• Molecules and Cells
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• v.38 no.7
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• pp.643-650
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• 2015

The use of conditioned medium from mesenchymal stem cells may be a feasible approach for regeneration of bone defects through secretion of various components of mesenchymal stem cells such as cytokines, chemokines, and growth factors. Mesenchymal stem cells secrete and accumulate multiple factors in conditioned medium under specific physiological conditions. In this study, we investigated whether the conditioned medium collected under hypoxic condition could effectively influence bone regeneration through enhanced migration and adhesion of endogenous mesenchymal stem cells. Cell migration and adhesion abilities were increased through overexpression of intercellular adhesion molecule-1 in hypoxic conditioned medium treated group. Intercellular adhesion molecule-1 was upregulated by microRNA-221 in mesenchymal stem cells because microRNAs are key regulators of various biological functions via gene expression. To investigate the effects in vivo, evaluation of bone regeneration by computed tomography and histological assays revealed that osteogenesis was enhanced in the hypoxic conditioned medium group relative to the other groups. These results suggest that behavioral changes of endogenous mesenchymal stem cells through microRNA-221 targeted-intercellular adhesion molecule-1 expression under hypoxic conditions may be a potential treatment for patients with bone defects.

• Journal of Periodontal and Implant Science
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• v.32 no.4
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• pp.769-779
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• 2002

This present study was carried out to find the effects of calcium aluminate cement($CaO\;{\cdot}\;Al_2O_3$, CAC), which has been developed with bio-compatibility and mechanical properties, in biological environments. Two different particle sizes of CAC - 3.5${\mu}m$ vs. 212${\sim}$250${\mu}m$ which is recommended in periodontal bone grafting procedures-were filled in 8mm calvarial defect in Sprague-Dawley rat. The specimens were examined histologically, especially the bone-cement interface and the response of surrounding tissues. The results are as follows; 1. In the control group, inflammatory cells were observed at 2 weeks. At 8 weeks, periosteum and dura mater were continuously joined together in the defect areas. But in the center of defect area were filled up with the loose connective tissues. 2. In the experimental group l($212{\mu}m{\sim}250{\mu}m$ particle), immature bone was formed and outermost layer was surrounded by osteoid layer at 2 weeks. Osteoblasts were arranged between immature bone and osteoid layer. And, osteoid layer was remained until 8 weeks after surgery. 3. In the experimental group 2, periosteum and dura mater lost its continuity at 2 weeks. Scattering of CAC particles and infiltration of inflammatory cells were observed, which this findings deepened at 8 weeks. The result of this study shows that when calvarial defects in white rats are filled with calcium aluminate cement of 212${\sim}$250${\mu}m$, the materials are to be bio-compatible in growth and healing on surrounding tissues. When further researches are fulfilled, such as direct bone adhesion and bone regeneration ability, it's possible that CAC could be applied to various periodontology fields in the future.

• Journal of Periodontal and Implant Science
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• v.28 no.4
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• pp.769-786
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• 1998

The purpose of the present study is to examine the effect of cell proliferation and alkaline phosphatase activity in osteoblastic cells and to compare the bone healing ability of rat calvarial defects between the control group and the safflower seed extract treated group. Osteoblastic cells were obtained from calvariae of a fetal rat. Cells were cultured containing DMEM and safflower seed extract ($10^{-6}g/ml$, $10^{-3}g/ml$) at $37^{\circ}$ with 5% $CO_2$ in 100% humidity for 3 days. MTT was performed to examine the viability of the cells, and alkaline phosphatase activity was analyzed to examine the mineralization in vitro. Rat calvarial defects($5{\times}5mm$) in 250g Sprague-Dawly were made using round bur. Rats were administrated with safflower seed extract(0.35g/kg/day) for experimental periods. Calvarial defects were studied histopathologically and immunohistochemically at 1,4, and 8 weeks. High concentration group($10^{-3}g/ml$) of safflower seed extract significantly increased in the cell proliferation and alkaline phos phatase synthesis in osteoblastic cells. The infiltration of inflammatory cells and osteoclastic activities were decreased in the safflower seed extract treated group as compared with control group. Bone maturation was accelerated in the safflower seed extract treated group as compared to control group. No difference in osteoinductive process was observed between the control and the safflower seed extract treated group. Immunohistochemical observation revealed that protein expression of TGF-$\beta$and osteonectin during early healing phase in the safflower seed extract treated group was slightly increased as compared to control group. These results indicate that safflower seed extract promotes the healing process in bony defect of rat calvariae, and retains a potential applicability as an adjuvant therapeutic modality for regeneration of periodontal bony defect.

