羊水間葉幹細胞培養於細胞外間質及材料硬度/彈性表面,其分化能力及多能性之研究

以作者查詢圖書館館藏

、以作者查詢臺灣博碩士

、以作者查詢全國書目

、勘誤回報

、線上人數：18

、訪客IP：18.222.115.217

姓名

許佳如(Chia-Ju Hsu) 查詢紙本館藏

畢業系所

化學工程與材料工程學系

論文名稱

羊水間葉幹細胞培養於細胞外間質及材料硬度/彈性表面,其分化能力及多能性之研究
(Pluripotency and Differentiation of Stem Cells Cultured on Biomaterials Grafted with Extracellular Matrix Having Different Elasticity)

相關論文

★ 人類幹細胞培養於熱敏感奈米片段材料之研究	★ 抗癌藥物的鑑定性用於分析大腸癌中癌幹細胞之研究
★ 利用具有奈米片段的生醫材料進行純化及去除癌症幹細胞	★ 人類脂肪幹細胞的膜純化法與分化能力研究
★ 人類羊水間葉幹細胞培養於具有奈米片段與最佳表面硬度的生醫材料，其增殖與成骨分化能力	★ 多能幹細胞在無異種條件下分化為間充質幹細胞的生物材料比較研究

檔案

[Endnote RIS 格式]

[Bibtex 格式]

[相關文章]

[文章引用]

[完整記錄]

[館藏目錄]

[檢視]

[下載]

本電子論文使用權限為同意立即開放。
已達開放權限電子全文僅授權使用者為學術研究之目的，進行個人非營利性質之檢索、閱讀、列印。
請遵守中華民國著作權法之相關規定，切勿任意重製、散佈、改作、轉貼、播送，以免觸法。

摘要(中)

間葉幹細胞 (MSCs) 是一種在臨床上被廣泛利用的自體幹細胞來源。從羊水中分離出來的幹細胞能夠分化成多種不同的細胞譜系，同時沒有道德上的問題。幹細胞的分化能力以及幹細胞的多能性不只取決於細胞本身，同時也會受到外在環境的影響。因此，使用天然的生物分子 (如細胞外間質) 模仿細胞的微環境，增加幹細胞在體外生長的數量以及調控幹細胞的分化對再生醫學是一件非常重要的研究。在這篇研究中，我們將羊水幹細胞(AFSCs)分別培養在不同軟硬度PVA-IA膜且有接枝collagen或fibronectin等細胞外間質 (ECM) 的膜上，以分析羊水幹細胞與細胞外間質的交互作用。可透過交聯時間(0.5小時-48小時)控制PVA-IA膜的軟硬度(3.7千帕-30.4千帕)。相對於AFSCs培養於較硬的膜上，AFSCs培養在較軟的膜上有較小的延展性、應力纖維和較慢的增殖率。測試AFSCs培養在含有接枝ECM不同軟硬度膜間的多能性基因表現。結果顯示，AFSCs培養在軟的膜(10.3千帕)有較好的多能性基因表現，也代表在此條件下可以維持AFSCs多功能，亦可拉長期培養時間。同時，也測試當細胞培養在不同軟硬度的膜上，基質對AFSCs的分化結果。結果發現，AFSCs分別培養在10.3千帕、19.6千帕、30.4千帕時，AFSCs會分化成神經細胞(Nestin)、平滑肌細胞（巢）（α-actin）、骨細胞（Runx2）。

摘要(英)

