(3.238.186.43) 您好!臺灣時間:2021/02/26 13:04
字體大小: 字級放大   字級縮小   預設字形  
回查詢結果

詳目顯示:::

我願授權國圖
: 
twitterline
研究生:柳青青
研究生(外文):Ching-Ching Liu
論文名稱:研發搭載Tideglusib之雙相型透明質酸膠體在牙本質牙髓組織再生的應用
論文名稱(外文):Development of Biphasic Hyaluronic Acid Gel with Tideglusib in Dentin-Pulp Tissue Regeneration
指導教授:李苑玲
指導教授(外文):Yuan-Ling Lee
口試委員:林峯輝李伯訓
口試委員(外文):Feng-Huei LinBor-Shiunn Lee
口試日期:2019-07-04
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:2019
畢業學年度:107
語文別:中文
論文頁數:91
中文關鍵詞:透明質酸膠體交聯反應環境粒徑大小物理性質Tideglusib生物相容性細胞分化動物模型纖維組織生成血管新生
DOI:10.6342/NTU201903468
相關次數:
  • 被引用被引用:0
  • 點閱點閱:19
  • 評分評分:系統版面圖檔系統版面圖檔系統版面圖檔系統版面圖檔系統版面圖檔
  • 下載下載:0
  • 收藏至我的研究室書目清單書目收藏:0
當牙髓發炎亦或感染時,清創根管後若能保留甚至再生活髓組織,維持牙齒感知外界刺激的功能,並使齒質持續生長,是現今牙髓疾病治療的目標。Tideglusib是FDA認證可於臨床治療阿茲罕默症的藥物,本身為肝醣合成酶激酶(glycogen synthase kinase-3, GSK-3)抑制劑,會參與牙齒生長發育過程並促進牙本質再生(dentinogenesis)。本研究目的是以人類牙髓幹細胞(human dental pulp stem cell, hDPSC)作為細胞來源,使用Tideglusib調控細胞的生長效應,同時以雙相型透明質酸膠體顆粒(biphasic hyaluronic acid granules, biHAG)作為支架,研發出可注射性的組織再生材料,誘導牙本質牙髓複合體(dentin-pulp complex)的再生。
支架材料是使用BDDE(1,4-butanediol diglycidyl ether)作為交聯劑製備出透明質酸膠體顆粒(hyaluronic acid granules, HAG),改變交聯反應環境和降低粒徑大小以改良製備流程,並與非交聯型透明質酸(non-crosslinked hyaluronic acid, ncHA)混合製備出biHAG,分別以HAG比例命名為HAG100、HAG80和HAG60,以流變性質及膨脹比例評估製程因素對物理性質的影響。同時使用hDPSC與材料共同培養,觀察材料生物相容性和細胞生長行為。也利用反轉錄聚合酶連鎖反應(reverse transcription-polymerase chain reaction, RT-PCR)檢測不同Tideglusib濃度對細胞分化之效應。動物實驗則將各組材料與hDPSC混合後進行免疫抑制鼠皮下注射,六周後犧牲,觀察材料在活體組織的代謝分解行為和組織反應。
物理性質分析發現,當交聯反應環境自烘箱改為水浴槽,材料的流體性質和均質性增加,但不影響交聯程度。而粒徑大小方面,結果顯示250 μm膠體材料的固態性質、流動性和可注射性較420 μm者高,其中250 μm HAG80、HAG60可通過27G的針頭進行注射,並且HAG80的結構維持性較HAG60穩定,因此選用HAG80進行後續研究。
細胞實驗方面,結果顯示Tideglusib濃度小於300 nM時,細胞存活率都超過八成。而搭載Tideglusib的Tide@HAG具備良好的生物相容性,並且使用萃取液模型檢測出透明質酸膠體顆粒並不會影響藥物釋放,亦不具備緩釋效果,約一天內會釋放所承載的藥物量。而與hDPSC共養的研究中,粒徑大小不會影響細胞的生長行為,所有組別的細胞形態正常無破孔,無論粒徑大小,添加100 nM Tideglusib有較好的細胞生長行為。檢測細胞分化發現,添加100 nM Tideglusib可提升DSPP及VEGF的基因表現,因此選用100 nM Tideglusib作為活體實驗的添加濃度。
而動物實驗結果顯示,所有組別的材料生物相容性良好,不會造成發炎反應。在不加藥的420 μm組別中,HAG100材料完整,沒有明顯纖維組織生成,HAG80和HAG60的材料間都有纖維組織和血管形成。粒徑大小方面,420 μm和250 μm材料其組織生長及血管增生情狀差異不大。而無論粒徑大小,添加100 nM Tideglusib組別相較於未加藥者,都有較多的組織生長量及血管形成,其中250 μm的組別在組織生成有顯著差異。
總結來說,此研發材料在改變交聯反應環境為水浴槽後,可提升流動性跟均質性;而降低粒徑大小可提升可注射性,其中以HAG80的結構維持性較佳。Tide@HAG生物相容性佳,搭載100 nM Tideglusib對細胞生長以及分化有提升的趨勢。活體組織反應觀察到,添加ncHA和Tideglusib都會促進纖維組織和血管生成,其中又以250 μm的組別較具備牙本質牙髓複合體再生的潛力。
Maintain the pulp vitality or regenerate the dentin-pulp complex to keep the tooth perception to external stimuli and root growth is the goal of modern endodontic treatment. Tideglusib, a FDA-approved drug for Alzheimer''s disease treatment, is a glycogen synthase kinase-3 (GSK-3) inhibitor that participates in tooth growth and promotes dentinogenesis. The purpose of this study was to develop an injectable, Tideglusib contained biphasic hyaluronic acid granules (biHAG) with human dental pulp stem cells (hDPSC) as cell sources for dentin-pulp complex regeneration.
