跳到主要內容

臺灣博碩士論文加值系統

(44.192.25.113) 您好!臺灣時間:2022/05/16 19:15
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

: 
twitterline
研究生:潘致良
研究生(外文):Chih-Liang Pan
論文名稱:利用蛋白質體探討樟芝萃取物處理在人類肺癌細胞株(A549)
論文名稱(外文):Proteomic analysis of human lung carcinoma A549 cells treated with ethanol-extracts of Antrodia camphorata by solid-state fermentation.
指導教授:吳定峰吳定峰引用關係
指導教授(外文):Ting-Feng Wu
學位類別:碩士
校院名稱:南台科技大學
系所名稱:生物科技系
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:2004
畢業學年度:92
語文別:中文
論文頁數:67
中文關鍵詞:樟芝二維電泳基質輔助雷射脫附游離法-飛行時間質譜儀高效能液相層儀儀-電噴灑游離
外文關鍵詞:Antrodia camphorataTwo-dimensional electrophoresisMALDI-TOFESI/MS/MS
相關次數:
  • 被引用被引用:9
  • 點閱點閱:1127
  • 評分評分:
  • 下載下載:167
  • 收藏至我的研究室書目清單書目收藏:3
中文摘要
樟芝又稱牛樟芝或牛樟菇,學名為Antrodia camphorata,是台灣特有一種真菌,僅生長於台灣特有之牛樟樹,生長緩慢、得之不易,故更顯其彌足珍貴。樟芝在台灣傳統醫藥歷史中始終被視為珍貴藥材,具有神奇的醫療保健功效。根據最近研究顯示,以2 % (v/v) 固態發酵樟芝乙醇萃取液處理人類肺癌A549細胞株72 小時,對A549細胞具有抑殺的效果。
在本實驗中,使用蛋白質體學技術來研究經固態發酵樟芝乙醇萃取液處理8小時的A549細胞和未經處理的A549細胞中表現量不同的蛋白質,利用PDQUEST軟體比對經處理及未經處理A549細胞的二維電泳圖譜,以找出表現差異2倍或2倍以上的蛋白質,在本論文中我們找到5個顯示統計上2倍或2倍以上顯著表現差異的蛋白質點(P<0.05)。
利用電灑游離式串連質譜儀鑑定這些表現差異蛋白質點,這五個蛋白質點分別被鑑定為 human galectin-1、human eukaryotic translation initiation factor 5A (eIf-5A)、Rho GDP dissociation inhibitor (Rho-GDI)、human calcium-dependent protease (CANP) and annexin V. 本論文在經固態發酵樟芝乙醇萃取液處理的A549細胞中找到五個表現差異的蛋白質,這些表現差異的蛋白質可以來幫助推測固態發酵樟芝乙醇萃取液如何影響腫瘤細胞的基因表達,以利未來進一步研究樟芝抑癌的機制。
Abstract
Antrodia camphorata (Chinese name, niu-chang-chih or niu-chang-ku) is a very rare and expensive mushroom because it slowly grows on the inner heart wood wall of the Cinnamomum kanehiraae only, which is an unique tree in Taiwan with limited amount. A. camphorata still has not been cultivated successfully. The potential health improvement and disease prevention of Antrodia camphorata have been emphasized in traditional Taiwanese herb medicine. Recent studies demonstrated 2 % (v/v) ethanol-extracts of A. camphorata cultivated by solid-state fermentation could effectively inhibit the proliferation of human lung carcinoma A549 cells in 72 hours.
In this study, we reported a proteomic experiment and showed the appreciable changes in protein synthesis after 8 hours treatment with 0.4% (v/v) ethanol-extracts of A. camphorata on A549 cells. After we compared two-dimension gel maps of non-treated with treated A549 cells using PDQUEST software, we have identified five protein spots,which were 2-fold or above differentially expressed with statistically significance(P<0.05).
These five significantly downregulated protein spots were further identificatied by high performance liquid chromatography-electrospray tandem mass spectrometry (HPLC-ESI/MS/MS). These five proteins were identified as human galectin-1、human eukaryotic translation initiation factor 5A (eIf-5A)、Rho GDP dissociation inhibitor (Rho-GDI)、human calcium-dependent protease (CANP) and annexin V respectively. The results of this study help us to get insight into the effect of ethanol-extracts of A. camphorata cultivated by solid-state fermentation on the gene expression in the tumor A549 cells.
