Skip to main content
SpringerLink
Log in

Human Embryonic Stem Cells for Tissue Engineering

  • Protocol
Tissue Engineering

Part of the book series: Methods in Molecular Medicine™ ((MIMM,volume 140))

summary

Human embryonic stem cells (HESCs) are characterized by their ability to self-renew and capacity to differentiate into almost every cell type. As a result, they have enormous potential for use in tissue engineering and transplantation therapy. If these cells can be induced to differentiate into a particular cell type, they may provide an almost unlimited source of cells for transplantation for treating certain diseases where normal cell function is impaired. The challenge lies in the development of techniques to induce differentiation into a specific cell type, to enrich for that population, and to isolate it. It is essential that the starting material, the undifferentiated embryonic stem cell line, is growing under optimal conditions that preserve its pluripotent potential and maintain a stable karyotype. This review will discuss methods for the growth, maintenance, and spontaneous differentiation of HESCs and methods to genetically manipulate them.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 129.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Evans, M. J. and Kaufman, M. H. (1981) Establishment in culture of pluripotential cells from mouse embryos. Nature 292, 154–156.

    Article  CAS  Google Scholar 

  2. Martin, G. R. (1981) Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells. Proc. Natl. Acad. Sci. U. S. A. 78, 7634–7638.

    Article  CAS  Google Scholar 

  3. Thomson, J. A., Itskovitz-Eldor, J., Shapiro, S. S, Waknitz, M. A., Swiergiel, J. J., Marshall, V. S., and Jones, J. M. (1998) Embryonic stem cell lines derived from human blastocysts. Science 282, 1145–1147.

    Article  CAS  Google Scholar 

  4. Amit, M., Carpenter, M. K., Inokuma, M. S., Chiu, C. P., Harris, C. P., Waknitz, M. A., Itskovitz-Eldor, J., and Thomson, J. A. (2000) Clonally derived human embryonic stem cell lines maintain pluripotency and proliferative potential for prolonged periods of culture. Dev. Biol. 227, 271–278.

    Article  CAS  Google Scholar 

  5. Maitra, A., Arking, D. E., Shivapurkar, N., Ikeda, M., Stastny, V., Kassauei, K., Sui, G., Cutler, D. J, Liu, Y, Brimble, S. N., Noaksson, K., Hyllner, J., Schulz, T. C., Zeng, X., Freed, W. J., Crook, J., Abraham, S., Colman, A., Sartipy, P., Matsui, S., Carpenter, M., Gazdar, A. F., Rao, M., and Chakravarti, A. (2005) Genomic alterations in cultured human embryonic stem cells. Nat. Genet. 37, 1099–1103.

    Article  CAS  Google Scholar 

  6. Brimble, S. N., Zeng, X., Weiler, D. A., Luo, Y., Liu, Y., Lyons, I. G., Freed, W. J., Robins, A. J., Rao, M. S., and Schulz, T. C. (2004) Karyotypic stability, genotyping, differentiation, feeder-free maintenance, and gene expression sampling in three human embryonic stem cell lines derived prior to August 9, 2001. Stem Cells Dev. 13, 585–597.

    Article  CAS  Google Scholar 

  7. Buzzard, J. J., Gough, N. M., Crook, J. M., and Colman, A. (2004) Karyotype of human ES cells during extended culture. Nat. Biotechnol. 22, 381–382; author reply 382.

    Article  CAS  Google Scholar 

  8. Hoffman, L. M. and Carpenter, M. K. (2005) Characterization and culture of human embryonic stem cells. Nat. Biotechnol. 23, 699–708.

    Article  CAS  Google Scholar 

  9. Reubinoff, B. E., Pera, M. F., Fong, C. Y., Trounson, A., and Bongso, A. (2000) Embryonic stem cell lines from human blastocysts: somatic differentiation in vitro. Nat. Biotechnol. 18, 399–404.

