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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 吳信志 | |
dc.contributor.author | Ya-Ping Yen | en |
dc.contributor.author | 顏雅萍 | zh_TW |
dc.date.accessioned | 2021-06-15T05:02:47Z | - |
dc.date.available | 2012-07-30 | |
dc.date.copyright | 2010-07-30 | |
dc.date.issued | 2010 | |
dc.date.submitted | 2010-07-28 | |
dc.identifier.citation | Alhadlaq, A., and J. J. Mao. 2004. Mesenchymal stem cells: isolation and therapeutics. Stem Cells Dev. 13(4): 436-448.
Aranda, P., X. Agirre, E. Ballestar, E. J. Andreu, J. Roman-Gomez, I. Prieto, J. I. Martin-Subero, J.C. Cigudosa, R. Siebert, M. Esteller, and F. Prosper. 2009. Epigenetic Signatures Associated with Different Levels of Differentiation Potential in Human Stem Cells. PLoS One 4(11): e7809. Baddoo, M., K. Hill, R. Wilkinson, D. Gaupp, C. Hughes, G.C. Kopen, and D.G. Phinney. 2003. Characterization of mesenchymal stem cells isolated from murine bone marrow by negative selection. J. Cell. Biochem. 89: 1235-1249. Barzilay, R., E. Melamed, and D. Offen. 2009. Introducing transcription factors to multipotent mesenchymal stem cells: making transdifferentiation possible. Stem Cells 27(10): 2509-2515. Beyer Nardi, N., and L. Silva Meirelles. 2006. Mesenchymal stem cells: isolation, in vitro expansion and characterization. Handb. Exp. Pharmacol. 174: 249-252. Bohrnsen, F., U. Lindner, M. Meier, A. Gadallah, P. Schlenke, H. Lehnert, J. Rohwedel, and J. Kramer. 2009. Murine mesenchymal progenitor cells from different tissues differentiated via mesenchymal microspheres into the mesodermal direction. BMC Cell Biol. 10: 92. Breyer, A., N. Estharabadi, M. Oki, F. Ulloa, M. Nelson-Holte, L. Lien, and Y. Jiang. 2006. Multipotent adult progenitor cell isolation and culture procedures. Exp. Hematol. 34(11): 1596-1601. Bruder, S.P., D. J. Fink, and A. I. Caplan. 1994. Mesenchymal stem cells in bone development, bone repair, and skeletal regeneration therapy. J. Cell. Biochem. 56(3): 283-294. Clarke, D.L., C.B. Johansson, J. Wilbertz, B. Veress, E. Nilsson, H. Karlstrom, U. Lendahl, and J. Frisen. 2000. Generalized potential of adult neural stem cells. Science 288(5471): 1660-1663. Conget, P.A., and J. J. Minguell. 1999. Phenotypical and functional properties of human bone marrow mesenchymal progenitor cells. J. Cell. Physiol. 181(1): 67-73. Deng, Y.B., Q. T. Yuan, X. G. Liu, X. L. Liu, Y. Liu, Z. G. Liu, and C. Zhang. 2005. Functional recovery after rhesus monkey spinal cord injury by transplantation of bone marrow mesenchymal-stem cell-derived neurons. Chin. Med. J. (Engl). 118(18): 1533-1541. Dexter, T. M., E. Spooncer, P. Simmons, and T. D. Allen. 1984. Long-term marrow culture: an overview of techniques and experience. Kroc. Found. Ser. 18: 57-96. D’Ippolito, G., S. Diabira, G. A. Howard, P. Menei, B. A. Roos, and P. C. Schiller. 