone density among 5 tested surfaces. Conclusion: We can conclude that rough surface implants coated with HA by IBAD are more biocompatible, and clinical, histological, and histometric analysis showed no differences when compared with the other established implant surfaces in normal bone.

Purpose: The aim of this study is to compare the healing response of various Hydroxyapatite(HA) coated dental implants by Ion-Beam Assisted Deposition(IBAD) placed in the surgically created circumferential gap in dogs. Materials and methods: In four mongrel dogs, all mandibular premolars and the first molar were extracted. After an 8 weeks healing period, six submerged type implants were placed and the circumferential cylindrical 2mm coronal defects around the implants were made surgically with customized step drills. Groups were divided into six groups : anodized surface, anodized surface with 150nm HA and heat treatment, anodized surface with 300nm HA and heat treatment, anodized surface with 150nm HA and no heat treatment, and anodized surface with 150nm HA, heat treatment and bone graft, anodized surface with bone graft. The dogs were sacrificed following 12 weeks healing period. Specimens were analyzed histologically and histomorphometrically. Results: During the healing period, healing was uneventful and implants were well maintained. Anodized surface with HA coating and $430^{\circ}C$ heat treatment showed an improved regenerative characteristics. Most of the gaps were filled with newly regenerated bone. The implant surface was covered with bone layer as base for intensive bone formation and remodeling. In case that graft the alloplastic material to the gaps, most of the coronal gaps were filled with newly formed bone and remaining graft particles. The bone-implant contact and bone density parameters showed similar results with the histological findings. The bone graft group presented the best bone-implant contact value which had statistical significance. Conclusion: Within the scope of this study, nano-scale HA coated dental implants appeared to have significant effect on the development of new bone formation. And additional bone graft is an effective method in overcoming the gaps around the implants.

Purpose: The aim of this study was to evaluate the factors affecting healing patterns of surgically created circumferential gap defects around implants in dogs. Materials and Methods: In four mongrel dogs, all mandibular premolars were extracted. After 8 weeks of healing periods, implants were submerged. According to the surface treatment, turned surface was designated as a group A and rough surface as a group B. In each dog, surgical defects on the left side were made with a customized tapered step drill and on the right with a customized paralleled drill. Groups were also divided according to the width of the coronal gaps: 1.0mm, 1.5mm, or 2.0mm. The dogs were sacrificed following 8 weeks and the specimens were analyzed histologically and histomorphometrically. Results: During the postoperative period, healing was uneventful and implants were well-maintained. As the size of the coronal gap was increased, the amount of bone-to-implant contact was decreased. The bone healing was greater in rough surface implants compared to the turned ones. About the defect morphology, tapered shape showed much bone healing and direct bone to implant contact even in the smooth surface implants. Conclusion: Healing of the circumferential defect around dental implant is influenced by the implant surface, defect width and the morphology of the defect. When using rough surface implants, circumferential gap defects within 2 mm do not need any kinds of regenerative procedures and the healing appeared to be faster in the tapered defect morphology than the paralleled one.

Purpose: Osteoprotegerin (OPG) is a secreted glycoprotein and a member of the tumor necrosis factor (TNF) receptor family that inhibits bone resorption by suppressing osteoclastogenesis. Gingival fibroblasts (GF) play a role in periodontal disease progression, and the purpose of this experiment was to evaluate influence of osteotropic factors on the expression of osteoprotegerin mRNA in these cells. Materials and Methods: In this experiment, the influence of osteoclastogenic factors, interleukin-1 beta (IL-$1{\beta}$), TNF-$\alpha$, prostanglandin E2 ($PEG_2$). parathyroid hormone (PTH) and 1$\alpha$, 25-dihydroxyvitamin $D_3$ on the expression of osteoprotegerin mRNA in GF was studied by Northern blot hybridization. Results: As expected, $PEG_2$ tended to inhibit OPG levels and this was most prominent at 24 hours of culture with $10^{-7}M$ of $PEG_2$. TNF-$\alpha$ at 10ng/ml and also at 25ng/ml decreased OPG levels to almost 30% of the control at 24 hours. This contrasts with reports of increased OPG levels from osteoblast/stromal cells and gingival fibroblasts stimulated by TNF-$\alpha$. Decrease of OPG levels with $PEG_2$ and TNF-$\alpha$ suggests a pathway whereby these mediators exert their resorptive effects. However, OPG levels were increased almost 3-fold at 24 hours with IL-1$\beta$(1 to 15ng/ml) and increased 1.4 fold with 24-hour treatment of $10^{-7}M$ PTH. Conclusion: Increase of OPG levels suggests that these 'osteoclastogenic' factors act in more complex ways and may act to inhibit bone resorption in inflammatory periodontitis. This result supports the role of OPG as a negative feedback mechanism in osteoclastic activity.

Purpose: Pathologic tooth migration (PTM) commonly occurs in the anterior region and is associated with periodontal disease. The treatment of PTM of anterior teeth can be complex and time consuming, and a multidisciplinary approach is often required. Materials and Methods: The patient was a 38-year-old woman with a chief complaint of saving and realigning her elongated maxillary left central incisor. This paper describes the successful combined periodontal regenerative (guided tissue regeneration) and prosthetic treatment and a 2-year follow-up of maxillary central incisor with pathologic tooth migration, deep intrabony defect, and poor prognosis. Results: The right maxillary central incisor was restored by laminate veneer and the left by all-ceramic crown. The patient had no pain and discomfort and was satisfied with the outcomes of her treatment for 2 years. She has maintained her recall program at the Department of Periodontology at 3 months interval. Conclusion: The key step in the successful treatment of PTM in anterior region is to obtain a high level of cooperation from the patient. Maintenance of the treatment result of PTM is dependent on the continuous preservation of periodontal health.