• Journal of Periodontal and Implant Science
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• v.34 no.2
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• pp.425-448
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• 2004

A bioactive and degradable poly(epsilon -caprolactone)/silica nanohybrid(PSH) was synthesized for the application as a bone substitute. PSH was manufactured by using silica and polycaprolacton. PSH was manufactured in some composition after low crystaline apatite had been formed in simulated body fluid and, was used this study. The safety of the PSH was established by test of acute, and subacute toxicity, sensitization cytotoxicity and sterility. In order to assess activity of osteoblast, the test for attaching osteoblast, proliferation test for osteoblast, differentiating gene expression test are performed in vitro. And bone substitutes were grafted in rabbit's calvarium, during 8 weeks for testing efficacy of bone substitutes. Degree of osteogenesis and absorption of substitutes were evaluated in microscopic level. In result, it was not appeared that acute and subacute toxicity, sensitization in intradermal induction phase, topical induction phase and challenge phase. It was shown that the test can not inhibit cell proliferation. adversely, it had some ability to accelerate cell proliferation. The result of sterility test described bacterial growth was not detected in most test tube. The attaching and proliferation test of osteoblast had good results. In the result of differentiating gene expression test for osteoblast, cbfa1 and, alkaline phosphatase, osteocalcin and GAPDH were detected with mRNA analysis. In the PSH bone formation test, ostgeoblastic activity would be different as material constitution but it had good new bone formation ability except group #218. futhermore, some material had been absorbed within 8 weeks. Above studies, PSH had bio-compatibility with human body, new bone formation ability and accelerate osteoblastic activity. So it would be the efficient bone substitute material with bio-active and biodegradable.

• Journal of the korean academy of Pediatric Dentistry
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• v.30 no.4
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• pp.643-653
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• 2003

Fibroblast growth factor (FGF) / FGF receptor (FGFR) mediated signaling is required for skeletogenesis in cluding intramembranous and endochondral ossifications Runx2 ($Cbfa1/Pebp2{\alpha}A/AML3$) is an essential transcription factor for osteoblast differentiation and bone formation. Murine calvaria and mandible are concurrently undergoing both intramembranous bone and cartilage formations in the early developmental stage. However the mechanism by which these cartilage formations are regulated remains unclear. To elucidate the effect of FGF signaling on development of cranial sutural cartilage and Meckel's cartilage and to understand the role of Runx2 in these process, we have done both in vivo and in vitro experiments. Alcian blue staining showed that cartilage formation in sagittal suture begins from embryonic stage 16 (E16), Meckel's cartilage formation in mandible from E12. We analyzed by in situ hybridization the characteristics of cartilage cells that type II collagen, not type X collagen, was expressed in sagittal sutural cartilage and Meckel's cartilage. In addition, Runx2 was not expressed in Meckel's cartilage as well as sagittal sutural cartilage, except specific expression pattern only surrounding both cartilages. FGF signaling pathway was further examined in vitro. Beads soaked in FGF2 placed on the sagittal suture and mandible inhibited both sutural and Meckel's cartilage formations. We next examined whether Runx2 gene lies in FGF siganling pathway during regulation of cartilage formation. Beads soaked in FGF2 on sagittal suture induced Runx2 gene expression. These results suggest that FGF signaling inhibits formations of sagittal sutural and Meckel's cartilages, also propose that FGF siganling is involved in the proliferation and differentiation of chondroblasts through regulating the transcription factor Runx2.

• Journal of Periodontal and Implant Science
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• v.35 no.2
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• pp.345-357
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• 2005

Prostaglandin plays a significant role in the local control of bone metabolism associated with periodontal disease. ${\Delta}^{12}-PGJ_2$ is a natural $PGD_2$ metabolite that is formed in vivo in the presence of plasma. It is known for ${\Delta}^{12}-PGJ_2$ to stimulate calcification in osteoblastic cells. Bone morphogenetic protein(BMP) stimulated osteoblastic differentiation in various types of cells and greatly enhanced healing of bony defects. The purpose of this study was to evaluate the effect of rhEMP-2 on ${\Delta}^{12}-PGJ_2$ induced osteoblastic differentiation and mineralization in vitro. A human osteosarcoma cells line Saos-2 were cultured. In the test groups, 10-7M of ${\Delta}^{12}-PGJ_2$ or mixture of 10-8M of ${\Delta}^{12}-PGJ_2$ and 100ng/ml of rhBMP-2 or 100ng/ml of rhEMP-2 were added to culture media. After 1 day, 2 days and 4 days of culture period, the cell number was measured. Alkaline phosphatase activity was measure at 3 days. Reverse transcription polymerase chain reaction(RT-PCR) was performed to determine the expression of mRNA of bone matrix protein at 8 hours, 1 day and 7 days. The ability to produce mineralized nodules in rat osteoblasts(MC3T3-E1) was evaluated at 21 days. The results were as follows : 1. rhEMP-2 or mixture of rhBMP-2 and ${\Delta}^{12}-PGJ_2$ inhibited cell proliferation of human osteosarcoma cells. 2. rhEMP-2 or mixture of rhBMP-2 and ${\Delta}^{12}-PGJ_2$ stimulated alkaline phosphatase activity significantly higher than ${\Delta}^{12}-PGJ_2$ alone. 3. rhBMP-2 or mixture of rhEMP-2 and ${\Delta}^{12}-PGJ_2$ stimulated mineralization compared to ${\Delta}^{12}-PGJ_2$ alone. 4. mRNA of alkaline phosphatase, BMP-2, cbfa 1, Type I collagen were detected in the group treated with ${\Delta}^{12}-PGJ_2$/rhBMP-2, rhBMP-2 alone, ${\Delta}^{12}-PGJ_2$ alone. These results show that mixture of ${\Delta}^{12}-PGJ_2$ and rhBMP-2 causes more bone formation than ${\Delta}^{12}-PGJ_2$ alone while the bone formation effects of mixture of ${\Delta}^{12}-PGJ_2$ and rhBMP-2 are less than those of rhBMP-2 alone. Further researches would be necessary to clarify the interactions of these agents.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.27 no.1
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• pp.1-8
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• 2005

