• Title/Summary/Keyword: Cell staining

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Effect of Pine (Pinus densiflora) Needle Extracts on Antioxidant Activity and Proliferation of Osteoclastic RAW 264.7 Cells (적송잎 추출물이 항산화 활성 및 파골세포의 증식에 미치는 영향)

  • Jeon, Min-Hee;Park, Mi-Ra;Park, Yong-Soo;Hwang, Hyun-Jung;Kim, Sung-Gu;Lee, Sang-Hyeon;Kim, Mi-Hyang
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.40 no.4
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    • pp.525-530
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    • 2011
  • Pine needles have long been used as a traditional health-promoting medicinal food in Korea. This study was carried out to investigate the effects of pine needle extracts on the antioxidant activity, and proliferation of osteoclastic RAW 264.7 cells. Pine needle extracts were examined using hot water, ethanol, hexane, hot water-ethanol, and hot water-hexane. The effects of the pine needle extracts were examined by comparing the results with that of a commercial agents, proanthocyanidin. Analysis of each extract indicated that hot water-ethanol and ethanol extracts contained the highest total polyphenol concentrations. The hot water-ethanol and ethanol extracts also showed relatively the highest SOD-like activity. The proliferation of osteoclastic RAW 264.7 cells treated with pine needle extracts was decreased by lower than 70%. In addition, the hot water and ethanol extracts of pine needle significantly reduced the number of tartrate-resistant acid phosphatase-positive ($TRAP^+$) multinucleated cells from osteoclatic RAW 264.7 cells. These results indicate that pine needle extracts had an anabolic effect on bone through the promotion of osteoclast differentiation, suggesting that they could be used for the treatment of common metabolic bone diseases.

Correlation between the Serum Leptin Level and the Expression of Leptin in Stomach Cancer Patients (위암에서 혈청 렙틴 레벨과 조직 내 렙틴 발현과의 상관관계)

  • Kim, Ji-Hyun;Jung, Hun;Jun, Kyong-Hwa;Kim, Sung-Keun;Chin, Hyung-Min;Jung, Ji-Han;Kim, Wook;Jeon, Hae-Myung;Park, Cho-Hyun;Park, Seung-Man;Park, Woo-Bae;Lim, Keun-Woo;Kim, Seung-Nam
    • Journal of Gastric Cancer
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    • v.8 no.4
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    • pp.176-181
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    • 2008
  • Purpose: The adipocyte-derived cytokine leptin plays a major role in the control of stable body weight by suppressing food intake and increasing energy metabolism. Leptin regulates the cell proliferation of various epithelial cells and it may be involved in the promotion of cancer. Leptin and its receptor are highly expressed in gastric adenocarcinoma, but the association between the serum leptin level and the tissue expression of leptin is uncertain. We evaluated the serum leptin level and the expressions of leptin and leptin receptor in gastric cancer, and we explore the possible mechanism and role of leptin in the carcinogenesis of gastric cancer. Materials and Methods: 72 carcinomas that were curatively resected at our hospital from October 2005 to March 2007 were included in this study. By immunoassay and immunohistochemical staining, we evaluated the serum leptin level and the expressions of leptin and its receptor, and we analyzed their relationship together with the clinicopathological variables. Results: The serum leptin level was increased as the patient's BMI increased and it was decreased in H. pylori infected patients. The expression of leptin was increased as the TNM stage increased (P=0.014), and the expression of leptin receptor in the intestinal type gastric adenocarcinoma was higher than that in the diffuse type gastric adenocarcinoma (71.4% vs 28.6%, respectively, P=0.033). Conclusion: There was no significant correlation between the serum leptin level and expression of leptin in gastric cancer patients. The expression of leptin was associated with the TNM stage, but its role in the pathogenesis of gastric cancer has to be elucidated.

