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http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/6877完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 林?輝 | |
| dc.contributor.author | Yung-Hsin Cheng | en |
| dc.contributor.author | 鄭詠馨 | zh_TW |
| dc.date.accessioned | 2021-05-17T09:20:07Z | - |
| dc.date.available | 2017-06-27 | |
| dc.date.available | 2021-05-17T09:20:07Z | - |
| dc.date.copyright | 2012-06-27 | |
| dc.date.issued | 2012 | |
| dc.date.submitted | 2012-06-13 | |
| dc.identifier.citation | [1] Urban JP, Roberts S. Degeneration of the intervertebral disc. Arthritis Research & Therapy 2003; 5:120-30.
[2] Goupille P, Jayson MI, Valat JP, Freemont AJ. Matrix metalloproteinases: The clue to intervertebral disc degeneration? Spine 1998; 23(14):1612-26. [3] Roughley PJ. Biology of intervertebral disc aging and degeneration. Spine 2004; 29(23):2691-9 [4] Hadjipavlou AG, Tzermiadianos MN, Bogduk N, Zindrick MR. The pathophysiology of disc degeneration: a critical review. J Bone Joint Surg Br. 2008; 90:1261-70. [5] Nerlich AG, Schleicher ED, Boos N. Immunohistologic markers for age-related changes of human lumbar intervertebral discs. Spine 1997; 22:2781-95. [6] Urban JP, Smith S, Fairbank JC. Nutrition of the intervertebral disc. Spine 2004; 29(23):2700-9. [7] Roberts S, Caterson B, Menage J, Evans EH, Jaffray DC, Eisenstein SM. Matrix metalloproteinases and aggrecanase: their role in disorders of the human intervertebral disc. Spine 2000; 25:3005-13. [8] Anderson DG, Tannoury C. Molecular pathogenic factors in symptomatic disc degeneration. Spine J. 2005; 5:260-66. [9] Peng B, Hao J, Hou S, Wu W, Jiang D, Fu X, Yang Y. Possible pathogenesis of painful intervertebral disc degeneration. Spine 2006; 31:560-6. [10] An H, Boden SD, Kang J, Sandhu HS, Abdu W, Weinstein J. Summary statement: emerging technique for treatment of degenerative lumbar disc disease. Spine 2003; 28:24-5. [11] Lee CK. Accelerated degeneration of the segment adjacent to a lumbar fusion. Spine 1988; 13:375-7. [12] Yoon ST. Molecular therapy of the intervertebral disc. Spine J. 2005; 5:280-6. [13] Paesold G, Nerlich AG, Boos N. Biological treatment strategies for disc degeneration: potentials and shortcomings. Eur Spine J. 2007; 16:447-68. [14] Zhang Y, An HS, Tannoury C, Thonar EJ, Freedman MK, Anderson DG. Biological treatment for degenerative disc disease: implications for the field of physical medicine and rehabilitation. Am J Phys Med Rehabil 2008; 87:694-702. [15] Alini M, Roughley PJ, Antoniou J, Stoll T, Aebi M. A biological approach to treating disc degeneration: not for today, but maybe for tomorrow. Eur Spine J. 2002; 11:S215-20. [16] Cassinelli EH, Hall RA, Kang JD. Biochemistry of intervertebral disc degeneration and the potential for gene therapy applications. Spine J 2001; 1:205-14. [17] Thompson JP, Pearce RH, Schechter MT, Adams ME, Tsang IK, Bishop PB. Preliminary evaluation of a scheme for grading the gross morphology of the human intervertebral disc. Spine 1990; 15:411-5. [18] Roughley PJ. Biology of intervertebral disc aging and degeneration: involvement of the extracellular matrix. Spine 2004; 29:2691-9. [19] Kanemoto M, Hukuda S, Komiya Y, Katsuura A, Nishioka J. Immunohistochemical study of matrix metalloproteinase-3 and tissue inhibitor of metalloproteinase-1 human intervertebral discs. Spine 1996; 21:1-8. [20] Kang JD, Stefanovic-Racic M, McIntyre LA, Georgescu HI, Evans CH. Toward a biochemical understanding of human intervertebral disc degeneration and herniation. Contributions of nitric oxide, interleukins, prostaglandin E2, and matrix metalloproteinases. Spine (Phila Pa 1976). 