In vitro culture and differentiation of normal mouse blastocysts

Abstract

WHEN preimplantation mouse embryos are transplanted to extrauterine sites, up to 20% develop almost normally to the egg cylinder stage (equivalent to about 6–7 d in utero). They then become disorganised and give rise to tumours, which are usually mature teratomas containing a wide range of differentiated tissues, but less frequently are progressively growing teratocarcinomas containing nests of undifferentiated pluripotent embryonal cells1,2. There are many reasons why it would be interesting to reproduce such normal and abnormal development efficiently in vitro. Although a number of techniques have been used to culture preimplantation embryos, none of them has proved entirely satisfactory3–5; in the best conditions only 5–20% of 3.5–4-d blastocysts develop in vitro to form differentiated, foetal tissues. Most only give rise to extraembryonic endoderm and its associated mesoderm; these cell types proliferate extensively in vitro, and consequently most of the continuous lines derived from blastocyst cultures have probably originated from extraembryonic tissues6. In this paper we describe a simple and reproducible method for culturing mouse blastocysts in vitro so that around 70% give rise to colonies which continue to grow for up to 4 weeks and produce a variety of differentiated tissues, including skin, nerve, beating muscle, cartilage and fibroblasts. The method relies on the technique of immunosurgery7 for killing and removing first the trophectoderm and then the hypoblast (primary endoderm) layers of the embryo, to yield clumps of isolated epiblast (embryonic ectoderm) (Fig. 1).

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References

  1. 1

    Stevens, L. C. J. Embryol. exp. Morph. 20, 329–341 (1968).

  2. 2

    Stevens, L. C. Devl Biol. 21, 364–382 (1970).

  3. 3

    Cole, R. J. & Paul, J., in Preimplantation Stages of Pregnancy, Ciba Foundation Symposium (ed. Wolstenholme, G. E. W. and O'Connor, M.) 82–112 (Churchill, London, 1965).

  4. 4

    Hsu, Y.-C. Nature 231, 100–102 (1971).

  5. 5

    Hsu, Y.-C., Baskar, J., Stevens, L. C. & Rash, J. E. J. Embryol. exp. Morph. 31, 235–245 (1974).

  6. 6

    Sherman, M. I. Cell 5, 343–349 (1975).

  7. 7

    Solter, D. & Knowles, B. Proc. natn. Acad. Sci. U.S.A. 72, 5099–5102 (1975).

  8. 8

    Mintz, B. in Methods in Mammalian Embryology (ed. Daniel, J. C.) 196–197 (Freeman, San Francisco, 1971).

  9. 9

    Hogan, B. L. M. Nature 263, 136–137 (1976).

  10. 10

    Spiegelman, M. & Bennett, D. J. Embryol. exp. Morph. 32, 723–738 (1974).

  11. 11

    Skreb, N., Svajger, A. & Levak-Svajger, B. in Embryogenesis in Mammals, Ciba Foundation Symposium, 40, 27–39 (Elsevier, Amsterdam, 1976).

  12. 12

    Diwan, S. & Stevens, L. C. J. natn. Cancer Inst. 57, 937–938 (1976).

  13. 13

    Grobstein, C. J. exp. Zool. 119, 355–379 (1952).

  14. 14

    Hunt, T. E. Anat. Rec. 68, 349–369 (1937).

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HOGAN, B., TILLY, R. In vitro culture and differentiation of normal mouse blastocysts. Nature 265, 626–629 (1977). https://doi.org/10.1038/265626a0

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