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The Immunohistochemical Expression of Nuclear Retinoid Receptor and CREB in Gastric Adenocarcinoma (위선암종에서 핵의 Retinoic Acid Receptor (RAR) 및 cAMP Response Element Binding Protein (CREB)의 면역조직화학적 발현양상)

  • Han, Ho-Sun;Kim, Yong-Seok;Park, Joong-Min;Choi, Yoo-Shin;Cha, Seong-Jae;Kim, Mi-Kyung;Chi, Kyung-Choun
    • Journal of Gastric Cancer
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    • v.8 no.4
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    • pp.182-188
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    • 2008
  • Purpose: Transcriptional factors of CREB (cAMP response element binding protein) are involved in regulating the gene expression in response to a variety of signaling pathways. The proteins produced by the CREB genes play key roles in many physiological processes, including memory and long-term potentiation. The retinoic acid receptor (RAR) axis mediates epithelial cell differentiation and proliferation in many tissues. This study examined the expressions of RAR and CREB and their relationship with the clinicopathologic factors and their significance. Materials and Methods: The levels of the RAR and CREB expressions were measured in 150 gastric adenocarcinomas by performing immunohistochemical staining. Results: 1. An RAR protein expression was found in 63.3% of the adenocarcinomas (95/150) and a CREB expression was found in 60.7% (91/150) of the adenocarcinomas. 2. An RAR protein expression was found in 72.2% (78/108) of the intestinal type adenocarcinomas and in 40.5% (17/42) of the diffuse type adenocarcinomas (P<0.05). Based on the depth of invasion, an RAR protein expression was found in 58.3% (14/24) of the T1 adenocarcinomas, in 61.9% (13/21) of the T2 adenocarcinomas, in 63.5% (61/96) of the T3 adenocarcinomas, in 77.8% (7/9) of the T4 adenocarcinomas and in 74.7% (62/83) of the adenocarcinomas with lymph node metastasis and in 49.2% (33/67) of the adenocarcinomas without lymph node metastasis (P<0.01). 3. A CREB expression was found in 69.4% (75/108) of the intestinal type and in 38.1% (16/42) of the diffuse type (P>0.05). Based on the depth of invasion, a CREB expression was found in 50% (12/24) of the T1 adenocarcinomas, in 52.4% (11/21) of the T2 adenocarcinomas, in 64.6% (62/96) of the T3 adenocarcinomas, in 66.6% (6/9) of the T4 adenocarcinomas, in 71.1% (59/83) of the adenocarcinomas with lymph node metastasis and in 47.8% (32/67) of the adenocarcinomas without lymph node metastasis (P<0.01). 4. The RAR protein and CREB expressions coincided in 71.4% of the gastric adenocarcinomas and a significant correlation between them was found (P<0.05). Conclusion: We found a significant relationship between the expression of RAR and CREB and the histology and lymph node metastasis of gastric cancer. Further studies are needed to confirm their biologic meaning in gastric carcinogenesis.

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THE EFFICACY OF PROGRAMMED CRYO-PRESERVATION UNDER PRESSURE IN RAT PERIODONTAL LIGAMENT CELLS (압력 저속 냉동 방법의 쥐 치아 치주인대세포 보존 효율 평가)

  • Lee, Young-Eun;Kim, Eui-Seong;Kim, Jin;Han, Seung-Hoon;Lee, Seung-Jong
    • Restorative Dentistry and Endodontics
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    • v.34 no.4
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    • pp.356-363
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    • 2009
  • The purpose of this study was to evaluate the viability of periodontal ligament cells in rat teeth using slow cryo-preservation method under pressure by means of MTT assay and WST-1 assay. Eighteen teeth of Sprague-Dawley white female rats of 4 week-old were used for each group. Both sides of the first and second maxillary molars were extracted as atraumatically as possible under Tiletamine anesthesia. The experimental groups were group 1 (Immediate control), group 2 (Cold preservation at $4^{\circ}C$for 1 week), group 3 (Slow freezing), group 4 (Slow freezing under pressure of 3 MPa). F-medium and 10% DMSO were used as preservation medium and cryo-protectant. For cryo-preservation groups, thawing was performed in $37^{\circ}C$water bath, then MTT assay and WST-1 assay were processed. One way ANOVA and Tukey method were performed at the 95% level of confidence. The values of optical density obtained by MTT assay and WST-1 were divided by the values of eosin staining for tissue volume standardization. In both MTT and WST-1 assay, group 4 showed significantly higher viability of periodontal ligament cells than group 2 and 3 (p < 0.05), but showed lower viability than immediate control group. By the results of this study, slow cryo-preservation method under pressure suggests the possibility for long term cryo-preservation of the teeth.

