• Journal of Periodontal and Implant Science
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• v.31 no.4
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• pp.833-846
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• 2001

Recently, use of natural medicine is getting more attention, and some of them are believed to be effective in　the treatment of periodontitis. Among them, the seeds of safflower(Carthamus tinctrorius L.) have been proven to be effective through its use in bone diseases such as fracture and osteoporosis. During the last few years, studies using the seeds of safflower gown in Korea have been active, and it has been reported that safflower seed extract increase the proliferation and the alkaline phosphatase(ALP) activity of human periodontal ligament fibroblast(hPDLF), osteoblast, and that they promote the mineralization process. In animal studies, when safflower seed extract were administered orally new bone formation was promoted. Recently, in an effort to find out the most effective osteogenic components, among many components of the safflower seed, various safflower seed fraction extracts were obtained by multistep extraction of the safflower components using various solvents. Among these, saf-M-W fraction extracted by methanol and water was most effective in increasing osteogenic potential of osteoblasts. In this study, the effect of safflower seed fraction extract, saf-M-W, on the growth and differentiation of hPDLF and MC3T3-E1 cell was investigated. The toxicity of saf-M-W on both cells was measured using M'IT(3-(4,5dimethylthiazol-2-y1)-2,5-diphenyl tetrazolium bromide) test, and ALP activity was measured using the colorimetric assay of hPDLF. In addition, in MC3T3-El cells, the expression of ALP, bone sialoprotein(BSP) mRNA was observed using Northern blot, and the mineralized nodule formation Was observed using von Kossa stain and phase-contrast microscope. 1. In concentrations below $10{\mu}g/ml$, saf-M-W didn't show any toxicity on hPDLF and MC3T3-El cell. 2. The change in saf-M-W concentration had no effect on the ALP activity of hPDLF. 3. In MC3T-E1 cells, mRNA expressions of ALP and BSP were greater in the experimental group treated with $10{\mu}g/ml$ concentration of saf-M-W compared with the control group. 4. In MC3T3-El cells, abundance of mineralized nodules were formed in the experimental group treated with $10{\mu}g/ml$ Concentration of saf-M-W, while no mineralized nodule was formed in the control group. These results suggest that safflower seed fraction extract, saf-M-W. didn't show any toxicity on hPDLF and MC3T3-E1 cell at concentrations below $10{\mu}g/ml$ and effectively enhanced the differentiation and osteogenic potential of MC3T3-El cell.

• Journal of Periodontal and Implant Science
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• v.30 no.4
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• pp.835-852
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• 2000

The ultimate goal of periodontal therapy is the regeneration of periodontal tissue and repair of function. For more than a decade there have been many efforts to develop materials and methods of treatment to promote periodontal wound healing. Recently many efforts are concentrated on the regeneration potential of material used in oriental medicine. In some in vitro and in vivo experiments, there have been many evidences that these materials have an effect on bone regeneration. The purpose of this study was to evaluate histologically and radiologically in Sprague-Dawley rats the effects of safflower seed extracts on the regeneration of the calvarial defects surgically produced. So in this study, the critical size defects were surgically produced in the calvarial bone of 30 Sprague-Dawley rats using the 8mm trephine bur. The safflower seed extract was applied into the defect of each rat in experimental group, whereas nothing was applied into the defect of each rat in control group. Rats were sacrificed at 2, 4, 8 weeks following operation and histomorphometric and radiodensitometric analysis were performed. 1. The newly formed bone length was $102.91{\pm}22.05$, $178.29{\pm}24.40$ at 2 week in the each control, experimental group, $130.95{\pm}39.24$, $242.62{\pm}50.33$ at 4 week and $181.53{\pm}76.35$, $240.36{\pm}22.00$ at 8 week($unit,{\mu}m$). In the 2, 4 week, there were statistically significant difference between control and experimental group(P<0.05). 2. The newly formed bone area was $2962.06{\pm}1284.48$, $10648.35{\pm}1284.48$ at 2 week, $5103.25{\pm}1375.88$, $9706.78{\pm}1481.81$ at 4 week, $8046.02{\pm}818.99$, $12057.06{\pm}740.47$ at 8 week($unit,{\mu}m^2$). In every week, there were statistically significant difference between control and experimental group(P<0.05). 3. The radiopacity was $14.26{\pm}.33$, $25.47{\pm}4.33$ at 2 week, $20.06{\pm}9.07$, $26.61{\pm}2.78$ at 4 week, $22.99{\pm}3.76$, $27.29{\pm}1.54$ at 8 week(unit, %). In the 2 week, there was statistically significant difference between control and experimental group(P<0.05). In conclusion, the results of the present study suggest that safflower seed extract initially has an effect on the newly formed bone area, length and radiopacity when it is applied to the calvarial defect of Sprague - Dawley rat. Then. the material has an effect on newly formed bone area and length.