    Article  CAS  Google Scholar 

  10. Smith, A. G., Heath, J. K., Donaldson, D. D., Wong, G. G., Moreau, J., Stahl, M., and Rogers, D. (1988) Inhibition of pluripotential embryonic stem cell differentiation by purified polypeptides. Nature 336, 688–690.

    Article  CAS  Google Scholar 

  11. Williams, R. L., Hilton, D. J., Pease, S., Willson, T. A., Stewart, C. L., Gearing, D. P., Wagner, E. F., Metcalf, D., Nicola, N. A., and Gough, N. M. (1988) Myeloid leukaemia inhibitory factor maintains the developmental potential of embryonic stem cells. Nature 336, 684–687.

    Article  CAS  Google Scholar 

  12. Humphrey, R. K., Beattie, G. M., Lopez, A. D., Bucay, N., King, C. C., Firpo, M. T., Rose-John, S., and Hayek, A. (2004) Maintenance of pluripotency in human embryonic stem cells is STAT3 independent. Stem Cells 22, 522–530.

    Article  CAS  Google Scholar 

  13. Daheron, L., Opitz, S. L., Zaehres, H., Lensch, W. M., Andrews, P. W., Itskovitz-Eldor, J., and Daley, G. Q. (2004) LIF/STAT3 signaling fails to maintain self-renewal of human embryonic stem cells. Stem Cells 22, 770–778.

    Article  CAS  Google Scholar 

  14. Xu, R. H., Peck, R. M., Li, D. S., Feng, X., Ludwig, T., and Thomson, J. A. (2005) Basic FGF and suppression of BMP signaling sustain undifferentiated proliferation of human ES cells. Nat. Methods 2, 185–190.

    Article  CAS  Google Scholar 

  15. Klimanskaya, I., Chung, Y., Meisner, L., Johnson, J., West, M. D., and Lanza, R. (2005) Human embryonic stem cells derived without feeder cells. Lancet 365, 1636–1641.

    Article  CAS  Google Scholar 

  16. Wang, L., Li, L., Menendez, P., Cerdan, C., and Bhatia, M. (2005) Human embryonic stem cells maintained in the absence of mouse embryonic fibroblasts or conditioned medium are capable of hematopoietic development. Blood 105, 4598–4603.

    Article  CAS  Google Scholar 

  17. Wang, G., Zhang, H., Zhao, Y., Li, J., Cai, J., Wang, P., Meng, S., Feng, J., Miao, C., Ding, M., Li, D., and Deng, H. (2005) Noggin and bFGF cooperate to maintain the pluripotency of human embryonic stem cells in the absence of feeder layers. Biochem. Biophys. Res. Commun. 330, 934–942.

    Article  CAS  Google Scholar 

  18. Levenstein, M. E., Ludwig, T. E., Xu, R. H., Llanas, R. A., Vandenheuvel-Kramer, K., Manning, D., and Thomson, J. A. (2006) Basic FGF support of human embryonic stem cell self-renewal. Stem Cells 24, 568–574.

    Article  CAS  Google Scholar 

  19. Itskovitz-Eldor, J., Schuldiner, M., Karsenti, D., Eden, A., Yanuka, O., Amit, M., Soreq, H., and Benvenisty, N. (2000) Differentiation of human embryonic stem cells into embryoid bodies comprising the three embryonic germ layers. Mol. Med. 6, 88–95.

    CAS  Google Scholar 

  20. Schuldiner, M., Yanuka, O., Itskovitz-Eldor, J., Melton, D. A., and Benvenisty, N. (2000) Effects of eight growth factors on the differentiation of cells derived from human embryonic stem cells. Proc. Natl. Acad. Sci. U. S. A. 97, 11307–11312.

    Article  CAS  Google Scholar 

  21. Dvash, T., Mayshar, Y., Darr, H., McElhaney, M., Barker, D., Yanuka, O., Kotkow, K. J., Rubin, L. L., Benvenisty, N., and Eiges, R. (2004) Temporal gene expression during differentiation of human embryonic stem cells and embryoid bodies. Hum. Reprod. 19, 2875–2883.