2004. Marrow-isolated adult multilineage inducible (MIAMI) cells, a unique population of postnatal young and old human cells with extensive expansion and differentiation potential. J. Cell. Sci. 117: 2971–2981. Eslaminejad, M.B., A. Nikmahzar, L. Taghiyar, S. Nadri, and M. Massumi. 2006. Murine mesenchymal stem cells isolated by low density primary culture system. Dev. Growth. Differ. 48: 361-370. Feng, B., J. H. Ng, J. C. Heng, and H. H. Ng. 2009. Molecules that promote or enhance reprogramming of somatic cells to induced pluripotent stem cells. Cell Stem Cell 4(4): 301-312. Friedenstein, A., R. Chailakhyan, and U. Gerasimov. 1987. Bone marrow osteogenic stem cells: in vitro cultivation and transplantation in diffusion chambers. Cell Tissue Kinet. 20: 263-272. Gang, E. J., D. Bosnakovski, C. A. Figueiredo, J. W. Visser, and R. C. Perlingeiro. 2007. SSEA-4 identifies mesenchymal stem cells from bone marrow. Blood 109(4): 1743-1751. Greco, S. J., K. Liu, and P. Rameshwar. 2007. Functional similarities among genes regulated by OCT4 in human mesenchymal and embryonic stem cells. Stem Cells 25(12): 3143-3154. Guo, G., J. Yang, J. Nichols, J. S. Hall, I. Eyres, W. Mansfield, and A. Smith. 2009. Klf4 reverts developmentally programmed restriction of ground state pluripotency. Development 136(7): 1063-1069. Hansis, C., J. A. Grifo, and L. C. Krey. 2000. Oct-4 expression in inner cell mass and trophectoderm of human blastocysts. Mol. Hum. Reprod. 6(11): 999-1004. Hermann, A., R. Gastl, S. Liebau, M. O. Popa, J. Fiedler, B. O. Boehm, M. Maisel, H. Lerche, J. Schwarz, R. Brenner, and A. Storch. 2004. Efficient generation of neural stem cell-like cells from adult human bone marrow stromal cells. J. Cell. Sci. 1;117(Pt 19): 4411-4422. In 't Anker, P.S., S. A. Scherjon, C. Kleijburg-van der Keur, G. M. Groot-Swings, F. H. Claas, W. E. Fibbe, and H. H. Kanhai. 2004. Isolation of mesenchymal stem cells of fetal or maternal origin from human placenta. Stem Cell 22(7): 1338-1345. Jaiswal, R. K., N. Jaiswal, S. P. Bruder, G. Mbalaviele, D. R. Marshak, and M. F. Pittenger. 2000. Adult human mesenchymal stem cell differentiation to the osteogenic or adipogenic lineage is regulated by mitogen-activated protein kinase. J. Biol. Chem. 275(13): 9645-9652. Jiang, Y., B. N. Jahagirdar, R. L. Reinhardt, R. E. Schwartz, C. D. Keene, X. R. Ortiz-Gonzalez, M. Reyes, T. Lenvik, T. Lund, M. Blackstad, J. Du, S. Aldrich, A. Lisberg, W. C. Low, D. A. Largaespada, and C. M. Verfaillie. 2002. Pluripotency of mesenchymal stem cells derived from adult marrow. Nature 418(6893): 41-49. Johnstone, B., T. M. Hering, and A. I. Caplan. 1998. In vitro chondrogenesis of bone marrow-derived mesenchymal progenitor cells. Exp. Cell. Res. 238: 265-272. Kang, X.Q., W. J. Zang, T. S. Song, X. L. Xu, X. J. Yu, D. L. Li, K. W. Meng, S. L. Wu, Z. Y. Zhao. 2005. Rat bone marrow mesenchymal stem cells differentiate into hepatocytes in vitro. World J. Gastroenterol. 11(22): 3479-3484. Kucia, M., W. Wu, and M. Z. Ratajczak. 