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研究生:傅康貴
研究生(外文):Kang-Kuei Fu
論文名稱:複合式雙鈣磷酸鹽類應用於植體周圍骨缺損之骨再生能力評估:動物試驗
論文名稱(外文):Effects of Dicalcium Phosphate Composite Graft in Bone Regeneration for Peri-Implant Bony Defect: Animal Study
指導教授:林俊彬林俊彬引用關係
指導教授(外文):Chun-Pin Lin
口試委員:郭英雄
口試日期:2013-06-26
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:2013
畢業學年度:101
語文別:中文
論文頁數:77
中文關鍵詞:雙鈣磷酸鹽骨塊
外文關鍵詞:Dicalcium phosphate dihydratebone block
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近年來,使用磷酸鈣鹽類作為生物合成的骨材在臨床上已經相當普遍,但此類骨移植材多為顆粒型式,顆粒大小自數十毫米到一毫米都有。雖然顆粒型的骨材比較容易填充縫隙,但是對於移植材料要能夠堅固撐出所需維持的空間以及要能穩定不動這兩點而言,顆粒型的骨材比較不適當。當應用在比較大之齒槽骨缺損時,可能會因為材料之擠壓變形而未能撐出空間,最後無法達到穩定良好的齒槽骨修復。因此,目前有許多複合式磷酸鹽類被發展出來並克服其強度不足的部分。在這些材料中,最近有一種主要由雙鈣磷酸鹽合併氫氧基磷灰石組成的新式骨塊被研發出來,這種材料除了具有足夠的機械強度之外,同時也具備了適當的降解率。本實驗的目的主要在研究此種新式複合代用骨塊用於植體周圍大範圍齒槽骨缺損重建之效能。
本實驗使用了3隻米格魯一歲成犬,體重介於7公斤到10公斤。主要分成3組,實驗組(n=7),使用的骨材為複合雙鈣磷酸鹽與氫氧基磷灰石之代用骨塊;對照組A(n=6),使用的骨材為添加了膠原蛋白的雙相磷酸鹽(複合氫氧基磷灰石與β型三鈣磷酸鹽);對照組B(n=5),沒有使用任何骨材。實驗的方式主要是將3隻米格魯獵犬的下顎骨左右4顆小臼齒與第一大臼齒拔除,在等待傷口癒合6週之後,會將本實驗3組不同材料合併植體依照不同的時間點隨機種植於3隻米格魯獵犬的下顎骨左右兩側,手術的方式主要是先在實驗動物的無牙齒槽脊上製備好一定大小的骨缺損,隨即利用預先準備好的骨材進行引導骨生成手術合併植牙手術。手術的時間點主要分成3個,分別是第0週、第4週與第8週,手術同時也會進行植體共振頻率的量測。另外在實驗的第8週與第10週分別會在3隻米格魯獵犬皮下注射骨標定染劑以供日後使用倒立螢光顯微鏡觀察植體周圍新生骨質沉積的速率與部位。最後,3隻米格魯獵犬統一於實驗的第12週犧牲,接著利用植體穩定商數分析、放射線影像分析、組織切片判讀、斷層掃描分析與骨頭螢光標定觀察等方式分析人工植體周圍的骨整合程度。
在結果的部分,實驗組複合雙鈣磷酸鹽與氫氧基磷灰石之代用骨塊對於幫助新骨生成的部分,不論是新骨的量與質地或是新骨出現的快慢,其結果都優於對照組A(複合氫氧基磷灰石、β型三鈣磷酸鹽與膠原蛋白的骨材)與對照組B(不含任何骨材)。在統計上有明顯差異的部分分別是12週組的植體穩定商數、12週組的x光片植體周圍骨覆蓋率、8週組與12週組的組織磨片植體周圍骨覆蓋率以及12週組的斷層掃描植體周圍骨密度。另外在骨頭螢光標定的部分也顯示出實驗組複合雙鈣磷酸鹽與氫氧基磷灰石之代用骨塊在骨頭成熟的速度是明顯優於其他兩組對照組。
最後結論的部分,透過本研究團隊所建立的動物試驗模式,可以證明此種新式複合雙鈣磷酸鹽與氫氧基磷灰石之代用骨塊具有作為骨填補材的潛力。另外透過骨標定染色技術,除了呼應本實驗的其他結果,也讓我們發現新骨在骨缺損處生成的模式。


