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산소 장력이 치주인대에서 유래한 세포의 활성과 기능에 미치는 영향
사명희,양원식 대한치과교정학회 1997 대한치과교정학회지 Vol.27 No.3
정치료를 위해 발거한 제 1 소구치에서 사람의 치주인대 세포를 채취하여 37℃, 5% CO₂, 100% 습도의 환경에서 배양한 후 5 내지 6 계대배양하여 실험에 사용하였다. Gaspack system에 0.2㎛ Millipore filter를 부착하여 세균 감염을 막았고, 저산소군에는 10% O₂, 5% CO₂, 85% N2 가스를, 과산소군에는 90% O₂, 5% CO₂, 5% N₂ 가스를 연결하여 산소 장력에 변화를 주었으며 대조군에는 5% CO₂, 95% 공기를 공급하였다. 37℃에서 2, 4, 6일간 배양한 후에 세포활성을 측정하기 위해 tetrazolium(MTT) assay를 시행하였고 sulforhodamine B(SRB) assay를 이용하여 총 단백질 합성을 알아보았으며, 4-hydroxyproline 측정으로 교원질 합성을 측정해 보았다. 또한 효소면역측정법으로 IL-1β, Il-6, TNF-α를 측정하여 다음과 같은 결과를 얻었다. 1. 저산소군에서 세포 활성과 총 단백질 합성은 대조군보다 다소 높거나 같은 정도이었다. 2. 과산소군에서 세포 활성은 대조군보다 낮았으며 총 단백질 합성도 다소 감소되었다. 3. 교원질 합성은 저산소군과 과산소군에서 대조군보다 유의하게 감소되었다가 배양기간이 길어짐에 따라 증가되어 대조군과 유사한 정도에 달했다. 4. 효소면역법 측정 결과 IL-6, TNF-α, IL-1β순으로 다량 검출되었다. 5. IL-6, TNF-α, IL-1β는 저산소군과 과산소군에서 배양기간이 길어짐에 따라 대조군에 비해 급격히 증가하였다. 6. IL-6, TNF-α는 과산소 조건에서 배양6일 후에는 대조군에 비해 유의하게 증가하였으며 이는 배양 2일 또는 4일 후보다도 유의하게 증가한 것이었다. This study was undertaken to investigate the effect of oxygen tension on the activity and function of the cells derived from human periodontal ligament by measuring cell activity, total protein synthesis, collagen synthesis, IL-1β, IL-6, TNF-α. Human periodontal ligament fibroblasts were collected from premolars extracted for orthodontic treatment and incubated in the environment of 37℃, 5% CO₂, 100% humidity. After the fifth to sixth passage they were used for the experiment. Gaspack system to which 0.2㎛ Millipore filter was attached was connected to mixed-gas tanks. The mixed gases were composed of 10% O₂, 5% CO₂, 85% N₂ in hypoxic group of 90% O₂, 5% CO₂, 5% N₂ in hyperoxic group and 5% CO₂, 95% air for control. After incubation in 37℃ for 2, 4, 6 days, cell activity was determined by tetrazolium(MTT) assay and total protein synthesis was assayed using sulforhodamine B(SRB). And measurement of 4-hydroxyproline was performed to assess collagen synthesis and IL-1β, IL-6 and TNF-α were measured by enzymeimmunoassay. The results were as follows. 1. The cell activity and total protein synthesis in hypoxic group were a little higher than or almost the same with those in control group. 2. In hyperoxic group, the cell activity was lower than that in control group and total protein synthesis was decreased. 3. Collagen synthesis was significantly decreased initially in both hypoxic and hyperoxic group and increased nearly to the level of control group as the duration of cell incubation was longer. 4. As a result of enzymeimmunoassay, the amount of cytokines was IL-6, TNF-α and IL-1β in order. 5. IL-6, TNF-α and IL-1β were increased more rapidly in both hypoxic and hyperoxic group than in control group as the duration of cell incubation was longer. 6. There were more Il-6 and TNF-α in hyperoxic group than in control group after 6 days, and there were more Il-6 and TNF-α after 6 days than after 2 or 4 days in hyperoxic group. These results suggested that oxygen tension might modulate the production of extracellular matrix and cytokines in the cells derived from human periodontal ligament.