    Article  CAS  Google Scholar 

  22. Dvash, T. and Benvenisty, N. (2004) Human embryonic stem cells as a model for early human development. Best Pract. Res. Clin. Obstet. Gynaecol. 18, 929–940.

    Article  Google Scholar 

  23. Zwaka, T. P. and Thomson, J. A. (2003) Homologous recombination in human embryonic stem cells. Nat. Biotechnol. 21, 319–321.

    Article  CAS  Google Scholar 

  24. Urbach, A., Schuldiner, M., and Benvenisty, N. (2004) Modeling for Lesch-Nyhan disease by gene targeting in human embryonic stem cells. Stem Cells 22, 635–641.

    Article  CAS  Google Scholar 

  25. Matin, M. M., Walsh, J. R., Gokhale, P. J., Draper, J. S., Bahrami, A. R., Morton, I., Moore, H. D., and Andrews, P. W. (2004) Specific knockdown of Oct4 and beta2-microglobulin expression by RNA interference in human embryonic stem cells and embryonic carcinoma cells. Stem Cells 22, 659–668.

    Article  CAS  Google Scholar 

  26. Vallier, L., Rugg-Gunn, P. J., Bouhon, I. A., Andersson, F. K., Sadler, A. J., and Pedersen, R. A. (2004) Enhancing and diminishing gene function in human embryonic stem cells. Stem Cells 22, 2–11.

    Article  CAS  Google Scholar 

  27. Zaehres, H., Lensch, M. W., Daheron, L., Stewart, S. A., Itskovitz-Eldor, J., and Daley, G. Q. (2005) High-efficiency RNA interference in human embryonic stem cells. Stem Cells 23, 299–305.

    Article  CAS  Google Scholar 

  28. Soria, B., Roche, E., Berna, G., Leon-Quinto, T., Reig, J. A., and Martin, F. (2000) Insulin-secreting cells derived from embryonic stem cells normalize glycemia in streptozotocin-induced diabetic mice. Diabetes 49, 157–162.

    Article  CAS  Google Scholar 

  29. Lavon, N., Yanuka, O., and Benvenisty, N. (2004) Differentiation and isolation of hepatic-like cells from human embryonic stem cells. Differentiation 72, 230–238.

    Article  CAS  Google Scholar 

  30. Menendez, P., Wang, L., and Bhatia, M. (2005) Genetic manipulation of human embryonic stem cells: a system to study early human development and potential therapeutic applications. Curr. Gene Ther. 5, 375–385.

    Article  CAS  Google Scholar 

  31. Robertson, E., Bradley, A., Kuehn, M., and Evans, M. (1986) Germ-line transmission of genes introduced into cultured pluripotential cells by retroviral vector. Nature 323, 445–448.

    Article  CAS  Google Scholar 

  32. Pfeifer, A., Ikawa, M., Dayn, Y., and Verma, I. M. (2002) Transgenesis by lentiviral vectors: lack of gene silencing in mammalian embryonic stem cells and preimplantation embryos. Proc. Natl. Acad. Sci. U. S. A. 99, 2140–2145.

    Article  CAS  Google Scholar 

  33. Gropp, M., Itsykson, P., Singer, O., Ben-Hur, T., Reinhartz, E., Galun, E., and Reubinoff, B. E. (2003) Stable genetic modification of human embryonic stem cells by lentiviral vectors. Mol. Ther. 7, 281–287.

    Article  CAS  Google Scholar 

  34. Ma, Y., Ramezani, A., Lewis, R., Hawley, R. G., and Thomson, J. A. (2003) High-level sustained transgene expression in human embryonic stem cells using lentiviral vectors. Stem Cells 21, 111–117.

    Article  CAS  Google Scholar 

  35. Xiong, C., Tang, D. Q., Xie, C. Q., Zhang, L., Xu, K. F., Thompson, W. E., Chou, W., Gibbons, G. H., Chang, L. J., Yang, L. J., and Chen, Y. E. (2005) Genetic engineering of human embryonic stem cells with lentiviral vectors. Stem Cells Dev. 14, 367–377.

