• Imaging Science in Dentistry
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• v.38 no.3
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• pp.125-132
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• 2008

Purpose : To investigate the effects of irradiation on transforming growth factor ${\beta}_1$ (TGF-${\beta}_1$) mRNA expression and calcific nodule formation in MC3T3-E1 osteoblastic cell line. Materials and Methods : Cells were cultured in alpha-minimum essential medium ($\alpha$-MEM) supplemented with 10% fetal bovine serum and antibiotics. When the cells reached the level of 70-80% confluence, culture media were changed with $\alpha$-MEM supplemented with 10% FBS, 5 mM $\beta$-glycerol phosphate, and $50\;{\mu}g/mL$ ascorbic acid. Thereafter the cells were irradiated with a single dose of 2, 4, 6, 8 Gy at a dose rate of 1.5 Gy/min. The expression pattern of TGF-${\beta}_1$ mRNA, calcium content and calcific nodule formation were examined on day 3, 7, 14, 21, 28, respectively, after the irradiation. Results : The amount of TGF-${\beta}_1$ mRNA expression decreased significantly on day 7 after irradiation of 4, 6, 8 Gy. It also decreased on day 14 after irradiation of 6, 8 Gy. and decreased on day 21 after irradiation of 8 Gy. The amount of calcium deposition decreased significantly on day 7 after irradiation of 4, 8 Gy (P < 0.01) and showed a decreased tendency on day 14, 21 after irradiation of 4, 6, 8 Gy. The number of calcific nodules was decreased on day 7 after irradiation of 4, 8 Gy. Conclusion: Irradiation with a single dose of 4, 6, 8 Gy influences negatively the bone formation at the molecular level by affecting the TGF-${\beta}_1$ mRNA expression that was associated with proliferation and the production of extracellular matrix in MC3T3-E1 osteoblastic cell line.

• Imaging Science in Dentistry
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• v.35 no.1
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• pp.1-8
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• 2005

Purpose : To investigate the effects of irradiation on the calcium content and calcific nodule formation in the MC3T3-E1 osteoblastic cell line. Materials and Methods : Cells were irradiated with a single dose of 2,4 and 8 Gy at a dose rate of 5.38 Gy/min using a Cs-137 irradiator. After irradiation, the calcium content and calcific nodule formation were examined on the 1 st, 2nd, 3rd and 4th week. Results : A decreasing dose-dependent tendency of the cell proliferation rate was found in all irradiated groups of this experiment when compared with the unirradiated control group. In accordance with the duration of culture, there was no significant difference in the cell proliferation rate after irradiation of 2 Gy when compared with the unirradiated group, however a decreasing tendency was found in 4 Gy- and 8 Gy-irradiated groups. While an increase in total calcium content after irradiation of 2 Gy was found at week 1, week 2, and week 4, there was a decrease in calcium content at week 1 through 4 in the 8 Gy- irradiated group. Calcific nodule formation was increased in irradiated experimental groups when compared with the unirradiated control group in the 2 Gy-irradiated group, but decreased in the 4 Gy- and 8 Gy-irradiated groups at the same stage. Conclusion : The results showed a mild increasing tendency of the calcific nodule formation after irradiation of 2 Gy. However, a decreased calcific nodule formation in 4 Gy- and 8 Gy-irradiated groups was found. Taken together, the irradiation of 2 Gy mildly activated bone formation, however 4 Gy or 8 Gy suppressed bone formation by decreasing cell numbers in the MC3T3-El osteoblastic cell line.

• International Journal of Oral Biology
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• v.33 no.3
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• pp.105-111
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• 2008

Thrombospondins (TSP-1, TSP-2) are secretory extracellular glycoproteins that are involved in a variety of physiological processes such as tumor cell adhesion, invasion, and metastasis. The present study was undertaken to elucidate the involvement of thrombospondins in the adhesion of osteoblast-like cells using the TSP-1 or TSP-2 antisense MG63 and MC3T3-E1 cell lines. For downregulation of TSPs expression, we prepared antisense constructs for TSP-1 and TSP-2 using the pREP4 an episomal mammalian expression vector, which be able to produce the specific antisense oligonucleotides around chromosome. MG63 and MC3T3-E1 osteoblast-like cells were transfected with the antisense constructs and nonliposomal Fugene 6, and then selected under hygromycin B (50 ${\mu}g/ml$) treatment for 2 weeks. Western blot analysis revealed that expression of the TSP proteins was downregulated in the antisense cell lines. The cell adhesion assay showed that adhesive properties of TSP-1 and TSP-2 antisense MG63 cells on the polystyrene culture plate were reduced to 17% and 21% of the control cells, respectively, and those of the TSP-1 and TSP-2 antisense MC3T3-E1 cells also decreased to 19% and 27% of control, respectively. Adhesion of TSP-1 and TSP-2 antisense MC3T3-E1 cells on Type I collagen-coated culture plate decreased to 27% and 76%, respectively. These results indicate that TSP-1 and TSP-2 proteins may have an important role in adhesion of osteoblast-like cells to extracellular matrix.