2007. Bone marrow-derived very small embryonic-like stem cells their developmental origin and biological significance. Dev. Dyn. 236(12): 3309-3320. Lee, K.H., C. K. Chuang, H. W. Wang, L. Stone, C. H. Chen, and C. F. Tu. 2007. An alternative simple method for mass production of chimeric embryos by coculturing denuded embryos and embryonic stem cells in Eppendorf vials. Theriogenology 67(2): 228-237. Lee, O. K., T. K. Kuo, W. M. Chen, K. D. Lee, S. L. Hsieh, and T. H. Chen. 2004. Isolation of multipotent mesenchymal stem cells from umbilical cord blood. Blood 103(5): 1669-1675. Lengner, C.J., F. D. Camargo, K. Hochedlinger, G. G. Welstead, S. Zaidi, S. Gokhale, H. R. Scholer, A. Tomilin, and R. Jaenisch. 2007. Oct4 expression is not required for mouse somatic stem cell self-renewal. Cell Stem Cell 1(4): 403-415. Liedtke, S., M. Stephan, and G. Kogler. Oct4 expression revisited: potential pitfalls for data misinterpretation in stem cell research. 2008. Biol. Chem. 389(7): 845-850. Li, Y., J. Chen, L. Wang, L. Zhang, M. Lu, and M. Chopp. 2001. Intracerebral transplantation of bone marrow stromal cells in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model of Parkinson's disease. Neurosci. Lett. 316(2): 67-70. Mackay, A. M., S. C. Beck, J. M. Murphy, F. P. Barry, C. O. Chichester, M. F. Pittenger. 1998. Chondrogenic differentiation of cultured human mesenchymal stem cells from marrow. Tissue Eng. 4(4): 415-428. Martin, D. R., N. R. Cox, and T. L. Hathcock. 2002. Isolation and characterization of multipotential mesenchymal stem cells from feline bone marrow. Exp. Hematol. 30: 879-886. Martin G. R. 1981. Isolationof a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells. Proc. Natl. Acad. Sci. 78: 7634-7638. Martin, G. R., M. J. Evans. 1975. Differentiation of clonal lines of teratocarcinoma cells: formation of embryoid bodies in vitro. Proc. Natl. Acad. Sci. 72(4): 1441-1445. Miyahara, Y., N. Nagaya, M. Kataoka, B. Yanagawa, K. Tanaka, H. Hao, K. Ishino, H. Ishida, T. Shimizu, K. Kangawa, S. Sano, T. Okano, S. Kitamura, and H. Mori. 2006. Monolayered mesenchymal stem cells repair scarred myocardium after myocardial infarction. Nat. Med. 12(4): 459-465. Moscoso, I., A. Centeno, E. Lopez, J.I. Rodriguez-Barbosa, and I. Filgueira. 2005. Differentiation in vitro of primary and immortalized porcine mesenchymal stem cells into cardiomyocytes for cell transplantation. Transplant. Proc. 37: 481-482. Nagai, A., W. K. Kim, H. J. Lee, H. S. Jeong, K. S. Kim, S. H. Hong, I. H. Park, and S. U. Kim. 2007. Multilineage potential of stable human mesenchymal stem cell line derived from fetal marrow. PLoS One 2(12): e1272. Nekanti, U., V. B. Rao, A. G. Bahirvani, M. Jan, S. Totey, and M. Ta. 2009. Long-term Expansion and Pluripotent Marker Array Analysis of Wharton's Jelly-Derived Mesenchymal Stem Cells. Stem Cells Dev. 19(1): 117-130. Ovitt, C. E., and H. R. Scholer. 1998. The molecular biology of Oct-4 in the early mouse embryo. Mol. Hum. Reprod. 4(11): 1021-1031. Peister, A., J. A. Mellad, B. L. Larson, B. M. Hall, L. F. Gibson, and D. J. Prockop. 