The 2% cross-linked hyaluronic acid granules (HAG) with granular size of 420 μm and 250 μm were synthesized by using BDDE (1,4-butanediol diglycidyl ether) as a crosslinking agent under oven or water bath environments. Then, HAG was mixing with non-crosslinked hyaluronic acid (ncHA) to produce biHAG, named HAG100, HAG80 and HAG60 according to the composition ratio of HAG. Rheology and swelling ratios of different biHAG were assessed . The biocompatibility and the induction of cell differentiation of biHAG and Tideglusib contained biHAG (Tide@HAG) were also evaluated. In addition, the tissue response to the biHAG and Tide@HAG with hDPSC mixtures were investigated using subcutaneous injection model on immunosuppressive mice, sacrificed at the sixth weeks.
The results showed biHAG cross-linked at water bath presented the similar swelling ratio, but more homogeneity and fluid properties than that at oven. biHAG with 250 μm not only had better injectability and fluidity but also structure maintenance property than those of 420 μm. 250 μm HAG80 and HAG60 could be injected through a 27G needle, and HAG80 could maintain its shape than HAG60 after injection. Therefore, HAG80 was selected for further in vitro and in vivo studies.
No cytotoxicity of Tideglusib with more than 80% cell viability was found when its concentration less than 300 nM. Tide@HAG presented good biocompatibility using extract model. In addition, almost all Tideglusib would release from HAG within 24 hours, indicating HAG did not have a sustained release effect. In all biHAG and Tide@HAG, hDPSC attached and growth on the materials were observed and granular sizes did not affect the cell growth behaviors, in which, HAG80 with 100 nM Tideglusib had best cell growth behavior and more expression of DSPP and VEGF in vitro. Therefore, 100 nM Tideglusib was used for in vivo experiment.
The histological findings of all biHAG and Tide@HAG confirmed they were all biocompatible and no inflammatory response were found. In the 420 μm groups without drug, HAG100 showed more materials retained with few fibrous tissue formation and both HAG80 and HAG60 presented more fibrous tissue and blood vessels formation. No obvious differences on fibrous tissue growth and angiogenesis were found between HAG80 with 420 μm and 250 μm. The addition of 100 nM Tideglusib did promote more tissue growth and angiogenesis than other groups, and the 250 μm groups had significant better tissue formation compare to 420 μm groups.
In summary, the biHAG cross-linked at water bath had more fluidity and homogeneity than those at oven condition and biHAG with 250 μm showed better injectability, in which HAG80 presented better properties for injection and structure maintenance. Tide@HAG had good biocompatibility and 100 nM Tideglusib could enhance cell growth and cell differentiation in vitro. In addition, the additon of ncHA and 100nM Tideglusib in biHAG would promote fibrous tissue and vessels formation in vivo. 250 μm biHAG with 100 nM Tideglusib presented the better fibrous tissue formation and angiogenesis, showing the highly potential in dentin-pulp complex regeneration.