    Article  Google Scholar 

  36. Volpers, C. and Kochanek, S. (2004) Adenoviral vectors for gene transfer and therapy. J. Gene Med. 6 (Suppl. 1), S164–S171.

    Article  CAS  Google Scholar 

  37. Eiges, R., Schuldiner, M., Drukker, M., Yanuka, O., Itskovitz-Eldor, J., and Benvenisty, N. (2001) Establishment of human embryonic stem cell-transfected clones carrying a marker for undifferentiated cells. Curr. Biol. 11, 514–518.

    Article  CAS  Google Scholar 

  38. Siemen, H., Nix, M., Endl, E., Koch, P., Itskovitz-Eldor, J., and Brustle, O. (2005) Nucleofection of human embryonic stem cells. Stem Cells Dev. 14, 378–383.

    Article  CAS  Google Scholar 

  39. Lakshmipathy, U., Pelacho, B., Sudo, K., Linehan, J. L., Coucouvanis, E., Kaufman, D. S., and Verfaillie, C. M. (2004) Efficient transfection of embryonic and adult stem cells. Stem Cells 22, 531–543.

    Article  Google Scholar 

  40. Vasquez, K. M., Marburger, K., Intody, Z., and Wilson, J. H. (2001) Manipulating the mammalian genome by homologous recombination. Proc. Natl. Acad. Sci. U. S. A. 98, 8403–8410.

    Article  CAS  Google Scholar 

  41. Hoffman, L. M., Hall, L., Batten, J. L., Young, H., Pardasani, D., Baetge, E. E., Lawrence, J., and Carpenter, M. K. (2005) X-Inactivation status varies in human embryonic stem cell lines. Stem Cells 23, 1468–1478.

    Article  CAS  Google Scholar 

  42. Abeyta, M. J., Clark, A. T., Rodriguez, R. T., Bodnar, M. S., Pera, R. A., and Firpo, M. T. (2004) Unique gene expression signatures of independently-derived human embryonic stem cell lines. Hum. Mol. Genet. 13, 601–608.

    Article  CAS  Google Scholar 

  43. Bhattacharya, B., Miura, T., Brandenberger, R., Mejido, J., Luo, Y., Yang, A. X., Joshi, B. H., Ginis, I., Thies, R. S., Amit, M., Lyons, I., Condie, B. G., Itskovitz-Eldor, J., Rao, M. S., and Puri, R. K. (2004) Gene expression in human embryonic stem cell lines: unique molecular signature. Blood 103, 2956–2964.

    Article  CAS  Google Scholar 

  44. Mountford, P., Nichols, J., Zevnik, B., O’Brien, C., and Smith, A. (1998) Maintenance of pluripotential embryonic stem cells by stem cell selection. Reprod. Fertil. Dev. 10, 527–533.

    Article  CAS  Google Scholar 

  45. Tan, S. M. and Droge, P. (2005) Comparative analysis of sequence-specific DNA recombination systems in human embryonic stem cells. Stem Cells 23, 868–873.

    Article  CAS  Google Scholar 

  46. Drukker, M., Katz, G., Urbach, A., Schuldiner, M., Markel, G., Itskovitz-Eldor, J., Reubinoff, B., Mandelboim, O., and Benvenisty, N. (2002) Characterization of the expression of MHC proteins in human embryonic stem cells. Proc. Natl. Acad. Sci. U. S. A. 99, 9864–9869.

    Article  CAS  Google Scholar 

  47. Drukker, M. (2004) Immunogenicity of human embryonic stem cells: can we achieve tolerance? Springer Semin. Immunopathol. 26, 201–213.

    Article  Google Scholar 

  48. Drukker, M. and Benvenisty, N. (2004) The immunogenicity of human embryonic stem-derived cells. Trends Biotechnol. 22, 136–141.

    Article  CAS  Google Scholar 

  49. Schuldiner, M., Itskovitz-Eldor, J., and Benvenisty, N. (2003) Selective ablation of human embryonic stem cells expressing a “suicide” gene. Stem Cells 21, 257–265.