This study was aimed to characterize osteogenic potential of rat bone marrow stromal cells (BMSC) isolated with standard flushing method and investigate the plasticity of transdifferentiation between osteoblastic and adipocytic lineage of cultured BMSC. Unlike aspiration method in human, rat bone marrow was extracted by means of irrigation with culture media that elevates the possibility of co-extraction of committed osteoprogenitor, or preosteoblast or other progenitor cells of several types present inside bone marrow. The cultured stromal cells showed high ALP activity which is representative marker of osteoblast without any treatment. Osteogenic inducers such as Dex and BMP-2 were examined for the evaluation of their effect on osteogenic and adipocytic differentiation of stromal cells, because they function as osteoinductive agent in stromal cells, but simultaneously induce adipogenic differentiation. Osteogenic differentiation was evaluated by measuring alkaline phosphatase activity or mRNA expression of osteoblast markers such as osteopontin, bone sialoprotein, collagen type I and CbfaI, and in vitro matrix mineralization by von Kossa staining. Oil red staining method was used to detect adipocyte and adipocytic marker, aP2 and $PPAR{\gamma}2$ expression was examined using RT-PCR. It can be supposed that irrigation procedure resulted in high portion of already differentiation-committed osteoprogenitor cell showing elevated ALP activity and strong mineralization only under the supplement of $100{\mu}M$ ascorbic 2-phosphate and 10mM ${\beta}$-glycerophosphate without any treatment of osteogenic inducers such as Dex and BMP-2. Dex and BMP-2 seemed to transdifferentiate osteoprogenitor cells having high ALP activity into adipocytes temporarily, but continuous treatment redifferentiated into osteoblast and developed in vitro matrix mineralization. This property must be considered either in tissue engineering for bone regeneration, or in research of characterization of osteogenic differentiation, with rat BMSC isolated by the standard irrigation method.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.32 no.2
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• pp.97-106
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• 2010

Aim of the study: An alternative source of adult stem cells that could be obtained in large quantities, under local anesthesia, with minimal discomfort would be advantageous. Adipose tissue could be processed to obtain a fibroblast-like population of cells or adipose tissue-derived stromal cells (ATSCs). This study was performed to confirm the availability of ATSCs in bone tissue engineering. Materials amp; Methods: In this study, adipose tissue-derived mesenchymal stem cell was extracted from the liposuctioned abdominal fat of 24-old human and cultivated, and the stem cell surface markers of CD 105 and SCF-R were confirmed by immunofluorescent staining. The proliferation of bone marrow mesenchymal stem cell and ATSCs were compared, and evaluated the osteogenic differentiation of ATSCs in a specific osteogenic induction medium. Osteogenic differentiation was assessed by von Kossa and alkaline phosphatase staining. Expression of osteocyte specific BMP-2, ALP, Cbfa-1, Osteopontin and osteocalcin were confirmed by RT-PCR. With differentiation of ATSCs, calcium concentration was assayed, and osteocalcin was evaluated by ELISA (Enzyme-linked immunosorbant assay). The bone formation by 5-week implantation of HA/TCP block loaded with bone marrow mesenchymal stem cells and ATSCs in the subcutaneous pocket of nude mouse was evaluated by histologic analysis. Results: ATSCs incubated in the osteogenic medium were stained positively for von Kossa and alkaline phosphatase staining. Expression of osteocyte specific genes was also detected. ATSCs could be easily identified through fluorescence microscopy, and bone formation in vivo was confirmed by using ATSC-loaded HA/TCP scaffold. Conclusions: The present results show that ATSCs have an ability to differentiate into osteoblasts and formed bone in vitro and in vivo. So ATSCs may be an ideal source for further experiments on stem cell biology and bone tissue engineering.