Mesenchymal stem cells (MSCs) are a valuable cell source for tissue engineering and regenerative medicine. Stem cells derived from amniotic fluid (AFSCs) are pluripotent fetal cells capable of differentiating into multiple lineages, containing representatives of all the three embryonic germ layers. AFSCs are categorized as the intermediate stage between the embryonic stem cells (ESCs) and adult stem cells, and AFSCs may have a distinct mechanism to choose their fate. Therefore, amniotic fluid represents a rich and more suitable source of stem cells in regenerative medicine and tissue engineering than ESCs and induced pluripotent stem cells (iPSCs) due to the lack of ethical concerns regarding use of ESCs and the lack of concerns about xenogenic contamination arising from the use of mouse embryonic fibroblasts as a feeder layer for iPSCs and ESCs. However, stem cells fate of decision is regulated not only by stem cells themselves, but also the effect of microenvironment of stem cells. Therefore, stem cell microenvironment surrounding specific matrix proteins should be important to decide the stem cell fate of differentiation. The optimal design of stem cell culture biomaterials will facilitate the in-vitro production of the large numbers of pluripotent stem cells and specifically more numbers of differentiated cells, which are demanded in the regenerative medicine.
In this study, we investigated whether the stiffness of cell adhesion substrates affect the maintenance of pluripotency of the stem cells and/or modulated the stem cell fate of differentiation. Stem cells from amniotic fluid were cultured on different stiffness of polyvinylalcohol-co-itaconic acid (PVA-IA) films grafted with extracellular matrix (ECM) dishes where collagen, and fibronectin were selected for further studies as ECM components (nanosegments). PVA-IA films were selected as the base matrix due to easy regulation of elasticity (3.7 kPa- 30.4 kPa) by changing the crosslinking time (0.5 hr – 48 hr). AFSCs on soft substrates (3.7 kPa-10.6 kPa) had less spreading, fewer stress fibers and lower proliferation rate than AFSCs on stiff substrate (25.3 kPa-30.4 kPa). The effects of interaction between AFSCs and ECMs(nanosegments) were investigated on the expression of pluripotent genes (e.g., Nanog, Sox2, and Oct4 ) of AFSCs and on the differentiation abilities of AFSCs into several lineage, which were cultured on PVA-IA films grafted with collagen, and fibronectin having different elasticity at passage 4. It was found that AFSCs on PVA-IA films grafted with ECM of elastic modulus (10.3 kPa-12.2 kPa) in the soft showed higher pluripotency genes than hard substrates. AFSCs could differentiate into neural (nestin), smooth muscle cell (α-actin), or osteogenic (Runx2) phenotypes depending on whether they were cultured on PVA-IA films grafted with ECM substrates of elastic modulus in the lower, intermediate or higher ranges. AFSCs cultured on soft substrates have weaker cell adhesion, and showed higher expression of early marker of neural cells (nestin) in expansion medium without addition of induction molecules.

關鍵字(中)

★ 羊水幹細胞
★ 基質軟硬
★ 多功能
★ 分化能力

關鍵字(英)

★ amniotic fluid stem cells
★ elasticity of cell culture matrix
★ pluripotency
★ differentiation ability

論文目次

CHAPTER ONE INTRODUCTION 1
1-1 Mesenchymal stem cells 1
1-2 Amniotic fluid-derived stem cells 3
1-2-1 Amniotic fluid cells 3
1-2-2 Amniotic fluid cell types 4
1-2-3 Isolation of amniotic fluid stem cells (AFSCs) 7
1-2-4 Characterization of amniotic fluid stem cells 8
1-2-5 Pluripotency of amniotic fluid stem cells 10
1-3 Microenvironment of stem cells 12
1-3-1 Soluble factors 13
1-3-2 Cell-cell interactions 13
1-3-3 Cell-biomaterials interactions 13
1-3-4 Physical factors 14
1-4 Type and classification of artificial ECMs 16
1-5 Markers of piuripotent gene 21
1-6 Markers of differentiation lineages of MSCs 23
1-7 Polymerase chain reaction (PCR) 25
1-7-1 Procedure of PCR 26
1-7-2 Reverse transcription polymerase chain reaction (RT-PCR) &Quantitative real time polymerase chain reaction (qRT-PCR) 28
1-7-3 Procedure of RT-PCR 29
CHAPTER TWO MATERIALS AND METHOD 30
2-1 Materials 30
2-1-1 PVA-IA film 30
2-1-2 Cultured medium 30
2-1-3 Serum 30
2-1-4 Antibiotic 31
2-1-5 Growth factor 31
2-1-6 ECM proteins 31
2-1-7 RNA extraction 31
2-1-8 Reverce transcriptase (RT) 31
2-1-9 Real - time (qPCR) 31
2-1-10 Probes 32
2-2 Method and Analysis 32
2-2-1 PVA-IA film preparation 32
2-2-2 Preparation of PVA-IA coating dish grafted with ECM 33
2-2-3 Elasticity measurement of PVA-IA 34
2-2-4 XPS analysis of dish surface 36
2-2-5 Phosphate buffer saline (PBS) preparation 36
2-2-6 Preparation of FGF-2 (b-FGF) protein stock solution 36
2-2-7 Preparation of cell cultured medium 37
2-2-8 Cell cultivation 37
2-2-9 Cell density measurement 38
2-2-10 Isolation of total RNA 39
2-2-11 Reverse Transcription of mRNA into cDNA 40
2-2-12 Quantitative real time polymerase chain reaction 41
CHAPTER THREE RESULTS & DISCUSSION 43
3-1 The elasticity of the PVA-IA films 43
3-2 The XPS measurements of the PVA-IA and ECM-grafted PVA-IA films having different elasticity 44
3-3 Morphology of AFSCs cultured on PVA-IA and ECM-grated PVA-IA films having different elasticity 60
3-4 Pluripotency of AFSCs cultured on PVA-IA and ECM-grafted PVA-IA films having different elasticity 65
3-5 PVA-IA and ECM-grafted PVA-IA films having different elasticity guide AFSCs into specific lineages of differentiation 72
CHAPTER FOUR CONCLUSION 80
CHAPTER FIVE REFERENCE 82