目 次
英文摘要 iv
中文摘要 v
誌謝 vi
目次 vii
表目錄 ix
圖目錄 x
符號與縮寫 xii
第一章 前言 1
1-1 背景的介紹 1
1-2 研究目的 3
第二章 文獻整理 4
2.1 樟芝的介紹 4
2-1-1 樟芝的特性 4
2-1-2 分類學 5
2-1-3 樟芝生理活性的研究 6
2-1-4 樟芝之藥理作用 7
2.2 蛋白質體學 9
2-2-1 蛋白質二維電泳 10
2-2-2 蛋白質鑑定技術 11
第三章 研究材料與方法 14
3-1 材料與藥品. 14
3-1-1 樟芝固態發酵乙醇萃取液 14
3-1-2 細胞株 14
3-1-3 藥品 14
3-1-3 實驗儀器 16
3-2 實驗方法 17
3-2-1 固體發酵樟芝乙醇萃取液對A549 細胞之處理 17
3-2-2 細胞蛋白質萃取液之製備 18
3-2-3 蛋白質液定量 18
3-2-4 蛋白質等電點聚焦電泳法 (isoelectric focusing, IEF) 18
3-2-5 SDS-聚丙烯膠電泳(SDS-polyacrylamide gel electrophoresis) 19
3-2-5 銀染 19
3-2-6 軟體比對 20
3-2-7 膠內蛋白質水解 20
3-2-7 電灑游離式串連質譜儀鑑定-Electrospray ionization tandem mass spectrometry (ESI/MS/MS) 20
3-2-8西方墨點法 21
第四章 結果 22
4-1 0.4 % (v/v)的固體發酵樟芝乙醇萃取液處理A549細胞存活測試 22
4-2 二維電泳圖譜條件之建立 22
4-3 以二維電泳圖譜分析固體發酵樟芝乙醇萃取液對人類A549肺癌細胞蛋白質圖譜的影響 23
4-4 受固體發酵樟芝乙醇萃取液影響而表現差異的蛋白質點 24
4-5 表現差異蛋白質點身份之確認 25
4-6以西方墨點法確認蛋白質表現的差異 25
第五章 討論 27
第六章 參考文獻 32
表目錄
表1 菇類的藥效成分 40
表 2. 未處理的人類肺癌細胞株(A549),經與固態樟芝發酵後的乙醇萃取液處理的人類肺癌細胞株細胞株中具有顯著性差異的蛋白質點 41
表3. 有差異之蛋白質點經LC/MS/MS所鑑定出的蛋白質資料 42
表4 、LC/MS/MS軟體與蛋白質資料庫比對的相關參數定義 43
圖目錄
圖2-1 樟芝(Antrodia camphorata) 44
圖2-2 樟芝子實體萃取物中發現新的三帖類化合物 45
圖2-3 樟芝子實體萃取物中發現新的三帖類化合物 46
圖2-4 由樟芝分離得的十二種化合物 47
圖2-6 固態樟芝發酵後的熱水及乙醇萃取物作用於 A549 細胞的實驗結果 48
圖2-7 固態樟芝發酵後的熱水及乙醇萃取物作用於 Hep3B 細胞的實驗結果 48
圖2-8 蛋白質體學簡化流程圖 49
圖4-1 0.4 % (v/v)的固態發酵樟芝乙醇萃取液處理0、2、4、6、8小時以及0.4% (v/v)固體培養基基材乙醇萃取液與100%乙醇(固體發酵樟芝是經乙醇萃取)處理 A549細胞8小時的細胞存活率 50
圖4-2. E7 cells 細胞二維電泳圖譜 51
圖4-3 二維電泳條件改進後之E7 細胞二維電泳圖譜 52
圖4-4 (A)為未處理的人類肺癌細胞株(A549) 53
圖4-4 (B)固態樟芝發酵後的乙醇萃取液處理的人類肺癌細胞株(A549) 8小時的代表性二維電泳圖譜 54
圖4-5 (A)未處理的人類肺癌細胞株中蛋白質點的二維電泳圖譜放大圖(B)同區域的固態樟芝發酵後的乙醇萃取液處理的人類肺癌細胞株的二維電泳圖譜放大圖 56
圖4-6 未處理的人類肺癌細胞株(A549),經與固態樟芝發酵後的乙醇萃取液處理的人類肺癌細胞株細胞株中具有顯著性差異的蛋白質點的平均volume數值 57
圖.4-7 (A)Spot 1:Human Galectin-1蛋白質點LC/MS/MS比對的結果。 58
圖.4-7 (B)Spot 2:Human translation initiation factor 5A蛋白質點LC/MS/MS比對的結果 59
圖.4-7 (C)Spot 3:Rho GDP dissociation inhibitor蛋白質點LC/MS/MS比對的結果 60
圖.4-7 (D)Spot 4:Human Calcium-dependent proteas 蛋白質點LC/MS/MS比對的結果 61
圖.4-7 (E)Spot 5:Human Annexin A5蛋白質點LC/MS/MS比對的結果。 62
圖4-8 (A)∼(D)有差異蛋白質點之西方墨點法的結果。 66
第六章參考文獻
1. 賴慶亮譯,1997。水野卓 川合正允原著,菇類的化學、生化學,國立編譯館。
2. 陳啟楨,2004,菇菇的世界,偉銘圖書出版。
3. Zang, M., and SU, Q. (1990) Ganoderma camphoratum, a new taxon in genus Ganoderma from Taiwan, China Acta Bot.Yunnanica. 12: 395-396.