誌謝 i
中文摘要 ii
英文摘要 iv
目錄 vii
圖目錄 xi
表目錄 xiv
縮寫表 xvi
第一章 前言 1
第二章 文獻回顧 3
2.1 現代根管治療的發展與限制 3
2.2 組織工程的再生技術 5
2.3 幹細胞 6
2.4支架 8
2.4.1 透明質酸的特性 8
2.4.2雙相型透明質酸膠體顆粒在牙科的應用 9
2.5生長因子 10
2.5.1 生長因子的選擇 10
2.5.2 Wnt訊息傳遞路徑在牙齒發育扮演的角色 10
2.5.3 GSK-3抑制劑的選用 11
第三章 動機與目的 14
第四章 材料與方法 15
4.1 儀器裝置 15
4.2 藥品材料 16
4.3 材料製備 16
4.3.1 製備交聯型透明質酸膠體顆粒(HAG)材料 16
4.3.2 製備雙相型透明質酸膠體顆粒(biHAG)材料 17
4.4 物理性質分析 17
4.4.1 流變性質分析 17
4.4.2 膨脹比例分析 18
4.5 Tideglusib濃度對細胞活性的作用 18
4.5.1 細胞選擇與培養 18
4.5.2 細胞活性之分析 19
4.5.3 細胞存活率 19
4.5.4 統計分析 20
4.6 Tide@HAG的生物相容性 20
4.6.1 製備搭載Tideglusib之雙相型透明質酸膠體(Tide@biHAG)材料 20
4.6.2 細胞選擇與培養 21
4.6.3 材料萃取液製備 21
4.6.4 細胞存活率 21
4.6.5 細胞致死率 22
4.6.6 統計分析 22
4.7 細胞與Tide@HAG共養之細胞生長行為 22
4.7.1 細胞生長行為之分析 23
4.8 不同濃度Tideglusib對細胞分化的影響 23
4.8.1 細胞選擇與培養 23
4.8.2 RNA萃取及反轉錄聚合酶鏈鎖反應 24
4.8.3 統計分析 25
4.9 小鼠背部皮下注射材料之組織反應 25
4.9.1 注射材料製備 25
4.9.2 小鼠背部皮下注射模型 25
4.9.3 檢體觀察及組織學切片分析 26
4.9.3 統計分析 26
第五章 結果 27
5.1 雙相型透明質酸膠體顆粒之物理性質分析 27
5.1.1 流變性質(rheology property)分析 27
5.1.2 膨脹比例(swelling ratio)分析 28
5.2 搭載Tideglusib之雙相型透明質酸膠體顆粒對細胞之影響 28
5.2.1 Tideglusib的細胞毒性分析 28
5.2.2 Tide@HAG的生物相容性分析 29
5.2.3 細胞生長行為之觀察分析 29
5.2.4 不同濃度Tideglusib對細胞分化之影響分析 30
5.3 小鼠背部皮下注射之活體實驗結果 30
5.3.1 檢體外觀及體積大小 30
5.3.2 組織切片觀察及半定量分析 31
第六章 討論 33
6.1 透明質酸膠體的物理性質探討 33
6.1.1 溫度反應環境對膠體影響之探討 33
6.1.2 粒徑大小對膠體影響之探討 34
6.2 Tide@HAG對於細胞影響之探討 36
6.2.1 Tideglusib的細胞毒性之探討 36
6.2.2 Tide@HAG的生物相容性探討 36
6.2.3 細胞和Tide@HAG共養的生長行為探討 37
6.2.4 不同濃度Tideglusib對細胞分化行為之探討 38
6.3 小鼠背部注射biHAG和Tide@HAG對活體組織影響之探討 39
6.3.1 實驗模型設計之探討 39
6.3.2 檢體外觀及尺寸之探討 40
6.3.3 組織學觀察之探討 41
第七章 結論 44
參考文獻 46
附錄 51
Aguilar, P., & Linsuwanont, P. (2011). Vital pulp therapy in vital permanent teeth with cariously exposed pulp: a systematic review. J Endod, 37(5), 581-587. doi:10.1016/j.joen.2010.12.004
Andre, P. (2008). New trends in face rejuvenation by hyaluronic acid injections. J Cosmet Dermatol, 7(4), 251-258. doi:10.1111/j.1473-2165.2008.00402.x
Annalisa La Gatta, C. S., Agata Papa, Mario De Rosa. (2011). Comparative analysis of commercial dermal fillers based on crosslinked hyaluronan: Physical characterization and in vitro enzymatic degradation. Polymer Degradation and Stability, 630-636. doi:10.1016/j.polymdegradstab.2010.12.025
Baksh, D., Song, L., & Tuan, R. S. (2004). Adult mesenchymal stem cells: characterization, differentiation, and application in cell and gene therapy. J Cell Mol Med, 8(3), 301-316. doi:10.1111/j.1582-4934.2004.tb00320.x
Banava, S., Fazlyab, M., Heshmat, H., Mojtahedzadeh, F., & Motahhary, P. (2015). Histological Evaluation of Single and Double-visit Direct Pulp Capping with Different Materials on Sound Human Premolars: A Randomized Controlled Clinical Trial. Iran Endod J, 10(2), 82-88.
