• 대한치과보철학회지
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• 제37권6호
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• pp.717-729
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• 1999

Although many studies on the cytotoxicity of the dental cast base metal alloys and their components have been carried out, the results are rather conflicting because of the different type of cells used and the various experimental procedures taken. Recently a number of scientists have claimed that it would be preferable to focus on the use of cells from relevant specific location of the human bodies. Consequently, the primary cultured oral keratinocyte derived from oral mucous along with nickel chloride and several of widely used dental cast base metal alloys(two Ni-Cr alloys and one Co-Cr alloy)in domestic were selected for this study, from which 1) The amounts of released metal ions were determined using atomic absorption spectrometry, 2) The cytotoxicity of nickel chloride and dental cast base metal alloys was evaluated via MTT assay, and finally, 3) The amounts of released metal ions and the cytotoxicity of nickel chloride were correlated with the cytotoxicity of dental cast base metal alloys And, the results were summarized as follows; 1. Nickel ion from Ni-Cr alloys and Cobalt ion from Co-Cr alloys resulted in maximum releasing rate during first 2h hours, followed by a decrease in releasing rate with time. Chromium ion were found to be minimal in all alloys. 2. In cytotoxic test. with $40{\mu}M,\;80{\mu}M$ of nickel chloride, there were observed an increase in the relative cell number compared to control samples after 24 hours. With $160{\mu}M$, there was found to be no difference in the relative cell number with control, except that 48 hour showed a increase in relative cell number. With $320{\mu}M$, the relative cell number remained constant and decreased after 48 hours, and with $640{\mu}M$, a continuing decrease in relative cell number was observed throughout test period. 3 The sensitivity of primary cultured oral epithelium to nickel was lower compared to the cells used in other studies. 4. CB-80 Soft and Regalloy showed no cytotoxicity to primary cultured oral epithelium and New crown resulted in a slight cytotoxicity. In conclusion, it was shown that the primary cultured oral keratinocytes could be applied successfully as testing cells in cytotoxicity test. Futhermore, the dental cast base metal alloys used in this study were found to be biocompatible.

• International Journal of Oral Biology
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• 제39권2호
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• pp.97-105
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• 2014

The aim of this study was to determine the beneficial effect of propofol on human keratinocytes that have undergone hypoxia reoxygenation (H/R) injury and to investigate whether autophagy is associated with the protective mechanism. Thus, we evaluated how propofol influences the intracellular autophagy and apoptosis during the H/R process in the HaCaT cells. The cultured human keratinocyte cells were exposed to 24 h of hypoxia (5% $CO_2$, 1% $O_2$, 94% $N_2$) followed by 12 h of reoxygenation (5% $CO_2$, 21% $O_2$, 74% $N_2$). The experiment was divided into 4 groups: (1) Control=Normoxia ; (2) H/R=Hypoxia Reoxygenation ; (3) PPC+H/R=Propofol Preconditioning+Hypoxia Reoxygenation; (4) 3-MA+PPC+ H/R=3-MA-Methyladenine+Propofol Preconditioning+ Hypoxia Reoxygenation. In addition, Western blot analysis was performed to identify the expression of apoptotic pathway parameters, including Bcl-2, Bax, and caspase 3 involved in mitochondrial-dependent pathway. Autophagy was determined by fluorescence microscopy, MDC staining, AO staining, and western blot. The H/R produced dramatic injuries in keratinocyte cells. In our study, the viability of Propofol in H/R induced HaCaT cells was first studied by MTT assay. The treatment with 25, 50, and $100{\mu}M$ Propofol in H/R induced HaCaT cells enhanced cell viability in a dose-dependent manner and $100{\mu}M$ was the most effective dose. The Atg5, Becline-1, LC3-II, and p62 were elevated in PPC group cells, but H/R-induced group showed significant reduction in HaCaT cells. The Atg5 were increased when autophagy was induced by Propofol, and they were decreased when autophagy was suppressed by 3-MA. These data provided evidence that propofol preconditioning induced autophagy and reduced apoptotic cell death in an H/R model of HaCaT cells, which was in agreement with autophagy playing a very important role in cell protection.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제30권4호
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• pp.308-315
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• 2004

