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論文中文名稱:以快速原型法製作幾丁聚醣複合材料之3D支架及其對於成骨組織修復之研究 [以論文名稱查詢館藏系統]
論文英文名稱:Using Rapid Prototyping System to Produce Chitosan Composite 3D Scaffolds for Bone Tissue Repair [以論文名稱查詢館藏系統]
院校名稱:臺北科技大學
學院名稱:工程學院
系所名稱:化學工程研究所
畢業學年度:100
出版年度:101
中文姓名:劉易昕
英文姓名:I-Hsin Liu
研究生學號:99738025
學位類別:碩士
語文別:中文
口試日期:2012-07-13
論文頁數:59
指導教授中文名:林忻怡
口試委員中文名:鍾仁傑;蔡偉博;王孟菊
中文關鍵詞:快速原型系統幾丁聚醣骨母細胞
英文關鍵詞:rapid prototyping systemchitosanosteoblast
論文中文摘要:快速原型系統(rapid prototyping (RP) system)可以製作出孔洞互連的多孔性支架,可使細胞均勻分布於支架中,並有利於細胞養分與代謝物運輸,刺激細胞生長、分化和形成細胞外間質(Extracellular Matrix, ECM)。幾丁聚醣可刺激骨細胞分化並促使骨骼生成,可應用於暫時性骨科補材或骨丁骨板。先前的研究少有以快速原型系統製作幾丁聚醣與其複合材料之多孔支架。本研究以快速原型技術,製作出幾丁聚醣(C)、幾丁聚醣與槴子素交聯(CG),以及幾丁聚醣與果膠交聯(CP)之多孔性支架,並與以冷凍乾燥法製成的幾丁聚醣支架(CFD)做對照組,比較不同樣品物理性質以及生物相容性之差異,來評估材料是否適合促進骨整合。
物理測試的結果顯示CFD這組的孔隙度較其它組來的大,所以造成其降解也較其它組快。CG、CP這兩組在拉伸及壓縮的強度都較其它組來得高。而生物相容性的部分,透過掃描式電子顯微鏡觀察骨母細胞的生長型態,以DNA、鹼性磷酸酶和膠原蛋白定量來測定細胞生長與分化,並利用Von kossa鈣質染色以及鈣質定量觀察礦化程度。實驗結果顯示,C,CG,CP等組在DNA、鈣質和膠原蛋白定量的測定都較CFD組為高。
由結果得知,在生物相容性的測試上,以快速原型技術製作的多孔性支架較冷凍乾燥法來得好,而複合材料的機械強度又較純幾丁聚醣好,因此,本研究所測試之樣品中,以快速原型技術製作CG和CP多孔性支架較為適合用於促進骨整合的材料。
論文英文摘要:Rapid prototyping (RP) systems produce porous scaffolds with interconnecting pores with evenly distributed cells. This is beneficial for nutritional and metabolic substance transportation to stimulate cell growth, differentiation, and production of extracellular matrix (ECM). Chitosan stimulates the differentiation of bone cells to promote bone growth and has application in temporary orthopedic filling materials, screws, and plates. Previous studies have neglected porous scaffolds made by RP systems with chitosan and composite materials. We have implemented RP technology to produce chitosan (C), chitosan-genipin cross-linked (CG), and chitosan-pectin cross-linked (CP) porous scaffolds. Further, we have compared them with lyophilization produced chitosan scaffolds (CFD) to identify the physical and biological compatibility characteristics differences to assess the suitability of the aforementioned materials to facilitate osseointegration.
Physical tests showed that the CFD material had the largest pore diameter when compared to other materials, which means that the degradation of CFD is comparatively faster. The CG and CP materials had the greatest elasticity, flexibility, and strength. Biocompatibility characteristics were determined through the observation of the osteoblast growth formed through a scanning electronic microscope, and growth and differentiation were obtained through the quantities of DNA, alkaline phosphatase, and collagen. Furthermore, the degree of pyritization was measured using Von Kossa calcium staining and calcium quantity. Results show that C, CG, and CP had higher measurements in DNA, calcium, and collagen when compared to CFD.
Results show that the biocompatibility of the materials made from RP techniques were better than by lyophilization and the mechanical strength of compound materials was better than pure chitosan. We have concluded that RP produced CG and CP porous scaffolds were optimal materials for the application in osseointegration.