Birger Nygaard‐Östby, O. H. (1971). Tissue formation in the root canal following pulp removal. Oral Science, 79(3), 17.
Caliari, S. R., & Burdick, J. A. (2016). A practical guide to hydrogels for cell culture. Nat Methods, 13(5), 405-414. doi:10.1038/nmeth.3839
Chang, S. W., Kim, J. Y., Kim, M. J., Kim, G. H., Yi, J. K., Lee, D. W., . . . Kim, E. C. (2016). Combined effects of mineral trioxide aggregate and human placental extract on rat pulp tissue and growth, differentiation and angiogenesis in human dental pulp cells. Acta Odontol Scand, 74(4), 298-306. doi:10.3109/00016357.2015.1120882
Chrepa, V., Austah, O., & Diogenes, A. (2017). Evaluation of a Commercially Available Hyaluronic Acid Hydrogel (Restylane) as Injectable Scaffold for Dental Pulp Regeneration: An In Vitro Evaluation. J Endod, 43(2), 257-262. doi:10.1016/j.joen.2016.10.026
Clevers, H. (2006). Wnt/beta-catenin signaling in development and disease. Cell, 127(3), 469-480. doi:10.1016/j.cell.2006.10.018
da Rosa, W. L. O., Cocco, A. R., Silva, T. M. D., Mesquita, L. C., Galarca, A. D., Silva, A. F. D., & Piva, E. (2018). Current trends and future perspectives of dental pulp capping materials: A systematic review. J Biomed Mater Res B Appl Biomater, 106(3), 1358-1368. doi:10.1002/jbm.b.33934
Eldar-Finkelman, H., & Martinez, A. (2011). GSK-3 Inhibitors: Preclinical and Clinical Focus on CNS. Front Mol Neurosci, 4, 32. doi:10.3389/fnmol.2011.00032
Friedman, S., & Mor, C. (2004). The success of endodontic therapy--healing and functionality. J Calif Dent Assoc, 32(6), 493-503.
Galler, K. M., D''Souza, R. N., Hartgerink, J. D., & Schmalz, G. (2011). Scaffolds for dental pulp tissue engineering. Adv Dent Res, 23(3), 333-339. doi:10.1177/0022034511405326
Gronthos, S., Brahim, J., Li, W., Fisher, L. W., Cherman, N., Boyde, A., . . . Shi, S. (2002). Stem cell properties of human dental pulp stem cells. J Dent Res, 81(8), 531-535. doi:10.1177/154405910208100806
Gronthos, S., Mankani, M., Brahim, J., Robey, P. G., & Shi, S. (2000). Postnatal human dental pulp stem cells (DPSCs) in vitro and in vivo. Proc Natl Acad Sci U S A, 97(25), 13625-13630. doi:10.1073/pnas.240309797
Hargreaves KM, Goodis HE. Seltzer and Bender''s Dental Pulp. 3rd ed. Chicago: Quintessence; 2002.
He, X., Semenov, M., Tamai, K., & Zeng, X. (2004). LDL receptor-related proteins 5 and 6 in Wnt/beta-catenin signaling: arrows point the way. Development, 131(8), 1663-1677. doi:10.1242/dev.01117
Huang, G. T. (2011). Dental pulp and dentin tissue engineering and regeneration: advancement and challenge. Front Biosci (Elite Ed), 3, 788-800.