4. Chang, T. T., and Chou, W. N. (1995). Antrodia cinnamomea sp.Nov. on Cinnmomum Kamehirai in Taiwan. Mycol. Res. 99: 76-758.
5. Wu, S. H., Leif, R., and Chang, T. T. (1997). Antrodia camphorata ("niu-chang-chih"), new combination of a medicinal fungus in Taiwan. Bot. Bull. Acad. Sin. 38: 73-75.
6. 黃鈴娟,樟芝與姬松茸芝抗氧化性質及其多醣組成分析,國立中興大學食品科學系碩士論文(2000)。
7. Cherng, I. H., and Chiang, H. C., (1995). “ Three new triterpenoids from Antrodia cinnamomea “. J Natural Products.58:365-371.
8. Cherng, I. H., Wu, D. P., and Chiang, H. C., (1996). “ Triterpenoids from Antrodia cinnamomea “. Phytochemistry. 41:263-267.
9. 楊書威,中藥樟菇活性成分之研究,國立台灣大學藥學研究所碩士論文(1991)。
10. 吳德鵬,樟芝微量成份的研究,國立臺灣師範大學化學研究所碩士論文(1995)。
11. Song, T. Y., and Yen, G. C. (2003). Protective effects of fermented filtrate from Antrodia camphorata in submerged culture against CCl4-induced hepatic toxicity in rats. J Agric. Food Chem. 51:1571-1577.
12. Hsiao, G., Shen, M.-Y., Lin, K.-H., Lan, M.-H., Wu, L.-Y., Chou, D.-S., Lin, C.-H., Su, C.-H., and Sheu, J.-R. (2003). Antioxidative and hepatoprotective effects of Antrodia camphorata extract. J Agric. Food Chem. 51: 3302-3308.
13. Song, T.Y.,and Yen, G. C. (2002). Antioxidant properties of Antrodia camphorata in submerged culture. J Agric Food Chem. 50:3322-3327.
14. Hseu, Y. C., Chang, W. C., Hseu, Y. T., Lee, C.Y., Yech, Y. J., Chen, P. C., Chen, J. Y., Yang, H. L. (2002). Protection of oxidative damage by aqueous extract from Antrodia camphorata mycelia in normal human erythrocytes. Life Sci. 71: 469-482.
15. Kataoka, K., Muta, T., Yamazak, S., and Takeshige, K. (2002). Activation of macrophage by linear (1®3)-b-D-glucans. J. Biol. Chem. 277: 36825-36831.
16. Olson, E. J., Standing, J. E., Griego-Happer, N., Hoffman, O. A., and Limper, A. H. (1996). Fungal b-glucan interacts with vitronectin and stimulates tumor necrosis factor alpha release from macrophage. Infect. Immun. 64: 3548-3554.
17. Brown, G.. D.,and Gordon, S. (2001). Immune recognition. A new receptor for beta-glucans. Nature 413: 36-37.
18. Harada, N., Kodama, N., and Nanba, H. (2003). Relationship between dendritic cells and the D-fraction-induced Th-1 dominant response in BALB/c tumor-bearing mice. Cancer Lett. 192:181-187.