• Journal of Acupuncture Research
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• v.21 no.4
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• pp.225-236
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• 2004

본 실험에서는 녹용 약침액의 항 골재흡수 속성을 조사하였다. PTH, $1,25(OH)_2D_3$와 IL-1을 각각 골재흡수 인자로 사용하여 생쥐의 두개골에서 osteoblast 세포를 격리, 배양, 그리고 자극시켰을 때 collagenolysis의 증가를 보였다. 두 가지를 동시에 사용한 결과, IL-1은 골재흡수성을 촉진시키고 재 흡수력을 생산하였다. In vitro에서의 세포독성 결과는 $1-200{\mu}g/ml$의 녹용 약침액 농도 분포에서 무세포독성을 보였다. 또한 녹용 약침액은 생쥐의 두개골 골아세포 내에서 PTH (2 unit/ml), IL-$1{\alpha}$ (1 ng/ml), $1,25(OH)_2D_3$ (10 ng/ml), IL-$1{\alpha}$ 및 IL-$1{\beta}$로 인해 유발된 collagenolysis에 대해서 대항하는 보호활동성을 나타내었다. 녹용약침액은 IL-$1{\alpha}$ 와 IL-$1{\beta}$로 인해 유발된 collagenolysis에 대항하는 보호활동성을 지녔다. DAS는 IL-$1{\alpha}$와 IL-$1{\beta}$로 인해 촉진된 골재 흡수력을 억제하는 효과를 보였다. 이와 같은 결과는 녹용약침액이 골다공증과 연관된 질환에 대해서 매우 안정적인 임상적 사용이 가능한 것을 관찰할 수 있으므로 추후 이와 관련한 지속적인 연구가 필요할 것으로 사료되었다.

• Journal of Periodontal and Implant Science
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• v.38 no.2
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• pp.125-134
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• 2008

Purpose: Bone morphogenetic protein (BMP) is a potent differentiating agent for cells of the osteoblastic lineage. It has been used in the oral cavity under a variety of indications and with different carriers. However, the optimal carrier for each indication is not known. This study evaluated the bone regenerative effect of rhBMP-2 delivered with different carrier systems. Materials and Methods: 8 mm critical-sized rat calvarial defects were used in 60 male Sprague-Dawley rats. The animals were divided into 6 groups containing 10 animals each. Two groups were controls that had no treatment and absorbable collagen membrane only. 4 groups were experimentals that contained rhBMP-2 only and applied with absorbable collagen sponge($Collatape^{(R)}$), $MBCP^{(R)}$, Bio-$Oss^{(R)}$ each. The histological and histometric parameters were used to evaluate the defects after 2- or 8-week healing period. The shape and total augmented area were stable in all groups over the healing time. Results: New bone formation was significantly greater in the rhBMP-2 with carrier group than control group. rhBMP-2/ACS was the highest in bone density but gained less new bone area than rhBMP-2/$MBCP^{(R)}$ and rhBMP-2/Bio-$Oss^{(R)}$. The bone density after 8 weeks was greater than that after 2 weeks in all groups. However, rhBMP-2 alone failed to show the statistically significant difference in new bone area and bone density compared to control group. Also $MBCP^{(R)}$ and Bio-$Oss^{(R)}$ particles remained after 8 weeks healing period. Conclusion: These results suggest that rhBMP-2 with carrier system is an excellent inductive agent for bone formation and we can use it as the predictable bone tissue engieering technique. Future study will likely focus on the kinetics of BMP release and development of carriers that is ideal for it.

