簡易檢索 / 詳目顯示

回結果列表
研究生： 翁育詩
Weng, Yu-Shih
論文名稱： 無細胞支架之肝組織晶片培養研究
Scaffold-free liver-on-a-chip with multiscale organotypic cultures
指導教授： 賴志煌
LAI, CHIH-HUANG
口試委員: 宋信文
SUNG, HSING-WEN
王兆麟
Wang, Jaw-Lin
葉明龍
Yeh, Ming-Long
張秀鳳
Chang, Shau-Feng
學位類別： 博士
Doctor
系所名稱： 工學院 - 材料科學工程學系
Materials Science and Engineering
論文出版年： 2018
畢業學年度： 106
語文別： 英文
論文頁數： 73
中文關鍵詞： 三維共培養 、仿生輻射流體 、無細胞支架 、肝組織晶片 、局部生理特性
外文關鍵詞： spatial co-culture, biomimetic radial flow, scaffold-free, liver-on-a-chip, zonal effect
相關次數： 點閱：1  下載：0 
分享至:
查詢本校圖書館目錄 查詢臺灣博碩士論文知識加值系統 勘誤回報

    • 儘管近年在來組織培養技術(organotypic cultures)有許多顯著的進展，但現有組織培養技術仍無法克服有效表現活體組織特有的功能與生理複雜性。尤其在一些需要多工與複雜生物處理程序的器官，例如肝臟。初代肝臟細胞(Primary liver cells)被認為是最理想的細胞來源，但在應用上卻高度受制於體外快速喪失生理活性。在本研究中，我們運用晶體成長 (lattice growth) 的原理，發展具有模擬生理環境的組織培養系統以控制初代肝臟細胞的三維結構類組織重建。相關模擬生理環境的因子包括在無細胞支架的輔助下，透過生物成長基板(biological growing template) ， 三維共培養(spatial co-culture) ，仿生輻射流體 (biomimetic radial flow) 與模擬體循環 (circulation) 達到體外重建生理微環境。本研究中，我們證實可以在所開發之長天期培養的肝組織培養系統下重現從細胞到組織層級上不同程度的生理複雜性，包括 組織結構(structural hierarchy) ，複雜的藥物代謝(complex drug clearance) ，與局部生理特性(zonal physiology)。我們的研究方法，提供未來在藥物動力與個人醫學上可行的新應用。

    The considerable advances that have been made in the development of organotypic cultures have failed to overcome the challenges of expressing tissue specific functions and complexities, especially for organs that require multitasking and complex biological processes, such as the liver. Primary liver cells are ideal biological building blocks for functional organotypic reconstruction, but are limited by their rapid loss of physiological integrity in vitro. Here, we apply the concept of lattice growth used in material science to develop a tissue incubator which provides physiological cues and controls the three dimensional assembly of primary cells. The cues include a biological growing template, spatial co-culture, biomimetic radial flow and circulation in a scaffold-free condition. We demonstrate the feasibility of recapitulating a multiscale physiological structural hierarchy, complex drug clearance, and zonal physiology from the cell to tissue level in long-term cultured liver-on-a-chip (LOC). Our methods are promising for future applications in pharmacodynamics and personal medicine.

    Table of contents 7 Chapter 1 Motivation 9 Chapter 2 Introduction 10 Chapter 3 Background 12 3.1. Introduction of liver 12 3.1.1. Cell Polarity & multitasking for drug clearance 15 3.1.2. Drug clearance and DILI 16 3.1.3. Zonation 18 3.2. Organoid culture 20 3.2.1. Biological hierarchy and physiological integrity 20 3.2.2. Cell 20 3.2.3. Scaffold 21 3.2.4. Spatial Co-culture 22 3.2.5. Fluidic system and Organ-on-a-chip 24 Chapter 4 Design principle of Liver-on-a-chip (LOC) 25 4.1. Introduction 25 4.2. Experimental procedures 26 4.3. Results and discussion 28 4.4. Summary 33 Chapter 5 CHARACTERIZATION OF LONG-TERM CULTURED LOC 34 5.1. Introduction 35 5.2. Experimental procedures 37 5.3. Results and discussion 40 5.3.1. Morphogenesis of LOC 40 5.3.2. Molecular polarization of LOC 44 5.3.3. Characterization and Spatial location of HSC in LOC 48 5.3.4. Characterization of LOC: Liver-specific functions in LOCs 51 5.4. Summary 53 5.4.1. Morphogenesis of LOC 53 5.4.2. Molecular polarization of LOC 54 5.4.3. Characterization and Spatial location of HSC in LOC 55 5.4.4. Liver Characterization of LOC: Liver-specific functions in LOCs 56 Chapter 6 HEPATOTOXICITY AND ZONATION IN LOC: Radial gradient and dose-associated zonal hepatotoxicity in LOC 58 6.1. Introduction 58 6.2. Experimental procedures 59 6.3. Results and discussion 60 6.4. Summary 65 Chapter 7. Conclusion 66 Chapter 8. Future Perspectives 67 Chapter 9. References 68