• Journal of Periodontal and Implant Science
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• v.35 no.4
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• pp.1019-1037
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• 2005

The major goals of periodontal therapy are the functional regeneration of periodontal supporting structures already destructed by periodontal disease as well as the reduction of signs and symptoms of progressive periodontal disease. There have been many efforts to develop materials and therapeutic methods to promote periodontal wound healing. There have been increasing interest on the chitosan made by chtin. Chitosan is a derivative of chitin made by deacetylation of side chains. Chitosan has been widely studied as bone substitution and membrane material in periodontology. Many experiments using chitosan in various animal models have proven its beneficial effects. Tetracycline has been considered for use in the treatment of chronic periodontal disease and gingivitis. The aim of this study is to evlauate the osteogenesis of tetracycline blended chitosan membranes on the calvarial critical size defect in Sprague Dawley rats. An 8mm surgical defect was produced with a trephine bur in the area of the midsagittal suture. The rats were divided into five groups: Untreated control group versus four experimental group. Four types of membranes were made and comparative study was been done. Two types of non-woven membranes were made by immersing non-woven chitosan into either the tetracycline solution or chitosan-tetracycline solution. Other two types of sponge membranes were fabricated by immersing chitosan sponge into the tetracycline solution, and subsequent freeze-drying. The animals were sacrificed at 2 and 8 weeks after surgical procedure. The specimens were examined by histologic analyses. The results are as follows: 1. Clinically the use of tetracycline blended chitosan membrane showed great healing capacity. 2. The new bone formations of all the experimental group, non-woven and sponge type membranes were greater than those of control group. But, there was no significant difference between the experimental groups. 3. Resorption of chitosan membranes were not shown in any groups at 2 weeks and 8 weeks. These results suggest that the use of tetracycline blended chitosan membrane on the calvarial defects in rats has significant effect on the regeneration of bone tissue in itself. And it implicate that tetracycline blended chitosan membrane might be useful for guided tissue regeneration.

• Journal of Periodontal and Implant Science
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• v.26 no.2
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• pp.469-490
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• 1996

The initial events required for periodontal regeneration is the attachment, spreading, and proliferation of appropriated cells at the healing sites. These have been reported that minocycline stimulates the attachment of periodontal ligament cells, and also $TGF-{\beta}1$ enhances the proliferation of periodontal ligament cells. The purpose of the present study was to evaluate the effects of $TGF-{\beta}1$ on the cellular activity of minocycline treated human periodontal ligament cells. Periodontal ligament cells were obtained from the explants of healthy periodontal ligaments of extracted 3rd molars or premolar teeth extracted from the patients for orthodontic treatment. The cells were cultured in minimal essential medium(${\alpha}-MEM$) supplemented with 10.000units/ml penicillin, $10,000{\mu}g/ml$ streptomycin and 10% FBS(fetal bovine serum) at $37^{\circ}C$ in a humidified atmosphere of 5% carbon dioxide and the 5th to the 8th passages of the cells were used. To evaluate the effect of minocycline on cell attachment, the cells were seeded at a cell density of $1.5{\times}10^4$ cells/well in 24-well culture plates and treated with $20{\mu}g/ml$ and $100{\mu}g/ml$ of minocycline for 1.5 h. After trypsinization, the cells were counted with hemocytometer and were taken photographs for observation of cellular morphology. To evaluate the effect of $TGF-{\beta}1$ on minocycline-pretreated periodontal ligament cells, the cells were seeded at a cell density of $1{\times}10^4$ cells/ well in 24-well culture plates and treated with $20{\mu}g/ml$ and $100{\mu}g/ml$ of minocycline for 1.5 h. After incubation, 1 and 10ng/ml of $rh-TGF-{\beta}1$ were also added to the each well and incubated for 1 and 2 days, respectively. Then, MTT assay, DNA synthesis($^3H-thymidine\;assay$), and protein and collagen assay(3H-proline assay) were carried out. In the MIT assay, after 200ul MTT solutionlconeentration of 5mg/ml) were added to the each well of the 24-well plates and incubated for 3 hours, and 200 ul DMSO were added so as to dissolve insoluble blue formazan crystals which was formed in incubated period. Then it read plates on a ELISA reader. For mitogenic assay, 1 uCi/ml $^3H-thymidine$ was added to each well for the final 2 hours of the incubation periods. After labeling, the wells were washed 3 times with ice cold PBS and 4 times with 5% TCA to remove unincorporated label and precipitate the cellular DNA. DNA, with the incorporated $^3H-thymidine$, was solubilized with 500 ul of 0.1% NaOH/0.1% SDS. A 250 ul aliquot was removed from each well and placed in a scintillation vial with 4ml of scintillation cocktail. Using an liguid scintillation counter, counts per minute(CPM) were determined for each samples. 3 uCi/ml $^3H-proline$ was added to each well for the final 4 hours of the incubation periods and total protein and percent collagen synthesis were carried out. The results indicate that minocycline treated group with $100{\mu}g/ml$ concentration for 1.5 hours significantly increased than that of control in cell attachment, and cell process is also evident compared with that of control in cell morphology, and the cellular activity and DNA synthesis rate of cells treated minocycline and $TGF-{\beta}1$ significantly increased than that of control values, but were below to values of the $TGF-{\beta}1$ only treated group in MIT assay and $^3H-thymidine\;assay$, and the total protein synthesis of minocycline and $TGF-{\beta}1$ treated group also significantly increased than that of control values, but the percent collagen synthesis of tested group significantly decreased to compared with control. On the above the findings, the tested group of minocycline and $TGF-{\beta}1$ did not increase the effect on the cell activity than $TGF-{\beta}1$ only tested group and the tested group of minocycline inhibited cell activity. This results indicate that minocycline was effective on cell attachment in early stage, but it is harmful to cell activity, that inhibitory effect of minocycline was compensated with stimulatory effect of $TGF-{\beta}1$.