Aim: The aim of this study was to investigate the mechanism of promoted skin wound healing in skin defects with primary cultured oral mucosal keratinocytes. Materials and methods: Thirty adult female nude mice weighing $20{\pm}2g$ were used for the experiment. Primary cultured and suspended oral mucosal keratinocytes, labeled with BrdU, were scattered onto $1.5cm{\times}1.5cm$ sized full thickness skin defects in the experimental group(N=15), and no grafts were placed the control group(N=15). They were sacrificed at 3 days, 1 week and 2 weeks after the treatment respectively. Histological examination of each wounds were performed to review the healing progress on measuring the length from the wound margin to regenerating epithelial front. The role of keratinocytes were assessed by double immunohistochemical staining with Anti-BrdU and Anti-cytokeratin AE1/3. Results: In the experimental group the wound was completely covered with regenerating epithelia in 2 weeks, but partially regenerated in the control group. The immunohistochemical studies unexpectedly reveal that most of regenerating epithelial cells were induced from marginal epithelium of the margin, not from the scattered keratinocytes. Conclusion: We could successfully confirm that graft of primary cultured oral mucosal keratinocytes promotes the regeneration of skin defects.

• 한국치위생학회지
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• 제15권4호
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• pp.719-725
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• 2015

Objectives: The purpose of this study was to determine the toxic effects of sodium lauryl sulfate(SLS) in human keratinocyte HaCaT cells and mouse fibroblast NIH-3T3 cells. Methods: The effect of sodium lauryl sulfate(SLS) cell viability and proliferation were determined by WST-1 assay and changes shape of nucleus were evaluated by Hoechst staining under fluorescence microscopy. Additionally, observation of cell morphological changes under light microscopy. Results: SLS induced cytotoxicity and a marked apoptosis in both HaCaT and NIH-3T3 cell lines. With the result of the WST-1 assay, SLS induced the cytotoxicity of 0.005% and 0.0075%, 0.01% SLS for 24 h after HaCaT and NIH-3T3 cells in time and dose-dependent manner(p<0.005). SLS inhibited cell growth and caused apoptosis as evidenced by nuclear fragmentation and condensation. Thus, determination of the morphological changes to define apoptosis was visualized using inverted phase contrast microscopy. Conclusions: SLS had toxicity of the human keratinocyte cells and mouse fibroblast cells and this study will provide the basic data for the development of proper SLS concentration in dentifrice.

• 한국콘텐츠학회논문지
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• 제22권3호
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• pp.586-592
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• 2022

본 연구는 기존의 치면열구전색재 ConciseTM에 세륨옥사이드 나노입자(Cerium oxide nano particles; CNPs)를 0-4.0 wt%를 첨가하여 새로운 치면열구전색재를 제조하여 물리적 성질과 세포독성을 평가하였다. 물리적 성질은 중합깊이, 물흡수도와 용해도를 측정하였고 세포독성평가는 불멸화된 구강점막세포(Immortalized human oral keratinocyte (IHOK))를 이용하여 MTT assay법으로 평가하였다. 실험 결과 중합 깊이는 CNPs 첨가량이 증가할수록 감소하였고, 용해도는 CNPs를 2.0 wt% 첨가된 실험군에서 가장 낮은 값을 보였으며 물흡수도는 각 실험군별 유의한 차이는 나타나지 않았다(p>0.05). 세포독성 실험 결과는 모든 실험군에서 높은 세포 생존율을 보였다. 이는 CNPs가 물리적 성질을 크게 약화 시키지 않으며 세포독성을 나타내지 않았으므로 생체 적합성을 입증하였다. CNPs의 특성을 고려하여 향후 CNPs의 효율적인 분산 기술에 대한 추가 연구가 필요할 것으로 사료된다.