2004. Adult stem cells from bone marrow (MSCs) isolated from different strains of inbred mice vary in surface epitopes, rates of proliferation, and differentiation potential. Blood 103:1662–1668. Phinney, D.G., G. Kopen, R.L. Isaacson, and D.J. Prockop. 1999. Plastic adherent stromal cells from the bone marrow of commonly used strains of inbred mice: variations in yield , growth and differentiation. J. Cell. Biochem. 72: 570-585. Pittenger, F., A. Mackay, and S. Beck. 1999. Multilineage potential of adult human mesenchymal stem cells. Science 284: 143-147. Reyes, M., A. Dudek, B. Jahagirdar, L. Koodie, P. H. Marker, and C. M. Verfaillie. 2002. Origin of endothelial progenitors in human postnatal bone marrow. J. Clin. Invest. 109(3): 337-346. Reyes, M., and C. M. Verfaillie. 2001. Characterization of multipotent adult progenitor cells, a subpopulation of mesenchymal stem cells. Ann. N. Y. Acad. Sci. 938:231-233. Riekstina, U., I. Cakstina, V. Parfejevs, M. Hoogduijn, G. Jankovskis, I. Muiznieks, R. Muceniece, and J. Ancans. 2009. Embryonic stem cell marker expression pattern in human mesenchymal stem cells derived from bone marrow, adipose tissue, heart and dermis. Stem Cell Rev. 5(4): 378-386. Roche, S., B. Delorme, R. A. Oostendorp, R. Barbet, D. Caton, D. Noel, K. Boumediene, H. A. Papadaki, B. Cousin, C. Crozet, O. Milhavet, L. Casteilla, J. Hatzfeld, C. Jorgensen, P. Charbord, and S. Lehmann. 2009. Comparative proteomic analysis of human mesenchymal and embryonic stem cells towards the definition of a mesenchymal stem cell proteomic signature. Proteomics 9(2): 223-232. Sanchez-Ramos, J., S. Song, F. Cardozo-Pelaez, C. Hazzi, T. Stedeford, A. Willing, T. B. Freeman, S. Saporta, W. Janssen, N. Patel, D. R. Cooper, and P. R. Sanberg. 2000. Adult bone marrow stromal cells differentiate into neural cells in vitro. Exp. Neurol. 164(2):247-256. Schuh, E. M., M. S. Friedman, D. D. Carrade, J. Li, D. Heeke, S. M. Oyserman, L. D. Galuppo, D. J. Lara, N. J. Walker, G. L. Ferraro, S. D. Owens, and D. L. Borjesson. 2009. Identification of variables that optimize isolation and culture of multipotent mesenchymal stem cells from equine umbilical-cord blood. Am. J .Vet. Res. 70(12): 1526- 1535. Schwartz, R.E., M. Reyes, L. Koodie, Y. Jiang, M. Blackstad, T. Lund, T. Lenvik, S. Johnson, W. S. Hu, and C. M. Verfaillie. 2002. Multipotent adult progenitor cells from bone marrow differentiate into functional hepatocyte-like cells. J .Clin. Invest. 109(10): 1291-1302. Spees, J. L., S. D. Olson, J. Ylostalo, P. J. Lynch, J. Smith, A. Perry, A. Peister, M. Y. Wang, and D. J. Prockop. 2007. Differentiation, cell fusion, and nuclear fusion during ex vivo repair of epithelium by human adult stem cells from bone marrow stroma. Proc. Natl. Acad. Sci. 100(5): 2397-2402. Shi, Q., S. Rafii, M. H. Wu, E. S. Wijelath, C. Yu, A. Ishida, Y. Fujita, S. Kothari, R .Mohle, L. R. Sauvage, M. A. Moore, R. F. Storb, and W. P. Hammond. 