1997; 22:1065-73. [21] Masuda K, Oegema TR Jr, An HS. Growth factors and treatment of intervertebral disc degeneration. Spine 2004; 29:2757-69. [22] Sowa G, Vadala G, Studer R, Kompel J, Iucu C, Georgescu H, et al. Characterization of intervertebral disc aging. Spine 2008; 33:1821-8. [23] Sobajima S, Shimer AL, Chadderdon RC, Kompel JF, Kim JS, Gilbertson LG et al. Quantitative analysis of gene expression in a rabbit model of intervertebral disc degeneration by real-time polymerase chain reaction. Spine J. 2005; 5:14-23. [24] Tertti M, Paajanen H, Laato M, Aho H, Komu M, Kormano M. Disc degeneration in magnetic resonance imaging. A comparative biochemical, histologic, and radiologic study in cadaver spines. Spine 1991; 16:629-34. [25] Zhao W, Zhao T, Chen Y, Ahokas RA, Sun Y. Zhao W, Zhao T, Chen Y, Ahokas RA, Sun Y. Oxidative stress mediates cardiac fibrosis by enhancing transforming growth factor-beta1 in hypertensive rats. Mol Cell Biochem. 2008; 317:43-50. [26] Sullivan DE, Ferris M, Pociask D, Brody AR. The latent form of TGFbeta(1) is induced by TNFalpha through an ERK specific pathway and is activated by asbestos-derived reactive oxygen species in vitro and in vivo. J Immunotoxicol. 2008; 5:145-9. [27] Bellocq A, Azoulay E, Marullo S, Flahault A, Fouqueray B, Philippe C, Cadranel J, Baud L. Reactive oxygen and nitrogen intermediates increase transforming growth factor-beta1 release from human epithelial alveolar cells through two different mechanisms. Am J Respir Cell Mol Biol. 1999; 21:128-36. [28] Liu RM, Gaston Pravia KA. Oxidative stress and glutathione in TGF-β-mediated fibrogenesis. Free Radic Biol Med. 2010; 48:1-15. [29] Allen RG, Tresini M. Oxidative stress and gene regulation. Free Radic Biol Med. 2000; 28:463-99. [30] Valko M, Leibfritz D, Moncol J, Cronin MT, Mazur M, Telser J. Free radicals and antioxidants in normal physiological functions and human disease. Int J Biochem Cell Biol. 2007; 39:44-84. [31] Finkel T, Holbrook NJ. Oxidants, oxidative stress and the biology of ageing. Nature. 2000; 408:239-47. [32] Ben-Porath I, Weinberg RA. The signals and pathways activating cellular senescence. Int J Biochem Cell Biol. 2005; 37:961-76. [33] Zhao CQ, Wang LM, Jiang LS, Dai LY. The cell biology of intervertebral disc aging and degeneration. Ageing Res Rev. 2007; 6:247-61. [34] Kim KW, Chung HN, Ha KY, Lee JS, Kim YY. Senescence mechanisms of nucleus pulposus chondrocytes in human intervertebral discs. Spine J. 2009; 9:658-66. [35] Gruber HE, Ingram JA, Norton HJ, Hanley EN Jr. Senescence in cells of the aging and degenerating intervertebral disc: immunolocalization of senescence-associated beta-galactosidase in human and sand rat discs. Spine 2007; 32:321-7. [36] Kim KW, Ha KY, Lee JS, Rhyu KW, An HS, Woo YK. The apoptotic effects of oxidative stress and antiapoptotic effects of caspase inhibitors on rat notochordal cells. Spine 2007; 32: 2443-48. [37] Earnshaw WC, Martins LM, Kaufmann SH. Mammalian caspases: structure, activation, substrates, and functions during apoptosis. Annu Rev Biochem. 