Antioxidant and Anticancer Activities of Euonymus porphyreus Extract in Human Lung Cancer Cells A549 (인체 폐암 세포주 A549에서 Euonymus porphyreus 추출물의 항산화 및 항암활성 분석)

  • Jin, Soojung;Oh, You Na;Son, Yu Ri;Bae, Soobin;Park, Jung-ha;Kim, Byung Woo;Kwon, Hyun Ju
    • Journal of Life Science
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    • v.31 no.2
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    • pp.199-208
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    • 2021
  • Euonymus porphyreus, a species of plant in the Celastraceae family, is widely distributed in East Asia, especially in Southern China. The botanical characteristics of E. porphyreus have been reported, but its antioxidative and anticancer activities remain unclear. In this study, we evaluated the antioxidative and anticancer effects of ethanol extracts of E. porphyreus (EEEP) and the molecular mechanism of its anticancer activity in human lung adenocarcinoma A549 cells. The total polyphenol and flavonoid compound contents from EEEP were 115.42 mg/g and 23.07 mg/g, respectively. EEEP showed significant antioxidative effects with a concentration at 50% of the inhibition (IC50) value of 11.09 ㎍/ml, as measured by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay. EEEP showed cytotoxic activity by increasing the SubG1 cell population of A549 cells in a dose-dependent manner. Apoptosis in A549 cells treated with EEEP was evident due to increased apoptotic cells and apoptotic bodies, as detected by Annexin V and 4,6-diamidino-2-phenylindole (DAPI) staining, respectively. EEEP-induced apoptosis resulted in increased expression of the First apoptosis signal (Fas), p53, and Bax, with decreased expression of Bcl-2 and subsequent activation of caspase-8, -9, and caspase-3, leading to cleavage of poly (ADP-ribose) polymerase (PARP). Collectively, these results suggest that EEEP may exert an anticancer effect by inducing apoptosis in A549 cells through both intrinsic and extrinsic pathways.

Antibacterial and Antibiofilm Activities of Leaf Extracts of Stewartia koreana against Porphyromonas gingivalis (Porphyromonas gingivalis에 대한 노각나무 잎 추출물의 항균활성 및 생물막 형성 억제 효과)

  • Kim, Hye Soo;Park, Min Jeong;Kim, Soo Jeong;Kim, Bu Kyung;Park, JunHo;Kim, DaeHyun;Cho, Soo Jeong
    • Journal of Life Science
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    • v.31 no.3
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    • pp.330-337
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    • 2021
  • This study was conducted to investigate the potential of Stewartia koreana as oral healthcare materials. The antibacterial activity of ethanol extracts from leaves and branches of S. koreana against oral bacteria was confirmed. The leaf and branch extracts (1 mg/disc) showed antibacterial activity against P. gingivalis only among several tested oral bacteria. The leaf extracts showed higher antibacterial activity, with values similar to those of chlorhexidine, which was used as a positive control. The MIC of the leaf extract against P. gingivalis was 0.4 mg/ml and showed bacteriostatic action. The inhibitory effects of the extract on biofilm formation and on gene expression related to biofilm formation by P. gingivalis were determined by biofilm biomass staining, scanning electron microscopy (SEM), and qRT-PCR analysis. The biofilm production rate and cell growth of P. gingivalis in the cultures treated with 0.2-2.0 mg/ml of S. koreana leaf extracts were significantly decreased in a concentration-dependent manner. The inhibitory effect on the formation of P. gingivalis biofilms at concentrations of 1 mg/ml was confirmed by SEM. The qRT-PCR analysis showed concentration-dependent suppression of the fimA and fimB gene expression associated with fimbriae formation in the cultures treated with 0.2-2.0 mg/ml S. koreana leaf extract. These results support the conclusion that S. koreana leaf extracts can be used as oral healthcare materials derived from natural materials, as demonstrated by the antibacterial action and inhibition of biofilm formation of P. gingivalis.