1998. Evidence for circulating bone marrowderived endothelial cells. Blood 92: 362–367. Silva J, and A. Smith. 2008. Capturing pluripotency. Cell 132(4): 532-536. Sun, S., Z. Guo, X. Xiao, B. Liu, X. Liu, P. H. Tang, and N. Mao. 2003. Isolation of mouse marrow mesenchymal progenitors by a novel and reliable method. Stem Cell 21: 527-535. Takahashi, K., and S. Yamanaka. 2006. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126(4): 663-676. Tao, X. R., W. L. Li, J. Su, C. X. Jin, X. M. Wang, J. X. Li, J. K. Hu, Z. H. Xiang, J. T. Lau, and Y. P. Hu. 2009. Clonal mesenchymal stem cells derived from human bone marrow can differentiate into hepatocyte-like cells in injured livers of SCID mice. J. Cell. Biochem. 108(3): 693-704. Tsai, M.S., J. L. Lee, Y. J. Chang, and S. M. Hwang. 2004. Isolation of human multipotent mesenchymal stem cells from second-trimester amniotic fluid using a novel two-stage culture protocol. Hum. Reprod. 19(6): 1450-1456. Tavassoli, M., and K. Takahashi. 1982. Morphological studies on long-term culture of marrow cells: characterization of the adherent stromal cells and their interactions in maintaining the proliferation of hemopoietic stem cells. Am. J. Anat. 164: 91-111. Temple S. 2001. Stem cell plasticity — building the brain of our dreams. Nat. Rev. Neurosci. 2(7): 513-520. Terada, N., T. Hamazaki, M. Oka, M. Hoki, D. M. Mastalerz, Y. Nakano, E. M. Meyer, L. Morel, B. E. Petersen, and E. W. Scott. 2002. Bone marrow cells adopt the phenotype of other cells by spontaneous cell fusion. Nature 416(6880): 542-545. Thomson, J.A., J. Kalishman, T. G. Golos, M. Durning, C. P. Harris, R. A. Becker, and J. P. Hearn. 1995. Isolation of a primate embryonic stem cell line. Proc. Natl. Acad. Sci. 92: 7844-7848. Toyooka, Y., N. Tsunekawa, R. Akasu, and T. Noce. 2003. Embryonic stem cells can form germ cells in vitro. Proc. Natl. Acad. Sci. 100: 11457-11462. Tropel, P., D. Noel, N. Platet, P. Legrand, A.L. Benabid, and F. Berger. 2004. Isolation and characterization of mesenchymal stem cells from adult mouse bone marrow. Exp. Cell. Res. 295: 395-406. Tsou, C. C., C. F. Tsai, Y. H. Tsui, P. R. Sudhir, Y. T. Wang, Y. J. Chen, J. Y. Chen, T. Y. Sung, and W. L. Hsu. 2009. IDEAL-Q, an automated tool for label-free quantitation analysis using an efficient peptide alignment approach and spectral data validation. Mol. Cell. Proteomics. 9(1): 131-144. Urban, V. S., J. Kiss, J. Kovacs, E. Gocza, V. Vas, E. Monostori, and F. Uher. 2008. Mesenchymal stem cells cooperate with bone marrow cells in therapy of diabetes. Stem Cells 26(1): 244-253. Wang, T., Z. Xu, W. Jiang, and A. Ma. 2006. Cell-to-cell contact induces mesenchymal stem cell to differentiate into cardiomyocyte and smooth muscle cell. Int. J. Cardiol. 109(1): 74-81. Woodbury, D., E. J. Schwarz, D. J. Prockop, and I. B. Black. 2000. Adult rat and human bone marrow stromal cells differentiate into neurons. J. Neurosci. Res. 61(4): 364-370. Young, R.G., D. L. Butler, W. Weber, A. I. Caplan, S. L. Gordon, and D. J. Fink. 1998. Use of mesenchymal stem cells in a collagen matrix for Achilles tendon repair. J. Orthop. Res. 16(4): 406-413. Zavan, B., C. Giorgi, G. P. Bagnara, V. Vindigni, G. Abatangelo, and R. Cortivo. 