• Maxillofacial Plastic and Reconstructive Surgery
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• 제27권3호
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• pp.226-237
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• 2005

Purpose : Extensive defect of oral and maxillofacial area is usually reconstructed with composite flap including skin paddle. However, if the defects are lined with only skin components, the mucosa's role in mastication and texture are not restored. Furthermore, stiffness and hair-growing prevent denture rehabilitation and good oral hygiene. This study was performed to overcome the disadvantages of composite soft tissue flaps including the skin and to make a model for myo-mucosal flaps. Materials and methods : Buccal mucosa sized $0.5\times1.0\;cm^2$ from New Zealand rabbit (around 1.5kg) was harvested and cultivated by the modification of Rheinwald and Green's keratinocyte culture method. Cultured mucosa was grafted on the fascia of latismus dorsi as form of mucosal sheet. After 7, 10, 14 days, the myomucosal flap was excised and evaluated under light microscope with H & E and immunohistochemical staining. As control group, harvested buccal mucosa from rabbit was transplanted to gracilis muscle(n=6). Results : From 7 days after prelamination, the basal layer of the grafted mucosa resembled that of normal mucosa. As control group, transplanted mucosa had original shape but there's slight inflammatory reaction. Prelaminated mucosa has 19.8$\pm$4.59 cell layers and some samples have more than 20 layers. The expression rate of PCNA was relatively strong (42.9%$\pm$14.1) at the basal layer of grafted mucosa and the laminin was found at the basal layer. On the contrary, prelaminated mucosa at 10 days showed moderate expression rate of PCNA(32.4%$\pm$4.62). We found the mucosal layer was somehow disappeared and there is strong inflammatory reaction. After 14 days prelamination, the grafted oral keratinocytes were almost disappeared and expression of PCNA was not observed. Conclusion : We can make 75 fold large mucosal($3850mm^2$) sheet from small samples of mucosa $(50mm^2)$. Epithelial sheet that grafted on the fascia of muscle underwent differentiation and proliferation. But after 10, 14 days, there was strong inflammatory reaction and the grafted mucosa was destroyed from surface layer. In rabbit model, transfer of fascio-mucosal flap should be done from 7 to 10 days after prelamination.

• 대한치과마취과학회지
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• 제14권3호
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• pp.157-165
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• 2014

Background: Incisional site of surgical operation become transient ischemic state and then occur reoxygenation due to vasodilatation by inflammatory reaction, the productive reactive oxygen species (ROS) give rise to many physiologic results. Apoptosis have major role on elimination of inflammatory cell and formation of granulation tissue in normal wound healing process. Remifentanil can prevent the inflammatory response and can suppress inducible nitric oxide synthase expression in a septic mouse model. After cardiopulmonary bypass for coronary artery surgery, remifentanil can also inhibit the release of biomarkers of myocardial damage. Here we investigated whether remifentanil pretreatment has cellular protective effect against hypoxia-reoxygenation in HaCaT human keratinocytes, if so, the role of apoptosis and autophagy on this phenomenon. Methods: The HaCaT human keratinocytes were exposed to various concentrations of remifentanil (0.01, 0.05, 0.1, 0.5 and 1 ng/ml) for 2 h before hypoxia (RPC/HR group). These cells were cultured under 1% oxygen tension for 24h at $37^{\circ}C$. After hypoxia, to simulate reoxygenation and recovery, the cells were reoxygenated for 12 h at $37^{\circ}C$. 3-MA/RPC/HR group was treated 3-methyladenine (3-MA), autophagy inhibitor for 1h before remifentanil treatment. Cell viability was measured using a quantitative colorimetric assay with thiazolyl blue tetrazoliumbromide (MTT, amresco), showing the mitochondrial activity of living cells. To investigate whether the occurrence of autophagy and apoptosis, we used fluorescence microscopy and Western blot analysis. Results: The viability against hypoxia-reoxygenation injury in remifentanil preconditioning keratinocytes were increased, and these cells were showed stimulated expression of autophagy 3-MA suppressed the induction of autophagy effectively and the protective effects on apoptosis. Atg5, Beclin-1, LC3-II and p62 were elevated in RPC/HR group. But they were decreased when autophagy was suppressed by 3-MA. Conclusions: Remifentanil preconditioning showed the protective effect in human keratinocytes, and we concluded that autophagy may take the major role in the recovery of wound from hypoxia-reoxygenation injury. We suggest that further research is needed about the cell protective effects of autophagy.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제31권3호
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• pp.219-227
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• 2005