1999; 68:383-424. [38] Nunez G, Benedict MA, Hu Y, Inohara N. Caspases: the proteases of the apoptotic pathway. Oncogene 1998; 17:3237-45. [39] Budihardjo I, Oliver H, Lutter M, Luo X, Wang X. Biochemical pathways of caspase activation during apoptosis. Annu Rev Cell Dev Biol. 1999; 15:269-90. [40] Higuchi Y. Chromosomal DNA fragmentation in apoptosis and necrosis induced by oxidative stress. Biochem Pharmacol. 2003; 66:1527-35. [41] Wei A, Brisby H, Chung SA, Diwan AD. Bone morphogenetic protein-7 protects human intervertebral disc cells in vitro from apoptosis. The spine journal 2008; 8: 466-474. [42] Pandey KB, Rizvi SI. Plant polyphenols as dietary antioxidants in human health and disease. Oxid Med Cell Longev. 2009; 2:270-8. [43] Kikuzaki H, Hisamoto M, Hirose K, Akiyama K, Taniguchi H. Antioxidant properties of ferulic acid and its related compounds. J Agric Food Chem. 2002; 50:2161-8. [44] Srinivasan M, Sudheer AR, Menon VP. Ferulic Acid therapeutic potential through its antioxidant property. J Clin Biochem Nutr. 2007; 40:92-100. [45] Rahman I, Biswas SK, Kirkham PA. Regulation of inflammation and redox signaling by dietary polyphenols. Biochem Pharmacol. 2006; 72:1439-52 [46] Kanski J, Aksenova M, Stoyanova A, Butterfield DA. Ferulic acid antioxidant protection against hydroxyl and peroxyl radical oxidation in synaptosomal and neuronal cell culture systems in vitro structure-activity studies. J Nutr Biochem. 2002; 13:273-281. [47] Sudheer AR, Muthukumaran S, Kalpana C, Srinivasan M, Menon VP. Protective effect of ferulic acid on nicotine-induced DNA damage and cellular changes in cultured rat peripheral blood lymphocytes: a comparison with N-acetylcysteine. Toxicol In Vitro. 2007; 21:576-85. [48] Prasad NR, Ramachandran S, Pugalendi KV, Menon VP. Ferulic acid inhibits UV-B–induced oxidative stress in human lymphocytes. Nutrition Research 2007; 27:559-64. [49] Srinivasan M, Sudheer AR, Pillai KR, Kumar PR, Sudhakaran PR, Menon VP. Influence of ferulic acid on γ-radiation induced DNA damage, lipid peroxidation and antioxidant status in primary culture of isolated rat hepatocytes. Toxicology. 2006; 228:249-58. [50] Balasubashini MS, Rukkumani R, Viswanathan P, Menon VP. Ferulic acid alleviates lipid peroxidation in diabetic rats. Phytother Res. 2004; 18:310-4. [51] Drury JL, Mooney DJ. Mooney. Hydrogels for tissue engineering: scaffold design variables and applications. Biomaterials 2003; 24:4337-51. [52] Ruel-Gariepy E, Leroux JC. In situ-formation hydrogels--review of temperature-sensitive systems. Eur J Pharm Biopharm. 2004; 58:409-26. [53] Ta HT, Dass CR, Dunstan DE. Injectable chitosan hydrogels for localised cancer therapy. J Control Release 2008; 126:205-16. [54] Bhattarai N, Ramay HR, Gunn J, Matsen FA, Zhang M. PEG-grafted chitosan as an injectable thermosensitive hydrogel for sustained protein release. J Control Release. 