樹脂改良玻璃離子體黏著劑（resin-modified glass-ionomer cement，RM-GICs）為玻璃離子體之改良型，發展已有數十年之歷史，至今於牙科臨床補綴上仍有廣泛地應用，RM-GIC其因樹脂的添加有效改良了原本玻璃離子體強度不足的缺點，但伴隨著亦降低了生物相容性；氧化釔安定氧化鋯（Yttrium stabilized Zirconia，YSZ）為具有良好生物相容性與機械強度的陶瓷材料，近年來被廣泛應用於生醫材料上。因此，本研究目的為探討藉由YSZ的添加對RM-GIC於機械強度與生物相容性的影響，期望藉由YSZ的優點讓RM-GIC擁有更良好的表現。
本實驗為添加不同比例之微米級YSZ至RM-GIC材料中，以光聚合製成試片進行機械強度以及生物相容性之評估實驗。以未添加YSZ之RM-GIC為控制組，機械強度為測量壓縮強度以及對徑壓縮強度之表現；生物相容性則將材料浸泡於不同時間點，選用L929老鼠纖維母細胞已浸泡液觀察細胞存活，以及浸泡後才料之表面細胞貼附型態，以及以觀察細胞內環氧化酶二型（COX-2）的表現與細胞外腫瘤壞子因子α（TNFα）的產量來評估材料與細胞發炎反應之相關性。
結果顯示，YSZ不論添加5%、10%或是20%，壓縮強度與對徑壓縮強度皆無統計上顯著性差異。而將細胞培養於添加5%與10% YSZ的RM-GIC 0-3天之浸泡液細胞存活率，相較於控制組有統計上顯著較多（p＜0.05）；從SEM觀察細胞貼附型態，於未浸泡與浸泡3天後之細胞貼附材料表面形態，添加YSZ之組別相較於控制組有較良好的細胞貼附型態；在發炎反應COX-2表現上，0-3天浸泡液培養細胞12小時時，添加10% YSZ之組別相較於控制組統計上有顯著較少的COX-2表現量（p＜0.05），但各組於細胞培養24小時後之TNFα產物則無顯著性差異。因此，本研究可得結論，添加10% YSZ之RM-GIC能在不影響機械強度之同時提高材料之生物相容性。

Resin-modified glass-ionomer cements (RM-GICs) containing a basic ion-leachable glass, a water soluble polymeric acid, organic monomer and an initiator system, was considered not as biocompatible as conventional GICs. This study, therefore, was focused on the effects, especially adverse cellular reactions, of yttria-stabilized zirconia (YSZ) addition on resin-modified GIC. YSZ powder was added into Fuji II LC (GC, Japan) in the ratios of 0%, 5%, 10%, and 20%, respectively, molded into columns (2 mm x 4 mm), and light-cured as instructed. There were no statistically significant differences between groups in their compressive strength and diametric tensile strengths, although the group of 20% YSZ addition showed slight reductions. When incubating L929 cells, a fibroblast cell line, with cured 0-3day RMGIC-YSZ eluates, YSZ addition showed a dose dependent improvement of cell vitality, with a 25% increase in 10% YSZ group compared with the control（p＜0.05）. Better cell attachment was observed under electronic microscope in the YSZ group than the control. The production of COX-2 by L929 cells when incubating with RM-GIC eluates, was curbed by the addition of YSZ, also in a dose-dependent pattern, but there were no statistically significant differences between groups in TNFα production . We concluded that the addition of YSZ is capable of improving biocompatibility of GIC without compromising mechanical strength.

