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研究生:鍾慧穎
研究生(外文):Chung, Huiying
論文名稱:熱可逆水膠Pluronic F127在組織工程上的應用
論文名稱(外文):Thermoreversible Hydrogel Pluronic F127 Application In Tissue Engineering
指導教授:官常慶官常慶引用關係
指導教授(外文):Kwan, Changchin
口試委員:張乃方林孝道張家寧楊順發
口試委員(外文):Chang, NaifangLin, ShawtaoChang, ChianingYang, Shunfa
口試日期:2012-06-29
學位類別:博士
校院名稱:靜宜大學
系所名稱:應用化學系
學門:自然科學學門
學類:化學學類
論文種類:學術論文
論文出版年:2012
畢業學年度:100
語文別:中文
論文頁數:68
中文關鍵詞:組織工程Pluronic F127熱可逆可注射性水膠
外文關鍵詞:Tissue EngineeringPluronic F127ThermoreversibleInjectable hydrogel
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組織工程(Tissue Engineering)的興起主要的原因在於為了解決臨床上因為器官或組織的缺損或衰竭所造成的問題,希望組織工程的發展最終能夠修復、維持、改善人體組織的功能。三十多年來世界各國在組織工程領域的研究成果是非常豐碩的,但是顯少一系列由小動物到大型動物不同物種的相關研究發表,來證實的相關成果在臨床應用的可行性。
骨髓間質幹細胞(Bone Marrow Mesenchymal stem cell, MSC)是組織工程常採用的細胞種類,本論文採用自體骨髓間質幹細胞,其特點為自體細胞不用考量免疫性問題,且可輕易體外培養出足夠的細胞量。將骨髓間質幹細胞利用重組人類第二型骨形態發生蛋白(Human recombinant bone morphogenetic protein-2, rhBMP-2)引導其分化為成骨細胞。
熱可逆水膠Pluronic F127(FP127)目前已被美國食品藥物管理局(FDA)核准用於醫藥上之高分子材料,其特點為在低溫中為液態,當溫度接近人體溫度37℃時會形成凝膠狀。在實際應用上可以非常簡便的方法,在低溫中,讓MSC與PF127充份混合後,將細胞與材料支架的混合物以18號針頭注射入生物體內,經過一定時間的觀察其組織恢復情況。
本研究的重點在以先前在免子上的研究實驗方法及結果為基礎,在不同物種(狗、豬)的生物體,使用相同的材料及實驗方法,經過一定時間的觀察組織恢復的情況,來評估PF127在組織工程領域正式落實在臨床應用的可行性。
由實驗結果知:在不同物種(狗、豬),經rhBMP-2基因組織工程之自體骨幹細胞來修復牙周骨缺陷時,皆可有效增加牙周骨與牙周組織重建與再生的質與量,並可得到較佳之牙周組織結構。此實驗證明此方法及材料可在不同物種之生物體上皆獲得正向之結果,故可以期待PF127及體外引導間質幹細胞分化後最終成為骨組織的研究方法可早日實際在臨床醫療上使用。
In recent years, the development of tissue engineering technique has enabled as to create functional tissues with biocompatible, biodegradable polymers seeded with living cells. Tissue engineering is the application of the principles and modalities of engineering and the life sciences toward the basic perceptive of structure-function relationships in normal and pathological mammalian tissues and the growth of biological substitutes to re-establish, retain or improve functions. During the four decades, the international products of tissue engineering research is rewarding, but htere is lack of series of publication by testing small to large mammalian animals to prove the reality in practical possible clinical application.
It has been proved that mesenchymal stem cells which may develop into various cellular lineage capable of producing bone, muscle, cartilage, fat, and fibrous tissue. Bone Marrow Mesenchymal stem cells (MSCs) have been widely employed in the field of tissue engineering as primary cell source. In this dissertation, we use autologous MSCs to avoid the immunological reactions. MSCs are easly to cultivate to the desired cellular amounts. Furthermore, we utilize human recombinant bone morphogenetic protein-2 (rhBMP-2) for its oseto-induction ability to regenerate bony tissues.
Thermoreversible Pluronic F127 (FP127) has been approve by the FDA as medical polymers. The characters FP127 are liquid form in lower temperature, while becoming gel-like near human body tempateraure of 37℃. In practise it is easy to mixed MSCs evenly with FP127 in lower temperature settings, then the construct composed of cell and biomaterials, was injected to the animal hosts. the observation is carried for 6 moths in vivo.
These researches were exploited and based on previous laboratory results to test the same constructs (MSCs mixed in FP127) on alveolar bony defects in different animals such as dogs and pigs. After 6 months in vivo, the regenerated periodontal attachment apparatus was evaluated histologically and the whole regenerated bone volume was scrutinized from three-dimensional computed tomography analysis. The constructs were analysed to investigate the possibilty of practical clinical applications.
Periodontal apparatus regeneration was significantly better in the BMP-2-expressing MSCs group. New cementum and Sharpey’s fibers were observed on the denuded root surfaces in the BMP-2-expressing MSCs group, whereas incomplete healing with localized root surface resorption was noted in the control group. Post implantation, BMP-2-expressing MSCs group showed significant increase in the bone regeneration than the control one. There were different biological regeneration in different animals.
Thus, the use of autologous BMP-2-expressing MSCs/ FP127 boosted bone and periodontal apparatus regeneration in aveolar periodontal defects. This de novo approach might be suitable for clinical maxilla or mandibular bone repair and periodontal apparatus repair. This modality of BMP-2-expressing MSCs/FP127 is successful in various animals. It is expectable that FP127 and BMP-2-expressing MSCs may be the final method in tissue engineering of bone and future clinical applications.
中文摘要 I
ABSTRACT III
目 錄 V
圖目錄 VII
表目錄 VIII
符號說明 IX
第一章、緒論 1
1-1 背景 1
1-2 組織工程 3
1-3 骨髓間質幹細胞 8
1-4 支架在組織工程中的作用 9
第二章、論文研究目的 15
第三章、研究方法 17
3-1 原料 17
3-2 儀器 18
3-3 耗材 18
3-4 實驗流程 20
3-5 實驗方法 21
第四章、結果與討論 33
4-1西方墨點法 33
4-2巨觀觀察 34
4-3 組織染色觀察 35
4-4 立體電腦斷層掃描 37
4-5 實驗討論 39
第五章、結論 42
參考文獻 43
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