2005; 103:609-24. [55] Chenite A, Chaput C, Wang D, Combes C, Buschmann MD, Hoemann CD, et al. Novel injectable neutral solutions of chitosan form biodegradable gels in situ. Biomaterial 2000; 21:2155-61. [56] Chenite A, Buschmann M, Wang D, Chaput C, Kandani N. Reheological characterisation of thermogelling chitosan/glycerol-phospate solution. Carbohydrate Polymers 2001; 46:39-47. [57] Berger J, Reist M, Chenite A, Felt-Baeyens O, Mayer JM, Gurny R. Pseudo-thermosetting chitosan hydrogels for biomedical application. International Journal of Pharmaceutics 2005; 288:197-206. [58] Roughley P, Hoemann C, DesRosiers E, Mwale F, Antoniou J, Alini M. The potential of chitosan-based gels containing intervertebral disc cells for nucleus pulposus supplementation. Biomaterials 2006; 27:388-96. [59] Cheng YH, Yang SH, Su WY, Chen YC, Yang KC, Lin FH et al. Thermosensitive chitosan-gelatin-glycerol phosphate hydrogels as a cell carrier for nucleus pulposus regeneration: an in-vitro study. Tissue Eng Part A. 2010; 16:695-703. [60] Tang YF, Du YM, Hu XW, Shi XW, Kennedy JF. Rheological characterisation of a novel thermosensitive chitosan/poly(vinyl alcohol) blend hydrogel. Carbohydrate Polymers 2007; 67:491-9. [61] Bhattarai N, Gunn J, Zhang M. Chitosan-based hydrogels for controlled, localized drug delivery. Adv Drug Deliv Rev. 2010; 62:83-99. [62] Ruel-Gariepy E, Chenite A, Chaput C, Guirguis S, Leroux J. Characterization of thermosensitive chitosan gels for the sustained delivery of drugs. Int J Pharm. 2000; 203:89-98. [63] Ruel-Gariepy E, Shive M, Bichara A, Berrada M, Le Garrec D, Chenite A, et al. A thermosensitive chitosan-based hydrogel for the local delivery of paclitaxel. Eur J Pharm Biopharm. 2004; 57:53-63. [64] Wu J, Su ZG, Ma GH. A thermo- and pH-sensitive hydrogel composed of quaternized chitosan/glycerophosphate. Int J Pharm. 2006; 315:1-11. [65] Komeima K, Rogers BS, Lu L, Campochiaro PA. Antioxidants reduce cone cell death in a model of retinitis pigmentosa. Proc Natl Acad Sci U S A. 2006; 103:11300-5. [66] Fiorito C, Rienzo M, Crimi E, Rossiello R, Balestrieri ML, Casamassimi A et al. Antioxidants increase number of progenitor endothelial cells through multiple gene expression pathways. Free Radical Research 2008; 42:754-62. [67] Rao GN, Corson MA, Berk BC. Uric acid stimulates vascular smooth muscle cell proliferation by increasing platelet-derived growth factor A-chain expression. J Biol Chem. 1991; 266:8604-8. [68] Blokhina O, Virolainen E, Fagerstedt KV. Antioxidants, oxidative damage and oxygen deprivation stress: a review. Ann Bot. 2003; 91:179-94. [69] Masuda K, An HS. Growth factors and the intervertebral disc. The Spine Journal 2004; 4:330-40. [70] Lipson SJ, Muir H. Vertebral osteophyte formation in experimental disc degeneration. Morphologic and proteoglycan changes over time. Arthritis Rheum. 