• Journal of Periodontal and Implant Science
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• v.38 no.2
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• pp.237-246
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• 2008

Purpose: If bone grafts and guided tissue regeneration are effective individually in treating osseous defects, then the questionis, what would happen when they are combined. Bone grafts using Calcium Carbonate(Biocoral) and Guided Tissue Regeneration using Calcium Sulfate(CALMATRIX) will maximize their advantages and show the best clinical results in intrabony defects. This study was to compare the effects of a combination of CS and CC with control treated only with modified widman flap in a periodontal repair of intrabony defects. Materials and Methods: 30 patients with chronic periodontitis were used in this study. 10 patients were treated with a combination of CS and CC as the experimental group II and another 10 patients were treated with CC as the experimental group I, and the remaining 10 patients, the control group were treated only with modified widman flap. Clinical parameters including probing depth, gingival recession, bone probing depth and loss of attachment were recorded 6 months later. Results: The probing depth changes were $3.30{\pm}1.34\;mm$ in the control group, $4.2{\pm}1.55\;mm$ in the experimental group I(CC) and $5.00{\pm}1.33\;mm$ in the experimental group II(CS+CC). They all showed a significant decrease 6 months after surgery(p<0.01). There was a significant difference(p<0.05) between the control and experimental group. However there were no significant difference(p<0.05) between the experimental group I and II. The gingival recession changes w $-1.30{\pm}1.25\;mm$ in the control group, This is a significant difference(p<0.01). However, there was a $-0.50{\pm}0.53\;mm$ change in the experimental group I(CC) and $-0.60{\pm}0.97\;mm$ in the experimental group II(CS+CC). In addition, in terms of gingival recession, there was a no significance difference(p<0.05) among the groups. The clinical attachment level changes were $2.00{\pm}1.33\;mm$ in the control group, $3.60{\pm}1.58\;mm$ in the experimental group I(CC) and $4.40{\pm}1.17\;mm$ in the experimental group II(CS+CC). They all showed a significant decrease 6 months after surgery(p<0.01). There was a significant difference(p<0.05) between the control and experimental group. However there was a no significance difference(p<0.05) between the experimental group I and II. The bone probing depth changes were $0.60{\pm}0.52\;mm$ in the control group, $3.20{\pm}1.48\;mm$ in the experimental group I(CC) and $4.60{\pm}1.43\;mm$ in the experimental group II(CS+CC). All of them showed a significant decrease 6 months after surgery(p<0.01), there was a significance difference(p<0.05) among the groups. Conclusion: Treatment using a combination of CS and CC have a potential to improve periodontal parameters in intrabony defects and More efficient clinical results can be expected in intrabony defects less than 2 walls grafted with CS and CC.