參考文獻

1. Jiang, Y.H., et al., Pluripotency of mesenchymal stem cells derived from adult marrow. Nature, vol. 418, pp. 41-49, 2002.
2. Thomson, J.A., et al., Embryonic stem cell lines derived from human blastocysts. Science, vol. 282, pp. 1145-1147, 1998.
3. Frankel, M.S., In search of stem cell policy. Science, vol. 287, pp. 1397-1397, 2000.
4. Higuchi, A., et al., Biomaterials for the Feeder-Free Culture of Human Embryonic Stem Cells and Induced Pluripotent Stem Cells. Chemical Reviews, vol. 111, pp. 3021-3035, 2011.
5. Okita, K., T. Ichisaka, and S. Yamanaka, Generation of germline-competent induced pluripotent stem cells. Nature, vol. 448, pp. 313-U1, 2007.
6. Takahashi, K. and S. Yamanaka, Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell, vol. 126, pp. 663-676, 2006.
7. Yu, J.Y., et al., Induced pluripotent stem cell lines derived from human somatic cells. Science, vol. 318, pp. 1917-1920, 2007.
8. Lin, S.L., et al., Mir-302 reprograms human skin cancer cells into a pluripotent ES-cell-like state. Rna-a Publication of the Rna Society, vol. 14, pp. 2115-2124, 2008.
9. Zhou, H.Y., et al., Generation of Induced Pluripotent Stem Cells Using Recombinant Proteins. Cell Stem Cell, vol, 4. pp. 381-384, 2009.
10. Higuchi, A., et al., Biomimetic Cell Culture Proteins as Extracellular Matrices for Stem Cell Differentiation. Chemical Reviews, vol, 112, pp. 4507-4540, 2012.
11. Brighton, C.T. and R.M. Hunt, Early histological and ultrastructural changes in medullary fracture callus. J Bone Joint Surg Am, vol, 73, pp. 832-47, 1991.
12. Friedenstein, A.J., J.F. Gorskaja, and N.N. Kulagina, Fibroblast precursors in normal and irradiated mouse hematopoietic organs. Exp Hematol, vol, 4, pp. 267-74, 1976.
13. Vaananen, H.K., Mesenchymal stem cells. Annals of Medicine, vol, 37, pp. 469-479, 2005.
14. Friedenstein, A.J., S. Piatetzky, II, and K.V. Petrakova, Osteogenesis in transplants of bone marrow cells. J Embryol Exp Morphol, vol. 16, pp. 381-90, 1966.
15. Friedenstein, A.J., et al., Heterotopic of bone marrow. Analysis of precursor cells for osteogenic and hematopoietic tissues. Transplantation, vol. 6, pp. 230-47, 1968.
16. Friedenstein, A.J., R.K. Chailakhjan, and K.S. Lalykina, The development of fibroblast colonies in monolayer cultures of guinea-pig bone marrow and spleen cells. Cell Tissue Kinet, vol. 3, pp. 393-403, 1970.
17. Friedenstein, A.J., R.K. Chailakhyan, and U.V. Gerasimov, Bone marrow osteogenic stem cells: in vitro cultivation and transplantation in diffusion chambers. Cell Tissue Kinet, vol. 20, pp. 263-72, 1987.
18. Gosden, C.M., Amniotic fluid cell types and culture. Br Med Bull, vol. 39, pp. 348-54, 1983.
19. Hoehn, H. and D. Salk, Morphological and biochemical heterogeneity of amniotic fluid cells in culture. Methods Cell Biol, vol. 26, pp. 11-34, 1982.
20. Tsai, M.S., et al., Isolation of human multipotent mesenchymal stem cells from second-trimester amniotic fluid using a novel two-stage culture protocol. Hum Reprod, vol. 19, pp. 1450-6, 2004.
21. Perrone, L., et al., Postnatal weight change is influenced by mother-newborn pair leptin levels. Nutrition Research, vol. 20, pp. 1531-1536, 2000.
22. Zambotti, F., et al., Monoamine metabolites and related compounds in human amniotic fluid: assay by gas chromatography and gas chromatography-mass spectrometry. Clin Chim Acta, vol. 61, pp. 247-56, 1975.
23. http://muta-tion.blogspot.tw/2011/09/amniocentesis.html.
24. De Coppi, P., et al., Isolation of amniotic stem cell lines with potential for therapy. Nat Biotechnol, vol. 25, pp. 100-6, 2007.