1980; 23:319-24. [71] Stern R, Kogan G, Jedrzejas MJ, Soltes L. The many ways to cleave hyaluronan. Biotechnol Adv. 2007; 25:537-57. [72] Walker MH, Anderson DG. Molecular basis of intervertebral disc degeneration. Spine J. 2004; 4:158-66. [73] Rodriguez E, Roughley P. Link protein can retard the degradation of hyaluronan in proteoglycan aggregates. Osteoarthritis Cartilage 2006; 14:823-9. [74] Hengartner MO. The biochemistry of apoptosis. Nature 2000; 407:770-6. [75] Li X, Phillips FM, An HS, Ellman M, Thonar EJ, Wu W et al. The Action of Resveratrol, a phytoestrogen found in grapes, on the intervertebral disc. Spine 2008; 33:2586-95. [76] Martindale JL, Holbrook NJ. Cellular response to oxidative stress: signaling for suicide and survival. J Cell Physiol. 2002; 192:1-15. [77] Barone E, Calabrese V, Mancuso C. Ferulic acid and its therapeutic potential as a hormetin for age-related diseases. Biogerontology 2009; 10:97-108. [78] Grimm T, Schafer A, Hogger P. Antioxidant activity and inhibition of matrix metalloproteinases by metabolites of maritime pine bark extract (pycnogenol). Free Radic Biol Med. 2004; 36:811-22. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/6877 | - |
| dc.description.abstract | 椎間盤退化與椎間盤突出及背痛有高度的相關性，這些病徵增加了健康照護的支出。椎間盤退化過程可分為五個階段，在退化的第一至二階段時，並沒有明顯的病徵出現，但可透過核磁共振或電腦斷層掃描檢查來追蹤，臨床上一般並不會在此階段給予治療。近來的文獻指出，活性氧自由基不僅會加速椎間盤退化的過程，且會造成髓核細胞的凋亡和細胞外基質的降解。阿魏酸是一種抗氧化物並且可以較穩定地存在於空氣中；阿魏酸被證實對於活性氧自由基所引起的相關疾病具有預防的效果。本研究的目的除了評估阿魏酸對於雙氧水所引起的氧化壓力導致髓核細胞傷害的可能治療效果外，並評估利用溫感性甲殼素/明膠/甘油磷酸水膠做為阿魏酸持續釋放早期治療椎間盤退化的可行性。
在本研究中的試驗結果指出，500 μM為阿魏酸對紐西蘭兔的髓核細胞的安全閥值，利用阿魏酸治療被雙氧水所引起的氧化壓力所傷害的髓核細胞，其aggrecan, type II collagen和BMP-7 的基因表現可以有顯著的提升，而MMP-3的表現量有顯著的下降，而硫酸化葡萄胺聚醣含量有顯著的上升，細胞凋亡的情形也有顯著的抑制。而以甲殼素/明膠/甘油磷酸水膠做為阿魏酸持續釋放的試驗中，阿魏酸可以從水膠中緩釋，包覆阿魏酸的水膠除了能提升被雙氧水所引起的氧化壓力所傷害的髓核細胞中aggrecan和type II collagen基因的表現量外，並能抑制MMP-3的表現量，而在硫酸化葡萄胺聚醣生成量及alcian blue的染色的結果指出，包覆阿魏酸的水膠可以使受傷害的髓核細胞恢復到正常髓核細胞的表現量，另外在caspase-3和TUNEL的染色結果上，也指出其細胞凋亡的情形可以被顯著抑制。在本研究中證明，阿魏酸可成功藉由N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide (EDC)/ N-hydroxysuccinimide (NHS)固定於甲殼素/明膠/甘油磷酸水膠上；在中性環境下，該水膠之成膠溫度為攝氏31.8度，而以阿魏酸固定於甲殼素/明膠/甘油磷酸上的水膠治療被雙氧水所引起的氧化壓力所傷害的髓核細胞，可顯著的提升受傷害細胞中aggrecan和type II collage的表現量，並抑制其MMP-3的表現量；此外，硫酸化葡萄胺聚醣生成量也能恢復到正常的水平，另外在caspase-3和TUNEL的染色結果中顯示，細胞凋亡的情形可以被有效的抑制。 綜合上述，本研究的試驗結果指出阿魏酸可做為髓核再生的治療分子，而溫感性甲殼素/明膠/甘油磷酸水膠可做為阿魏酸長期釋放的良好載體，將阿魏酸固定於甲殼素/明膠/甘油磷酸水膠上可有效延長釋放的時間；結合阿魏酸及溫感性甲殼素/明膠/甘油磷酸水膠顯然可以有效治療因氧化壓力所傷害的髓核細胞，在未來更可應用於髓核再生的微創手術中。 | zh_TW |
| dc.description.abstract | Disc degeneration is strongly associated with back pain and herniation that increase the costs of health care. The degeneration of intervertebral disc (IVD) could be divided into 5 stages. In the first and second stages, there are no significant symptoms but could be traced by magnetic resonance imaging or computed tomography-scan. Generally, no aggressive treatment would be processed in the clinics. Recent studies indicated that overproduction of reactive oxygen species (ROS) may accelerate the degenerative process of IVD and associate with apoptosis of nucleus pulposus (NP) cells and degradation of extracellular matrix. Ferulic acid (FA) is an excellent antioxidant and relatively stable in air. FA has been proven to have ability to prevent ROS-induced diseases. The object of the study was aimed to evaluate the possible therapeutic effect of FA on hydrogen peroxide (H2O2)-induced oxidative stress NP cells and the feasibility of use the thermosensitive chitosan/gelatin/glycerophosphate (C/G/GP) hydrogel as a sustained release system of FA for early treatment in IVD degeneration.