致謝 ....................................................................... I
中文摘要 ................................................................ II
Abstract ................................................................ III
總目錄 .................................................................. IV
表目錄 ................................................................. VII
圖目錄 ................................................................. VIII

第一章 前言 .................................................................. 1
1-1 文獻回顧 ................................................................. 1
1-1-1 玻璃離子體 ......................................................... 1
1-1-2 牙科樹脂之生物相容性 ........................................... 3
1-1-3 氧化鋯 ............................................................... 6
1-2 研究目的 ................................................................. 8
第二章 材料與方法 ........................................................ 9
2-1 材料備製 ................................................................. 9
2-2 機械性質測試 ......................................................... 11
2-2-1 壓縮強度測試 ................................................... 11
2-2-2 對徑壓縮強度測試 ............................................. 11
2-3 材料晶相與表面觀察 ............................................... 12
2-3-1 材料晶相分析 ................................................... 12
2-3-2 材料表面觀察 ................................................... 12
2-4 生物相容性分析 ...................................................... 14
2-4-1 浸泡液配製 ...................................................... 14
2-4-2 pH值測量 ......................................................... 14
2-4-3 細胞培養 ......................................................... 14
2-4-4 細胞貼附 ......................................................... 14
2-4-5 細胞存活率 ...................................................... 15
2-4-6 蛋白質萃取與定量 ............................................. 15
2-4-7 環氧化酶二型（COX-2）表現 .................................. 17
2-4-8腫瘤壞死因子α（TNFα）Cytokine 釋放量 ................. 19
2-5統計分析 ................................................................. 20
第三章 結果 ................................................................. 21
3-1 機械性質 ................................................................ 21
3-1-1 壓縮強度 ......................................................... 21
3-1-2 對徑壓縮強度 ................................................... 22
3-2 材料性質與型態 ...................................................... 24
3-2-1 材料晶相分析 ................................................... 24
3-2-2 材料表面形態 ................................................... 26
3-3 生物相容性 ............................................................. 28
3-3-1 材料浸泡液pH值 ............................................... 28
3-3-2 細胞存活率 ...................................................... 30
3-3-3 細胞貼附觀察 ................................................... 32
3-3-4 環氧化酶二型（COX-2）表現 .................................. 36
3-3-5 腫瘤壞子因子α（TNFα）表現 ................................. 36
第四章 討論 ................................................................. 39
4-1添加YSZ對RM-GIC之機械強度影響 ............................ 39
4-2各組材料對細胞存活率與貼附之影響 .......................... 40
4-3 各組材料於發炎反應之探討 ...................................... 42
第五章 結論 ................................................................. 44
第六章 參考文獻 ........................................................... 45

1.Wilson, A.D. and B.E. Kent, A new translucent cement for dentistry. The glass ionomer cement. Br Dent J, 1972. 132(4): p. 133-5.
2.Mathis, R.S. and J.L. Ferracane, Properties of a glass-ionomer/resin-composite hybrid material. Dent Mater, 1989. 5(5): p. 355-8.
3.Bala, O., M. Uctasli, H. Can, E. Turkoz, and M. Can, Flouride release from various restorative materials. J Nihon Univ Sch Dent, 1997. 39(3): p. 123-7.
4.Shaw, A.J., T. Carrick, and J.F. McCabe, Fluoride release from glass-ionomer and compomer restorative materials: 6-month data. J Dent, 1998. 26(4): p. 355-9.
5.Wilson, A.D., Resin-modified glass-ionomer cements. Int J Prosthodont, 1990. 3(5): p. 425-9.
6.Musa, A., G.J. Pearson, and M. Gelbier, In vitro investigation of fluoride ion release from four resin-modified glass polyalkenoate cements. Biomaterials, 1996. 17(10): p. 1019-23.
7.Wilson, A.D., Developments in glass-ionomer cements. Int J Prosthodont, 1989. 2(5): p. 438-46.
8.Zhang, Y.H., [New dental filling material: silver alloy glass ionomer cement]. Zhonghua Hu Li Za Zhi, 1993. 28(9): p. 541-3.
9.Moshaverinia, A., S. Ansari, M. Moshaverinia, N. Roohpour, J.A. Darr, and I. Rehman, Effects of incorporation of hydroxyapatite and fluoroapatite nanobioceramics into conventional glass ionomer cements (GIC). Acta Biomater, 2008. 4(2): p. 432-40.
10.Gu, Y.W., A.U. Yap, P. Cheang, and K.A. Khor, Effects of incorporation of HA/ZrO(2) into glass ionomer cement (GIC). Biomaterials, 2005. 26(7): p. 713-20.
