• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.34 no.4
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• pp.419-427
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• 2008

The present study aimed to investigate the osteogenic potentials of differentiated osteoblast-like cells (DOCs) induced from bone marrow-derived mesenchymal stem cells (MSCs) on ${\beta}-tricalcium$ phosphate (${\beta}-TCP$) with recombinant human bone morphogenetic protein (rhBMP-2) in vitro. Osteoblast differentiation was induced in confluent cultures by adding 100 nM dexamethasone, 10 mM ${\beta}$-glycerophosphate, 50 mM L-ascorbic acid. The Alizarin red S staining and reverse transcriptase-polymerase chain reaction (RT-PCR) were perfomed to examine the mRNA expression of alkaline phosphatase (ALP), bone sialoprotein (BSP), osteocalcin (OCN), receptor activator for nuclear factor ${\kappa}B$ ligand (RANKL), runt-related transcription factor 2 (RUNX2), collagen-Ⅰ (COL-Ⅰ). There were no significant differences in the osteogenic potentials of DOCs induced from MSCs on ${\beta}-TCP(+/-)$. According to the incubation period, there were significant increasing of Alizadin red S staining in the induction 3 weeks. The mRNA expression of ALP, RUNX2, and RANKL were higher in DOCs/${\beta}-TCP(-)$ than DOCs/${\beta}-TCP(+)$. According to rhBMP-2 concentrations, the mRNA expression of BSP was significantly increased in DOCs/${\beta}-TCP(+)$ compared to that of DOCs/${\beta}-TCP(-)$ on rhBMP 10 ng/ml. Our study presented the ${\beta}-TCP$ will have the possibility that calcium phosphate directly affect the osteoblastic differentiation of the bone marrowderived MSCs.

• Journal of Korean Foot and Ankle Society
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• v.18 no.3
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• pp.100-107
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• 2014

Purpose: The purpose of this study is to evaluate the efficacy of mesenchymal stem cell (MSC) isolation by the magnetic-activated cell sorting (MACS) method in tendon tissue-derived cells compared to the colony picking method for isolation of MSCs by picking colony-forming cells. Materials and Methods: Human tendon-derived cells were isolated by enzyme digestion using normal tendon tissues from three donors. We used the magnetic kit and well-known MSC markers (CD90 or CD105) to isolate MSCs in tendon-derived cells using MACS. Cloning cylinders were used to isolate colony-forming cells having MSC characteristics in tendon-derived cells. Colony-forming unit-fibroblast (CFU-F) assay was used to evaluate the self-renewal capacity of cells isolated using the colony picking method or MACS. For comparison of differentiation potentials into osteogenic or adipogenic lineage between two groups, alizarin red S and oil red O staining were performed at 14 days after induction of differentiation in vitro. Results: Flow cytometry results showed that early passage tendon-derived cells expressed CD44 in 99.13%, CD90 in 56.51%, and CD105 in 86.19%. In the CFU-F assay, CD90+ or CD105+ cells isolated with MACS showed larger colony formation in size than cells isolated using the colony picking method. We also observed that CD90+ or CD105+ cells were constantly differentiated into both osteogenic and adipogenic lineages in cells from all donors, whereas cells isolated using the colony picking method were heterogeneous in differentiation potentials to the osteogenic and adipogenic lineages. Conclusion: CD90+ or CD105+ cells isolated using MACS showed superior MSC characteristics in the self-renewal and multi-differentiation capacities compared with cells isolated using the colony picking method.

• Journal of Society of Preventive Korean Medicine
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• v.14 no.3
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• pp.1-12
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• 2010

Objective : Samulatang (herbal description) is much used for women's disease in Korean Traditional Medicine. The aim of this study is to evaluate reproductive toxic effect by Samultang in pregnant rats and fetuses, and ascertain a dose-response relationship Method : Pregnant Sprague-Dawley rats were administered with the Samultang at single, double and quadruple dose for 20 days, orally. Pregnant rats were sacrificed at 20th day of gestation, and observed internal and reproductive organs. Live fetuses of gestation were randomly selected and fixed in 95% ethanol. Fetuses were stained with alcian blue and alizarin red S. We observe maternal body weight,, index associated pregnancy, and skeletal malformations in fetus Result : Maternal body weight of Samultang treated group has increased, side effect was not found in maternal body compared to that of control group. There were no significant difference in internal and reproductive organs. Double concentration administered group had lowest value in number of implantation, live fetuses, implantation rate and delivery rate, Also double concentration administered group showed higher early and late resorption rate than the other group. But, these are not significant. In the sex ratio, number of females, bigger than number of males in all Samultang administered groups. The fetuses of dams treated with Samultang didn't showed external and skeletal malformation. Vertebral and sternal variations were observed in single, double and quadruple concentration administered group but, compared to the control, those variations were insignificant. There were no significant changes in number of ribs, cervical, thoracic, lumber, sacral and caudal vertebrae Conclusion : Samultang is not expected to affect on pregnant rats and fetus about maternal body weight and number of live fetuses. There were no significant changes in organ weight, reproductive organs. Although skeletal variations were showed in vertebrae and sternum, treated groups were shown insignificant changes in skeletal variation