論文目次:中文摘要.................................................i
英文摘要.................................................iii
誌謝.....................................................v
目錄.....................................................vi
表目錄...................................................ix
圖目綠...................................................x
第一章 緒論........................................1
1.1 前言..............................................1
1.2 研究動機..........................................1
第二章 文獻回顧....................................3
2.1 幾丁聚醣..........................................3
2.1.1 幾丁聚醣介紹..................................3
2.1.2 幾丁聚醣在骨組織之應用........................5
2.1.3 幾丁聚醣之交聯材料............................6
2.1.3.1 共價鍵交聯................................6
2.1.3.2 離子鍵交聯................................7
2.2 骨組織............................................9
2.2.1 骨母細胞......................................9
2.2.2 骨骼修復......................................11
2.3 快速原型系統......................................13
第三章 實驗材料與方法..............................15
3.1 實驗材料..........................................15
3.1.1 細胞來源......................................15
3.1.2 實驗藥品......................................15
3.1.3 實驗儀器......................................17
3.1.4 實驗溶液配置..................................19
3.2 實驗方法..........................................20
3.2.1 實驗設計......................................20
3.2.2 多孔性支架物理性質測試........................23
3.2.2.1 掃描式電子顯微鏡(SEM).....................23
3.2.2.2 纖維大小與纖維間距........................23
3.2.2.3 孔隙度測試(Porosity)......................23
3.2.2.4 降解測試(Degradation).....................23
3.2.2.5 壓縮測試(Compression).....................24
3.2.3 骨母細胞在支架上的活性測試....................24
3.2.3.1 種入細胞前樣本的前置處理..................24
3.2.3.2 細胞接種..................................25
3.2.3.3 掃描式電子顯微鏡 (SEM)...................25
3.2.3.4 鹼性磷酸酶活性 (ALP activity)............25
3.2.3.5 DNA 定量...................................26
3.2.3.6 膠原蛋白定量(Collagen)....................27
3.2.3.7 鈣質含量(Calcium).........................27
3.2.3.8 鈣質染色(Von Kossa).......................28
3.2.3.9 統計分析..................................28
第四章 結果與討論..................................30
4.1 多孔性支架物理性質測試............................30
4.1.1 掃描式電子顯微鏡 (SEM).......................30
4.1.2 纖維與纖維之間的大小..........................32
4.1.3 孔隙度測試(Porosity)..........................34
4.1.4 降解測試(Degradation).........................35
4.1.5 壓縮測試(Compression).........................36
4.2 骨母細胞在支架上的活性測試........................37
4.2.1 掃描式電子顯微鏡 (SEM).......................37
4.2.2 鹼性磷酸酶活性 (ALP activity)................42
4.2.3 DNA 定量.......................................43
4.2.4 膠原蛋白定量(Collagen)........................44
4.2.5 鈣質含量(Calcium).............................45
4.2.6 鈣質染色(Von Kossa)...........................46
第五章 結論........................................49
參考文獻.................................................50
論文參考文獻:[1] Khor, E. and L.Y. Lim, Implantable applications of chitin and chitosan.
Biomaterials, 2003. 24(13): p. 2339-49.
[2] Harish Prashanth, K.V. and R.N. Tharanathan, Chitin/chitosan: modifications and
their unlimited application potential - an overview. Trends in Food Science &
Technology 2007. 18: p. 117-131.
[3] Kim, I.Y., et al., Chitosan and its derivatives for tissue engineering applications.
Biotechnol Adv, 2008. 26(1): p. 1-21.
[4] Madihally, S.V. and H.W. Matthew, Porous chitosan scaffolds for tissue
engineering. Biomaterials, 1999. 20(12): p. 1133-42.
[5] Seol, Y.J., et al., Chitosan sponges as tissue engineering scaffolds for bone
formation. Biotechnol Lett, 2004. 26(13): p. 1037-41.
[6] Seeherman, H., R. Li, and J. Wozney, A review of preclinical program
development for evaluating injectable carriers for osteogenic factors. J Bone Joint
Surg Am, 2003. 85-A Suppl 3: p. 96-108.
[7] Di Martino, A., M. Sittinger, and M.V. Risbud, Chitosan: a versatile biopolymer
for orthopaedic tissue-engineering. Biomaterials, 2005. 26(30): p. 5983-90.
[8] Zhang, Y. and M. Zhang, Synthesis and characterization of macroporous
chitosan/calcium phosphate composite scaffolds for tissue engineering. J Biomed
Mater Res, 2001. 55(3): p. 304-12.
[9] Zhang, Y. and M. Zhang, Calcium phosphate/chitosan composite scaffolds for
controlled in vitro antibiotic drug release. J Biomed Mater Res, 2002. 62(3): p.