Using particulate form calcium phosphates as the biosynthetic materials has become popular in clinical practice. Although the particulate form calcium phosphates are easily to fill small bony defect, however, their insufficiency in strength lead to instability of the graft and poor space maintenance of defect which result compromised efficacy in alveolar bone regeneration, especially in large alveolar bone defect. Thus many combinations of different forms of calcium phosphates have been proposed to overcome insufficiency of strength in grafts. Recently, a new bone block constitutes with mainly dicalcium phosphates and partially hydroxyapatites with sufficient strength and adequate resorption rate has been developed. The purpose of this study is to evaluate efficacy of new bone formation of such new developed bone block in large peri-implant alveolar bony defect in animal model.
In this study, we use three beagle dogs, weighing between 7 kg to 10 kg, distribute to three groups as following: 1. experimental group (n = 7) which using the bone block containing dicalcium phosphates and hydroxyapatite (DCPD+ HA) as graft in defect; 2.control group A (n = 6) which using the collagen enhanced particulate biphasic calcium phosphate(hydroxyapatite and β-tricalcium phosphate with collagen (HA+β-TCP+collagen)) for defect repair; 3.control group B (n = 5) which without any bone graft (blood clot only) in defect. In this experiment, the extraction of four mandibular premolars and one first molar at bilateral mandible was done in the beginning. Following six weeks healing, the implantation over the mandible with bony defect preparation and guide bone regeneration would be preceded with test bone grafts or without any grafts randomly. The operations are performed at different time points (4-week, 8-week and 12-week) before the animal’s sacrifices. At the time of operation, the implantation was performed simultaneously with the measurements of implant stability using resonance frequency detector. We also injected the bone labeling fluorescence subcutaneously at the time point 4-w and 2-w before the animal’s sacrificed for evaluation of the area and amounts of new bone deposition with inverted fluorescence microscope. After sacrificing, the use of implant stability quotient analysis, radiographic analysis, histological analysis, CT scan analysis and bone labeling technique were performed to evaluate the new bone formation and osseointegration at the bony defect around the eighteen implants.
Results of this study revealed that the efficacy of new bone formation and osseointegration of the experimental group (DCPD+HA) are better than control group A (HA+β-TCP+collagen) and control group B (blood clot only), whether in the quantity and the quality of new bone or the speed of new bone formation. Statistical significant differences among different groups can be shown in peri-implant bone coverage ratios at 8-w following surgery. Moreover, significant statistical differences can also be shown in implant stability quotient values, radiographic peri-implant bone coverage ratios, histological peri-implant bone coverage ratios, CT bone mineral density at 12-w following surgery. In addition, the bone labeling technique proved similar pattern of healing among the three groups, however, the speed of new bone formation of the experimental group (DCPD+HA) is significantly higher than the other two control groups.
In present animal study, healing pattern and clinical efficacy in the new dicalcium phosphates and hydroxyapatite bone block has been throughout investigated and evaluated, furthermore, such a new dicalcium phosphates and hydroxyapatite bone block has showed promising efficacy in bone regeneration as compare to commercial calcium phosphates.


目錄
口試委員會審定書 I
誌謝 II
中文摘要 III
英文摘要 V
目錄 VII
圖目錄 X
表目錄 XII
第 一 章 緒論 1
1.1 前言 1
1.2 研究動機 3
1.3 論文架構 4
第 二 章 文獻回顧 5
2.1 骨移植材料的種類 5
2.2 磷酸鈣鹽類 6
2.2.1 歷史沿革 6
2.2.2 分類 7
2.2.3 引導骨生成機制 8
2.2.4 製備方式 9
2.3 骨水泥 10
2.3.1 特性 10
2.3.2 分類 11
2.4 磷酸鹽添加膠原蛋白 12
2.5 關於非侵入性測量的方法 12
2.6 關於骨頭與植體接觸面積的計算方法 13
2.7 斷層掃描(micro-CT) 14
第 三 章 材料與方法 16
3.1 實驗動物的選擇 16
3.2 實驗材料與實驗設計 16
3.2.1 實驗植體種類 16
3.2.2 實驗骨材種類 16
3.2.3 實驗設計 19
3.3 手術過程及術後照顧 20
3.3.1 實驗動物的麻醉 20
3.3.2 第一階段的手術步驟 21
3.3.3 第二階段的手術步驟 21
3.4 非侵入性臨床實驗觀察 25
3.4.1 臨床觀察及照顧 25
3.4.2 共振頻率測定方法 25
3.5 動物的犧牲及標本的取得 26
3.5.1 福馬林藥水的製備 26
3.5.2 動物的犧牲 26
3.5.3 標本的取得 27
3.6 標本製作與染色 28
3.6.1 標本的初步切割 28
3.6.2 不含植體的脫鈣標本製備 29
3.6.3 含植體磨片標本的製備 30
3.7 骨頭與植體接觸面積的計算方法 32
3.8 放射線影像分析與斷層掃描 34
3.8.1 放射線影像分析 34
3.8.2 斷層掃描 36
3.9 骨頭螢光標定 36
3.9.1 標定方法 36
3.9.2 本實驗所使用的螢光染劑 36
3.9.3 標定步驟 37
3.9.4 螢光顯微鏡觀察 37
3.10 本實驗所使用的統計方法 37
第 四 章 實驗結果 38
4.1 植體存活率與樣本排除 38
4.2 植體穩定商數(ISQ value) 39
4.3 放射線影像分析 42
4.4 植體周圍齒槽骨再生率 47
4.5 斷層掃描分析 54
4.6 骨頭螢光標定 59
第 五 章 討論 63
5.1 存活率與排除樣本之探討 63
5.2 植體穩定商數(ISQ)之探討 63
5.3 放射線影像分析之探討 65
5.4 植體周圍齒槽骨再生之探討 66
5.5 斷層掃描分析之探討 67
5.6 骨頭螢光標定的探討 67
5.7 實驗設計限制之探討 69
第 六 章 結論 70
參考文獻 71



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