Leaves of Cudrania tricuspidata on the Shoot Positional Sequence Show Different Inhibition of Adipogenesis Activity in 3T3-L1 Cells (꾸지뽕 신초 엽위별 잎 추출물의 항비만 효과)

  • Park, Ju Ha;Guo, Lu;Kang, He Mi;Son, Beung Gu;Kang, Jum Soon;Lee, Yong Jae;Park, Young Hoon;Je, Byoung Il;Choi, Young Whan
    • Journal of Life Science
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    • v.31 no.2
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    • pp.209-218
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    • 2021
  • This study aimed to evaluate the anti-obesity effects of Cudrania tricuspidata leaf extract in the order of leaf development on the shoot (L0, L1, L2, L3, L4, and L5). The leaves at the apex of a Cudrania tricuspidata shoot were classified as L0; the next leaves of the apex were classified as L1, L2, L3, and L4 from highest to lowest; and the lowest leaf was classified as L5. A series of 70% ethyl alcohol leaf extracts were screened for the inhibitory effects of adipogenesis in 3T3-L1 preadipocytes. We found that the apical leaf extract of Cudrania tricuspidata (CTL0) was the most effective. Next, a study was conducted on the inhibitory action mechanism of CTL0. Treatment with CTL0 significantly suppressed the differentiation of 3T3-L1 preadipocytes in a dose-dependent manner, as confirmed by the decrease in lipid droplet content observed with Oil Red O staining. Treatment with 12.5 ㎍/ml, 25 ㎍/ml, and 50 ㎍/ml of CTL0 significantly reduced the lipid droplet content. Glucose and cellular triglyceride concentrations were reduced in the 3T3-L1 cells on the CTL0-treated medium compared to the differentiation medium (DM control, DMEM + insulin + dexamethasone + rosiglitazone). Compared with DM, CTL0 significantly inhibited the expression of key pro-adipogenic transcription factors, including peroxisome proliferator-activated receptor γ (PPARγ), LPL, A-FABP, and Glut4. These findings show that CTL0 extract has potent anti-obesity effects.

Clinical Analysis According to $p21^{Waf1/Cip1}\;and\;p27^{kip1}$ Expression in Gastric Cancer (위암에서의 $p21^{Waf1/Cip1}\;and\;p27^{kip1}$ 단백 발현)

  • Kim, Sin-Sun;Park, Yong-Geun;Jun, Kyong-Hwa;Jung, Hun;Song, Gyo-Young;Kim, Jin-Joo;Chin, Hyung-Min;Kim, Wook;Park, Cho-Hyun;Park, Seung-Man;Lim, Keun-Woo;Kim, Seung-Nam;Jeon, Hae-Myung
    • Journal of Gastric Cancer
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    • v.6 no.1
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    • pp.36-42
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    • 2006
  • Purpose: The $p21^{Waf1/Cip1}$ protein Inhibits the cell cycle by Inhibiting the phosphorylation at the $G1{\rightarrow}S$ check point, and the $p27^{kip1}$ protein similarly performs the suppressor function by controlling the p27-mediated G1 arrest. In this study, we analysed the clinical status and survival rates in correlations with p21 and p27 expression patterns in gastric cancer. Materials and Methods: Between 1993 and 1997, 192 patients who underwent surgeries in Catholic Medical Center were analysed retrospectively in this study. Immunohistochemical staining was performed and if the nuclei of the tumor cells were stained, we assumed those as positive results. Statistical analysis was based on clinicopathological findings and differences in survival rates. Results: The expression rate of p27 was 28.1% and 15.6% in p21 each. The ratio of T1-2(80.0%) was significantly high in p21 (+), but the ratio of T3-4 (50.6%) was slightly high in p21 (-). There was no statistical significance regarding other factors. The results in p27 was not much different from expression rate of p21 in T-stage. In addition, p27 expression in diffuse type (91.3%) was higher than in intestinal type (62.7%) by Lauren's classification (P<0.05). Also, there was no statistical significance in other factors. In the correlation of p21 and p27, p27 was positive when p21 was positive (53.5%). Conversely, p27 was negative when p21 was negative (76.5%, p<0.05). In the p21 and p27 combination test, there was higher rate of T1-2 (87.5%) in p21 (+)/p27 (+), and higher rate of T3-4 (58.1%) in p21 (-)/p27 (-) (P<0.05). Results showed higher rate of intestinal type (100%) in p21 (+)/p27 (+), and diffuse type (87.0%) was dominant in p21 (-)/p27 (-) (P<0.05) by Lauren's classification. Moreover, there was no statistical significance in the 5-year survival rate in the expression of p21 and p27, and the 5-year survival rate was highest in the case of p21 (+)/p27 (+) without statistical significance. Conclusion: In our study, $p21^{Waf1/Cip1}\;and\;p27^{kip1}$ expressed similar patterns. The expression of $p21^{Waf1/Cip1}\;and\;p27^{kip1}$ affected the degree of invasiveness of the tumor, and. Combined examination result revealed the correlation of $p21^{Waf1/Cip1}\;and\;p27^{kip1}$ with Lauren's classification and depth of invasion of the tumor. However, we assumed that little difference between the survival rates depending on expression of $p21^{Waf1/Cip1}\;and\;p27^{kip1}$ has limited their value as predictable prognostic indicators.