• International Journal of Industrial Entomology and Biomaterials
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• v.45 no.2
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• pp.78-83
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• 2022

There are several subspecies of Bombyx mori, whose silk sericin variants differ. Silk sericin can induce bone morphogenic protein-2 (BMP-2) in macrophages, and silk sericin from different species may have different levels of BMP-2 induction ability. In this study, silk sericin from three B. mori subspecies (Baegokjam, Yeonnokjam, and Goldensilk) was prepared. They were administered to RAW264.7 cells and BMP-2 expression level was studied. Bone regeneration was evaluated using a rat calvarial defect model. BMP-2 expression level was the highest in the Baegokjam group. The bone volume in the Baegokjam group was significantly higher than that in the Yeonnokjam group (P = 0.003). In conclusion, sericin from Baegokjam showed higher levels of BMP-2 expression and bone regeneration than those from Yeonnokjam and Goldensilk.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.31 no.5
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• pp.440-453
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• 2005

Objectives : To develop a bioactive membrane for guided bone regeneration (GBR), the biocompatibility and bone regenerating capacity of the cellulose membrane obtained from the Ascidians squirt skin were evaluated. Materials and methods : After processing the pure cellulose membrane from the squirt skin, the morphological study, amino acid analysis and the immunoreactivity of the cellulose membrane were tested. Total eighteen male Spraque-Dawley rats (12 weeks, weighing 250 to 300g) were divided into two control (n=8) and another two experimental groups (n=10). In the first experimental group (n=5), the cellulose membrane was applicated to the 8.0 mm sized calvarial bone defect and the same sized defect was left without cellulose membrane in the first control group (n=4). In the another experimental group (n=5), the cellulose membrane was applicated to the same sized calvarial bone defect after femoral bone graft and the same sized defect with bone graft was left without cellulose membrane in the another control group (n=4). Each group was sacrificed after 6 weeks, the histological study with H&E and Masson trichrome stain was done, and immunohistochemical stainings of angiogenin and VEGF were also carried out. Results : The squirt skin cellulose showed the bio-inductive effect on the bone and mesenchymal tissues in the periosteum of rat calvarial bone. This phenomenon was found only in the inner surface of the cellulose membrane after 6 weeks contrast to the outer surface. Bone defect covered with the bioactive cellulose membrane showed significantly greater bone formation compared with control groups. Mesenchymal cells beneath the inner surface of the bioactive cellulose membrane were positive to the angiogenin and VEGF antibodies. Conclusion : We suppose that there still remains extremely little amount of peptide fragment derived from the basement membrane matrix proteins of squirt skin, which is a kind of anchoring protein composed of glycocalyx. This composition could prevent the adverse immunological hypersensitivity and also induce bioactive properties of cellulose membrane. These properties induced the effective angiogenesis with rapid osteogenesis beneath the inner surface of cellulose membrane, and so the possibilities of clinical application in dental field as a GBR material will be able to be suggested.

• Journal of Periodontal and Implant Science
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• v.41 no.5
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• pp.218-226
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• 2011