• Journal of Periodontal and Implant Science
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• v.37 no.1
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• pp.115-124
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• 2007

Silica is known as a promising osteoconductive material, and polycaprolactone is a bioactive and degradable material. The purpose of this study was to monitor the differentiation of MC3T3-E1 cells cultured on the layer-built silica/poly caprolactone non-woven fabric produced by electrospinning. Non-woven fabric (silica, polycaprolactone, PSP, SPS) was made by electrospinning and they were inserted in the 48 well cell culture plate. MC3T3-E1 cells were prepared by subculture. Cells were seeded to each well $1{\times}10^5$ concentration per well. Dulbecco's modified eagle medium with 10% FBS and 1% antibiotic-antimycotic solution was used. Confocal laser scanning microscope was taken 4 hours after incubation (95% air. 5% $CO_2$, $37^{\circ}C$). Cell proliferation was monitored by spectrophotometer on 1, 7, 14 days, and the morphology of the growing cells was observed by field emission scanning electron microscope. To monitor the differentiation of osteoblasts on the materials, MC3T3-E1 cells were incubated in 48 well culture plate after seeding with the density of $1{\times}10^5$ concentration. Then ELISA kit & EIA kit were used on to assess osteocalcin and osteopontin expression respectively. The other conditions were the same as above. MC3T3-E1 cells were proliferated well on all of the materials. There were no statistical differences among them. The osteopontin expression of silica, PSP, SPS was significantly higher than other groups on day 3 (p/0,05), but after that time, there were no statistically signigicant differences. The osteocalcin expression was significantly higher in silica and PSP than other groups on day 14. These findings show that PSP was as good as silica on the effect of osteoblast differentiation. The PSP non-woven fabric may have the possibility as bone graft materials.

• Natural Product Sciences
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• v.11 no.2
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• pp.103-108
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• 2005

Bioflavone quercetin is thought to have an important role to inhibit bone loss by affecting osteoclastogenesis and regulating a number of systemic and local factors such as hormones and cytokines. In this study, we examined how quercetin acts on cytokine production and mineralization of osteoblast in the presence of tumor necrosis factor-alpha $(TNF-{\alpha})$ which has been known to play a pivotal role in bone metabolic diseases. Quercetin inhibited $TNF-{\alpha}-induced$ secretion of $IFN-{\gamma}$ and IL-6 in differentiated MC3T3-E1 cells. As indicated by the markers that are characteristics of the osteoblast phenotype, such as alkaline phosphatase (ALP) activity and calcium deposition, quercetin treatment slightly prevented the $TNF-{\alpha}-induced$ dramatic inhibition of differentiation and mineralization of MC3T3-E1 cells. Further, quercetin inhibited the production of nitric oxide induced by $TNF-{\alpha}$ in the cells. Collectively, our findings indicate that quercetin inhibites $TNF-{\alpha}-induced$ secretion of inflammatory cytokines in differentiated MC3T3-E1 cells without any cytotoxic effects.

• Korean Journal of Food Science and Technology
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• v.44 no.1
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• pp.82-88
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• 2012

Chrysanthemum indicum L. (Asteraceae) is a traditional herbal medicine that has been used for the treatment of inflammation, hypertension, and respiratory diseases due to its strong antagonistic activity against inflammatory cytokines. The effects of Chrysanthemum indicum L. Extract (CIE) for increasing cell growth, alkaline phosphatase (ALP) activity, and collagen content were totally inhibited, suggesting that the effect of CIE might be partly involved with estrogen activity. Furthermore, the protective effects of CIE on the response of osteoblasts to oxidative stress were evaluated. Osteoblastic MC3T3-E1 cells were incubated with hydrogen peroxide and/or CIE, and markers of osteoblast function and oxidative damage were examined. CIE significantly increased cell survival, ALP activity, and calcium deposition, and decreased the production of Reactive Oxygen Species (ROS) and Tumor Necrosis Factor-${\alpha}$ (TNF-${\alpha}$) in osteoblasts. Taken together, these results indicate that the enhancement of osteoblast function by CIE may prevent osteoporosis and inflammatory bone diseases.

• Korean Journal of Plant Resources
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• v.33 no.4
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• pp.227-236
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• 2020

Panax ginseng C.A. Meyer (P. ginseng) is known to exert a wide range of pharmacological effects both in vitro and in vivo. Although studies on ginsenoside, antioxidant activity, and anticancer effect of the cultivated wild Panax ginseng (CWP) have been conducted, there is little research on the effect of CWP extract on bone metabolism. In this study, we investigated the potential anti-osteoporotic properties of CWP on the growth and differentiation of MC3T3-E1 cells. CWP significantly increased the viability and proliferation of MC3T3-E1 cells. CWP activated intracellular alkaline phosphatase (ALP) activity in MC3T3-E1 cells. In addition, CWP increased the mineralized nodules in MC3T3-E1 cells. Furthermore, CWP increased the expression of genes such as Runx2, ALP, OPN and OCN associated with osteoblast growth and differentiation in a dose-dependent manner.