11.Ana, I.D., S. Matsuya, M. Ohta, and K. Ishikawa, Effects of added bioactive glass on the setting and mechanical properties of resin-modified glass ionomer cement. Biomaterials, 2003. 24(18): p. 3061-7.
12.Hurrell-Gillingham, K., I.M. Reaney, I. Brook, and P.V. Hatton, In vitro biocompatibility of a novel Fe2O3 based glass ionomer cement. J Dent, 2006. 34(8): p. 533-8.
13.Sasanaluckit, P., K.R. Albustany, P.J. Doherty, and D.F. Williams, Biocompatibility of glass ionomer cements. Biomaterials, 1993. 14(12): p. 906-16.
14.Six, N., J.J. Lasfargues, and M. Goldberg, In vivo study of the pulp reaction to Fuji IX, a glass ionomer cement. Journal of Dentistry, 2000. 28(6): p. 413-22.
15.Souza, P.P., A.M. Aranha, J. Hebling, E.M. Giro, and C.A. Costa, In vitro cytotoxicity and in vivo biocompatibility of contemporary resin-modified glass-ionomer cements. Dent Mater, 2006. 22(9): p. 838-44.
16.Nicholson, J.W. and B. Czarnecka, The biocompatibility of resin-modified glass-ionomer cements for dentistry. Dent Mater, 2008. 24(12): p. 1702-8.
17.de Souza Costa, C.A., J. Hebling, F. Garcia-Godoy, and C.T. Hanks, In vitro cytotoxicity of five glass-ionomer cements. Biomaterials, 2003. 24(21): p. 3853-8.
18.Aranha, A.M., E.M. Giro, P.P. Souza, J. Hebling, and C.A. de Souza Costa, Effect of curing regime on the cytotoxicity of resin-modified glass-ionomer lining cements applied to an odontoblast-cell line. Dent Mater, 2006. 22(9): p. 864-9.
19.Limapornvanich, A., S. Jitpukdeebodintra, C. Hengtrakool, and U. Kedjarune-Leggat, Bovine serum albumin release from novel chitosan-fluoro-aluminosilicate glass ionomer cement: stability and cytotoxicity studies. J Dent, 2009. 37(9): p. 686-90.
20.Kanerva, L., R. Jolanki, T. Leino, and T. Estlander, Occupational allergic contact dermatitis from 2-hydroxyethyl methacrylate and ethylene glycol dimethacrylate in a modified acrylic structural adhesive. Contact Dermatitis, 1995. 33(2): p. 84-9.
21.Lindstrom, M., K. Alanko, H. Keskinen, and L. Kanerva, Dentist''s occupational asthma, rhinoconjunctivitis, and allergic contact dermatitis from methacrylates. Allergy, 2002. 57(6): p. 543-5.
22.Bouillaguet, S., M. Virgillito, J. Wataha, B. Ciucchi, and J. Holz, The influence of dentine permeability on cytotoxicity of four dentine bonding systems, in vitro. J Oral Rehabil, 1998. 25(1): p. 45-51.
23.Gerzina, T.M. and W.R. Hume, Diffusion of monomers from bonding resin-resin composite combinations through dentine in vitro. J Dent, 1996. 24(1-2): p. 125-8.
24.Olgart, L., L. Edwall, and B. Gazelius, Involvement of afferent nerves in pulpal blood-flow reactions in response to clinical and experimental procedures in the cat. Arch Oral Biol, 1991. 36(8): p. 575-81.
25.Bergenholtz, G., S. Nagaoka, and M. Jontell, Class II antigen expressing cells in experimentally induced pulpitis. Int Endod J, 1991. 24(1): p. 8-14.
26.Huang, F.M. and Y.C. Chang, Induction of cyclooxygenase-2 mRNA and protein expression by dentin bonding agents in human gingival fibroblasts. J Biomed Mater Res B Appl Biomater, 2004. 70(2): p. 297-302.
27.Huang, F.M., C.H. Tsai, S.J. Ding, and Y.C. Chang, Induction of cyclooxygenase-2 expression in human pulp cells stimulated by dentin bonding agents. Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 2005. 100(4): p. 501-6.
