DOI QR코드

DOI QR Code

The Investigation of Cell Culture Conditions to Maintain Chicken Embryonic Stem Cells as Totipotent Cells

  • Du, Lixin (Lab of Animal Biotech, College of Animal Science Technolojgy, Shandong Agriculture University) ;
  • An, Jing (Lab of Animal Biotech, College of Animal Science Technolojgy, Shandong Agriculture University)
  • Received : 2002.11.14
  • Accepted : 2003.04.04
  • Published : 2003.08.01

Abstract

The ES cell can provide a useful system for studying differentiation and development in vitro and a powerful tool for producing transgenic animalds. To investigate the culture condition of chicken embryonic stem (CES) cells which can retain their multipotentiality or totipotency, three kinds of feeder layer cells, SNL cells, primary mice embryonic fibroblasts (PMEF) cells and primary chicken embryonic fibroblasts (PCEF) cells, were used as the feeder cells in media of DMEM supplemented with leukemia inhibitory factor (LIF), basic fibroblast growth factor (bFGF) and stem cell factor (SCF) for co-culture with blastoderm cells from stage X embryos of chicken. The alkaline phosphatase (AKP) test, differentiation experiment in vitro and chimeric chicken production were carried out. The results showed that culture on feeder layer of PMEF yielded high quality CES cell colonies. The typical CES cells clone shape revealed as follows: nested aggregation (clone) with clear edge and round surface as well as close arrangement within the clone. Strong alkaline phosphatase (AKP) reactive cells were observed in the fourth passage cells. On the other hand, the fourth passage CES cells could differentiate into various cells in the absence of feeder layer cells and LIF in vitro. The third and fourth passage cells were injected into the subgerminal cavity of recipient embryos at stage X. Of 269 Hailan embryos injected with CES cells of Shouguang Chickens, 8.2% (22/269) survived to hatching, 5 feather chimeras had been produced. This suggests that an effective culture system established in this study can promote the growth of CES cells and maintain them in the state of undifferentiated and development, which lays a solid foundation for the application of CES cells and may provide an alternative tool for genetic modification of chickens.