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사명희,양원석 대한치과교정학회 1992 대한치과교정학회지 Vol.22 No.1
This study was aimed to compare the relative cytotoxicity of the five common orthodontic bonding materials (Concise, Mon-lok, Ortho-One, Super C, Transbond) using cell culture technique. DNA synthesis of the fibroblasts was assessed by ³H-thymidine uptake to evaluate the effect of the bonding materials on the growth of the cells. The human gingival fibroblasts were explanted from the buccal gingiva of 10 year-old girl and cultured in α-MEM/10% FBS/1% antibiotics medium, 37℃, 5% CO₂incuvator. The gingival fibroblasts were tested with the medium into which the bonding materials gad been soaked for 1 week. Or the bonding materials were placed on the cells immediately or 2 weeks after polymerization. After 22 hours, ³H-thymidine was added into the microtest wells and after 24 hours, the uptake of ³H-thymidine was determined by liquid scintilation counter. The results of this study were as follows. 1. DNA synthesis was significantly decreased with Super C and Transbond than Ortho-One, when treated with medium into which the bonding materials had been soaked for 1 week. 2. DNA synthesis was significantly decreased with Concise, Super C and Transbond than control, when treated immediatedly after polymerization. 3. DNA synthesis was significantly decreased with Concise, Super C and Transbond than Ortho-One, when immediatedly after polymerization. 4. There was no significant difference in DNA synthesis between the bonding materials, when treated 2 weeks after polymerization.
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산소배양환경이 치주인대에서 유래한 세포에 미치는 영향에 관한 형태학적 연구
김기영,유수경,김강주,사명희,양원식 원광대학교 생체재료·매식연구소 1996 원광생체재료·매식 Vol.5 No.1
This study was undertaken to investigate the effect of oxygen culture condition on the cells derived from human periodontal ligament by observation of morphological change. Human periodontal ligament fibroblasts were collected from premolars extracted for orthhodontic treatment and incubated in the environment of 37℃, 5% CO_2, 100% humidity. For the change of culture condition, O_2, CO_2 and N_2 gases were mixed with 10%, O_2, 5% CO_2 and 85% N_2 in hypoxic group, 90% O_2, 5% CO_2 and 5% N_2 in hyperoxic group, and 5% CO_2 and 95% air for control, and the cells were incubated in 37℃ for 2 and 4 days. The cells after incubation were measured for the quantity of total collagen by colorimetry and were stained for the observation of morphological change by direct immunfluoroscence method. After 2 days, the cells showed the decreased level of total collagen in the 10% O_2 and 90% O_2 cultured cells in contrast cells (Control : 9.33 ± 1.49, 10% O_2 : 5.32 ± 0.51, 90% O_2 : 5.82 ± 0.94). After 4 days, the cells increased the amount of total collagen in 10% O_2 and 90% O_2 groups but this value was not overtaken in control cells. In the morphological change, cells showed the 10% O_2 cultured cells after 2days atrophy of nucleus, hypertrophy of cytoplasm, and weakly stimulation of differentiation and growth in compared to control cells. After 4 days, cells showed cell necrosis and moderate hypertrophy of cytoplasm. The 90% O_2 cultured cells in 2 day groups, the nucleus was changed to oval and cells showed cellular swelling and partial death. At 4 days after culture, dead cells were increased and the nucleus became long and thin. These results suggest that periodontal ligament cell might induce irreversible cellular adaption by hypoxia and hyperoxia condition, and consequently rapid irreversible cellular adaption might be induced by hyperoxia condition.
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