Purpose: Abnormalities in the p53 gene are regarded as the most consistent genetic abnormalities detected in head and neck squamous cell carcinogenesis. Two new members of the p53 gene family, p73 and p63 have recently been identified. They share considerable sequence homology with p53 in the transactivation, DNA binding, and oligomerization domains, indicating possible involvement in carcinogenesis. Disruption of the homeostatic balance between proliferation and apoptosis is widely believed to contribute to human oral carcinogenesis. The aim of this study was to analyze expression of p63 in squamous cell carcinogenesis and to compare with immunochemical markers representing cell proliferation and apoptosis. Materials and Methods: Using the Syrian hamster oral cancer model, the fraction of apoptotic (apoptotic index-AI), proliferating (mitotic index-MI) and p63 expressing keratinocytes were examined at normal, dysplastic and malignant oral epithelium using the TUNEL assay, PCNA and p63 immunostaining. Results: p63 significantly increased between normal and dysplastic epithelium and between dysplastic and malignant epithelium. PCNA significantly increased between normal and dysplastic epithelium and between normal and malignant epithelium. However, increase between dysplastic and malignant epithelium, though still increasing, was not statistically significant. The percentage of TUNEL positive cells increased from normal to dysplastic epithelium and returned to normal keratinocyte level in the malignant epithelium. However, differences between tissue types were not significant. The ratio of MI:AI increased significantly only in the dysplastic-malignant epithelial transition. The increase of p63 expression closely reflected the change in the MI:AI ratio during oral carcinogenesis. Conclusion: The p63 may be associated with the regulation of epithelial proliferation and apoptosis in DMBA-induced hamster buccal pouch squamous cell carcinogenesis. Further study is required to investigate which p63 isoforms are involved in hamster buccal pouch carcinogenesis.

• Maxillofacial Plastic and Reconstructive Surgery
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• 제23권3호
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• pp.258-262
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• 2001

Distraction osteogenesis(DO) can be performed in the bony defect associated with trauma, anomaly, and various kinds of disease. The gap generated by DO is filled with growing callus : during the period of distraction, the osteogenesis is continued. However, there have been few reports about expression pattern of growth factors in newly formed bone during the consolidation periods. We performed DO in the mandibular defect case and studied the expressed pattern of growth factors. Its pattern was compared to that of the same patient. BMP-2 and -4 were strongly expressed in the DO site. Particularly, BMP-4 was not expressed in the normal mature bone, but expressed in new bone in DO. However, there was no difference in the FGF-7 expression between the sites. Therefore, strong expression of BMP-4 are related to new bone formation in DO and they may not be related to the normal homeostasis in human bone. Though FGF-7 is related to the growth of keratinocyte, it may have minimal role in the DO and normal mature bone.

• Mycobiology
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• 제51권1호
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• pp.1-15
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• 2023

Wound care has become increasingly important over the years. Various synthetic products for wound care treatment have been reported to cause toxic side effects and therefore natural products are in significant demand as they have minimal side effects. The presence of bioactive compounds in medicinal mushrooms contributes to various biological activities which assist in the early inflammatory phase, keratinocyte proliferation, and its migration enhancement which are pertinent to wound rehabilitation. Lignosus rhinocerus (tiger milk mushroom) can reduce the inflammation phase in wound healing by fighting off bacterial infection and modulating pro-inflammatory cytokines expression in the early stage to avoid prolonged inflammation and tissue damage. The antibacterial, immunomodulating, and anti-inflammatory activities exhibited by most macrofungi play a key role in enhancing wound healing. Several antibacterial and antifungal compounds sourced from traditional botanicals/-products may prevent further complications and reoccurrence of injury to a wounded site. Scientific studies are actively underway to ascertain the potential use of macrofungi as a wound healing agent.