25. Barry, F.P., et al., The monoclonal antibody SH-2, raised against human mesenchymal stem cells, recognizes an epitope on endoglin (CD105). Biochem Biophys Res Commun, vol. 265, pp. 134-9, 1999.
26. Barry, F., et al., The SH-3 and SH-4 antibodies recognize distinct epitopes on CD73 from human mesenchymal stem cells. Biochem Biophys Res Commun, vol. 289, pp. 519-24, 2001.
27. Tsai, M.S., et al., Clonal amniotic fluid-derived stem cells express characteristics of both mesenchymal and neural stem cells. Biol Reprod, vol. 74, pp. 545-51, 2006.
28. Baksh, D., L. Song, and R.S. Tuan, Adult mesenchymal stem cells: characterization, differentiation, and application in cell and gene therapy. J Cell Mol Med, vol. 8, pp. 301-16, 2004.
29. Ririe, K.M., R.P. Rasmussen, and C.T. Wittwer, Product differentiation by analysis of DNA melting curves during the polymerase chain reaction. Anal Biochem, vol. 245, pp. 154-60, 1997.
30. Shamblott, M.J., et al., Human embryonic germ cell derivatives express a broad range of developmentally distinct markers and proliferate extensively in vitro. Proc Natl Acad Sci U S A, vol. 98, pp. 113-8, 2001.
31. Pittenger, M.F., et al., Multilineage potential of adult human mesenchymal stem cells. Science, vol. 284, pp. 143-7, 1999.
32. Zuk, P.A., et al., Human adipose tissue is a source of multipotent stem cells. Molecular Biology of the Cell, vol. 13, pp. 4279-4295, 2002.
33. Ter Brugge, P.J. and J.A. Jansen, In vitro osteogenic differentiation of rat bone marrow cells subcultured with and without dexamethasone. Tissue Engineering, vol. 8, pp. 321-331, 2002.
34. Cooper, G.M., The Cell: A Molecular Approach. 2000-2009.
35. Lu, Z., et al., Collagen Type II Enhances Chondrogenesis in Adipose Tissue-Derived Stem Cells by Affecting Cell Shape. Tissue Engineering Part A, vol. 16, pp. 81-90, 2010.
36. Kikkawa, Y., et al., Integrin binding specificity of laminin-10/11 : laminin-10/11 are recognized by alpha 3 beta 1, alpha 6 beta 1 and alpha 6 beta 4 integrins. Journal of Cell Science, vol. 113, pp. 869-876, 2000.
37. Jiang, X.S., et al., Surface-immobilization of adhesion peptides on substrate for ex vivo expansion of cryopreserved umbilical cord blood CD34(+) cells. Biomaterials, vol. 27, pp. 2723-2732, 2006.
38. Gelain, F., et al., Designer Self-Assembling Peptide Nanofiber Scaffolds for Adult Mouse Neural Stem Cell 3-Dimensional Cultures. Plos One, vol. 1, 2006.
39. Gilbert, P.M., et al., Substrate Elasticity Regulates Skeletal Muscle Stem Cell Self-Renewal in Culture. Science, vol. 329, pp. 1078-1081, 2010.
40. Georges, P.C., et al., Matrices with compliance comparable to that of brain tissue select neuronal over glial growth in mixed cortical cultures. Biophysical Journal, vol. 90, pp. 3012-3018, 2006.
41. Flanagan, L.A., et al., Neurite branching on deformable substrates. Neuroreport, vol. 13, pp. 2411-2415, 2002.
42. Kondo, T., et al., Sonic hedgehog and retinoic acid synergistically promote sensory fate specification from bone marrow-derived pluripotent stem cells. Proceedings of the National Academy of Sciences of the United States of America, vol. 102, pp. 4789-4794, 2005.
43. Hofstetter, C.P., et al., Marrow stromal cells form guiding strands in the injured spinal cord and promote recovery. Proceedings of the National Academy of Sciences of the United States of America, vol. 99, pp. 2199-2204, 2002.