• Reproductive and Developmental Biology
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• v.37 no.4
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• pp.219-224
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• 2013

Muscular satellite cell (SC), which is stem cell of postnatal pig, is an important for study of differentiation into adipogenesis, myogenesis, and osteoblastogenesis. In this study, we isolated and examined from pig muscle tissue to determine capacity in proliferate, differentiate, and expression of various genes. Porcine satellite cells (PSC) were isolated from semimembranosus (SM) muscles of 90~100 days old pigs according to standard conditions. The cell proliferation increased in multi-potent cell by Masson's, oil red O, and Alizarin red staining respectively. We performed the expression levels of differentiation related genes using real-time PCR. We found that the differentiation into adipocyte increased expression levels of both fatty acid binding protein 4 (FABP4) and peroxisome proliferator-activated receptor gamma ($PPAR{\gamma}$) genes (p<0.01). Myocyte increased the expression levels of the myosin heavy chain (MHC), myogenic factor 5 (Myf5), myogenic regulatory factor (MyoD), and Myogenic factor 4 (myogenin) (p<0.01). Osteoblast increased the expression levels of alkaline phosphatase (ALP) (p<0.01). Finally, porcine satellite cells were induced to differentiate towards adipogenic, myogenic, and osteoblastogenic lineages. Our results suggest that muscle satellite cell in porcine may influence cell fate. Understanding the progression of PSC may lead to improved strategies for augmenting meat quality.

• Journal of Periodontal and Implant Science
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• v.51 no.5
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• pp.352-363
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• 2021

Purpose: Some systemic conditions, especially diabetes mellitus (DM), adversely affect dental implant success. This study aimed to investigate the effects of ibuprofen-loaded TiO₂ nanotube (ILTN) dental implants in alloxan-induced diabetic rabbits. Methods: Twenty-six New Zealand white rabbits were treated with alloxan monohydrate to induce DM. At 2 weeks following DM induction, 3 types of implants (sandblasted, large-grit, and acid-etched [SLA], ILTN, and machined) were placed into the proximal tibia in the 10 rabbits that survived following DM induction. Each type of implant was fitted randomly in 1 of the holes (round-robin method). The animals were administered alizarin (at 3 weeks) and calcein (at 6 weeks) as fluorescent bone markers, and were sacrificed at 8 weeks for radiographic and histomorphometric analyses. Results: TiO₂ nanotube arrays of ~70 nm in diameter and ~17 ㎛ in thickness were obtained, and ibuprofen was loaded into the TiO₂ nanotube arrays. A total of 26 rabbits were treated with alloxan monohydrate and only 10 rabbits survived. The 10 surviving rabbits showed a blood glucose level of 300 mg/dL or higher, and the implants were placed in these diabetic rabbits. The implant stability quotient (ISQ) and bone-to-implant contact (BIC) values were significantly higher in the ILTN group (ISQ: 61.8, BIC: 41.3%) and SLA group (ISQ: 62.6, BIC: 46.3%) than in the machined group (ISQ: 53.4, BIC: 20.2%), but the difference in the BIC percentage between the SLA and ILTN groups was not statistically significant (P=0.628). However, the bone area percentage was significantly higher in the ILTN group (78.0%) than in the SLA group (52.1%; P=0.000). Conclusions: The: ILTN dental implants showed better stability (ISQ) and BIC than the machined implants; however, these values were similar to the commercially used SLA implants in the 2-week diabetic rabbit model.