2007. Osteogenic and chondrogenic differentiation: comparison of human and rat bone marrow mesenchymal stem cells cultured into polymeric scaffolds. Eur. J. Histochem. 51(1): 1-8. Zscharnack, M., C. Poesel, J. Galle, and A. Bader. 2009. Low oxygen expansion improves subsequent chondrogenesis of ovine bone-marrow-derived mesenchymal stem cells in collagen type I hydrogel. Cells Tissues Organs 190(2): 81-93. Zuckerman, K. S., and M. S. Wicha. 1983. Extracelluar matrix production by the adherent cells of long-term murine bone marrow cultures. Blood 61: 5 | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/46307 | - |
dc.description.abstract | 近年來幹細胞及再生醫學相關研究蓬勃發展,尤其是「胚幹細胞」(embryonic stem cells, ESCs) 及諸多不同組織來源之「成體幹細胞」(adult stem cells, ASCs)已被成功分離、純化與定性,且大多已證明具備多分化潛能之特性。目前並有研究指稱將成體幹細胞藉由顯微操作方式送入囊胚腔後,確可成功分化為各胚層細胞。有鑑於此,本研究室選用源自小鼠骨髓之成體幹細胞作為試驗處理組,並以源自小鼠胚幹細胞作為本研究之對照組,觀察比較源自成體幹細胞與胚幹細胞二者分別於早期胚中嵌入及分化之潛能。
研究中所使用之綠色螢光小鼠之骨髓間葉幹細胞 (enhanced green fluorescent protein-mesenchymal stem cells, EGFP-mMSCs; type I and II) 乃分離自攜帶β-actin啟動子之綠色螢光蛋白 (enhanced green fluorescent protein, EGFP) 之轉基因小鼠骨髓液或骨頭,並分別以兩種純化方式,分離獲得之小鼠骨髓及骨間葉幹細胞。在完成初代培養並經純化獲得之EGFP-mMSCs type I及II,以不同細胞數為組別,分別送入受精後第3.5日齡小鼠胚之囊胚腔中,再經胚移置於代理孕母子宮中,俟胚在體內發育達第6.5及9.5日之際,分別進行觀察各胎小鼠對EGFP之進一步表現情形,藉由螢光顯微鏡觀察外並透過聚合酶連鎖反應 (polymerase chain reaction, PCR) 分析,試驗結果確認EGFP於胚胎6.5及9.5日之表現,顯示EGFP-mMSCs以囊胚注射方式於早期胚胎不具有分化能力。試驗二中,乃以8細胞至桑椹胚 (morula) 時期胚細胞與EGFP-mMSCs type I及II進行細胞聚合方式,並於體外培養至3.5天囊胚時期及4.5天孵化 (hatch) 時期,再透過免疫螢光染色(immuno-fluorescence stain) 進行偵測EGFP-mMSCs type I及II於胚胎內之表現情形。試驗結果亦顯示,前述方法聚合之成體幹細胞,亦未達順利於早期胚胎發育中進行分化及增生。 由以上結果顯示,小鼠胚幹細胞以囊胚注射方式將細胞送入囊胚後可與囊胚共同增生分化至成體,然而骨髓及骨間葉幹細胞卻無此能力,與原先假設骨髓及骨間葉幹細胞具有多分化潛能有所出入,同時為探究為何EGFP-mMSCs type I及II於胚胎發育過程中即不再具有表現,初步懷疑是否此種細胞於胚胎發育環境下會進行細胞凋亡作用 (apoptosis),因此試驗過程中以半胱天冬酶-3 (caspase-3) 與EGFP進行免疫螢光複染,然而目前尚無任何證據可以證明EGFP-mMSCs於胚胎體內消失的原因為何。 骨髓間葉幹細胞雖於體外不同條件培養後可分化為各個胚層的細胞,堪稱為具有多能性之幹細胞。但在體內試驗中,因其生長環境不符合其分化條件,且此種細胞乃為成體分離之幹細胞,與早期囊胚內之未分化微環境 (niche) 相異,且內細胞群細胞 (inner cell mass) 與骨髓間葉幹細胞之膜蛋白質組成或許有極大差別,以至於它們在細胞之間的聚合作用亦有很大的阻礙。綜合以上所述,在本試驗研究條件下,骨髓間葉幹細胞之多能性表現只受限於適當的體外分化條件下始具有此特性。 | zh_TW |
dc.description.abstract | In recent years, many tissue specific stem cells had been successfully isolated and transdifferentiated into lineages other than the tissue of origin. Several studies have reported that bone marrow derived mesenchymal stem cells (MSCs) possess the potency to differentiate into various lineages. Recent study also indicated that a tissue specific stem cell, neural stem cells could contribute to many tissues of the chimera mouse when injected into a blastocyst. Therefore, the object of this study was to investigate the differentiation potential of MSCs in vivo. The ability to contribute to the embryogenesis was determined by introducing them into the early embryonic environment and observing the fate of their progeny.