19. Muto, S., Vetvicka, V., and Ross, G.. D. (1993). CR3 (CD11b/CD18) expressed by cytotoxic T cells and natural killer cells is upregulated in a manner similar to neutrophil CR3 following stimulation with various activating agents. J Clin Immunol. 13: 175-184.
20. 張紹欣,天然食材的樟芝發酵產物對抑制腫瘤細胞之評估,南台科技大學生物科技研究所碩士論文(2003)。
21. Klose, J., and Kobalz, U.Two-dimensional electrophoresis of proteins: an updated protocol and implications for a functional analysis of the genome. Electrophoresis 16:1034-1059,1995.
22. Costello, C. E. Time, life and mass spectrometry. New techniques to address biological questions.Biophys. Chem. 68:173-188,1997.
23. Gevaert, K., and Vandekerckhove, J.(2000) Protein identification methods in proteomics. Electrophoresis 21:1145-1154.
24. Kratzer, R., Eckerskorn, C., Karas, M. and Lottspeich, F. (1998) Suppression effects in enzymatic peptide ladder sequencing using ultraviolet - matrix assisted laser desorption/ionization - mass spectormetry. Electrophoresis 19:1910-1919.
25. Klose, J.,and Kobalz, U. (1995) Two-dimensional electrophoresis of proteins: an updated protocol and implications for a functional analysis of the genome. Electrophoresis 16:1034-1059.
26. Roger, J.B. King (2000).Nature Biotechnology Proteomics. Nature Biotechnology 18: supplement IT45-IT46.
27. Ostergaard, M., Wolf, H., Orntoft, T.F., and Celis, J.E. (1999)Psoriasin (S100A7): a putative urinary marker for the follow-up of patients with bladder squamous cell carcinomas. Electrophoresis. 20(2):349-54.
28. Seow, T. K., Ong, S. E., Liang, R. C., Ren, E. C., Chan, L., Ou, K. and Chung, M. C. (2000) Two-dimensional electrophoresis map of the human hepatocellular carcinoma cell line, HCC-M, and identification of the separated proteins by mass spectrometry. Electrophoresis 21:1781-1813.
29. Osada, T., Sakamoto, M., Ino, Y., Iwamatsu, A., Matsuno, Y., Muto, T., and Hirohashi, S. (1996)E-cadherin is involved in the intrahepatic metastasis of hepatocellular carcinoma. Hepatology. 24(6):1460-7.
30. Schwalke, MA., Doremus, CM., Bleday, R., Wanebo, HJ., and Vezeridis, M.P. (1990)Protein differences in human pancreatic cancer cell lines with diverse metastatic potential. Arch Surg. 125(4):469-71.
31. Pasquali, C., Vilbois, F., Curchod, ML, Hooft van Huijsduijnen, R., and Arigoni, F. 2000. Mapping and identification of protein-protein interactions by two-dimensional far-Western immunoblotting.Electrophoresis 21: 3357—3368.
32. Tureci, O., Schmitt, H., Fadle, N., Pfreundscuh, M. and Sahin, U. (1997). Molecular definition of a noval human galectin which is immunogenic in patients with Hdgkin''s disease. J. Biol. Chem. 272:6416-6422.
33. Hughtes, R. C. (1999).Secretion of the galectin family of mammalian carbonhydrate-binding protein.Biochim. Biophys. Acta 1473:172-185
34. Yang, R.Y. and Liu, F.T. (2003). Galectins in cell growth and apoptosis.. Cell Mol Life Sci. 60(2):267-76.
35. Barondes, S. H., Cooper, D. N. W., Gitt, M. A. and Leffler, H. (1994). Galectins: structure and function of a large family of animal lectins. Galectins: structure and function of a large family of animal lectins.J. Biol. Chem. 269:20807-20810.
36. Perillo, N.L., Marcus, M.E., and Baum, L.G.(1998) Galectins: versatile modulators of cell adhesion, cell proliferation, and cell death. J Mol Med. 76(6):402-12.