• Molecules and Cells
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• v.41 no.12
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• pp.1016-1023
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• 2018

Regenerative orthopedics needs significant devices to transplant human stem cells into damaged tissue and encourage automatic growth into replacements suitable for the human skeleton. Soft biomaterials have similarities in mechanical, structural and architectural properties to natural extracellular matrix (ECM), but often lack essential ECM molecules and signals. Here we engineer mineralized polysaccharide beads to transform MSCs into osteogenic cells and osteoid tissue for transplantation. Bone morphogenic proteins (BMP-2) and indispensable ECM proteins both directed differentiation inside alginate beads. Laminin and collagen IV basement membrane matrix proteins fixed and organized MSCs onto the alginate matrix, and BMP-2 drove differentiation, osteoid tissue self-assembly, and small-scale mineralization. Augmentation of alginate is necessary, and we showed that a few rationally selected small proteins from the basement membrane (BM) compartment of the ECM were sufficient to up-regulate cell expression of Runx-2 and osteocalcin for osteoid formation, resulting in Alizarin red-positive mineral nodules. More significantly, nested BMP-2 and BM beads added to a non-union skull defect, self-generated osteoid expressing osteopontin (OPN) and osteocalcin (OCN) in a chain along the defect, at only four weeks, establishing a framework for complete regeneration expected in 6 and 12 weeks. Alginate beads are beneficial surgical devices for transplanting therapeutic cells in programmed (by the ECM components and alginate-chitosan properties) reaction environments ideal for promoting bone tissue.

• Journal of dental hygiene science
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• v.23 no.4
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• pp.282-295
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• 2023

Background: Secretory leukocyte protease inhibitor (SLPI) protects tissues from proteases and promotes cell proliferation and healing. SLPI also reduces periodontal inflammation and alveolar bone resorption by inhibiting proinflammatory cytokine expression in rat periodontal tissues and osteoblasts. However, little is known of the role of SLPI in the expression of osteoclast regulatory factors from osteoblasts, which are crucial for the interaction between osteoblasts and osteoclasts. Therefore, we aimed to determine the effects of SLPI on the regulation of osteoclasts and osteoblasts in LPS-treated alveolar bone and osteoblasts. Methods: Periodontitis was induced in rats using LPS. After each LPS injection, SLPI was injected into the same area. Immunohistochemical analysis was performed with antibodies against SLPI, RANKL, OPG, and Runx2 in the periodontal tissue. RT-PCR and western blotting were performed to determine the expression levels of SLPI, RANKL, OPG, and Runx2 in LPS- and SLPI/LPS-treated MC3T3-E1 cells. SLPI/LPS-treated MC3T3-E1 cells were also stained with Alizarin Red S. Results: Immunohistochemical analysis showed that the expression levels of SLPI, OPG, and Runx2 were higher while that of RANKL was lower in the LPS/SLPI group relative to those in the LPS group. The mRNA and protein expression of SLPI, OPG, and Runx2 was higher in SLPI/LPS/MC3T3-E1 cells than in LPS/MC3T3-E1 cells, and RANKL expression was lower. During differentiation, OPG and Runx2 protein levels were higher whereas RANKL levels were lower in SLPI/LPS/MC3T3-E1 than in LPS/MC3T3-E1 cells on days 0, 4, 7, and 10. In addition, mineralization and matrix deposition were higher in SLPI/LPS/MC3T3-E1 than in LPS/MC3T3-E1 on days 7 and 10. SLPI decreased RANKL expression in LPS-treated alveolar bone and osteoblasts but increased the expression of OPG and Runx2. Conclusion: SLPI can be considered as a regulatory molecule that indirectly regulates osteoclast activation via osteoblasts and promotes osteoblast differentiation.

• Journal of the korean academy of Pediatric Dentistry
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• v.24 no.1
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• pp.1-17
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• 1997

The purpose of this study was to investigate the aspects of proliferation and bone nodule formation of osteogenic precursor cells. To determine the effects of ascorbic acid and dexamethasone upon capacity of osteoblast proliferation and bone nodule formation, cells were maintained in the presence of one or some of these additives for up to 30 days. Group I culture was maintained in standard medium(DMEM plus 10% plus antibiotics), group II was maintained in supplemented medium containing dexamethasone, group III was maintained in supplemented medium containing ascorbic acid and sodium-${\beta}$-glycerophosphate, and group IV was maintained in supplemented containing ascorbic acid, sodium-${\beta}$-glycerophosphate and dexamethasone. Morphology of bone nodules was observed with light microscope and electron microscope. The results were as follows: ${\bullet}$ Proliferation capacity of osteoblasts was not affected by single use of dexamethasone, but it was chiefly affected by ascorbic acid. ${\bullet}$ Cellular morphology was fibroblastic appearance initially, but, it was gradually changed to polygonal shape accompanied by confluency stage. ${\bullet}$ Pluripotent mesenchymal cells existed during primary culture, they were differentiated to adipocyte, chondrocyte, osteocyte according to culture condition. ${\bullet}$ Dexamethasone increased bone nodule formation under the condition that the culture was maintained with supplemented medium ascorbic acid and sodium-${\beta}$-glycerophosphate. ${\bullet}$ when the cultures were stained with alizarin red, the group supplemented with dexamethasone, ascorbic acid and sodium-${\beta}$-glycerophosphate showed the marked increase of bone nodule formation, but the group supplemented with ascorbic acid and sodium-${\beta}$-glycerophosphate revealed only small amounts of bone nodules. And the groups cultured without ascorbic acid showed no observed any of bone-like mass independent of dexamethasone addition.

