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研究生:劉佳明
研究生(外文):Chia-ming, Liu
論文名稱:植體粒子引發成骨細胞凋零機制之探討
論文名稱(外文):The mechanism of apoptosis in bone cell after implant particle induction
指導教授:王若松王若松引用關係洪志遠洪志遠引用關係
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:臨床牙醫學研究所
學門:醫藥衛生學門
學類:牙醫學類
論文種類:學術論文
論文出版年:2002
畢業學年度:90
語文別:中文
中文關鍵詞:植體粒子細胞凋零機制成骨細胞
外文關鍵詞:implant particlesapoptosisosteoblast cell line
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目標: 具有生物相容性的植體 (Implants)已經廣泛的應用在牙科和骨科的領域上。雖然成功率極高,但是植體因為感染或是非感染性鬆脫 (aseptic loosening) 仍然是臨床上討論的重點。長期的臨床研究也顯示植體的磨耗是無法避免的,卻少有研究把焦點放在成骨細胞 (Osteoblasts) 被植體粒子 (Implant particles) 所誘發的反應。所以本實驗的目的是為了釐清植體粒子所誘發的細胞反應,並進一步找出細胞反應可能的機制。
實驗材料與方法: 四種金屬粒子被應用在本實驗:純鈦 (cpTi), 鎳化鈦 (TiN), 鈦合金(Ti-6Al-4V), and 鈷鉻錳(Co-Cr-Mo)。兩種人體成骨細胞株 (MG-63 and U-2 OS) 也被採用在本實驗中。首先我們利用電子顯微鏡 (SEM) 和 Auger electron analysis 來做粒子的定性觀察,再利用 TUNEL 染色法來觀察細胞凋零 (Apoptosis) 的現象。最後,並利用西方墨點染色法 (western blotting) 來觀察細胞內蛋白濃度的表現量。
結果: 以電子顯微鏡觀察,金屬粒子在大小以及分布方面都不相同。Auger electron analysis 也表現出金屬粒子表面化學物質的不純性。而 TUNEL染色法則顯示出只有純鈦 (cpTi), 鎳化鈦 (TiN)兩種金屬粒子會造成細胞凋零現象。我們進一步利用西方墨點法來觀察被植體金屬粒子所引發細胞凋零現象與細胞內蛋白濃度之間的關係, 結果顯示不論在wild type p53 細胞株或是mutant type p53細胞株中,p53基因與其下游的基因- Bax 以及Bcl-2表現量都沒有增加,反而是Fas基因表現量在植體粒子 (純鈦以及鎳化鈦) 作用後有明顯增加。
結論: 實驗結果顯示植體粒子(純鈦以及鎳化鈦)引起的成骨細胞凋零現象的機制可能是經由Fas 基因的路徑,不需p53 基因及其相關的路徑 。
Objectives: Biomaterial implant are widely used in dental and orthopedic fields.The purpose of this study is to clarify particles-induced cellular responses and find out the possible mechanism.
Materials & Methods: Four kinds of particles are used in this study─commercially pure titanium (cpTi), titanium nitride (TiN), titanium alloy (Ti-6Al-4V), and Co-Cr-Mo. Two human cell lines (MG-63 and U-2 OS) were employed in the experiments. Particle characterization was done by SEM observation and auger electron analysis. TdT-mediated dUTP Nick End Labeling (TUNEL) staining were used for observation cell apoptosis. Besides, we use western blot to evaluate the protein expression level after particle induction.
Results: Particles were variations in size and distribution under SEM observations. Auger electron analysis showed that particles surface element impurity. TUNEL staining indicates that only cpTi and TiN induce bone cell apoptosis. Western blot showed that p53 expression level was not increased after particle treatment. Fas expression level was increased after particle treatment.
Conclusions: The cpTi and TiN particles induce bone cell apoptosis may via Fas-ligand pathway independent with p53 gene.
Abstract………………………………………………………………….i
中文摘要………………………………………………………………..ii
Table of Contents………………………………………………………..iv
List of Figures……………………………………………………………v
Background
I. Biomaterials and clinical implication…………………………….1
II. Apoptosis……………………………………………………….12
III. Genetic and biochemistry of apoptosis………………………..19
Specific Aims…………………………………………………………...32
Materials and Methods………………………………………………….33
Results…………………………………………………………………..39
Discussion………………………………………………………………61
Conclusion………………………………………………………………68
References………………………………………………………………69
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