44. Engler, A.J., et al., Myotubes differentiate optimally on substrates with tissue-like stiffness: pathological implications for soft or stiff microenvironments. Journal of Cell Biology, vol. 166, pp. 877-887, 2004.
45. Ferrari, G., et al., Muscle regeneration by bone marrow derived myogenic progenitors. Science, vol. 279, pp. 1528-1530, 1998.
46. Andrades, J.A., et al., Selection and amplification of a bone marrow cell population and its induction to the chondro-osteogenic lineage by rhOP-1: an in vitro and in vivo study. International Journal of Developmental Biology, vol. 45, pp. 689-693, 2001.
47. Holmbeck, K., et al., MT1-MMP-deficient mice develop dwarfism, osteopenia, arthritis, and connective tissue disease due to inadequate collagen turnover. Cell, vol. 99, pp. 81-92, 1999.
48. Morinobu, M., et al., Osteopontin expression in osteoblasts and osteocytes during bone formation under mechanical stress in the calvarial suture in vivo. Journal of Bone and Mineral Research, vol. 18, pp. 1706-1715, 2003.
49. Deng, J., et al., Mesenchymal stem cells spontaneously express neural proteins in culture and are neurogenic after transplantation. Stem Cells, vol. 24, pp. 1054-1064, 2006.
50. Pittenger, M.F., et al., Multilineage potential of adult human mesenchymal stem cells. Science, vol. 284, pp. 143-147, 1999.
51. McBeath, R., et al., Cell shape, cytoskeletal tension, and RhoA regulate stem cell lineage commitment. Developmental Cell, vol. 6, pp. 483-495, 2004.
52. LaIuppa, J.A., et al., Culture materials affect ex vivo expansion of hematopoietic progenitor cells. Journal of Biomedical Materials Research, vol. 36, pp. 347-359, 1997.
53. Engler, A.J., et al., Matrix elasticity directs stem cell lineage specification. Cell, vol. 126, pp. 677-689, 2006.
54. Rosso, F., et al., Smart materials as scaffolds for tissue engineering. J Cell Physiol, vol. 203, pp. 465-70, 2005.
55. Moroni, L., J.R. de Wijn, and C.A. van Blitterswijk, Integrating novel technologies to fabricate smart scaffolds. J Biomater Sci Polym Ed, vol. 19, pp. 543-72, 2008.
56. Mano, J.F., et al., Natural origin biodegradable systems in tissue engineering and regenerative medicine: present status and some moving trends. J R Soc Interface, vol. 4, pp. 999-1030, 2007.
57. Lee, C.H., A. Singla, and Y. Lee, Biomedical applications of collagen. International Journal of Pharmaceutics, vol. 221, pp. 1-22, 2001.
58. Pankov, R. and K.M. Yamada, Fibronectin at a glance. Journal of Cell Science, vol. 115, pp. 3861-3863, 2002.
59. Mao, Y. and J.E. Schwarzbauer, Fibronectin fibrillogenesis, a cell-mediated matrix assembly process. Matrix Biology, vol. 24, pp. 389-399, 2005.
60. http://www.sigmaaldrich.com/catalog/product/sigma/V8379?lang=en®ion=TW.
61. Ogawa, T., et al., The short arm of laminin gamma2 chain of laminin-5 (laminin-332) binds syndecan-1 and regulates cellular adhesion and migration by suppressing phosphorylation of integrin beta4 chain. Mol Biol Cell, vol. 18, pp. 1621-33, 2007.
62. Klees, R.F., et al., Laminin-5 induces osteogenic gene expression in human mesenchymal stem cells through an ERK-dependent pathway. Molecular Biology of the Cell, vol. 16, pp. 881-890, 2005.
63. Suzuki, S., et al., Effects of Extracellular Matrix on Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells into Smooth Muscle Cell Lineage: Utility for Cardiovascular Tissue Engineering. Cells Tissues Organs, vol. 191, pp. 269-280, 2010.
64. van Dijk, A., et al., Differentiation of human adipose-derived stem cells towards cardiomyocytes is facilitated by laminin. Cell and Tissue Research, vol. 334, pp. 457-467, 2008.