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The Effects of Proinflammatory Cytokines and TGF-beta, on The Fibroblast Proliferation (Proinflammatory Cytokines과 TGF-beta가 섬유모세포의 증식에 미치는 영향)

  • Kim, Chul;Park, Choon-Sik;Kim, Mi-Ho;Chang, Hun-Soo;Chung, Il-Yup;Ki, Shin-Young;Uh, Soo-Taek;Moon, Seung-Hyuk;Kim, Yong-Hoon;Lee, Hi-Bal
    • Tuberculosis and Respiratory Diseases
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    • v.45 no.4
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    • pp.861-869
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    • 1998
  • Backgrounds: The injury of a tissue results in the infalmmation, and the imflammed tissue is replaced by the normal parenchymal cells during the process of repair. But, constitutional or repetitive damage of a tissue causes the deposition of collagen resulting in the loss of its function. These lesions are found in the lung of patients with idiopathic pulmonary fibrosis, complicated fibrosis after diffuse alveolar damage (DAD) and inorganic dust-induced lung fibrosis. The tissue from lungs of patients undergoing episodes of active and/or end-stage pulmonary fibrosis shows the accumulation of inflammatory cells, such as mononuclear cells, neutrophils, mast cells and eosinophils, and fibroblast hyperplasia. In this regard, it appears that the inflammation triggers fibroblast activation and proliferation with enhanced matrix synthesis, stimulated by inflammatory mediators such as interleukin-1 (IL-1) and/or tumor necrosis factor (TNF). It has been well known that TGF-$\beta$ enhance the proliferation of fibroblasts and the production of collagen and fibronectin, and inhibit the degradation of collagen. In this regard, It is likely that TGF-$\beta$ undergoes important roles in the pathogenesis of pulmonary fibrosis. Nevertheless, this single cytokine is not the sole regulator of the pulmonary fibrotic response. It is likely that the balance of many cytokines including TGF-$\beta$, IL-1, IL-6 and TNF-$\alpha$ regulates the pathogenesis of pulmonary fibrosis. In this study, we investigate the interaction of TGF-$\beta$, IL-1$\beta$, IL-6 and TNF-$\alpha$ and their effect on the proliferation of fibroblasts. Methods: We used a human fibroblast cell line, MRC-5 (ATCC). The culture of MRC-5 was confirmed by immunofluorecent staining. First, we determined the concentration of serum in cuture medium, in which the proliferation of MRC-5 is supressed but the survival of MRC-5 is retained. Second, we measured optical density after staining the cytokine-stimulated cells with 0.5% naphthol blue black in order to detect the effect of cytokines on the proliferation of MRC-5. Result: In the medium containing 0.5% fetal calf serum, the proliferation of MRC-5 increased by 50%, and it was maintained for 6 days. IL-1$\beta$, TNF-$\alpha$ and IL-6 induced the proliferation of MRC-5 by 45%, 160% and 120%, respectively. IL-1$\beta$ and TNF-$\alpha$ enhanced TGF-$\beta$-induced proliferation of MRC-5 by 64% and 159%, but IL-6 did not affect the TGF-$\beta$-induced proliferation. And lNF-$\alpha$-induced proliferation of MRC-5 was reduced by IL-1$\beta$ in 50%. TGF-$\beta$, TNF-$\alpha$ and both induced the proliferation of MRC-5 to 89%, 135% and 222%, respectively. Conclusions: TNF-$\alpha$, TGF-$\beta$ and IL-1$\beta$, in the order of the effectiveness, showed the induction of MRC-5 proliferation of MRC-5. TNF-$\alpha$ and IL-1$\beta$ enhance the TGF-$\beta$-induced proliferation of MRC-5, but IL-6 did not have any effect TNF-$\alpha$-induced proliferation of MRC-5 is diminished by IL-1, and TNF-$\alpha$ and TGF-$\beta$ showed a additive effect.