Huang, G. T., Gronthos, S., & Shi, S. (2009). Mesenchymal stem cells derived from dental tissues vs. those from other sources: their biology and role in regenerative medicine. J Dent Res, 88(9), 792-806. doi:10.1177/0022034509340867
Huang, G. T., Shagramanova, K., & Chan, S. W. (2006). Formation of odontoblast-like cells from cultured human dental pulp cells on dentin in vitro. J Endod, 32(11), 1066-1073. doi:10.1016/j.joen.2006.05.009
Huang, G. T., Yamaza, T., Shea, L. D., Djouad, F., Kuhn, N. Z., Tuan, R. S., & Shi, S. (2010). Stem/progenitor cell-mediated de novo regeneration of dental pulp with newly deposited continuous layer of dentin in an in vivo model. Tissue Eng Part A, 16(2), 605-615. doi:10.1089/ten.TEA.2009.0518
Hulsart-Billstrom, G., Yuen, P. K., Marsell, R., Hilborn, J., Larsson, S., & Ossipov, D. (2013). Bisphosphonate-linked hyaluronic acid hydrogel sequesters and enzymatically releases active bone morphogenetic protein-2 for induction of osteogenic differentiation. Biomacromolecules, 14(9), 3055-3063. doi:10.1021/bm400639e
Iohara, K., Zheng, L., Ito, M., Ishizaka, R., Nakamura, H., Into, T., . . . Nakashima, M. (2009). Regeneration of dental pulp after pulpotomy by transplantation of CD31(-)/CD146(-) side population cells from a canine tooth. Regen Med, 4(3), 377-385. doi:10.2217/rme.09.5
Kablik, J., Monheit, G. D., Yu, L., Chang, G., & Gershkovich, J. (2009). Comparative physical properties of hyaluronic acid dermal fillers. Dermatol Surg, 35 Suppl 1, 302-312. doi:10.1111/j.1524-4725.2008.01046.x
Karbanova, J., Soukup, T., Suchanek, J., Pytlik, R., Corbeil, D., & Mokry, J. (2011). Characterization of dental pulp stem cells from impacted third molars cultured in low serum-containing medium. Cells Tissues Organs, 193(6), 344-365. doi:10.1159/000321160
Kim, S. G., Malek, M., Sigurdsson, A., Lin, L. M., & Kahler, B. (2018). Regenerative endodontics: a comprehensive review. Int Endod J, 51(12), 1367-1388. doi:10.1111/iej.12954
Kim, S. G., Zhou, J., Solomon, C., Zheng, Y., Suzuki, T., Chen, M., . . . Mao, J. J. (2012). Effects of growth factors on dental stem/progenitor cells. Dent Clin North Am, 56(3), 563-575. doi:10.1016/j.cden.2012.05.001
Kisby, L. (2016). Vital Pulp Therapy in Primary Teeth: An Update. Dent Today, 35(5), 112-113.
Langer, R., & Vacanti, J. P. (1993). Tissue engineering. Science, 260(5110), 920-926.
Liu, Y. P., Seckin, H., Izci, Y., Du, Z. W., Yan, Y. P., & Baskaya, M. K. (2009). Neuroprotective effects of mesenchymal stem cells derived from human embryonic stem cells in transient focal cerebral ischemia in rats. J Cereb Blood Flow Metab, 29(4), 780-791. doi:10.1038/jcbfm.2009.1
Liu CC, Tsai PH & Lee YL, J Dent Res 2018; 97(spec B): 1158, (Poster paper)
Lo, B., & Parham, L. (2009). Ethical issues in stem cell research. Endocr Rev, 30(3), 204-213. doi:10.1210/er.2008-0031
Logan, C. Y., & Nusse, R. (2004). The Wnt signaling pathway in development and disease. AnnuRevCellDevBiol,20,781-810. doi:10.1146/annurev.cellbio.20.010403.113126
MacDonald, B. T., Tamai, K., & He, X. (2009). Wnt/beta-catenin signaling: components, mechanisms,anddiseases.Dev Cell, 17(1), 9-26. doi:10.1016/j.devcel.2009.06.016
Mansbridge, J. (2008). A “Selection Model” of Political Representation. KSG Faculty Research Working Paper Series.
