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. 1983 Jun 1;96(6):1690–1696. doi: 10.1083/jcb.96.6.1690

Epithelial layer formation in differentiating aggregates of F9 embryonal carcinoma cells

PMCID: PMC2112462  PMID: 6343402

Abstract

F9 embryonal carcinoma (EC) cells, cultured in suspension in medium containing 5 X 10(-8) M retinoic acid, aggregate and differentiate into embryoid bodies with an outer layer of visceral endoderm cells that synthesize and secrete alphafetoprotein (AFP) (Hogan, B. L. M., A. Taylor, and E. Adamson, 1981, Nature (Lond.). 291:235-237). Here we analyze the formation of the outer layer of cells as a model for epithelial differentiation. Three morphological phases are described, but analyses of cell numbers and the synthetic rates of some proteins, as well as the appearance of markers of visceral endoderm and basement membrane, show that the formation of the outer layer occurs as an orderly progression of multiple events. The markers used to follow the ontogeny of epithelial layer formation include SSEA-1, l, and i blood group antigens, laminin, fibronectin, type IV collagen, cytoskeletal intermediate filament proteins (vimentin, Endo A, and B), and AFP. The onset of epithelium formation occurs between the third and fourth day of culture, but its function is maximally expressed only when it is well organized. We found the rate of AFP secretion to be a measure of the proper alignment and maturity of the epithelium which occurs at the seventh or eighth day. This model of epithelium formation may help to explain how similar processes occur during embryogenesis.