In this study, type I and type II EGFP-mMSCs were isolated from the femur of transgenic mice carrying s-actin promoter constructed with enhanced green fluorescent protein (EGFP) cDNA serving as a tracing marker. Grouping by cell numbers, EGFP-mMSCs were injected to C57BL/6JNarl blastocysts and injected embryos were transferred to the uterus of the pseudo-pregnant foster mothers to assess the potency of EGFP-mMSCs to integrate into inner cell mass (ICM) during early embryonic development. The results of fluorescence microscopy and polymerase chain reaction analysis showed that EGFP-mMSCs injected embryos failed to give rise to chimeras and EGFP signal could not be observed in conceptuses day 9.5 and day 6.5. In contrast, EGFP embryonic stem (ESCs) could proliferate spontaneously with the ICM and contribute to all cell lineages in vivo. Compared with ESCs, mMSCs don’t have the ability to differentiate in vivo. To test whether mMSCs have the stem cell plasticity via cell aggregation, type I and II EGFP-mMSCs were aggregated with eight-cell or morula stage embryos and further cultured to blastocyst and hatching stages. Analysis on preimplantation embryos by immunofluorescence stain showed that type I and II EGFP-mMSCs could not aggregate with early embryos, suggesting that these somatic stem cells are unable to participate in early embryogenesis. Results in this thesis revealed that the hypothesis may be false that mMSCs could not integrate into the host embryos during early embryogenesis. The reason injected mMSCs disappear in day 6.5 embryos is required to be clarified, but cell death is a likely answer. Putatively, mMSCs could not aggregate with eight-cell or morula stage embryos which may due to the incompatible cell niche or microenvironment. In addition, the composition of membranous protein of mMSCs and inner cell mass may be enormously different which could be the reason why these cells would not aggregate with each other. Taken together, although mMSCs can differentiate to tri-lineage tissues via in vitro induction, the microenvironment in early embryos might not support for mMSCs survival and differentiation. | en |
dc.description.provenance | Made available in DSpace on 2021-06-15T05:02:47Z (GMT). No. of bitstreams: 1 ntu-99-R97626018-1.pdf: 2097621 bytes, checksum: 3f715f0392693dbe80d44d1e32133b2d (MD5) Previous issue date: 2010 | en |
dc.description.tableofcontents | 國立臺灣大學碩士學位論文口試委員會審定書.................I