• Journal of Periodontal and Implant Science
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• v.27 no.3
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• pp.603-619
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• 1997

The use of laser in the treatment of soft tissue minimizes hemorrhage, provides better view of the operating field, and thereby minimizes operating time. Also, there will be far less post-operative swelling, pain and scar formation, and sterilizing effect are shown in some portions of the wound site. All these advantages of laser therapy contribute to its widespread use in the field of medicine and dentistry. Regarding such facts, we used CO2 laser of different watts in gingivectomy for white rats to compare initial healing process. For the control group, the least amount of output in performing gingivectomy(4watts) was offered, and for the experimental group, 6watts was given. Animals were sacrificed on the second, third days, 1 weeks, 2 weeks, and 3 weeks after operation, and their specimens were histologically analyzed. The following results were obtained: 1. Blood clot of small size was observed in both the control and experimental groups after two days, and no more thereafter. 2. In both the control and experimental groups, the inflammation zone size was the greatest after two days, and it decreased gradually to become almost invisble by the second week. The experimental group showed larger size of inflammation zone during second and third days: however, there was no difference after one week. 3. Granulation tissue in both the control and experimental groups showed gradual maturation with time, and by the second week, it was almost replaced by normal connective tissue. By the third week, complete healing pattern was observed. The experimental group showed larger granulation tissue than the control group until the third day, but there was no significant difference after one week. 4. In both the control and experimental groups, gingival epithelialization began on the second day. After one week, regeneration of rete peg and partial formation of junctional epithelium were observed: by the second week, keratinization of oral sulcular epithelium began, and it was completed by the third week.

• Development and Reproduction
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• v.13 no.1
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• pp.1-11
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• 2009

Implantation itself is governed by an array of endocrine, paracrine and autocrine modulators, of embryonic and maternal origin. Window of implantation is the unique temporal and spatial expression of factors allows the embryo to implant via signaling, appositioning, attachment, and invasion in a specific time frame of $2{\sim}4$ days. When the embryo has arrived in the uterine cavity, a preprogrammed sequence of events occurs, which involves the production and secretion of a multitude of biochemical factors such as cytokines, growth factors, and adhesion molecules by the endometrium and the embryo, thus leading to the formation of a receptive endometrium. Cytokines such as LIF, CSF-1, and IL-1 have all been shown to play important roles in the cascade of events that leads to implantation. Integrin, L-selectin ligands, glycodelin, mucin-1, HB-EGF and pinopodes are involved in appositioning and attachment. The embryo also produces cytokines and growth factors (ILs, VEGF) and receptors for endometrial signals such as LIF, CSF-1, IGF and HB-EGF. The immune system and angiogenesis play an important role. The usefulness of these factors to assess endometrial receptivity and to estimate the prognosis for pregnancy in natural and artificial cycles remains to be proven. Integrins, pinopodes, glycodelin and LIF (from biopsies) are promising candidates; from uterine flushings, glycodelin and LIF are also candidates. The ideal serum marker is not available, but VEGF, glycodelin and CSF have some clinical implications. Further evaluation that includes larger groups of infertile women and fertile controls are needed to elucidate whether their presence in plasma, flushing fluid, or endometrial samples can be used as some kind of a screening tool to assess endometrial function and prognosis for pregnancy before and after artificial reproductive therapy. A better understanding of their function in human implantation may lead to therapeutic intervention, thereby improving the success rate in reproduction treatment. New molecular techniques are becoming available for measuring both embryonic and endometrial changes prior to and during implantation. The use of predictive sets of markers may prove to be more reliable than a single marker. Ultimately, the aim is to use these tools to increase implantation in artificial cycles and consequently improve live-birth rates.