    1. E.R. Shamir, A.J. Ewald, Nat. Rev. Mol. Cell Biol. 2014, 15, 647.
    2. E.W Esch, A. Bahinski, D Huh, Nat. Rev. Drug Discov. 2015, 14, 248.
    3. S. Dimmeler, S. Ding, T.A. Rando, A. Trounson, Nat. Med.2014, 20, 814.
    4. M.G. Lerapetritou, P.G. Georgopoulos, C.M. Roth, L.P. Androulakis, 2009 Clin. Transl. Sci., 2, 228.
    5. K. Yum, S.G. Hong, K.E. Healy, L.P. Lee, Biotechnol. J. 2014, 9, 16.
    6. S.N. Bhatia, D.E. Ingber, Nat. Biotechnol. 2014, 32, 760-72.
    7. R Gebhardt, Pharmacol. Ther. 1992, 53, 275.
    8. W.M. Lee, N. Engl. J. Med. 2003, 349, 474.
    9. N. Kaplowitz, Nat. Rev. Drug Discov. 2005, 4, 489.
    10. S. Tujios, R.J. & Fontana, Nat. Rev. Gastroenterol. Hepatol. 2011, 8, 202
    11. P. Godoy, N. J. Hewitt, U. Albrecht, M. E. Andersen, N. Ansari, S. Bhattacharya, J. G. Bode, J.Bolleyn, C. Borner, J. Böttger, A. Braeuning, R. A. Budinsky, B. Burkhardt, N. R. Cameron, G. Camussi, C.-S. Cho, Y.-J. Choi, J. C. Rowlands, U. Dahmen, G. Damm, O. Dirsch, M. T. Donato, J. Dong, S. Dooley, D. Drasdo, R. Eakins, K. S. Ferreira, V. Fonsato, J. Fraczek, R. Gebhardt, A. Gibson, M. Glanemann, C. E. P. Goldring, M. J. Gómez-Lechón, G. M. M. Groothuis, L. Gustavsson, C. Guyot, D. Hallifax, S. Hammad, A. Hayward, D. Häussinger, C. Hellerbrand, P. Hewitt, S. Hoehme, H.-G. Holzhütter, J. B. Houston, J. Hrach, K. Ito, H. Jaeschke, V. Keitel, J. M. Kelm, B. K. Park, C. Kordes, G. A. Kullak-Ublick, E. L. LeCluyse, P. Lu, J. Luebke-Wheeler, A. Lutz, D. J. Maltman, M. Matz-Soja, P. McMullen, I. Merfort, S. Messner, C. Meyer, J. Mwinyi, D. J. Naisbitt, A. K. Nussler, P. Olinga, F. Pampaloni, J. Pi, L. Pluta, S. A. Przyborski, A. Ramachandran, V. Rogiers, C. Rowe, C. Schelcher, K. Schmich, M. Schwarz, B. Singh, E. H. K. Stelzer, B. Stieger, R. Stöber, Y. Sugiyama, C. Tetta, W. E. Thasler, T. Vanhaecke, M. Vinken, T.S. Weiss, A. Widera, C. G. Woods, J. J. Xu, K.M. Yarborough, J. G. Hengstler. Arch. Toxicol. 2013, 87, 1315.
    12. E.L. LeCluyse, R.P. Witek, M.E. Andersen, M.J. Powers, Crit. Rev. Toxicol.. 2012, 42, 501-548.
    13. F.Berthiaume , P.V. Moghe, M. Toner, M.L. Yarmush, FASEB J. 1996,10,1471.
    14. D. Farkas, S.R. Tannenbaum, Toxicol. Sci. 2005, 85, 927
    15. J.C. Dunn, R.G. Tompkins, M.L. Yarmush. Biotechnol. Prog. 1991, 7, 237.
    16. Y.C. Toh, T.C. Lim, D. Lim, G. Xiao, N.D. Van, H. Yu, Lab Chip 2009, 2026.
    17. S.V. Murphy, A. Atala, Nat. Biotechnol. 2014, 32, 773.
    18. O. Jeon, D.S. Alt, S.W. Linderman, E. Alsberg, Adv. Mater. 2013, 25, 6366
    19. A.D. Celiz, J. G. W. Smith, R. Langer, D. G. Anderson, D. A. Winkler, D. A. Barrett, M. C. Davies, L. E. Young, C. Denning, M. R. Alexander, Nat. Mater. 2007, 13, 570.
    20. J. Yeh, Y. Ling, J.M. Karp, J. Gantz, A. Chandawarkar, G. Eng, J. Blumling, R. Langer, A. Khademhosseini, Biomaterials 2006, 27, 5391
    21. M.M. Stevens, J.H. George, Science 2005, 310, 1135.
    22. A. Leal-Egaña, A. Díaz-Cuenca, A.R. Boccaccini, Adv. Mater. 2013, 25, 4049.
    23. P. Horvath, N. Aulner, M. Bickle, A.M. Davies, E.D. Nery, D. Ebner, M.C. Montoya, P. Östling P, V. Pietiäinen, L.S. Price, S.L. Shorte, G. Turcatti, C. von Schantz, N.O. Carragher. Nat. Rev. Drug Discov. 2015, 15, 751.