378-86.
[10] Zhang, Y. and M. Zhang, Three-dimensional macroporous calcium phosphate 51
bioceramics with nested chitosan sponges for load-bearing bone implants. J
Biomed Mater Res, 2002. 61(1): p. 1-8.
[11] Zhang, Y., et al., Calcium phosphate-chitosan composite scaffolds for bone tissue
engineering. Tissue Eng, 2003. 9(2): p. 337-45.
[12] Ge, Z., et al., Hydroxyapatite-chitin materials as potential tissue engineered
bone substitutes. Biomaterials, 2004. 25(6): p. 1049-58.
[13] Hu, Q., et al., Preparation and characterization of biodegradable
chitosan/hydroxyapatite nanocomposite rods via in situ hybridization: a potential
material as internal fixation of bone fracture. Biomaterials, 2004. 25(5): p.
779-85.
[14] Cai, K., et al., Surface modification of poly (D,L-lactic acid) with chitosan and
its effects on the culture of osteoblasts in vitro. J Biomed Mater Res, 2002. 60(3):
p. 398-404.
[15] Zhang, Y. and M. Zhang, Microstructural and mechanical characterization of
chitosan scaffolds reinforced by calcium phosphate. Non-crystalline solids, 2001.
282(2-3): p. 159-164.
[16] Hsieh, W.C., C.P. Chang, and S.M. Lin, Morphology and characterization of 3D
micro-porous structured chitosan scaffolds for tissue engineering. Colloids Surf
B Biointerfaces, 2007. 57(2): p. 250-5.
[17] 朱怡靜, 幾丁聚醣接枝胺基酸之多孔薄膜製備及其應用. 國立聯合大學化學
工程研究所 碩士論文, 1996.
[18] Bigi, A., et al., Stabilization of gelatin films by crosslinking with genipin.
Biomaterials, 2002. 23(24): p. 4827-32.
[19] Sung, H.W., et al., Feasibility study of a natural crosslinking reagent for
biological tissue fixation. J Biomed Mater Res, 1998. 42(4): p. 560-7.52
[20] Muzzarelli, R.A.A., Genipin-crosslinked chitosan hydrogels as biomedical and
pharmaceutical aids. Carbohydrate Polymers, 2009. 77: p. 1-9.
[21] Wang, L., et al., Chitosan-alginate PEC membrane as a wound dressing:
Assessment of incisional wound healing. J Biomed Mater Res, 2002. 63(5): p.
610-8.
[22] Dumitriu, S., Polysaccharide Book for Medicinal Application. Marcel Dekker
Inc, New York, 1998.
[23] 蔡政翰, 以化學修飾法改進幾丁聚醣之溶解度. 國立台灣大學食品科技研究
所 碩士論文, 1996.
[24] 葉志宗, 以幾丁聚醣為基質製備應用於藥物釋放之組織工程多孔性支架. 國
立台北科技大學化學工程研究所 碩士論文, 2009.
[25] Sriamornsak, P. and S. Puttipipatkhachorn, Chitosan-pectin composite gel
spheres: Effect of some formulation variables on drug release. Macromolecular
Symposia, 2004. 216: p. 17-21.
[26] Elsabee, M.Z., et al., Surface modification of polypropylene films by chitosan and
chitosan/pectin multilayer. Carbohydrate Polymers, 2008. 71(2): p. 187-195.
[27] Aubin, J.E., et al., Osteoblast and chondroblast differentiation. Bone, 1995. 17(2
Suppl): p. 77S-83S.
[28] Lian, J.B. and G.S. Stein, Development of the osteoblast phenotype: molecular
mechanisms mediating osteoblast growth and differentiation. Iowa Orthop J,
1995. 15: p. 118-40.
[29] Intan Zarina, Z.A., et al., Osteoclast and osteoblast development of Mus
musculus haemopoietic Mononucleated cells. Biological Science, 2008. 8(3): p.
506-516.
[30] Phan, T.C., J. Xu, and M.H. Zheng, Interaction between osteoblast and 53
osteoclast: impact in bone disease. Histol Histopathol, 2004. 19(4): p. 1325-44.
[31] Lian, J.B. and G.S. Stein, Osteoporosis:chapter 6. Osteoblast Biology, 2008: p.
93-150.
[32] van't Hof, R.J. and S.H. Ralston, Nitric oxide and bone. Immunology, 2001.
103(3): p. 255-61.