• Journal of Periodontal and Implant Science
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• v.24 no.3
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• pp.581-596
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• 1994

This in vitro study was undertaken to obtain optimal tetracycline concentration that aids proliferation and spreading of human periodontal ligament cells, for clinical application in root surfaces of periodontally diseased teeth. Periodontal ligament cells used in this study were obtained from explants of periodontal ligament of 1st premolar teeth which were extracted for the purpose of orthodontic treatment. The cells were cultured in Dulbecco's Modified Eagle Medium(DMEM) supplemented with 100 U/ml penicillin, $100\;{\mu}g/ml$ streptomycin and 10% FBS at $37^{\circ}C$, 100% humidity, 5% $CO_2-95%$ air. Cells were used between the third to 4th passage. After root planing of periodontally extracted teeth, the root slabs were cut with carborundum disk. In the cell proliferation experiment, experimental groups were root planing only group, immersed groups in 25, 50, 75, 100, 150mg/ml aqueous solution of Tetracycline HCl followed by a vigorous rinse in PBS. Human PDL cells at concentration of $1{\times}10^5\;cells/ml$ were seeded in each culture well which contained root slabs and incubated for 6 hours. Then, all of the root slabs were moved into new 24 culture well and incubated 24, 48 and 72 hours. The cell counting was done by inverted phase contrast microscope after trypsinization. The following results were obtained. The cell number was increased in order root planing only group, 25, 150, 50, 75, 100mg/ml of Tetracycline HCl treated group in 24, 48 and 72 hours. The maximal cell number was obtained when the root slabs were immersed in solution with 100mg/ml of Tetracycline HCl. There were statistically significant between the root planing only group and 75, 100 mg/ml of Tetracycline HCl treated group in 24 hours, between the root planing only group and 100mg/ml of Tetracycline HCl treated group in 48 hours, between the root planing only group and 50, 75, 100mg/ml of Tetracycline HCl treated group, between 25 and 100mg/ml of Tetracycline HCl treated group in 72 hours(p<0.05). In the cell spreading experiment, after 30 minutes of incubated, in the root planing only group, the cells were generally round in shape. The cell surface was mostly covered with blebs. The cells started to attach to root surface by cytoplasmic extension in 50, 100mg/ml of Tetracycline HCl treated groups, more numerous cells attached to root surface than root planing only group. Many orifices of dentinal tubule were exposed, cells showed radially spreaded cytoplasm and unspreaded central region of the cell was covered with blebs. After 6 hours of incubation, in the root planing only group, cells showed radially spreaded cytoplasm and were attached flat appearance. In 50, 100mg/ml of Tetracycline HCl treated groups, cellular margin was concaved and cytoplasm showed elongated appearance with polarity. After 24 hours of incubation, in the root planing group, cells showed characteristic polarity. In 50, 100mg/ml of Tetracycline HCl treated groups, cells showed more elongated and spindle - like appearance.

• Journal of Periodontal and Implant Science
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• v.32 no.3
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• pp.457-473
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• 2002

The purposes of this study is to evaluate the combination effects of TGF-${\beta}_1$ and PDGF-BB on the periodontal ligament cells to use as a regeneration promoting agent of periodontal tissue. Human periodontal ligament cells were prepared from the first premolar tooth extracted for the orthodontic treatment and were cultured in DMEM/100% FBS at the $37^{\circ}C$, 5% $CO_2$ incubator. Authors measured the DNA synthesis, total protein, collagen and noncollagenous protein synthesis according to the concentration of TGF-${\beta}_1$,(1,5ng/ml) and PDGF-BB (1,10 ng/ml) in combination. To explore further this delayed effect of TGF-${\beta}_1$, we preincubated human periodontal ligament cells with TGF-${\beta}_1$ for 4 or 24 hours before PDGF-BB stimulation. The results were as follows: The DNA synthetic activity was increased dose dependently by TGF-${\beta}_1$, PDGF-BB. The combination of TGF-${\beta}_1$ and PDGF-BB consistently enhanced the DNA synthetic activity to PDGF-BB alone. The ability of TGF-${\beta}_1$ to enhance DNA synthetic activity in PDGF-BB treated periodontal ligament cells was dose dependent. The maximum mitogenic effect was at the 5ng/ml of TGF-${\beta}_1$ and l0ng/ml of PDGF-BB. Preincubation of cell with TGF-${\beta}_1$ resulted in significantly greater response to PDGF-BB at all TGF-${\beta}_1$ concentration studied, and may be useful for clinical application in periodontal regenerative procedures. The total protein, collagen and noncollagen synthesis was increased dose pendently by TGF-${\beta}_1$, PDGF-BB. The % of collagen was slightly decreased according to the concentration of TGF-${\beta}_1$, PDGF-BB. The effect of TGF-${\beta}_1$, PDGF-BB were not specific for collagen synthesis since it also increased noncollagenous protein synthesis. This study demonstrates that PDGF-BB is major mitogens for human periodontal ligament cells in vitro, and supports a role for TGF-${\beta}_1$ as a regulation of the mitogenic and total protein formation to PDGF-BB in these cells.