    24. K.S. Vasanthan, A. Subramanian, U.M. Krishnan, S. & Sethuraman, Biotechnol Adv. 30, 742-752 (2012)
    25. H. Tekin, J.G. Sanchez, C. Landeros, K. Dubbin, R. Langer, A. Khademhosseini, Adv. Mater. 2012, 24, 5543.
    26. A.L. Olsen, S. A. Bloomer, E. P. Chan, M. D. A. Gaça, P. C. Georges, B. Sackey, M. Uemura, P. A. Janmey, R. G. Wells, Am. J. Physiol. Gastrointest. Liver Physiol. 2011, 301, G110.
    27. A. J. Hwa, R. C. Fry, A. Sivaraman, P. T. So, L. D. Samson, D. B. Stolz, L. G. Griffith, FASEB J. 2007, 1, 2564.
    28. M.C. Shih, S.H. Tseng, Y.S. Weng, I.M. Chu, C.H. Liu, Biomed Microdevices 2013,15, 767.
    29. M.J., Powers, K. Domansky, M.R. Kaazempur-Mofrad, A. Kalezi , A. Capitano, A. Upadhyaya, P, Kurzawski, K.E. Wack, D.B. Stolz , R, Kamm, L.G. Griffith. Biotechnol. Bioeng. 2002, 78, 257.
    30. J.W. Allen, S.N. Bhatia, Biotechnol. Bioeng. 2003, 82, 253.
    31. S. Kidambi, R. S. Yarmush, E. Novik, P. Chao, M. L. Yarmush, & Y. Nahmias, Proc. Natl. Acad. Sci. USA 2009, 106, 15714.
    32. A. Lamberti, S. L. Marasso, M. Cocuzza, RSC Adv. 2014, 106, 61415.
    33. J.C. McDonald, G.M. Whitesides, Acc. Chem. Res. 2002, 35, 491.
    34. Junqueira's Basic Histology: Text and Atlas, 13th Edition
    35. D. H. Adams and B. Eksteen. Nat Rev Immunol. 2006, 6, 244-251
    36. P Gissen, IM Arias. J Hepatol. 2015, 63, 1023-1037.
    37. W. Birchmeier. Nat. Cell Biol. 2016, 18, 463–465
    38. LP James, PR Mayeux, JA Hinson. Drug Metab Dispos. 2003, 31, 1499-506
    39. G.D. Block, J. Locker, W. C. Bowen, B. E. Petersen, S. Katyal, S. C. Strom, T. Riley, T. A. Howard, G. K. Michalopoulos, J. Cell Biol. 1996,132,1133-1149
    40. W. Liu, Y. Hou, H. Chen, H. Wei, W. Lin, J. Lai, M. Zhang, F. He, Y. Jiang. Proteomics. 2011, 11, 3556
    41. K.M. Giacomini, S.-M. Huang, D. J. Tweedie, L. Z. Benet, K. L.R. Brouwer, X. Chu, A. Dahlin, R. Evers, V. Fischer, K. M. Hillgren, K. A. Hoffmaster, T. Ishikawa, D. Keppler, R. B. Kim, C. A. Lee, M. Niemi, J. W. Polli, Y. Sugiyama, P. W. Swaan, J. A. Ware, S. H. Wright, S. Wah Yee, M. J. Zamek-Gliszczynski, L. Zhang, Nat. Rev. Drug Discov. 2010, 9, 215.
    42. E.M. Huisman, T. V. Dillen, P.R. Onck, E. V. Giessen, Phys. Rev. Lett. 2007, 99, 208103
    43. J. D. Humphrey, E.R. Dufresne, M. A. Schwartz, Nat. Rev. Mol. Cell Biol. 2015, 15, 802.
    44. L. Lanzetti, Curr. Opin. Cell Biol. 2007, 19, 453.
    45. M. Trauner, J.L. Boyer, Physiol. Rev. 2003, 83, 633-671.
    46. M. J. Zamek-Gliszczynski, K. A. Hoffmaster, X. Tian, R. Zhao, J. W. Polli, J. E. Humphreys, L. O. Webster, A. S. Bridges, J. C. Kalvass, K. L. R. Brouwer. Drug Metab. Dispos. 2005, 33, 1158
    47. A. Esteller. World J. Gastroenterol. 2008, 14, 5641.
    48. S. Hoehmea, M. Brulportb, A. Bauerb, E. Bedawyb, W. Schormannb, M. Hermesb, V. Puppeb, R. Gebhardtc, S. Zellmerc, M. Schwarzd, E. Bockampe, T. Timmelf, J. G. Hengstlerb,D. Drasdoa, Proc. Natl. Acad. Sci. USA 2010, 107, 10371.
    49. C. Yin, K. J. Evason, K. Asahina, D. Y. Stainier, J. Clin. Invest. 2013,123,1902
    50. S.L. Friedman, Physiol. Rev. 2008, 88, 125.

    QR CODE