Purpose: Micro-computed tomography (micro-CT) has been widely used in the evaluation of regenerated bone tissue but the reliability of micro-CT has not yet been established. This study evaluated the correlation between histomorphometric analysis and micro-CT analysis in performing new bone formation measurement. Methods: Critical-size calvarial defects were created using a 8 mm trephine bur in a total of 24 Sprague-Dawley rats, and collagen gel mixed with autogenous rat bone marrow stromal cells (BMSCs) or autogenous rat BMSCs transduced by adenovirus containing bone morphogenic protein-2 (BMP-2) genes was loaded into the defect site. In the control group, collagen gel alone was loaded into the defect. After 2 and 4 weeks, the animals were euthanized and calvaria containing defects were harvested. Micro-CT analysis and histomorphometric analysis of each sample were accomplished and the statistical evaluation about the correlation between both analyses was performed. Results: New bone formation of the BMP-2 group was greater than that of the other groups at 2 and 4 weeks in both histomorphometric analysis and micro-CT analysis (P=0.026, P=0.034). Histomorphometric analysis of representative sections showed similar results to histomorphometric analysis with a mean value of 3 sections. Measurement of new bone formation was highly correlated between histomorphometric analysis and micro-CT analysis, especially at the low lower threshold level at 2 weeks (adjusted $r^2=0.907$, P<0.001). New bone formation of the BMP-2 group analyzed by micro-CT tended to decline sharply with an increasing lower threshold level, and it was statistically significant (P<0.001). Conclusions: Both histomorphometric analysis and micro-CT analysis were valid methods for measurement of the new bone in rat calvarial defects and the ability to detect the new bone in micro-CT analysis was highly influenced by the threshold level in the BMP-2 group at early stage.

• Journal of Periodontal and Implant Science
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• v.35 no.3
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• pp.747-760
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• 2005

Periodontal therapy has dealt primarily with attempts at arresting progression of disease. however, more recent techniques have focused on regenerating the periodontal ligament having the capacity to regenerate the periodontium. Recombinant human bone morphogenetic protein-7(rhBMP-7) can differentiate the osteoprogenitor cells and induce bone formation. The purpose of this study was to evaluate the effect of BMP-7 on rat periodontal ligament cells differentiation, in vitro. In the control group, cells was cultured with DMEM media. In the experimental groups, cells were cultured with rhBMP-7 in concentration of 10, 25, 50 and 100 ng/ml. Each group was characterized by examining alkaline phosphatase activity at 3 and 5 days of culture and the ability to produce mineralized nodules of rat calvarial cells at 14 days of culture. Synthesis of type I collagen(COL-I), osteocalcin(OCN), and bone sialoprotein(BSP) was evaluated by RT-PCR at 7 days of culture. Activation of Smad proteins and p38 MAP kinase was determined by western blot analysis of the cell lysates. Alkaline phosphatase activity was significantly increased in the concentration of BMP-7 50 ng/ml and 100 ng/ml compared to the control(p<0.05). The mineralized bone nodule formation was greater with addition of 50 ng/ml and 100 ng/ml BMP-7 than the control(p<0.01). In 7 days' culture, the expressions of COL-I, BSP, and OCN was increased by BMP-7 in concentration of 10 $ng/ml{\sim}100$ ng/ml. In western blot analysis, BMP-7 treated culture cells expressed Smad 1,5,8 in dose-dependent manner, whereas BMP-7 did not activate phosphorylated form of p38 MAP kinase. These result suggested that BMP-7 stimulate rat periodontal ligament cells to differentiate toward osteoblast phenotype and increase bone matrix production by activation of BMP-Smad pathway.

• Journal of Periodontal and Implant Science
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• v.34 no.4
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• pp.771-780
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• 2004

Chronic exposure to high levels of manganese (Mn) leads a pronounced and debilitating disorder known as manganism. Research on the toxic manifestation of manganese have focused primarily on its neurological effects because exposure to high levels of the metal produces a distinct and irreversible extrapyramidal dysfunction resembling the dystonic movements associated with Parkinson's physiological and biochemical systems in the body. The purpose of this study is to determine the effects of Mn on mineralization in primary rat calvarial cells. The experimental groups were in concentration of 0, 10, 30 and 60 ${\mu}M$. The results were as follows: 1. ALP activity was decreased in concentration of 30 and 60 ${\mu}M$ (p<0.01). 2. Bone nodule formation was depressed in concentration of 30 and 60 ${\mu}M$ at day 14 and 21 (p<0.01). 3. RT-PCR results showed an altered expression of bone matrix proteins. These result suggested that manganese might decrease or alter the expression of the osteoblast phenotype.