Meijer, L., Flajolet, M., & Greengard, P. (2004). Pharmacological inhibitors of glycogen synthasekinase3.TrendsPharmacolSci,25(9),471-480. doi:10.1016/j.tips.2004.07.006
Miura, M., Gronthos, S., Zhao, M., Lu, B., Fisher, L. W., Robey, P. G., & Shi, S. (2003). SHED: stem cells from human exfoliated deciduous teeth. Proc Natl Acad Sci U S A, 100(10), 5807-5812. doi:10.1073/pnas.0937635100
Moussa, D. G., & Aparicio, C. (2019). Present and future of tissue engineering scaffolds for dentin-pulp complex regeneration. J Tissue Eng Regen Med, 13(1), 58-75. doi:10.1002/term.2769
Mozaffari, M. S., Emami, G., Khodadadi, H., & Baban, B. (2019). Stem cells and tooth regeneration: prospects for personalized dentistry. EPMA J, 10(1), 31-42. doi:10.1007/s13167-018-0156-4
Neves, V. C., Babb, R., Chandrasekaran, D., & Sharpe, P. T. (2017). Promotion of natural tooth repair by small molecule GSK3 antagonists. Sci Rep, 7, 39654. doi:10.1038/srep39654
Odorico, J. S., Kaufman, D. S., & Thomson, J. A. (2001). Multilineage differentiation from human embryonic stem cell lines. Stem Cells, 19(3), 193-204. doi:10.1634/stemcells.19-3-193
Pardue, E. L., Ibrahim, S., & Ramamurthi, A. (2008). Role of hyaluronan in angiogenesis and its utility to angiogenic tissue engineering. Organogenesis, 4(4), 203-214. doi:10.4161/org.4.4.6926
Rui Yang, a. L. T., †a Lian Cen*a and Zhibing Zhang*b. (2016). An injectable scaffold based on crosslinked hyaluronic acid gel for tissue regeneration. The Royal Society of Chemistry, 13. doi:10.1039/c5ra27870h
Rutherford, R. B., & Gu, K. (2000). Treatment of inflamed ferret dental pulps with recombinant bone morphogenetic protein-7. Eur J Oral Sci, 108(3), 202-206.
Sonoyama, W., Liu, Y., Fang, D., Yamaza, T., Seo, B. M., Zhang, C., . . . Shi, S. (2006). Mesenchymal stem cell-mediated functional tooth regeneration in swine. PLoS One, 1, e79. doi:10.1371/journal.pone.0000079
Tamura, M., & Nemoto, E. (2016). Role of the Wnt signaling molecules in the tooth. Jpn Dent Sci Rev, 52(4), 75-83. doi:10.1016/j.jdsr.2016.04.001
Wang, L., & Stegemann, J. P. (2010). Extraction of high quality RNA from polysaccharide matrices using cetyltrimethylammonium bromide. Biomaterials, 31(7), 1612-1618. doi:10.1016/j.biomaterials.2009.11.024
Wang, X., Thibodeau, B., Trope, M., Lin, L. M., & Huang, G. T. (2010). Histologic characterization of regenerated tissues in canal space after the revitalization/revascularization procedure of immature dog teeth with apical periodontitis. J Endod, 36(1), 56-63. doi:10.1016/j.joen.2009.09.039
Wei, X., Ling, J., Wu, L., Liu, L., & Xiao, Y. (2007). Expression of mineralization markers in dental pulp cells. J Endod, 33(6), 703-708. doi:10.1016/j.joen.2007.02.009
Witherspoon, D. E. (2008). Vital pulp therapy with new materials: new directions and treatment perspectives--permanent teeth. J Endod, 34(7 Suppl), S25-28. doi:10.1016/j.joen.2008.02.030
Yeom, J., Bhang, S. H., Kim, B. S., Seo, M. S., Hwang, E. J., Cho, I. H., . . . Hahn, S. K. (2010). Effect of cross-linking reagents for hyaluronic acid hydrogel dermal fillers on tissue augmentation and regeneration. Bioconjug Chem, 21(2), 240-247. doi:10.1021/bc9002647
QRCODE
 
 
 
 
 
                                                                                                                                                                                                                                                                                                                                                                                                               
第一頁 上一頁 下一頁 最後一頁 top
系統版面圖檔 系統版面圖檔