37. Lahm, H., Andre, S., Hoeflich, A., Fischer, J.R., Sordat, B., Kaltner, H., Wolf, E., and Gabius, H.J.(2001) Comprehensive galectin fingerprinting in a panel of 61 human tumor cell lines by RT-PCR and its implications for diagnostic and therapeutic procedures. J Cancer Res Clin Oncol. 127(6):375-86.
38. Dodd, J. and Jesell, T M. (1986).Cell surface glycoconjugates and carbohydrate-binding proteins: possible recognition signals in sensory neuron development.J. Exp. Biol. 124:225-238.
39. Hynes, M. A., Buck, L. B., Gitt, M., Barondes, S., Dodd, J. and Jesell, T. M. (1989).Carbonhydrate recognition in neuronal development: structure, expression of surface oligosaccarides and beta-galactoside- binding lectins. Clin Symp 145:189-210.
40. Joubert, R., Caron, M. and Bladier, D. (1987).Brain lectin-mediated agglutinability of dissociated cells from embryonic and postnatal mouse brain.Brain Res. 433:146-150.
41. Allen, H. J., Sucato, D., Woynarowska B., Gottstine, S., Sharma, A. and Bernacki R. J. (1990).Role of galaptin in ovarian carcinoma adhesion to extracellular matrix in vitro.J. Cell Biolchem. 43:43-57.
42. Cooper, D. N. W., Massa, S. M. and Barondes S. H. (1991).Endogenous muscle lectin inhibits myoblast adhesion to laminin.J. Cell Biol. 115:1437-1448.
43. Van den Brule, F.A., Buicu, C., Baldet, M., Sobel, M.E., Cooper, D.N., Marschal, P.,and Castronovo, V.(1995) Galectin-1 modulates human melanoma cell adhesion to laminin. Biochem Biophys Res Commun. 17;209(2):760-7.
44. Moiseeva, E. P., Javed, Q., Spring, E. L. and Bono, D. P. de (2000). Galectin1 is involved in vascular smooth muscle cell proliferation. Cardiovasc. Res. 45:493-502.
45. Sanford, G. L. and Harris-Hooker S. (1990).Stimulation of vascular cellproliferation by β-galactoside lectins. FASEB J. 4:2912-2918.
46. Yamanaka, K., Hesegawa, A. K. and Lotan, R. (1991).Cellular respose tooxidative damage in lung induced by the administration of dimethylarsenicacid, a major metabolite of inorganic arsenics, in mice. Toxicol. Appl. Pharmacol. 108:205-213.
47. Camby, I., Belot, N., Rorive, S., Lefranc, F., Maurage, C.A., Lahm, H., Kaltner, H., Hadari, Y., Ruchoux, M.M., Brotchi, J., Zick, Y., Salmon, I., Gabius, H.J.,and Kiss, R. (2001)Galectins are differentially expressed in supratentorial pilocytic astrocytomas, astrocytomas, anaplastic astrocytomas and glioblastomas, and significantly modulate tumor astrocyte migration. Brain Pathol. 11(1):12-26.
48. Perillo, N. L., Marcus, M. E. and Baum, L. G. (1998).Galectins: versatilemodulators of cell adhesion, cell proliferation, and cell death. J. Mol. Med. 76:402-412.
49. Raz, A. and Lotan, R. (1987).Endogenous galactoside-binding: a new class of functional tumor cell surface molecules related to metastasis.CancerMetastasis Rev. 6:433-452.
50. Rubinstein, N., Alvarez, M., Zwirner, N.W., Toscano, M.A., Ilarregui, J.M., Bravo, A., Mordoh, J., Fainboim, L., Podhajcer, O.L.,and Rabinovich G.A.(2004) Targeted inhibition of galectin-1 gene expression in tumor cells results in heightened T cell-mediated rejection; A potential mechanism of tumor-immune privilege. Cancer Cell.5(3):241-51.
51. Schnier J., Schwelberger H.G., Smit-McBride Z., Kang H.A. and Hershey J.W. (1991) Translation initiation factor 5A and its hypusine modification are essential for cell viability in the yeast Saccharomyces cerevisiae. Mol. Cell Biol. 11: 3105-3114.
52. Benne, R., Brown-Luedi M.L., and Hershey, J.W. (1978) Purification and characterization of protein synthesis initiation factors eIF-1, eIF-4C, eIF-4D, and eIF-5 from rabbit reticulocytes. J. Biol. Chem. 253: 3070-7.