65. Delcroix, G.J.R., et al., The therapeutic potential of human multipotent mesenchymal stromal cells combined with pharmacologically active microcarriers transplanted in hemi-parkinsonian rats. Biomaterials, vol. 32, pp. 1560-1573, 2011.
66. Ma, W., et al., Cell-extracellular matrix interactions regulate neural differentiation of human embryonic stem cells. Bmc Developmental Biology, 2008. 8.
67. Mruthyunjaya, S., et al., Laminin-1 induces neurite outgrowth in human mesenchymal stem cells in serum/differentiation factors-free conditions through activation of FAK-MEK/ERK signaling pathways. Biochemical and Biophysical Research Communications, vol. 391, pp. 43-48, 2010.
68. Tate, C.C., et al., Laminin and fibronectin scaffolds enhance neural stem cell transplantation into the injured brain. Journal of Tissue Engineering and Regenerative Medicine, vol. 3, pp. 208-217, 2009.
69. Hayman, M.W., et al., Growth of human stem cell-derived neurons on solid three-dimensional polymers. Journal of Biochemical and Biophysical Methods, vol. 62, pp. 231-240, 2005.
70. Martinez-Ramos, C., et al., Differentiation of postnatal neural stem cells into glia and functional neurons on laminin-coated polymeric substrates. Tissue Engineering Part A, vol. 14, pp. 1365-1375, 2008.
71. Li, S., et al., Matrix assembly, regulation, and survival functions of laminin and its receptors in embryonic stem cell differentiation. Journal of Cell Biology, vol. 157, pp. 1279-1290, 2002.
72. Hall, P.E., et al., Laminin enhances the growth of human neural stem cells in defined culture media. Bmc Neuroscience, 2008. 9.
73. Delcroix, G.J.R., et al., Adult cell therapy for brain neuronal damages and the role of tissue engineering. Biomaterials, vol. 31, pp. 2105-2120, 2010.
74. Kleinman, H.K. and G.R. Martin, Matrigel: Basement membrane matrix with biological activity. Seminars in Cancer Biology, vol. 15, pp. 378-386, 2005.
75. Donovan, P.J. and J. Gearhart, The end of the beginning for pluripotent stem cells. Nature, vol. 414, pp. 92-97, 2001.
76. Rosner, M.H., et al., A Pou-Domain Transcription Factor in Early Stem-Cells and Germ-Cells of the Mammalian Embryo. Nature, vol. 345, pp. 686-692, 1990.
77. Scholer, H.R., et al., Oct-4 - a Germline-Specific Transcription Factor Mapping to the Mouse T-Complex. Embo Journal, vol. 9, pp. 2185-2195, 1990.
78. Scholer, H.R., et al., New Type of Pou Domain in Germ Line-Specific Protein Oct-4. Nature, vol. 344, pp. 435-439, 1990.
79. Avilion, A.A., et al., Multipotent cell lineages in early mouse development depend on SOX2 function. Genes & Development, vol. 17, pp. 126-140, 2003.
80. Chambers, I., et al., Functional expression cloning of Nanog, a pluripotency sustaining factor in embryonic stem cells. Cell, vol. 113, pp. 643-655, 2003.
81. Mitsui, K., et al., The homeoprotein Nanog is required for maintenance of pluripotency in mouse epiblast and ES cells. Cell, vol. 113, pp. 631-642, 2003.
82. Carlin, R., et al., Expression of early transcription factors Oct-4, Sox-2 and Nanog by porcine umbilical cord (PUC) matrix cells. Reproductive Biology and Endocrinology, 2006. 4.
83. Darr, H., Y. Mayshar, and N. Benvenisty, Overexpression of NANOG in human ES cells enables feeder-free growth while inducing primitive ectoderm features. Development, vol. 133, pp. 1193-1201, 2006.
84. Zaehres, H., et al., High-efficiency RNA interference in human embryonic stem cells. Stem Cells, vol. 23, pp. 299-305, 2005.