致謝..................................II 目錄..................................III 摘要..................................IX Abstract................................XI 第壹章、緒論..............................1 第貳章、文獻檢討............................2 2.1 幹細胞(Stem cells).......................2 2.1.1 幹細胞介紹.......................2 2.1.2 胚幹細胞(embryonic stem cells, ESCs)...........8 2.1.3 骨髓間葉幹細胞 (bone marrow mesenchymal stem cells, BMMSCs)........................8 2.2 幹細胞之應用研究......................11 2.2.1 胚幹細胞之應用研究..................11 2.2.2 骨髓間葉幹細胞分化為多分化潛能幹細胞之應用研究......12 2.2.3 應用成體幹細胞分化為多分化潛能幹細胞之研究........13 第參章、試驗研究............................15 試驗一:綠色螢光小鼠間葉幹細胞之分離和純化...............15 一、前言............................15 二、材料與方法..........................16 2.1 實驗動物........................16 2.1.1 綠色螢光蛋白質基因轉殖小鼠的產製..........16 2.1.2 小鼠來源.....................16 2.2 小鼠骨髓間葉幹細胞(Type I及II)...............16 2.2.1 綠色螢光小鼠骨髓間葉幹細胞的取得與純化(Type I)...16 2.2.2 綠色螢光小鼠骨間葉幹細胞的取得與純化(Type II).....19 2.2.3 綠色螢光小鼠骨髓間葉幹細胞培養液之配置.......21 2.2.4 綠色螢光小鼠骨髓間葉幹細胞的培養與保存.......21 2.2.5 細胞計數.....................21 三、結果與討論..........................22 3.1 綠色螢光小鼠骨髓間葉幹細胞type I及II之分離.........22 3.2 綠色螢光小鼠骨髓間葉幹細胞之純化..............22 試驗二 : 綠色螢光小鼠骨髓間葉幹細胞囊胚注射..............25 一、前言............................25 二、材料方法.........................26 2.1 試驗動物........................26 2.2 綠色螢光小鼠之骨髓間葉幹細胞來源............26 2.3 綠色螢光小鼠胚幹細胞來源.................26 2.4 囊胚顯微注射操作流程...................27 2.5 螢光解剖顯微鏡觀察EGFP-mMSCs經由囊胚注射後於體內之表現..29 2.6 藉由聚合酶連鎖反應 (Polymerase chain reaction, PCR)偵測EGFP-mMSCs於囊胚注射後之表現.........29 2.6.1 總DNA萃取.....................29 2.6.2 聚合酶連鎖反應(Polymerase chain reaction, PCR)...29 2.6.3 瓊膠電泳分析...................30 三、結果討論.........................31 3.1 EGFP-mMSCs type I及II經囊胚注射之結果...........31 3.2 PCR偵測EGFP-mMSCs type I及II於囊胚注射後之表現......38 試驗三 : 以8細胞時期之胚葉細胞與綠色螢光小鼠骨髓間葉幹細胞進行聚合(Aggregation)......................43 一、前言...........................43 二、材料與方法........................45 2.1 實驗動物.......................45 2.2 綠色螢光小鼠之骨及骨髓間葉幹細胞來源...........45 2.3 綠色螢光小鼠胚幹細胞來源................45 2.4 8細胞期胚與小鼠骨髓幹細胞之聚合作用..........45 2.5 免疫螢光染色 (Immunofluorescence)............48 三、結果與討論........................49 3.1 EGFP-ESCs與8細胞時期胚進行聚合作用...........50 3.2 EGFP-mMSCs type I及II與8細胞時期胚進行聚合作用......51 第肆章、綜合討論............................57 第伍章 結論...............................59 參考文獻...............................60 附錄.................................70 | |
dc.language.iso | zh-TW | |
dc.title | 綠色螢光小鼠間葉幹細胞於早期胚胎發育之分化潛能 | zh_TW |
dc.title | The Differentiation Potential of Mesenchymal Stem Cells Isolated from EGFP Transgenic Mice in Early Embryogenesis | en |
dc.type | Thesis | |
dc.date.schoolyear | 98-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 鄭登貴,林淑華,唐品琦,劉逸軒 | |
dc.subject.keyword | 成體幹細胞,胚幹細胞,骨髓間葉幹細胞,多能性,囊胚, | zh_TW |
dc.subject.keyword | adult stem cells,embryonic stem cells,mesenchymal stem cells,pluripotent,blastocys, | en |
dc.relation.page | 86 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2010-07-28 | |
dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
dc.contributor.author-dept | 動物科學技術學研究所 | zh_TW |
顯示於系所單位: | 動物科學技術學系 |
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