• The korean journal of orthodontics
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• v.27 no.4 s.63
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• pp.623-632
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• 1997

The purpose of this study was to evaluate the effects of magnetic field on cellular activity of MC3T3-E1 cells. The cellular activity was monitored by alkaline phosphatase and DNA synthetic activity in control, static magnetic field and pulsed electromagnetic field groups. A static magnetic field was applied to the cell by placing one, two, three, foue, and five samarium-cobalt magnets above and below each cell plate for 24hours per day. A pulsed electromagnetic field with a frequency of 100 herz was applied for 10 hours per day. After 10 days of magnetic field exposure, there were increase of alkaline phosphatase activity in static magnetic field groups consisted of one, two and three magnetic groups. Alkaline phosphatase activities were not significantly increased in four and five magnetic groups. Application of pulsed electromagnetic field did not result in significant increase in alkaline phosphatase activity compared to control. DNA synthetic activity in both static and pulsed electromagnetic field group were not significantly different from that in control group. The result of this study suggest that magnetic field could have effect on the metabolism of bone cells related to the cellular metabolic process.

• Molecules and Cells
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• v.40 no.6
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• pp.393-400
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• 2017

Type 2 diabetes mellitus (T2DM) is a chronic metabolic disease characterized by lack of insulin and high glucose levels. T2DM can cause bone loss and fracture, thus leading to diabetic osteoporosis. Promoting osteogenic differentiation of osteoblasts may effectively treat diabetic osteoporosis. We previously reported that Sirtuin 1 (Sirt1), a $NAD^+$-dependent deacetylase, promotes osteogenic differentiation through downregulation of peroxisome proliferator-activated receptor (PPAR) ${\gamma}$. We also found that miR-132 regulates osteogenic differentiation by downregulating Sirt1 in a $PPAR{\beta}/{\delta}$-dependent manner. The ligand-activated transcription factor, $PPAR{\alpha}$, is another isotype of the peroxisome proliferator-activated receptor family that helps maintain bone homeostasis and promot bone formation. Whether the regulatory role of $PPAR{\alpha}$ in osteogenic differentiation is mediated via Sirt1 remains unclear. In the present study, we aimed to determine this role and the underlying mechanism by using high glucose (HG) and free fatty acids (FFA) to mimic T2DM in MC3T3-E1 cells. The results showed that HG-FFA significantly inhibited expression of $PPAR{\alpha}$, Sirt1 and osteogenic differentiation, but these effects were markedly reversed by $PPAR{\alpha}$ overexpression. Moreover, siSirt1 attenuated the positive effects of $PPAR{\alpha}$ on osteogenic differentiation, suggesting that $PPAR{\alpha}$ promotes osteogenic differentiation in a Sirt1-dependent manner. Luciferase activity assay confirmed interactions between $PPAR{\alpha}$ and Sirt1. These findings indicate that $PPAR{\alpha}$ promotes osteogenic differentiation via the Sirt1-dependent signaling pathway.

• The korean journal of orthodontics
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• v.26 no.3
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• pp.291-299
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• 1996

Orthodontic force is a mechanical stress controlling both of tooth movement and skeletal growth. The mechanical stress stimulate bone cells that may exert some influence on bone remodeling. The purpose of this study was to evaluate the difference in cellular activity depending on mechanical stresses such as compressive and tensile force by determining the alkaline phosphatase(ALP) activity. A clonal osteogenic cell line MC3T3-E1 was seeded into a 24-well plate($2{\times}10^4/well$). At the confluent phase, a continuous compressive hydrostatic pressure($25g/cm^2$, $300g/cm^2$) and continuous tensile hydrostatic pressure($-25g/cm^2$, $-300g/cm^2$) were applied for 4, 6, 10, 14, 18, 20 days respectively by a diaphgragm pump. At the end of the stimulation period, cell layers were prepared for ALP activity assay. The ALP activity of the compressive group increased more than that of the tensile group at same force magnitude, whereas the cells responded to a similar pattern regardless of the type of mechanical stress The ALP activity of the compressive and tensile group turned into the level of the control group as the length of time increased. These results indicated that a mechanical stress may be more effective on cellular activity during active cellular proliferation and differentiation periods. The time to achieve maximum ALP activity was delayed as the mechanical stress increased in both the compressive and the tensile group. Accordingly, the magnitude of the stress rather than the type of mechanical stress may have more influence on cellular activity.