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THE EFFECTS OF THE PLATELET-DERIVED GROWTH FACTOR-BB ON THE PERIODONTAL TISSUE REGENERATION OF THE FURCATION INVOLVEMENT OF DOGS (혈소판유래성장인자-BB가 성견 치근이개부병변의 조직재생에 미치는 효과)

  • Cho, Moo-Hyun;Park, Kwang-Beom;Park, Joon-Bong
    • Journal of Periodontal and Implant Science
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    • v.23 no.3
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    • pp.535-563
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    • 1993
  • New techniques for regenerating the destructed periodontal tissue have been studied for many years. Current acceptable methods of promoting periodontal regeneration alre basis of removal of diseased soft tissue, root treatment, guided tissue regeneration, graft materials, biological mediators. Platelet-derived growth factor (PDGF) is one of polypeptide growth factor. PDGF have been reported as a biological mediator which regulate activities of wound healing progress including cell proliferation, migration, and metabolism. The purposes of this study is to evaluate the possibility of using the PDGF as a regeneration promoting agent for furcation involvement defect. Eight adult mongrel dogs were used in this experiment. The dogs were anesthetized with Pentobarbital Sodium (25-30 mg/kg of body weight, Tokyo chemical Co., Japan) and conventional periodontal prophylaxis were performed with ultrasonic scaler. With intrasulcular and crestal incision, mucoperiosteal flap was elevated. Following decortication with 1/2 high speed round bur, degree III furcation defect was made on mandibular second(P2) and fourth(P4) premolar. For the basic treatment of root surface, fully saturated citric acid was applied on the exposed root surface for 3 minutes. On the right P4 20ug of human recombinant PDGF-BB dissolved in acetic acid was applied with polypropylene autopipette. On the left P2 and right P2 PDGF-BB was applied after insertion of ${\beta}-Tricalcium$ phosphate(TCP) and collagen (Collatape) respectively. Left mandibular P4 was used as control. Systemic antibiotics (Penicillin-G benzathine and penicillin-G procaine, 1 ml per 10-25 1bs body weight) were administrated intramuscular for 2 weeks after surgery. Irrigation with 0.1% Chlorhexidine Gluconate around operated sites was performed during the whole experimental period except one day immediate after surgery. Soft diets were fed through the whole experiment period. After 2, 4, 8, 12 weeks, the animals were sacrificed by perfusion technique. Tissue block was excised including the tooth and prepared for light microscope with H-E staining. At 2 weeks after surgery, therer were rapid osteogenesis phenomenon on the defected area of the PDGF only treated group and early trabeculation pattern was made with new osteoid tissue produced by activated osteoblast. Bone formation was almost completed to the fornix of furcation by 8 weeks after surgery. New cementum fromation was observed from 2 weeks after surgery, and the thickness was increased until 8 weeks with typical Sharpey’s fibers reembedded into new bone and cementum. In both PDGF-BB with TCP group and PDGF-BB with Collagen group, regeneration process including new bone and new cementum formation and the group especially in the early weeks. It might be thought that the migration of actively proliferating cells was prohibited by the graft materials. In conclusion, platelet-derived growth factor can promote rapid osteogenesis during early stage of periodontal tissue regeneration.

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