Keywords

Embryonic Stem Cells;Feeder Layer Cells;Chicken ES Cells;Chimeras

Acknowledgement

Supported by : Shandong Provincial Natural Science Foundation

References

  1. Sun, L., C. S. Bradford, C. Ghosh, P. Collodi and D. W. Barnes. 1995. ES-like cells cultures derived from early zebrafish embryos. Mol. Mar. Biol. Biotechnol. 4,193-199.
  2. 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. USA. 92:7844-7848. https://doi.org/10.1073/pnas.92.17.7844
  3. Etches, R. J., M. E. Clark, L. Aajchowski, B. Speksijder, A. M. Verrinder Gibbins, K. Kino, B. Pain and J. Samarut. 1997. Manipulation of blastodermal cells. Poult. Sci. 76:1075-1083. https://doi.org/10.1093/ps/76.8.1075
  4. Roger, A. P. 1994. Studies of in vitro differentiation with embryonic stem cells. Rep. Fertil. Dev. 6:543-552. https://doi.org/10.1071/RD9940543
  5. Uchida,M., T.Tokunaga, K.Niwa, and H.Imai, 1995 Effects of feeder cells and growth factors on the proliferation of mouse primordial germ cells. Theriogenology.
  6. Resnick, J. L., M. Ortiz, J. R. Keller and P. J. Donovan. 1998 Role of fibroblast growth factors and their receptors in mouse primordial germ cell growth. Biology of Reproduction. 59:1224-1229. https://doi.org/10.1095/biolreprod59.5.1224
  7. Pesce, M., M. G. Farrace, M. Piacentini, S. Dola and M. De Felici. 1993. Stem cell factor and leukemia inhibitory factor promote primordial germ cell survival by suppressing programmed cell death (apoptosis). Development 118:1089-1094.
  8. Shamblott, M. J., J. Axelman, S. Wang, E. M. Bugg, J. W. Littlefield, P. J. Donovan, P. D. Blumenthal, G. R. Huggins and J. D. Gearhart. 1998. Derivation of pluripotent stem cells from cultured human primordial germ cells. Proc. Natl. Acad. Sci. 95:13726-13731. https://doi.org/10.1073/pnas.95.23.13726
  9. Pain, B., M. E. Clark, M. Shen, H. Nakazawa, M. Sakurai, J. Samarut and R. J. Etches. 1996. Long-term in vitro culture and characterization of avian embryonic stem cells with multiple morphogenetic potentialities. Development. 122:2339-2348.
  10. Thomson, J. A., J. Itskovita-Eldor, S. S. Shapiro, M. A. Waknitz, J. J. Swiergiel, V. S. Marshall and J. M. Jones. 1998. Embryonic stem cell lines derived from human blastocysts. Science. 282:1145-1147. https://doi.org/10.1126/science.282.5391.1145
  11. Petitte, J. N., L. Karagenc. 1996. Growth factors during early events in avian embryo-development. Poult. Avian. Biol. Rev. 7:75-87.
  12. Thoraval, P., F. Lasserre, F. Coudert and G. Dambrine. 1994. Somatic and germline chicken chimeras obtained from brown and white leghorns by transfer of early blastodermal cells. Poult. Sci. 73:1897-1905. https://doi.org/10.3382/ps.0731897
  13. Yang, Z., J. N. Petitte. 1994. Use of avian cytokines in mammalian embryonic stem cell culture. Poult. Sci. 73:965-974. https://doi.org/10.3382/ps.0730965
  14. Du, L. X. and S. G. Li. 1998. Make chimeric chicken by transfer of early balstodermal cells. J. Shandong Agriculture University. 29(2):231-232.
  15. Kino, K., B. Pain, S. P. Leibo. 1997. Production of chicken chimeras from injection of frozen-thawed blastodermal cells. Poult. Sci. 76:753-760.
  16. First, N. L., M. M. Sins, S. P. Park and M. J. Kent-First. 1994. Systems for production of calves from cultured bovine embryonic cells. Rep. Fertil Dev. 6:553-562. https://doi.org/10.1071/RD9940553
  17. Petitte, J. N., M. E. Clark, G. Liu, A. M. Verrinder Gibbins and R. J. Etches. 1990. The production of somatic and germline chimeras in the chick by transfer of early blastodermal cell. Development. 108:185-190.
  18. Natio, M., M. Watanabe, M. Kinutani, K. Nirasaxa and T. Oishi. 1991. Production of quail-chick chimaeras by blastodermal cell transfer. Br. Poult. Sci. 32:79-86. https://doi.org/10.1080/00071669108417329
  19. Du, L. X. and C. G. Yin. 2002. Culture of chicken of stage X blastoderm cells. Acta. Zool. Sin. 48(4):549-553
  20. Piedrahita, J. A., D. J. Murray, M. M. McGloughlin, G. B. Anderson and A. M. Oberbauer. 1999. Isolation and genetic modification of pluripotent/totipotent cell lines. 10. Transgenic animals in research.
  21. Williams, R. L., D. J. Hilton, S. Pease, T. A. Willson, C. L. Stewart, D. P. Gearing, E. F. Wagner, D. Metcalr, N. A. Nicola, and N. M. Gough. 1988. Myeloid leukaemia inhibitory factor mainjtains the developmental potential of embryonic stem cells. Nature 336:684-687. https://doi.org/10.1038/336684a0
  22. Evans, M. J., M. H. Kaufman. 1981. Establishment in culture of pluripotential cells from mouse embryos. Nature 292:154-156. https://doi.org/10.1038/292154a0
  23. Wheeler, M. B. 1994 Development and validation of swine embryonic stem cells: a review. Rep. Fertil. Dev. 6:563-568. https://doi.org/10.1071/RD9940563
  24. Giles, J. R., X. Yang, W. Mark and R H. Foote. 1993. Pluritotency of cultured rabbit inner cell mass cells detected by isozyme analysis and eye pigmentation of fetuses following injection into blastocyst or morulae. Mol. Rep. Dev. 36:130-138. https://doi.org/10.1002/mrd.1080360203
  25. Nichols, J., E. P. Evans and A. G. Smith. 1990. Establishment of germ-line competent embryonic stem (ES) cells using differentiation inhibiting activity. Development. 110:1341-1348.
  26. Eyal-Giladi, H., S. Korak. 1976. From cleavage to primitive streak formation: a complementary normal table and a new look at the first stages of the development of the chick; I. General Morphology Dev. Biol. 49:321-337.
  27. Aajchowski, L. D., R. J. Etches. 2000. Transgenic chickens: past, present, and future. Avian and Poult. Biology Rev. 11(2):63-80.
  28. Ono, T., S. L. Muto and I. M. Mizutan. 1994. Production of quail chimera by transfer of early blastodermal cells and its use for transgenesis. J. Poult. Sci. 31(2):119-129. https://doi.org/10.2141/jpsa.31.119
  29. Gao, S. P., W. H. Shi and Y. Qin. 2000. Production of feeder layer used for isolation and culturing of embryonic stem cells, J. Lab. Anim. Sci. Sinica 10(2):78-81.