    Article  CAS  Google Scholar 

  50. Martin, M. J., Muotri, A., Gage, F., and Varki, A. (2005) Human embryonic stem cells express an immunogenic nonhuman sialic acid. Nat. Med. 11, 228–232.

    Article  CAS  Google Scholar 

  51. Richards, M., Fong, C. Y., Chan, W. K., Wong, P. C., and Bongso, A. (2002) Human feeders support prolonged undifferentiated growth of human inner cell masses and embryonic stem cells. Nat. Biotechnol. 20, 933–936.

    Article  CAS  Google Scholar 

  52. Richards, M., Tan, S., Fong, C. Y., Biswas, A., Chan, W. K., and Bongso, A. (2003) Comparative evaluation of various human feeders for prolonged undifferentiated growth of human embryonic stem cells. Stem Cells 21, 546–556.

    Article  CAS  Google Scholar 

  53. Amit, M., Margulets, V., Segev, H., Shariki, K., Laevsky, I., Coleman, R., and Itskovitz-Eldor, J. (2003) Human feeder layers for human embryonic stem cells. Biol. Reprod. 68, 2150–2156.

    Article  CAS  Google Scholar 

  54. Hovatta, O., Mikkola, M., Gertow, K., Stromberg, A. M., Inzunza, J., Hreinsson, J., Rozell, B., Blennow, E., Andang, M., and Ahrlund-Richter, L. (2003) A culture system using human foreskin fibroblasts as feeder cells allows production of human embryonic stem cells. Hum. Reprod. 18, 1404–1409.

    Article  Google Scholar 

  55. Amit, M., Shariki, C., Margulets, V., and Itskovitz-Eldor, J. (2004) Feeder layer- and serum-free culture of human embryonic stem cells. Biol. Reprod. 70, 837–845.

    Article  CAS  Google Scholar 

  56. Ludwig, T. E., Levenstein, M. E., Jones, J. M., Berggren, W. T., Mitchen, E. R., Frane, J. L., Crandall, L. J., Daigh, C. A., Conard, K. R., Piekarczyk, M. S., Llanas, R. A., and Thomson, J. A. (2006) Derivation of human embryonic stem cells in defined conditions. Nat. Biotechnol. 24, 185–187.

    Article  CAS  Google Scholar 

  57. Koivisto, H., Hyvarinen, M., Stromberg, A. M., Inzunza, J., Matilainen, E., Mikkola, M., Hovatta, O., and Teerijoki, H. (2004) Cultures of human embryonic stem cells: serum replacement medium or serum-containing medium and the effect of basic fibroblast growth factor. Reprod. Biomed. Online 9, 330–337.

    Article  CAS  Google Scholar 

  58. Heng, B. C., Liu, H., and Cao, T. (2004) Feeder cell density–a key parameter in multicolor FISH probe sets and their applications human embryonic stem cell culture. In Vitro Cell Dev. Biol. Anim. 40, 255–257.

    Article  Google Scholar 

  59. Sjogren-Jansson, E., Zetterstrom, M., Moya, K., Lindqvist, J., Strehl, R., and Eriksson, P. S. (2005) Large-scale propagation of four undifferentiated human embryonic stem cell lines in a feeder-free culture system. Dev. Dyn. 233, 1304–1314.

    Article  Google Scholar 

  60. Ezashi, T., Das, P., and Roberts, R. M. (2005) Low O2 tensions and the prevention of differentiation of hES cells. Proc. Natl. Acad. Sci. U. S. A. 102, 4783–4788.

    Article  CAS  Google Scholar 

  61. Bayani, J. and Squire, J. A. (2002) Spectral karyotyping. Methods Mol. Biol. 204, 85–104.

    Google Scholar 

  62. Liehr, T., Starke, H., Weise, A., Lehrer, H., and laussen, U. (2004) Multicolor FISH probe sets and their applications. Histol. Histopathol. 19, 229–237.