28.Huang, F.M. and Y.C. Chang, Prevention of the epoxy resin-based root canal sealers-induced cyclooxygenase-2 expression and cytotoxicity of human osteoblastic cells by various antioxidants. Biomaterials, 2005. 26(14): p. 1849-55.
29.Cohen, J.S., A. Reader, R. Fertel, M. Beck, and W.J. Meyers, A radioimmunoassay determination of the concentrations of prostaglandins E2 and F2alpha in painful and asymptomatic human dental pulps. J Endod, 1985. 11(8): p. 330-5.
30.Miyauchi, M., T. Takata, H. Ito, I. Ogawa, J. Kobayashi, H. Nikai, and N. Ijuhin, Immunohistochemical demonstration of prostaglandins E2, F2 alpha, and 6-keto-prostaglandin F1 alpha in rat dental pulp with experimentally induced inflammation. J Endod, 1996. 22(11): p. 600-2.
31.Huang, G.T., A.P. Potente, J.W. Kim, N. Chugal, and X. Zhang, Increased interleukin-8 expression in inflamed human dental pulps. Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 1999. 88(2): p. 214-20.
32.Newton, R., L.M. Kuitert, M. Bergmann, I.M. Adcock, and P.J. Barnes, Evidence for involvement of NF-kappaB in the transcriptional control of COX-2 gene expression by IL-1beta. Biochem Biophys Res Commun, 1997. 237(1): p. 28-32.
33.Darville, M.I. and D.L. Eizirik, Regulation by cytokines of the inducible nitric oxide synthase promoter in insulin-producing cells. Diabetologia, 1998. 41(9): p. 1101-8.
34.Smith, W.L., D.L. DeWitt, and R.M. Garavito, Cyclooxygenases: structural, cellular, and molecular biology. Annu Rev Biochem, 2000. 69: p. 145-82.
35.Issa, Y., D.C. Watts, P.A. Brunton, C.M. Waters, and A.J. Duxbury, Resin composite monomers alter MTT and LDH activity of human gingival fibroblasts in vitro. Dent Mater, 2004. 20(1): p. 12-20.
36.Ratanasathien, S., J.C. Wataha, C.T. Hanks, and J.B. Dennison, Cytotoxic interactive effects of dentin bonding components on mouse fibroblasts. J Dent Res, 1995. 74(9): p. 1602-6.
37.Yoshii, E., Cytotoxic effects of acrylates and methacrylates: relationships of monomer structures and cytotoxicity. J Biomed Mater Res, 1997. 37(4): p. 517-24.
38.Geurtsen, W., W. Spahl, and G. Leyhausen, Residual monomer/additive release and variability in cytotoxicity of light-curing glass-ionomer cements and compomers. J Dent Res, 1998. 77(12): p. 2012-9.
39.Geurtsen, W., W. Spahl, K. Muller, and G. Leyhausen, Aqueous extracts from dentin adhesives contain cytotoxic chemicals. J Biomed Mater Res, 1999. 48(6): p. 772-7.
40.Palmer, G., H.M. Anstice, and G.J. Pearson, The effect of curing regime on the release of hydroxyethyl methacrylate (HEMA) from resin-modified glass-ionomer cements. J Dent, 1999. 27(4): p. 303-11.
41.Thonemann, B., G. Schmalz, K.A. Hiller, and H. Schweikl, Responses of L929 mouse fibroblasts, primary and immortalized bovine dental papilla-derived cell lines to dental resin components. Dent Mater, 2002. 18(4): p. 318-23.
42.Geurtsen, W., Biocompatibility of resin-modified filling materials. Crit Rev Oral Biol Med, 2000. 11(3): p. 333-55.
43.About, I., J. Camps, T.A. Mitsiadis, M.J. Bottero, W. Butler, and J.C. Franquin, Influence of resinous monomers on the differentiation in vitro of human pulp cells into odontoblasts. J Biomed Mater Res, 2002. 63(4): p. 418-23.
44.Kaga, M., M. Noda, J.L. Ferracane, W. Nakamura, H. Oguchi, and H. Sano, The in vitro cytotoxicity of eluates from dentin bonding resins and their effect on tyrosine phosphorylation of L929 cells. Dent Mater, 2001. 17(4): p. 333-9.
45.Schweikl, H., G. Schmalz, and T. Spruss, The induction of micronuclei in vitro by unpolymerized resin monomers. J Dent Res, 2001. 80(7): p. 1615-20.
46.Geurtsen, W., F. Lehmann, W. Spahl, and G. Leyhausen, Cytotoxicity of 35 dental resin composite monomers/additives in permanent 3T3 and three human primary fibroblast cultures. J Biomed Mater Res, 1998. 41(3): p. 474-80.
47.Hanks, C.T., S.E. Strawn, J.C. Wataha, and R.G. Craig, Cytotoxic effects of resin components on cultured mammalian fibroblasts. J Dent Res, 1991. 70(11): p. 1450-5.
48.Chang, M.C., Y.S. Ho, P.H. Lee, C.P. Chan, J.J. Lee, L.J. Hahn, Y.J. Wang, and J.H. Jeng, Areca nut extract and arecoline induced the cell cycle arrest but not apoptosis of cultured oral KB epithelial cells: association of glutathione, reactive oxygen species and mitochondrial membrane potential. Carcinogenesis, 2001. 22(9): p. 1527-35.
49.Eastman, A. and J.R. Rigas, Modulation of apoptosis signaling pathways and cell cycle regulation. Semin Oncol, 1999. 26(5 Suppl 16): p. 7-16; discussion 41-2.
50.Schweikl, H., G. Spagnuolo, and G. Schmalz, Genetic and cellular toxicology of dental resin monomers. J Dent Res, 2006. 85(10): p. 870-7.
51.Spagnuolo, G., V. D''Anto, C. Cosentino, G. Schmalz, H. Schweikl, and S. Rengo, Effect of N-acetyl-L-cysteine on ROS production and cell death caused by HEMA in human primary gingival fibroblasts. Biomaterials, 2006. 27(9): p. 1803-9.
52.Clutton, S., The importance of oxidative stress in apoptosis. Br Med Bull, 1997. 53(3): p. 662-8.
53.Schnelldorfer, T., S. Gansauge, F. Gansauge, S. Schlosser, H.G. Beger, and A.K. Nussler, Glutathione depletion causes cell growth inhibition and enhanced apoptosis in pancreatic cancer cells. Cancer, 2000. 89(7): p. 1440-7.
54.Chang, H.H., M.K. Guo, F.H. Kasten, M.C. Chang, G.F. Huang, Y.L. Wang, R.S. Wang, and J.H. Jeng, Stimulation of glutathione depletion, ROS production and cell cycle arrest of dental pulp cells and gingival epithelial cells by HEMA. Biomaterials, 2005. 26(7): p. 745-53.
55.Eckhardt, A., N. Gerstmayr, K.A. Hiller, C. Bolay, C. Waha, G. Spagnuolo, C. Camargo, G. Schmalz, and H. Schweikl, TEGDMA-induced oxidative DNA damage and activation of ATM and MAP kinases. Biomaterials, 2009. 30(11): p. 2006-14.
56.Urcan, E., H. Scherthan, M. Styllou, U. Haertel, R. Hickel, and F.X. Reichl, Induction of DNA double-strand breaks in primary gingival fibroblasts by exposure to dental resin composites. Biomaterials, 2010. 31(8): p. 2010-4.
57.Niki, E., Free radical pathology and antioxidants: overview. J Nutr Sci Vitaminol (Tokyo), 1992. Spec No: p. 538-40.
58.Kelly, J.R. and I. Denry, Stabilized zirconia as a structural ceramic: an overview. Dent Mater, 2008. 24(3): p. 289-98.
59.Piconi, C. and G. Maccauro, Zirconia as a ceramic biomaterial. Biomaterials, 1999. 20(1): p. 1-25.
60.Hulbert, S.F., S.J. Morrison, and J.J. Klawitter, Tissue reaction to three ceramics of porous and non-porous structures. J Biomed Mater Res, 1972. 6(5): p. 347-74.
61.Harms, J. and E. Mausle, Tissue reaction to ceramic implant material. J Biomed Mater Res, 1979. 13(1): p. 67-87.