53. Park M.H., Wolff E.C. and Folk J.E. (1993) Is hypusine essential for eukaryotic cell proliferation? Trends Biochem. Sci. 18:475-479.
54. Li-En J. D., and Yu Chen K(2002)Subcellular localization of the hypusine-containing eukaryotic initiation factor 5A by immunofluorescent staining and green fluorescent protein tagging J Cell Biochem. 86(3):590-600.
55. Chen, G., Gharib, T.G., Thomas, D.G., Huang, C.C., Misek, D.E., Kuick, R.D., Giordano,T.J., Iannettoni, M.D., Orringer, M.B., Hanash, S.M., and Beer, D.G.(2003)Proteomic analysis of eIF-5A in lung adenocarcinomas. Proteomics. 3(4):496-504.
56. Sordella, R. (2002)Modulation of CREB activity by the Rho GTPase regulates cell and organism size during mouse embryonic development. Developmental Cell. 2, 553 — 565.
57. Sasaki T., and Takai, Y.(1998) The Rho small G protein family---Rho GDI system as a temporal and spatial determinant for cytoskeletal control. Biochemical and Biophysical Research communication 254 : 641-645.
58. Wan, J., Wang, J., Cheng, H., Yu, Y., Xing, G., Oiu, Z., Qian, X.,and He, F. (2001)Proteomic analysis of apoptosis initiation induced by all-trans retinoic acid in human acute promyelocytic leukemia cells. Electrophoresis. 22(14):3026-37.
59. Kovarova, H., Hajduch, M., Korinkova, G., Halada, P., Krupickova, S., Gouldsworthy, A., Zhelev, N.,and Strnad, M. (2003)Proteomic profiling drug-induced apoptosis in non-small cell lung carcinoma: identification of RS/DJ-1 and RhoGDIalpha. Cancer Res.15;63(20):6928-34.
60. Popp, O., Heidinger, M., Ruiz-Heinrich, L., Ries, C., Jochum, M., and Gil-Parrado, S.(2003) The calpastatin-derived calpain inhibitor CP1B reduces mRNA expression of matrix metalloproteinase-2 and -9 and invasion by leukemic THP-1 cells. Biol Chem. Jun. 384(6):951-8.
61. Carragher, N.O., and Frame, M.C.,(2002) Calpain: a role in cell transformation and migrationInt J Biochem Cell Biol.34(12):1539-43.
62. Crumpton, M.J., and Dedman, J.R.(1990)Protein terminology tangle Nature. 17;345(6272):212.
63. Vriens, P.W., Blankenberg, F.G., Stoot, J.H., Ohtsuki ,K., Berry, G.J., Tait, J.F., Strauss, H.W., and Robbins, R.C.(1998)The use of technetium Tc 99m annexin V for in vivo imaging of apoptosis during cardiac allograft rejectionJ Thorac Cardiovasc Surg. ;116(5):844-53.
64. Schlaepfer, D.D., Jones, J., and Haigler,H.T.(1992)Inhibition of protein kinase C by annexin V Biochemistry. 18;31(6):1886-91.
65. Williams, K., Chubb, C., Huberman, E., and Giometti, C.S. (1998)Analysis of differential protein expression in normal and neoplastic human breast epithelial cell lines Electrophoresis. 19(2):333-43.
66. Celis, A., Rasmussen, H.H., Celis, P., Basse, B., Lauridsen, J.B., Ratz, G., Hein, B., Ostergaard, M., Wolf, H., Orntoft, T., and Celis, J.E.(1999)Short-term culturing of low-grade superficial bladder transitional cell carcinomas leads to changes in the expression levels of several proteins involved in key cellular activities. Electrophoresis. 20(2):355-61.
67. 廖錦鳳,利用蛋白質體學技術尋找移形上皮細胞癌之分子標記,嘉南藥理科技大學生物科技研究所碩士論文(2003)。
連結至畢業學校之論文網頁點我開啟連結
註: 此連結為研究生畢業學校所提供,不一定有電子全文可供下載,若連結有誤,請點選上方之〝勘誤回報〞功能,我們會盡快修正,謝謝!
QRCODE
 
 
 
 
 
                                                                                                                                                                                                                                                                                                                                                                                                               
第一頁 上一頁 下一頁 最後一頁 top
無相關期刊