[33] Suda, T., et al., Regulation of osteoclast function. J Bone Miner Res, 1997. 12(6):
p. 869-79.
[34] Raisz, L.G., Physiology and pathophysiology of bone remodeling. Clin Chem,
1999. 45(8 Pt 2): p. 1353-8.
[35] Bone remodeling. Encyclopæ dia Britannica, Inc
[36] Sachlos, E., et al., Novel collagen scaffolds with predefined internal morphology
made by solid freeform fabrication. Biomaterials, 2003. 24(8): p. 1487-97.
[37] Sachlos, E. and J.T. Czernuszka, Making tissue engineering scaffolds work.
Review: the application of solid freeform fabrication technology to the
production of tissue engineering scaffolds. Eur Cell Mater, 2003. 5: p. 29-39;
discussion 39-40.
[38] Li, J.P., et al., Porous Ti6Al4V scaffold directly fabricating by rapid prototyping:
preparation and in vitro experiment. Biomaterials, 2006. 27(8): p. 1223-35.
[39] Woodfield, T.B., et al., Design of porous scaffolds for cartilage tissue
engineering using a three-dimensional fiber-deposition technique. Biomaterials,
2004. 25(18): p. 4149-61.
[40] Yeong, W.Y., et al., Rapid prototyping in tissue engineering: challenges and
potential. Trends Biotechnol, 2004. 22(12): p. 643-52.
[41] Mikos, A.G., et al., Preparation and characterization of poly(L-lactic acid) foam.
Polymer 1994. 35(5): p. 1068-1077.54
[42] Mooney, D.J., et al., Novel approach to fabricate porous sponges of
poly(D,L-lactic-co-glycolic acid) without the use of organic solvents.
Biomaterials, 1996. 17(14): p. 1417-22.
[43] Freed, L.E., et al., Biodegradable polymer scaffolds for tissue engineering.
Biotechnology (N Y), 1994. 12(7): p. 689-93.
[44] Lo, H., M.S. Ponticiello, and K.W. Leong, Fabrication of controlled release
biodegradable foams by phase separation. Tissue Eng, 1995. 1(1): p. 15-28.
[45] Thomson, R.C., et al., Fabrication of biodegradable polymer scaffolds to
engineer trabecular bone. J Biomater Sci Polym Ed, 1995. 7(1): p. 23-38.
[46] Whang, K., et al., A novel method to fabricate bioabsorbable scaffolds. Polymer
1995. 36(4): p. 837-842.
[47] Hsu, Y.Y., et al., Effect of polymer foam morphology and density on kinetics of in
vitro controlled release of isoniazid from compressed foam matrices. J Biomed
Mater Res, 1997. 35(1): p. 107-16.
[48] Zein, I., et al., Fused deposition modeling of novel scaffold architectures for
tissue engineering applications. Biomaterials, 2002. 23(4): p. 1169-85.
[49] Tan, K.H., et al., Scaffold development using selective laser sintering of
polyetheretherketone-hydroxyapatite biocomposite blends. Biomaterials, 2003.
24(18): p. 3115-23.
[50] Kim, S.S., et al., Survival and function of hepatocytes on a novel
three-dimensional synthetic biodegradable polymer scaffold with an intrinsic
network of channels. Ann Surg, 1998. 228(1): p. 8-13.
[51] Landers, R., et al., Fabrication of soft tissue engineering scaffolds by means of
rapid prototyping techniques. Materials Science, 2002. 37: p. 3107-3116.
[52] Rao, S.B. and C.P. Sharma, Use of chitosan as a biomaterial: studies on its safety and hemostatic potential. J Biomed Mater Res, 1997. 34(1): p. 21-8.
[53] Mi, F.L., et al., In vivo biocompatibility and degradability of a novel
injectable-chitosan-based implant. Biomaterials, 2002. 23(1): p. 181-91.
[54] Rosa, A.L. and M.M. Beloti, Development of the osteoblast phenotype of serial
cell subcultures from human bone marrow. Braz Dent J, 2005. 16(3): p. 225-30.
[55] Yao, C.H., et al., Biocompatibility and biodegradation of a bone composite
containing tricalcium phosphate and genipin crosslinked gelatin. J Biomed
Mater Res A, 2004. 69(4): p. 709-17.
[56] Braccini, I. and S. Perez, Molecular basis of C(2+)-induced gelation in alginates
and pectins: the egg-box model revisited. Biomacromolecules, 2001. 2(4): p.
1089-96.
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