62.Li, J., Y. Liu, L. Hermansson, and R. Soremark, Evaluation of biocompatibility of various ceramic powders with human fibroblasts in vitro. Clin Mater, 1993. 12(4): p. 197-201.
63.Warashina, H., S. Sakano, S. Kitamura, K.I. Yamauchi, J. Yamaguchi, N. Ishiguro, and Y. Hasegawa, Biological reaction to alumina, zirconia, titanium and polyethylene particles implanted onto murine calvaria. Biomaterials, 2003. 24(21): p. 3655-61.
64.Roualdes, O., M.E. Duclos, D. Gutknecht, L. Frappart, J. Chevalier, and D.J. Hartmann, In vitro and in vivo evaluation of an alumina-zirconia composite for arthroplasty applications. Biomaterials, 2010. 31(8): p. 2043-54.
65.Yelamanchili, A. and B.W. Darvell, Network competition in a resin-modified glass-ionomer cement. Dent Mater, 2008. 24(8): p. 1065-9.
66.Gu, Y.W., Zirconia–glass ionomer cement––a potential substitute for Miracle Mix. Scripta Materialia, 2005: p. 113-116.
67.Woolford, M.J. and R.G. Chadwick, Surface pH of resin-modified glass polyalkenoate (ionomer) cements. J Dent, 1992. 20(6): p. 359-64.
68.Bourke, A.M., A.W. Walls, and J.F. McCabe, Light-activated glass polyalkenoate (ionomer) cements: the setting reaction. J Dent, 1992. 20(2): p. 115-20.
69.Kakaboura, A., G. Eliades, and G. Palaghias, An FTIR study on the setting mechanism of resin-modified glass ionomer restoratives. Dent Mater, 1996. 12(3): p. 173-8.
70.Wan, A.C., A.U. Yap, and G.W. Hastings, Acid-base complex reactions in resin-modified and conventional glass ionomer cements. J Biomed Mater Res, 1999. 48(5): p. 700-4.
71.Lee, D.H., N.R. Kim, B.S. Lim, Y.K. Lee, and H.C. Yang, Effects of TEGDMA and HEMA on the expression of COX-2 and iNOS in cultured murine macrophage cells. Dent Mater, 2009. 25(2): p. 240-6.
72.Krifka, S., C. Petzel, K.A. Hiller, E.M. Frank, C. Bosl, G. Spagnuolo, F.X. Reichl, G. Schmalz, and H. Schweikl, Resin monomer-induced differential activation of MAP kinases and apoptosis in mouse macrophages and human pulp cells. Biomaterials, 2010. 31(11): p. 2964-75.
73.Noda, M., J.C. Wataha, P.E. Lockwood, K.R. Volkmann, M. Kaga, and H. Sano, Sublethal, 2-week exposures of dental material components alter TNF-alpha secretion of THP-1 monocytes. Dent Mater, 2003. 19(2): p. 101-5.
74.Lewis, T.S., P.S. Shapiro, and N.G. Ahn, Signal transduction through MAP kinase cascades. Advances in Cancer Research, 1998. 74: p. 49-139.
75.Davis, R.J., Signal transduction by the JNK group of MAP kinases. Cell, 2000. 103(2): p. 239-52.
76.Zhang, W. and H.T. Liu, MAPK signal pathways in the regulation of cell proliferation in mammalian cells. Cell Res, 2002. 12(1): p. 9-18.
77.Chang, M.C., L.D. Lin, C.P. Chan, H.H. Chang, L.I. Chen, H.J. Lin, H.W. Yeh, W.Y. Tseng, P.S. Lin, C.C. Lin, and J.H. Jeng, The effect of BisGMA on cyclooxygenase-2 expression, PGE2 production and cytotoxicity via reactive oxygen species- and MEK/ERK-dependent and -independent pathways. Biomaterials, 2009. 30(25): p. 4070-7.
78.Samuelsen, J.T., J.E. Dahl, S. Karlsson, E. Morisbak, and R. Becher, Apoptosis induced by the monomers HEMA and TEGDMA involves formation of ROS and differential activation of the MAP-kinases p38, JNK and ERK. Dent Mater, 2007. 23(1): p. 34-9.