In the study, NP cells were harvested from the IVD of New Zealand rabbits. The results showed that 500 μM of FA might be the threshold to treat NP cells without cytotoxicity. Post-treatment of FA on H2O2-induced oxidative stress NP cells significantly up regulated the expression of aggrecan, type II collagen and BMP-7 and down regulated the expression of MMP-3 in mRNA level. Post-treatment of FA on H2O2-induced oxidative stress NP cells could restore the production of sulfated glycosaminoglycans (GAGs) and inhibit the apoptosis caused by H2O2. The results showed that the release of FA from C/G/GP hydrogel could decrease the H2O2-induced oxidative stress. Post-treatment of FA-incorporated C/G/GP hydrogel on H2O2-induced oxidative stress NP cells showed up-regulation of aggrecan and type II collagen and down-regulation of MMP-3 in mRNA level. The results of sulfated GAGs to DNA ratio and alcian blue staining revealed that the GAGs production of H2O2-induced oxidative stress NP cells could reach to normal level. The results of caspase-3 activity and TUNEL staining indicated that FA-incorporated C/G/GP hydrogel decreased the apoptosis of H2O2-induced oxidative stress NP cells. The results showed that FA was successfully immobilized on C/G/GP hydrogel by N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide (EDC) and N-hydroxysuccinimide (NHS) crosslinking method. The gelation temperature of the FA-immobilized C/G/GP hydrogel was 31.80 degree celsius under neutral pH. Post-treatment of FA-immobilized C/G/GP hydrogel on H2O2-induced oxidative stress NP cells showed down-regulation of MMP-3 and up-regulation aggrecan and type II collagen in mRNA level. The sulfated GAGs production of H2O2-induced oxidative stress NP cells could be increased to the normal level in the post-treatment of FA-immobilized C/G/GP hydrogel group. The results of caspase-3 activity and TUNEL staining showed that the apoptosis of H2O2-induced oxidative stress NP cells could be inhibited by post-treatment of FA-immobilized C/G/GP hydrogel. From the results of the study, FA could be used as a therapeutic molecule for NP regeneration and FA-incorporated C/G/GP hydrogel might be potentially applied as a long-term release system. The immobilization of FA on C/G/GP hydrogel could significantly prolong the release period of FA. These results suggest that combination of FA and thermosensitive C/G/GP hydrogel can treat NP cells from the damage caused by oxidative stress and may apply in minimally invasive surgery for NP regeneration in the future. | en |
| dc.description.provenance | Made available in DSpace on 2021-05-17T09:20:07Z (GMT). No. of bitstreams: 1 ntu-101-D97548005-1.pdf: 2679438 bytes, checksum: bae51fb24b925a08e156575e8886c33c (MD5) Previous issue date: 2012 | en |
| dc.description.tableofcontents | 中文摘要 i
ABSTRACT iii TABLE OF CONTENTS vi CHAPTER 1 INTRODUCTION 1 1.1 Structure and function of intervertebral disc 1 1.2 Intervertebral disc degeneration 5 1.3 Current treatment options for disc degeneration 7 CHAPTER 2 THEORETICAL BASIS 10 2.1 The pathophysiology of disc degeneration 10 2.2 Reactive oxygen species and oxidative stress 14 2.3 Polyphenol: ferulic acid 19 2.4 In situ forming hydrogel 22 2.5 The purpose of study 27 CHAPTER 3 MATERIALS AND METHODS 29 3.1 Isolation of nucleus pulposus cells 29 3.2 Cytotoxicity of ferulic acid on nucleus pulposus cells 30 3.3 Chemiluminescence assay for reactive oxygen species production 32 3.4 The effects of ferulic acid on hydrogen peroxide-induced oxidative stress nucleus pulposus cells 33 3.4.1 Induction of oxidative stress and ferulic acid treatment 33 3.4.2 RNA extraction and gene expression 33 3.4.3 Total DNA quantification 35 3.4.4 Caspase-3 activity 35 3.4.5 TUNEL staining 36 3.5 Thermosensitive chitosan/gelatin/β-glycerol phosphate (C/G/GP) hydrogel as a controlled release system of ferulic acid (FA) for nucleus