• The Journal of Korean Academy of Prosthodontics
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• v.49 no.3
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• pp.245-253
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• 2011

Purpose: The aim of this in vitro study was to estimate surface characteristic after peptide coating and investigate biological response of human mesenchymal stem cell to anodized titanium discs coated with RGD peptide by physical adhesion and chemical fixation. Materials and methods: Fluorescence isothiocyanate (FITC) modified RGD-peptide was coated on the anodized titanium discs (diameter 12 mm, height 3 mm) using two methods. One was physical adhesion method and the other was chemical fixation method. Physical adhesion was performed by dip and dry procedure, chemical fixation was performed by covalent bond via silanization. In this study, human mesenchymal stem cell was used for experiments. The experiments consisted of surface characteristic evaluation after peptide coating, analysis about cell adhesion, proliferation, differentiation, and mineralization. Obtained data are statistically treated using Kruskal-Wallis test and Bonferroni test was performed as post hoc test (P=.05). Results: The evaluation of FE-SEM images revealed no diffenrence at micro-surfaces between each groups. Total coating dose was higher at physical adhesion experimental group than at chemical fixation experimental group. In cell adhesion and proliferation, RGD peptide coating did not show a statistical significance compared with control group (P>.05). In cell differentiation and mineralization, physical adhesion method displayed significantly increased levels compared with control group and chemical fixation method (P<.05). Conclusion: RGD peptide coating seems to enhance osseointegration by effects on the response of human mesenchymal stem cell. Especially physical adhesion method showed more effective than chemical fixation method on response of human mesenchymal stem cell.

• Development and Reproduction
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• v.15 no.2
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• pp.87-98
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• 2011

The present experiment was performed to evaluate the osteogenic differentiation of human adipose tissue derived mesenchymal stem cells (ATMSCs) seeded in bioceramic-poly D,L-latic-co-glycolic acid (PLGA) scaffold. Osteogenic differentiation of ATMSCs were induced using the osteogenic induction (OI) medium. ATMSCs were cultured with OI medium during 28 days in well plate. The proliferation of ATMSCs in OI medium group was significantly increased for 14 days of plate culture but slowed after 21 days. On the other hand, proliferation in the control group showed constant increase for 28 days of culturing. The alkaline phosphatase (ALP) activity of ATMSCs in OI medium group increased during the 21 days of culture but decreased on 28 days. However, in control group ALP activity of ATMSCs was continuously decreased as time goes. Nodule was observed at 21 days of culture in OI medium group and confirmed accumulation of calcium in cell by alizarin red staining. ATMSCs were seeded in PLGA scaffold or in Bioceramic-PLGA scaffold, and cultured with OI medium. ALP activity of ATMSCs by osteoblast differentiation in each scaffold increased on 21 days of culture and decreased rapidly on 28 days. ALP activity of ATMSCs was increased highly in Bioceramic-PLGA scaffold compared to PLGA scaffold on 21 days of culturing. SEM-EDS analysis demonstrated that calcium and phosphate content and Ca/P ratio in Bioceramic-PLGA scaffold increased higher than in PLGA scaffold. Biodegradability of scaffold at 56 days after implantation showed that Bioceramic-PLGA scaffold was more biodegradable than PLGA scaffold. The results demonstrated that the differentiation of ATMSCs to osteoblast were more effective in scaffold culture than well plate culture. Bioceramic increased cell adhesion rate on scaffold and ALP activity by osteoblast differentiation. Also, bioceramic was considered to increase the calcium and phosphate in scaffold when ATMSCs was mineralized by osteogenic differentiation. Bioceramic-PLGA scaffold enhanced the osteogenesis of seeded ATMSCs compared to PLGA scaffold.