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Selected References

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  1. Adamson E. D., Ayers S. E. The localization and synthesis of some collagen types in developing mouse embryos. Cell. 1979 Apr;16(4):953–965. doi: 10.1016/0092-8674(79)90110-7. [DOI] [PubMed] [Google Scholar]
  2. Adamson E. D., Gaunt S. J., Graham C. F. The differentiation of teratocarcinoma stem cells is marked by the types of collagen which are synthesized. Cell. 1979 Jul;17(3):469–476. doi: 10.1016/0092-8674(79)90254-x. [DOI] [PubMed] [Google Scholar]
  3. Albrechtsen R., Nielsen M., Wewer U., Engvall E., Ruoslahti E. Basement membrane changes in breast cancer detected by immunohistochemical staining for laminin. Cancer Res. 1981 Dec;41(12 Pt 1):5076–5081. [PubMed] [Google Scholar]
  4. Artzt K., Dubois P., Bennett D., Condamine H., Babinet C., Jacob F. Surface antigens common to mouse cleavage embryos and primitive teratocarcinoma cells in culture. Proc Natl Acad Sci U S A. 1973 Oct;70(10):2988–2992. doi: 10.1073/pnas.70.10.2988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Berstine E. G., Hooper M. L., Grandchamp S., Ephrussi B. Alkaline phosphatase activity in mouse teratoma. Proc Natl Acad Sci U S A. 1973 Dec;70(12):3899–3903. doi: 10.1073/pnas.70.12.3899. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chamberlain J. P. Fluorographic detection of radioactivity in polyacrylamide gels with the water-soluble fluor, sodium salicylate. Anal Biochem. 1979 Sep 15;98(1):132–135. doi: 10.1016/0003-2697(79)90716-4. [DOI] [PubMed] [Google Scholar]
  7. Dziadek M. A. Use of glycine as a non-enzymic procedure for separation of mouse embryonic tissues and dissociation of cells. Exp Cell Res. 1981 Jun;133(2):383–393. doi: 10.1016/0014-4827(81)90331-1. [DOI] [PubMed] [Google Scholar]
  8. Dziadek M., Adamson E. Localization and synthesis of alphafoetoprotein in post-implantation mouse embryos. J Embryol Exp Morphol. 1978 Feb;43:289–313. [PubMed] [Google Scholar]
  9. Engvall E. Enzyme immunoassay ELISA and EMIT. Methods Enzymol. 1980;70(A):419–439. doi: 10.1016/s0076-6879(80)70067-8. [DOI] [PubMed] [Google Scholar]
  10. Fox N., Damjanov I., Martinez-Hernandez A., Knowles B. B., Solter D. Immunohistochemical localization of the early embryonic antigen (SSEA-1) in postimplantation mouse embryos and fetal and adult tissues. Dev Biol. 1981 Apr 30;83(2):391–398. doi: 10.1016/0012-1606(81)90487-5. [DOI] [PubMed] [Google Scholar]
  11. Freeman A. E., Engvall E., Hirata K., Yoshida Y., Kottel R. H., Hilborn V., Ruoslahti E. Differentiation of fetal liver cells in vitro. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3659–3663. doi: 10.1073/pnas.78.6.3659. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hogan B. L., Taylor A., Adamson E. Cell interactions modulate embryonal carcinoma cell differentiation into parietal or visceral endoderm. Nature. 1981 May 21;291(5812):235–237. doi: 10.1038/291235a0. [DOI] [PubMed] [Google Scholar]
  13. Hogan B. L., Tilly R. Cell interactions and endoderm differentiation in cultured mouse embryos. J Embryol Exp Morphol. 1981 Apr;62:379–394. [PubMed] [Google Scholar]
  14. Kapadia A., Feizi T., Evans M. J. Changes in the expression and polarization of blood group I and i antigens in post-implantation embryos and teratocarcinomas of mouse associated with cell differentiation. Exp Cell Res. 1981 Jan;131(1):185–195. doi: 10.1016/0014-4827(81)90418-3. [DOI] [PubMed] [Google Scholar]
  15. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  16. Martin G. R., Evans M. J. Differentiation of clonal lines of teratocarcinoma cells: formation of embryoid bodies in vitro. Proc Natl Acad Sci U S A. 1975 Apr;72(4):1441–1445. doi: 10.1073/pnas.72.4.1441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Oshima R. G. Developmental expression of murine extra-embryonic endodermal cytoskeletal proteins. J Biol Chem. 1982 Apr 10;257(7):3414–3421. [PubMed] [Google Scholar]
  18. Oshima R. G. Identification and immunoprecipitation of cytoskeletal proteins from murine extra-embryonic endodermal cells. J Biol Chem. 1981 Aug 10;256(15):8124–8133. [PubMed] [Google Scholar]
  19. Oshima R., Linney E. Identification of murine extra-embryonic endodermal cells by reaction with teratocarcinoma basement membrane antiserum. Exp Cell Res. 1980 Apr;126(2):485–490. doi: 10.1016/0014-4827(80)90294-3. [DOI] [PubMed] [Google Scholar]
  20. Ruoslahti E., Vuento M., Engvall E. Interaction of fibronectin with antibodies and collagen in radioimmunoassay. Biochim Biophys Acta. 1978 Jun 21;534(2):210–218. doi: 10.1016/0005-2795(78)90003-x. [DOI] [PubMed] [Google Scholar]
  21. Sell S., Skelly H., Leffert H. L., Muller-Eberhard U., Kida S. Relationship of the biosynthesis of alpha-fetoprotein, albumin, hemopexin, and haptoglobin to the growth state of fetal rat hepatocyte cultures. Ann N Y Acad Sci. 1975 Aug 22;259:45–58. doi: 10.1111/j.1749-6632.1975.tb25401.x. [DOI] [PubMed] [Google Scholar]
  22. Solter D., Knowles B. B. Monoclonal antibody defining a stage-specific mouse embryonic antigen (SSEA-1). Proc Natl Acad Sci U S A. 1978 Nov;75(11):5565–5569. doi: 10.1073/pnas.75.11.5565. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Strickland S., Mahdavi V. The induction of differentiation in teratocarcinoma stem cells by retinoic acid. Cell. 1978 Oct;15(2):393–403. doi: 10.1016/0092-8674(78)90008-9. [DOI] [PubMed] [Google Scholar]
  24. Strickland S., Smith K. K., Marotti K. R. Hormonal induction of differentiation in teratocarcinoma stem cells: generation of parietal endoderm by retinoic acid and dibutyryl cAMP. Cell. 1980 Sep;21(2):347–355. doi: 10.1016/0092-8674(80)90471-7. [DOI] [PubMed] [Google Scholar]
  25. Sugrue S. P., Hay E. D. Response of basal epithelial cell surface and Cytoskeleton to solubilized extracellular matrix molecules. J Cell Biol. 1981 Oct;91(1):45–54. doi: 10.1083/jcb.91.1.45. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Thomas J. O., Kornberg R. D. An octamer of histones in chromatin and free in solution. Proc Natl Acad Sci U S A. 1975 Jul;72(7):2626–2630. doi: 10.1073/pnas.72.7.2626. [DOI] [PMC free article] [PubMed] [Google Scholar]

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