pulposus regeneration 37 3.5.1 Preparation of thermosensitive C/G/GP hydrogel 37 3.5.2 Reheological characterization 38 3.5.3 In vitro FA release study 38 3.5.4 Induction of oxidative stress and FA treatment 39 3.5.5 RNA extraction and gene expression 40 3.5.6 Total DNA quantification 41 3.5.7 Sulfated glycosaminoglycan content 41 3.5.8 Alcian blue staining 42 3.5.9 Caspase-3 activity 42 3.5.10 TUNEL staining 43 3.6 The effects of thermosensitive ferulic acid-immobilized chitosan/gelatin/β-glycerol phosphate (FA-immobilized C/G/GP) hydrogel on nucleus pulposus cells under hydrogen peroxide-induced oxidative stress 44 3.6.1 Preparation of thermosensitive FA-immobilized C/G/GP hydrogel 44 3.6.2 Characterization of FA-immobilized gelatin 45 3.6.3 Reheological characterization 46 3.6.4 Cytotoxicity of thermosensitive FA-immobilized C/G/GP hydrogel on NP cells 46 3.6.5 In vitro FA release study 47 3.6.6 Induction of oxidative stress and FA-immobilized C/G/GP hydrogel treatment 48 3.6.7 RNA extraction and gene expression 49 3.6.8 Analysis of cell numbers 50 3.6.9 Sulfated glycosaminoglycan content 50 3.6.10 Caspase-3 activity 51 3.6.11 TUNEL staining 52 3.7 Statistical analysis 53 CHAPTER 4 RESULTS 54 4.1 Cytotoxicity of ferulic acid on nucleus pulposus cells 54 4.2 Reactive oxygen species scavenging effect 57 4.3 The effects of ferulic acid on hydrogen peroxide-induced oxidative stress nucleus pulposus cells 59 4.3.1 Gene expression 59 4.3.2 Sulfated glycosaminoglycan production 63 4.3.3 Caspase-3 activity 64 4.3.4 TUNEL staining 65 4.4 Thermosensitive chitosan/gelatin/β-glycerol phosphate (C/G/GP) hydrogel as a controlled release system of ferulic acid for nucleus pulposus regeneration 67 4.4.1 The release of FA from C/G/GP hydrogel 67 4.4.2 Gene expression 68 4.4.3 Sulfated glycosaminoglycan production 71 4.4.4 Alcian blue staining 72 4.4.5 Caspase-3 activity 73 4.4.6 TUNEL staining 74 4.5 The effects of thermosensitive ferulic acid-immobilized chitosan/gelatin/β-glycerol phosphate (FA-immobilized C/G/GP) hydrogel on nucleus pulposus cells under hydrogen peroxide-induced oxidative stress 76 4.5.1 TNBS assay 76 4.5.2 Reheological characterization 78 4.5.3 Cytotoxicity of thermosensitive FA-immobilized C/G/GP hydrogel on NP cells 80 4.5.4 The release of FA from FA-immobilized C/G/GP hydrogel 82 4.5.5 Gene expression 84 4.5.6 Sulfated glycosamninoglycan production 89 4.5.7 Caspase-3 activity 90 4.5.8 TUNEL staining 91 CHAPTER 5 DISCUSSIONS 93 5.1 The effects of ferulic acid on hydrogen peroxide-induced oxidative stress nucleus pulposus cells 95 5.2 Thermosensitive chitosan/gelatin/β-glycerol phosphate hydrogel as a controlled release system of ferulic acid for nucleus pulposus regeneration 99 5.3 The effects of thermosensitive ferulic acid-immobilized chitosan/gelatin/β-glycerol phosphate hydrogel on nucleus pulposus cells under hydrogen peroxide-induced oxidative stress 104 CHAPTER 6 CONCLUSSION 110 References 113 Curriculum Vitae 125 | |
| dc.language.iso | en | |
| dc.title | 溫感性甲殼素/明膠/甘油磷酸水膠做為阿魏酸持續釋放於髓核再生之應用 | zh_TW |
| dc.title | Thermosensitive Chitosan/Gelatin/Glycerol Phosphate Hydrogel as a Sustained Release System of Ferulic Acid for Nucleus Pulposus Regeneration | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 100-2 | |
| dc.description.degree | 博士 | |
| dc.contributor.oralexamcommittee | 鄭登貴,劉華昌,陳克紹,楊禎明,吳信志 | |
| dc.subject.keyword | 髓核,阿魏酸,氧化壓力,溫感性水膠,抗氧化劑, | zh_TW |
| dc.subject.keyword | nucleus pulposus,ferulic acid,oxidative stress,thermosensitive hydrogel,antioxidant, | en |
| dc.relation.page | 127 | |
| dc.rights.note | 同意授權(全球公開) | |
| dc.date.accepted | 2012-06-14 | |
| dc.contributor.author-college | 工學院 | zh_TW |
| dc.contributor.author-dept | 醫學工程學研究所 | zh_TW |
| 顯示於系所單位： | 醫學工程學研究所 | |
文件中的檔案：
| 檔案 | 大小 | 格式 | |
|---|---|---|---|
| ntu-101-1.pdf | 2.62 MB | Adobe PDF | 檢視/開啟 |
系統中的文件，除了特別指名其著作權條款之外，均受到著作權保護，並且保留所有的權利。