    CAS  Google Scholar 

  63. Cowan, C. A., Klimanskaya, I., McMahon, J., Atienza, J., Witmyer, J., Zucker, J. P., Wang, S., Morton, C. C., McMahon, A. P., Powers, D., and Melton, D. A. (2004) Derivation of embryonic stem-cell lines from human blastocysts. N. Engl. J. Med. 350, 1353–1356.

    Article  CAS  Google Scholar 

  64. Reubinoff, B. E., Pera, M. F., Vajta, G., and Trounson, A. O. (2001) Effective cryopreservation of human embryonic stem cells by the open pulled straw vitrification method. Hum. Reprod. 16, 2187–2194.

    Article  CAS  Google Scholar 

  65. Tedder, R. S., Zuckerman, M. A., Goldstone, A. H., Hawkins, A. E., Fielding, A., Briggs, E. M., Irwin, D., Blair, S., Gorman, A. M., Patterson, K. G., Linch, D., Heptonstall, J., and Brink, N. (1995) Hepatitis B transmission from contaminated cryopreservation tank. Lancet 346, 137–140.

    Article  CAS  Google Scholar 

  66. Hovatta, O. (2003) Cryobiology of ovarian and testicular tissue. Best Pract. Res. Clin. Obstet. Gynaecol. 17, 331–342.

    Article  Google Scholar 

  67. Richards, M., Fong, C. Y., Tan, S., Chan, W. K., and Bongso, A. (2004) An efficient and safe xeno-free cryopreservation method for the storage of human embryonic stem cells. Stem Cells 22, 779–789.

    Article  Google Scholar 

  68. Kurosawa, H., Imamura, T., Koike, M., Sasaki, K., and Amano, Y. (2003) A simple method for forming embryoid body from mouse embryonic stem cells. J. Biosci. Bioeng. 96, 409–411.

    CAS  Google Scholar 

  69. Dang, S. M., Kyba, M., Perlingeiro, R., Daley, G. Q., and Zandstra, P. W. (2002) Efficiency of embryoid body formation and hematopoietic development from embryonic stem cells in different culture systems. Biotechnol. Bioeng. 78, 442–453.

    Article  CAS  Google Scholar 

  70. Wartenberg, M., Donmez, F., Ling, F. C., Acker, H., Hescheler, J., and Sauer, H. (2001) Tumor-induced angiogenesis studied in confrontation cultures of multicellular tumor spheroids and embryoid bodies grown from pluripotent embryonic stem cells. FASEB J. 15, 995–1005.

    Article  CAS  Google Scholar 

  71. Gerecht-Nir, S., Cohen, S., and Itskovitz-Eldor, J. (2004) Bioreactor cultivation enhances the efficiency of human embryoid body (hEB) formation and differentiation. Biotechnol. Bioeng. 86, 493–502.

    Article  CAS  Google Scholar 

  72. Tucker, K. L., Wang, Y., Dausman, J., and Jaenisch, R. (1997) A transgenic mouse strain expressing four drug-selectable marker genes. Nucleic Acids Res. 25, 3745–3746.

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Stem Cell Technologies (SCT) Ltd, Jerusalem, Israel

    Daniel Kitsberg

Authors
  1. Daniel Kitsberg
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Department of Gene Regulation and Differentiation, National Research Center for Biotechnology GBF, Braunschweig, Germany

    Hansjörg Hauser

  2. Institute for Chemical and Bio-Engineering, ETH Zurich, Zurich, Switzerland

    Martin Fussenegger

Rights and permissions

Reprints and permissions

Copyright information

© 2007 Humana Press Inc.

About this protocol

Cite this protocol

Kitsberg, D. (2007). Human Embryonic Stem Cells for Tissue Engineering. In: Hauser, H., Fussenegger, M. (eds) Tissue Engineering. Methods in Molecular Medicine™, vol 140. Humana Press. https://doi.org/10.1007/978-1-59745-443-8_3

Download citation

  • DOI: https://doi.org/10.1007/978-1-59745-443-8_3

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-756-3

  • Online ISBN: 978-1-59745-443-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation