• Journal of Periodontal and Implant Science
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• v.23 no.1
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• pp.97-108
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• 1993

One of the important initial events required for periodontal regeneration is the attachment and subsequent spreading of periodontal ligament cells on the root surface. The purposes of this study is to investigate the attachment and spreading pattern of human periodontal ligament cell on the surface of glass slides. After establishment of a cell line of the primary cell culture from the periodontal ligament of 1st premolar teeth which were extracted for the purpose of orthodontic treatment, author dispersed the cells at $5{\times}10^3\;cells/ml$ into the each 35mm culture petri-dish containing 2 glass slides. To observe the morphological changes of the cells which attached to the surfaces of glasses at every designed time schedule, author used the inverted phase contrast microscope and scanning electron microscope. During the whole experiment culture condition was at $37^{\circ}C$, 100% Humidity, 5% $CO_2$ gas incubator. The following results were obtained. Periodontal ligament cells showed spherical outline and started to attach to glass surface by basal sytoplasmic extension after 10min in culture. After 30min in culture, periodontal ligament cells were attached to glass surface by well - developed filopodia which protruded from the lamellipodia. The cell surface is covered with bubble-like structures and occasional microvillus can be seen with diffculty among these structures. After 1.5hr in culture, peridontal ligament cells shhowed radially well-spread cytoplasm and the nucleus was centered on its cytoplasm. Unspread central region of the cell was covered with numerous microvilli. The change of cell attachment and spreading pattern was manifest at 6hr in culture. At this time, periodontal ligament cell showed elongated outline and an oval-shaped nucleus. After 12hr in culture, periodontal ligament cells showed more stretched fibroblast-like appearance with polarity. Two long lamellipodia can be seen around the both terminal ends of cells. After 24hr in culture, periodontal ligament cells showed spindle shapes and an oval-shaped nucleus was slanted toward one side of the cell.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.31 no.3
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• pp.199-218
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• 2005

Purpose of Study: Peripheral nerve regeneration depends on neurotrophism of distal nerve stump, recovery potential of neuron, supporting cell like Schwann cell and neurotrophic factors such as BDNF. Peripheral nerve regeneration can be enhanced by the conduit which connects the both sides of transected nerve. The conduit maintains the effects of neurotrophism and BDNF produced by Schwann cells which can be made by gene therapy. In this study, we tried to enhance the peripheral nerve regeneration by using calcium phosphate coated porous conduit and BDNF-Adenovirus infected Schwann cells in sciatic nerve of rats. Materials and Methods: Microporous filter which permits the tissue fluid essential for nerve regeneration and does not permit infiltration of fibroblasts, was made into 2mm diameter and 17mm length conduit. Then it was coated with calcium phosphate to improve the Schwann cell adhesion and survival. The coated filter was evaluated by SEM examination and MTT assay. For effective allogenic Schwann cell culture, dorsal root ganglia of 1-day old rat were extracted and treated with enzyme and antimitotic Ara-C. Human BDNF cDNA was obtained from cDNA library and amplified using PCR. BDNF gene was inserted into adenovirus shuttle vector pAACCMVpARS in which E1 was deleted. We infected the BDNF-Ad into 293 human mammary kidney cell-line and obtained the virus plaque 2 days later. RT-PCR was performed to evaluate the secretion of BDNF in infected Schwann cells. To determine the most optimal m.o.i of BDNF-Ad, we infected the Schwann cells with LacZ adenovirus in 1, 20, 50, 75, 100, 250 m.o.i for 2 hours and stained with ${\beta}$-galactosidase. Rats(n=24) weighing around 300g were used. Total 14mm sciatic nerve defect was made and connected with calcium phosphate coated conduits. Schwann cells$(1{\times}10^6)$ or BDNF-Ad infected Schwann cells$(1{\times}10^6)$ were injected in conduit and only media(MEM) was injected in control group. Twelve weeks after surgery, degree of nerve regeneration was evaluated with gait analysis, electrophysiologic measurements and histomorphometric analysis. Results: 1. Microporous Millipore filter was effective conduit which permitted the adhesion of Schwann cells and inhibited the adhesion of fibroblast. We could enhance the Schwann cell adhesion and survival by coating Millipore filter with calcium phosphate. 2. Schwann cell culture technique using repeated treatment of Ara-C and GDNF was established. The mean number of Schwann cells obtained 1 and 2 weeks after the culture were $1.54{\pm}4.0{\times}10^6$ and $9.66{\pm}9.6{\times}10^6$. 3. The mRNA of BDNF in BDNF-Ad infected Schwann cells was detected using RT-PCR. In Schwann cell $0.69\;{\mu}g/{\mu}l$ of DNA was detected and in BDNF-Adenovirus transfected Schwann cell $0.795\;{\mu}g/{\mu}l$ of DNA was detected. The most effective infection concentration was determined by LacZ Adenovirus and 75 m.o.i was found the most optimal. Conclusion: BDNF-Ad transfected Schwann cells successfully regenerated the 14mm nerve gap which was connected with calcium phosphate coated Millipore filter. The BDNF-Ad group showed better results compared with Schwann cells only group and control group in aspect to sciatic function index, electrophysiologic measurements and histomorphometric analysis.

• The Journal of Korean Academy of Prosthodontics
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• v.45 no.3
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• pp.382-388
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• 2007

Statement of problem. The role of calcium sulfate in stimulating the growth of gingival soft tissue has been reported in few studies. Such a unique property of calcium sulfate could serve as a trouble-solving broker in compensating for the lack of soft tissues in various oral surgeries. Purpose. The purpose of this study was to compare the proliferating activities of human gingival fibroblasts seeded on various bone graft barrier materials of calcium sulfate, collagen, and polytetrafluorethylene (PTFE). Material and methods. Two calcium sulfates ($CAPSET^{(R)}$. and $CalForma^{(R)}$, Lifecore Biomedical Inc., St. Paul, Minnesota, USA), a resorbable natural collagen ($Bio-Gide^{(R)}$, Geistlich Pharma Ag., Wolhusen, Switzerland), and a non-resorbable PTFE ($TefGen-FD^{(R)}$, Lifecore Biomedical Inc., St. Paul, Minnesota, USA) served as the human gingival fibroblasts' substrates and comprised the four experimental groups, whereas the untreated floors of culture plastics were used in the control group, in this study. Cells were trypsinized, seeded, and incubated for 48 h. The proliferating activities of fibroblasts were determined by XTT and SRB assay and absorbance (optical density, OD) was measured. One-way ANOVA was used to analyze the differences in the mean OD values between the groups of CAPSET, CalForma, Bio-Gide, TefGen, and the control (p<0.05). Results. From the XTT assay, the mean OD value of the control group, the highest, was significantly greater than that of any of the four experimental groups followed by CalForma, CAPSET, TefGen, and Bio-Gide. Further, the mean OD value of CalForma, was significantly greater compared to that of Bio-Gide. From the SRB assay, Calforma showed the highest mean OD value, which was significantly greater than that of any other groups, followed by the control, CAPSET, Bio-Gide, and TefGen. The mean OD values of both the control and CAPSET were significantly greater compared to that of TefGen (p<0.05). Conclusion. Assessment of the viability and proliferation of cultured fibroblasts seeded and incubated for 48 h on various barrier-material substrates using XTT and SRB assay showed that calcium sulfate $CalForma^{(R)}$ promotes the proliferating activity of human gingival fibroblasts.

• Journal of Food Hygiene and Safety
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• v.33 no.2
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• pp.140-145
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• 2018

The skin of the human body occupies the largest surface area of the body and acts as a protection for the person's internal organs. As such, the skin is a major target of oxidative stressors, and these oxidative stressors are known to contribute to skin aging over the course of time. For the most part, an antioxidant is an effective approach to utilize to prevent symptoms related to the reactive oxygen species (ROS)-induced aging of the skin. Therefore, we investigated the antioxidant and anti-aging activity of the leaves of the Quercusaliena Blume extract (QBE). In our study, we confirmed that the cell viability tested with XTT {2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide innersalt} assay was not affected up to a concentration of $100{\mu}g/mL$. In addition, the cell viability of HDF cells induced by hydrogen peroxide was recovered from 81% to 104% after treatment with QBE, which showed the greater protective effect than that of ascorbic acid. Treatments of QBE dose-dependently inhibited reactive oxygen species (ROS) production in HDF cells induced by hydrogen peroxide, which correlated with their protective effects on cell viability. Since QBE treatment exhibited the suppression effect of skin aging by decreasing the ROS production, QBE could be used as a not only natural anti-aging but also antioxidant resource.

• Development and Reproduction
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• v.12 no.3
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• pp.221-229
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• 2008

For the clinical application, it is needed to keep characteristics of stem cells after storage for a long time. In the present study, we examined stem cell properties of human cord-derived stem cells (HUC) after cryopreservation. Cells were isolated from human umbilical cord and cultured in vitro. At passage 2 or 3, HUC were suspended at a concentration of $1.0{\times}10^6/m{\ell}$ in cryomedium consisting of DMSO and FBS. After freezing at $-80^{\circ}C$ overnight, HUC were cryopreserved at $-196^{\circ}C$ nitrogen gas. After 6 months, HUC were thawed and cultured in vitro. Assessment for the stem cell properties was made upon the morphology, population doubling time, and expression profiles of genes and various proteins. Cryopreserved HUC showed more than 70% viability and maintained fibroblast-like morphology similar to HUC before cryopreservation. Throughout the culture, they underwent average 42.8 doublings and produced $6.75{\times}{10^{18}}$ cells. RT-PCR analyses showed that cryopreserved HUC expressed Oct-4, nanog, SCF, NCAM, nestin, GATA-4, BMP4, and HLA-1 genes. They did not express Brachyury and HLA-DR genes. Immunocytochemical studies showed that cryopreserved HUC reacted with antibodies against SSEA-3, -4, Thy-1, vimentin, fibronectin, HCAM, ICAM, HLA-1 proteins. They did not react with antibody against HLA-DR protein. Theses genes and proteins expression patterns of cryopresserved HUC were similar to those of HUC before cryopreservation. These results suggest that cryopreserved HUC could retain proliferative potential and they expressed various genes and proteins similar to HUC before cryopreservation. Thus, cryopreservation might be useful for HUC for future research and clinical application.

• Reproductive and Developmental Biology
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• v.35 no.1
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• pp.123-129
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• 2011

The pig has been considered to serve as an appropriate model of human disease. Therefore, establishment of porcine embryonic stem cell lines is important. The purpose of the present study was to further work in this direction. We produced porcine parthenogenetic embryos, and separately aggregated two of each of two-cell ($2{\times}2$), four-cell ($2{\times}4$), and eight-cell ($2{\times}8$) embryos derived by parthenogenesis. After culture for 4 days, the developmental ability of the aggregates and total blastocyst cell numbers were evaluated. The percentage of blastocysts was significantly higher in both $2{\times}4$- and $2{\times}8$-aggregated embryos ($58.3{\pm}1.9%$ and $37.2{\pm}2.8%$, respectively) than in the control or $2{\times}2$-aggregated embryos ($23.6{\pm}1.1%$ and $12.5{\pm}2.4%$, respectively). Total blastocyst cell numbers were increased in the $2{\times}4$- and $2{\times}8$-aggregated embryos (by $44{\pm}3.0%$ and $45{\pm}3.3%$, respectively) compared with those of control or $2{\times}2$-aggregated embryos ($30.5{\pm}2.1%$ and $30.7{\pm}2.6%$, respectively; p<0.05). The levels of mRNA encoding Oct-4 were higher in both the $2{\times}4$- and $2{\times}8$-aggregated embryos than in the control. When blastocysts derived from $2{\times}4$- aggregated embryos or intact normal embryos were cultured on mouse embryonic fibroblast feeder cells to obtain porcine stem cells, blastocysts from aggregated embryos formed colonies that were better in shape compared with those derived from intact blastocysts. Together, the data show that aggregation of porcine embryos not only improves blastocyst quality but also serves as an efficient procedure by which porcine embryonic stem cells can become established.

• Journal of Korean society of Dental Hygiene
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• v.16 no.3
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• pp.471-477
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• 2016

Objectives: Fluoride is widely used in the prevention and control of dental caries. The purpose of this study is to examine the biological effects of Sodium fluoride on the proliferation of oral normal cell in vitro(MDPC-23, HaCaT, HGF-1 cells). Methods: The proliferation of normal cells and the cyto-skeletal change of normal cells were assessed by WST-1 assay and F-actin stain assay. The statistical significances of the resulting data were analyzed using SPSS(Window 12.0). Results: The sodium fluoride(0-12 mM) treatment decreased the cell viability in a dose and time dependent manner: HaCaT(6 h): $100{\pm}0$, $98{\pm}0.39$, $82{\pm}2.68$, $75{\pm}0.83$, $69{\pm}1$, $67{\pm}1.42%$(p<0.005); HaCaT(24 h): $100{\pm}0$, $98{\pm}1.85$, $54{\pm}0.64$, $43{\pm}0.4$, $38{\pm}0.32$, $36{\pm}0.13%$(p<0.006), MDPC-23(6 h): $100{\pm}0$, $93{\pm}1.48$, $85{\pm}0.28$, $82{\pm}1.58$, $79{\pm}1.48$, $76{\pm}1.93%$(p<0.009); MDPC-23(24 h): $100{\pm}0$, $91{\pm}1.26$, $58{\pm}0.65$, $49{\pm}1$, $44{\pm}0.74$, $2{\pm}0.05%$(p<0.005), HGF-1(6 h): $100{\pm}0$, $97{\pm}2.93$, $89{\pm}5$, $71{\pm}5.42$, $58{\pm}4.82$, $43{\pm}3.47%$(p<0.009); HGF-1(24 h): $100{\pm}0$, $97{\pm}2.05$, $73{\pm}1.73$, $22{\pm}1.61$, $14{\pm}1.73$, $7{\pm}0.85%$(p<0.005). Thus, changes in cell morphology and disruption of filamentous(F)-actin organization were observed in higher concentration. Conclusions: These results suggest that higher concentrations of fluoride lead to a reduce the number of cells and morphology change of normal cell.

• Korean Journal of Medicinal Crop Science
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• v.11 no.1
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• pp.31-39
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• 2003

The antioxidative activities and biological properties in the EtOAc extracts and purified extracts of Orostachys japonicus were measured by assay of DPPH, xanthine/xanthine oxidase and mammalian cells(2-12). Scavenging of DPPH radical and inhibition of xanthine/xanthine oxidase of EtOAc extracts were showed the highest activity in the arable land and in September. The S-4 fraction showed the highest activity among the silica-gel column chromatography fractions. LH-4 fraction showed higher activity than the other fractionsins in assay of DPPH and xanthine/xanthine oxidase. Fatty acids and phenolic compounds were identified by GC/MS and main compounds were 1,2,3-benzenetriol, alpha-androsta-7,14-diene in LH-4 fraction. The activities of POD and SOD in samples havested on different habitats were high such as arable land> intermountain> seashore. That of POD and SOD in crude extracts of late stage were higher than early stage. Isozyme bands of crude extracts samplinged in all habitats and all growing stages showed two bands and the signal intensity showed strongly according to passage of growing stage. The purified extracts of LH-4 fraction showed excellent inhibition effect in proliferation of HL-60 cells and markedly suppressed colony formation in mouse fibroblast cells. Dose response between partially purified extracts(400ppm) and negative control did not produced statistically significant reduction in colony formation.

• Journal of Life Science
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• v.31 no.4
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• pp.389-399
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• 2021

This study determined the effects of Centella asiatica leaf on H₂O₂ induced cell cycle arrest, mitochondrial activity, and proinflammatory cytokine production in human dermal fibroblast (HDF) cells. We used an 80% methanol extract of C. asiatica, its ethyl acetate fraction, and asiaticoside, the major constituent of C. asiatica. The C. asiatica extract, its ethyl acetate fraction, and asiaticoside attenuated G1 cell cycle-arrest and the apoptotic effect caused by H₂O₂-induced oxidative stress. The cells treated with C. asiatica extract, its ethyl acetate fraction, and asiaticoside secreted lower levels of TNF-α and IL-6. The antioxidant effect of asiaticoside was higher than that of C. asiatica extract and its ethyl acetate fraction. Treatment with C. asiatica extract, its ethyl acetate fraction, and asiaticoside also increased the mitochondrial membrane potential and restored normal mitochondrial morphology. Following H₂O₂ stress induction, cells treated with C. asiatica extract, its ethyl acetate fraction, and asiaticoside showed increased mitochondrial oxygen consumption rates and decreases in the TLR4-MyD88-TRAF6-p65 pathway activity. These findings suggest that C. asiatica extract, its ethyl acetate fraction, and asiaticoside have antioxidant and anti-inflammatory effects, as well as the ability to control the mitochondrial activities of HDF cells.

• The Journal of Korean Obstetrics and Gynecology
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• v.22 no.3
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• pp.51-65
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• 2009

Purpose: The purpose of this study is to investigate the anti-inflammatory effects of three types of Yongdamsagantanggamibang(YSTG) which has been medicated the patient with inflammatory disease of female genitourinary system. Methods: To verify the anti-inflammatory mechanism of YSTGs, expressions of IL-1${\beta}$, IL-6, MCP-1, COX-2 and TNF-${\alpha}$ mRNA in THP-1 cells were examined. And we investigated the production levels of IL-1${\beta}$, IL-6 and TNF-${\alpha}$ in mouse following LPS co-treatment. Results: 1. YSTG1, YSTG2 and YSTG3 extract did not show any cytotoxic effect on human fibroblast cells at any of the concentrations evaluated(500, 250, 125, 62.5, 37.25 ${\mu}g/m{\ell}$) 2. YSTG1, YSTG2 and YSTG3 extract showed scavenging activity on DPPH free radical and SOD-like activity. 3. YSTG1, YSTG2 and YSTG3 extract decreased production levels of IL-1${\beta}$, IL-6, IL-8, TNF-${\alpha}$ and MCP-1 in LPS-treated THP-1 cells. 4. YSTG1, YSTG2 and YSTG3 extract decreased expressions of IL-1${\beta}$, IL-6, MCP-1, COX-2 and TNF-${\alpha}$ mRNA in LPS-treated THP-1 cells. 5. YSTG1, YSTG2 and YSTG3 extract decreased production levels of IL-1${\beta}$, IL-6 and TNF-${\alpha}$ in serum of LPS-treated mouse. Conclusion: Based on results above, it is revealed three types of YSTG have the anti-inflammatory effect, and may be effective in the treatment for inflammatory disease of female genitourinary system.