85. Chambers, I. and S.R. Tomlinson, The transcriptional foundation of pluripotency. Development, vol. 136, pp. 2311-2322, 2009.
86. Masui, S., et al., Pluripotency governed by Sox2 via regulation of Oct3/4 expression in mouse embryonic stem cells. Nature Cell Biology, vol. 9, pp. 625-U26, 2007.
87. Takeda, J., S. Seino, and G.I. Bell, Human Oct3 Gene Family - Cdna Sequences, Alternative Splicing, Gene Organization, Chromosomal Location, and Expression at Low-Levels in Adult Tissues. Nucleic Acids Research, vol. 20, pp. 4613-4620, 1992.
88. Boyer, L.A., et al., Core transcriptional regulatory circuitry in human embryonic stem cells. Cell, vol. 122, pp. 947-956, 2005.
89. Looijenga, L.H.J., et al., POU5F1 (OCT3/4) identifies cells with pluripotent potential in human germ cell tumors. Cancer Research, vol. 63, pp. 2244-2250, 2003.
90. Rodda, D.J., et al., Transcriptional regulation of Nanog by Oct4 and Sox2. Journal of Biological Chemistry, vol. 280, pp. 24731-24737, 2005.
91. Otto, F., et al., Cbfa1, a candidate gene for cleidocranial dysplasia syndrome, is essential for osteoblast differentiation and bone development. Cell, vol. 89, pp. 765-771, 1997.
92. Komori, T., Regulation of osteoblast differentiation by transcription factors. Journal of Cellular Biochemistry, vol. 99, pp. 1233-1239, 2006.
93. Otto, F., H. Kanegane, and S. Mundlos, Mutations in the RUNX2 gene in patients with clelidocranial dysplasia. Human Mutation, vol. 19, pp. 209-216, 2002.
94. Kumagai, K., et al., The extent of degeneration of cruciate ligament is associated with chondrogenic differentiation in patients with osteoarthritis of the knee. Osteoarthritis and Cartilage, vol. 20, pp. 1258-1267, 2012.
95. http://www.ncbi.nlm.nih.gov/gene/6662.
96. Ng, L.J., et al., SOX9 binds DNA, activates transcription, and coexpresses with type II collagen during chondrogenesis in the mouse. Dev Biol, vol. 183, pp. 108-21, 1997.
97. Zhao, Q., et al., Parallel expression of Sox9 and Col2a1 in cells undergoing chondrogenesis. Dev Dyn, vol. 209, pp. 377-86, 1997.
98. Wright, E., et al., The Sry-Related Gene Sox9 Is Expressed during Chondrogenesis in Mouse Embryos. Nature Genetics, vol. 9, pp. 15-20, 1995.
99. Murakami, S., et al., Up-regulation of the chondrogenic Sox9 gene by fibroblast growth factors is mediated by the mitogen-activated protein kinase pathway. Proceedings of the National Academy of Sciences of the United States of America, vol. 97, pp. 1113-1118, 2000.
100. Kent, J., et al., A male-specific role for SOX9 in vertebrate sex determination. Development, vol. 122, pp. 2813-2822, 1996.
101. Aubin, J.E., et al., Intermediate filaments of the vimentin-type and the cytokeratin-type are distributed differently during mitosis. Exp Cell Res, vol. 129, pp. 149-65, 1980.
102. Chou, Y.H., et al., Intermediate Filament Reorganization during Mitosis Is Mediated by P34cdc2 Phosphorylation of Vimentin. Cell, vol. 62, pp. 1063-1071, 1990.
103. Chou, Y.H., et al., The relative roles of specific N- and C-terminal phosphorylation sites in the disassembly of intermediate filament in mitotic BHK-21 cells. Journal of Cell Science, vol. 109, pp. 817-826, 1996.
104. Chou, Y.H., et al., Nestin promotes the phosphorylation-dependent disassembly of vimentin intermediate filaments during mitosis. Molecular Biology of the Cell, vol. 14, pp. 1468-1478, 2003.
105. Eliasson, C., et al., Intermediate filament protein partnership in astrocytes. Journal of Biological Chemistry, vol. 274, pp. 23996-24006, 1999.
106. Kachinsky, A.M., J.A. Dominov, and J.B. Miller, Intermediate Filaments in Cardiac Myogenesis - Nestin in the Developing Mouse Heart. Journal of Histochemistry & Cytochemistry, vol. 43, pp. 843-847, 1995.
107. Michalczyk, K. and M. Ziman, Nestin structure and predicted function in cellular cytoskeletal organisation. Histology and Histopathology, vol. 20, pp. 665-671, 2005.
108. Otterbein, L.R., P. Graceffa, and R. Dominguez, The crystal structure of uncomplexed actin in the ADP state. Science, vol. 293, pp. 708-711, 2001.
109. Doherty, G.J. and H.T. McMahon, Mediation, modulation, and consequences of membrane-cytoskeleton interactions. Annual Review of Biophysics, vol. 37, pp. 65-95, 2008.
110. surface analysis methods in materials science. Berlin: Springer-Verlag. 1992.
111. Chadwick, K., et al., Cytokines and BMP-4 promote hematopoietic differentiation of human embryonic stem cells. Blood, vol. 102, pp. 906-915, 2003.
112. Lawyer, F.C., et al., High-level expression, purification, and enzymatic characterization of full-length Thermus aquaticus DNA polymerase and a truncated form deficient in 5’ to 3’ exonuclease activity. PCR Methods Appl, vol. 2, pp. 275-87, 1993.
113. Freeman, W.M., S.J. Walker, and K.E. Vrana, Quantitative RT-PCR: pitfalls and potential. Biotechniques, vol. 26, pp. 112-22, 124-5, 1999.
114. Chien, A., D.B. Edgar, and J.M. Trela, Deoxyribonucleic acid polymerase from the extreme thermophile Thermus aquaticus. J Bacteriol, vol. 127, pp. 1550-7, 1976.
115. Simmons, P.J. and B. Torokstorb, Identification of Stromal Cell Precursors in Human Bone-Marrow by a Novel Monoclonal-Antibody, Stro-1. Blood, vol. 78, pp. 55-62, 1991.
116. Gilliland, G., et al., Analysis of cytokine mRNA and DNA: detection and quantitation by competitive polymerase chain reaction. Proc Natl Acad Sci U S A, vol. 87, pp. 2725-9, 1990.
117. Rappolee, D.A., et al., Wound macrophages express TGF-alpha and other growth factors in vivo: analysis by mRNA phenotyping. Science, vol. 241, pp. 708-12, 1988.
118. Becker-Andre, M. and K. Hahlbrock, Absolute mRNA quantification using the polymerase chain reaction (PCR). A novel approach by a PCR aided transcript titration assay (PATTY). Nucleic Acids Res, vol. 17, pp. 9437-46, 1989.
119. Zhang, S.C., et al., In vitro differentiation of transplantable neural precursors from human embryonic stem cells. Nat Biotechnol, vol. 19, pp. 1129-33, 2001.
120. Cheng, S., et al., Effective amplification of long targets from cloned inserts and human genomic DNA. Proc Natl Acad Sci U S A, vol. 91, pp. 5695-9, 1994.
121. Holland, P.M., et al., Detection of specific polymerase chain reaction product by utilizing the 5’----3’ exonuclease activity of Thermus aquaticus DNA polymerase. Proc Natl Acad Sci U S A, vol. 88, pp. 7276-80, 1991.
122. Chen, C.F., et al., Real-time quantification of microRNAs by stem-loop RT-PCR. Nucleic Acids Research, vol. 33, 2005.
123. Niesters, H.G.M., Quantitation of viral load using real-time amplification techniques. Methods, vol. 25, pp. 419-429, 2001.
124. Livak, K.J. and T.D. Schmittgen, Analysis of relative gene expression data using real-time quantitative PCR and the 2(T)(-Delta Delta C) method. Methods, vol. 25, pp. 402-408, 2001.
125. Schmittgen, T.D. and K.J. Livak, Analyzing real-time PCR data by the comparative C-T method. Nature Protocols,vol. 3, pp. 1101-1108, 2008.
126. Higuchi, A., et al., Physical cues of biomaterials guide stem cell differentiation fate. Chem Rev, vol. 113, pp. 3297-328, 2013.

指導教授

樋口亞绀(Akon Higuchi)

審核日期

2013-7-8

推文