Summary
An improved method for the attachment and growth of normal human colonic epithelial cells from minute 1 to 3-mm3 biopsies has been developed. This yields four times as many cultured cells per biopsy than older methods, with a success rate of 97% in a series of 29 biopsies. Fetal bovine serum was eliminated from the medium, the medium pH was decreased to 6.7, the oxygen tension in the incubator was decreased to 3%, and the NCTC 168 medium was supplemented with ethanolamine, phosphoethanolamine, hydrocortisone, ascorbic acid, transferrin, glutamine, insulin, epidermal growth factor, pentagastrin, and deoxycholic acid. The best substrate for cell attachment was a mixture of ungelled collagen I and bovine serum albumin. This substrate was better than the identical mixture with fibronectin added, fibronectin alone, a thin gelatin film, collagen IV with or without fibronectin, and basement membrane preparations from four different cell lines.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
References
Bertolero, F.; Kaighn, M. E.; Camalier, R. F., et al. Effects of serum and serum-derived factors on growth and differentiation of mouse keratinocytes. In Vitro 22:423–428; 1986.
Buset, M.; Lipkin, M.; Winawer, S., et al. Inhibition of human colonic epithelial cell proliferation in vivo and in vitro by calcium. Cancer Res. 46:5426–5430; 1986.
Burtin, P.; Chavanel, G.; Foidart, J. M., et al. Antigens of the basement membrane and the peritumoral stroma in human colonic adenocarcinomas: an immunofluorescence study. Int. J. Cancer 30:13–20; 1982.
Carr, B. I.; Hayashi, I.; Branum, E. L., et al. Inhibition of DNA synthesis in rat hepatocytes by platelet-derived type beta transforming growth factor. Cancer Res. 46:2330–2334; 1986.
DeClerck, Y. A.; Bogenmann, E.; Jones, P. A. Collagen synthesis by short-term explants of pediatric tumors. Cancer Res. 45:1229–1238; 1985.
Ethier, S. P. Serum-free culture conditions for the growth of normal rat mammary epithelial cells in primary culture. In Vitro 22:485–490; 1986.
Friedman, E.; Gillin, S.; Lipkin, M. 12-0-Tetradecanoylphorbol-13-acetate stimulation of DNA synthesis in cultured preneoplastic familial polyposis colonic epithelial cells but not in normal colonic epithelial cells. Cancer Res. 44:4078–4086; 1984.
Friedman, E.; Higgins, P. J.; Lipkin, M., et al. Tissue culture of human epithelial cells from benign colonic tumors. In Vitro 17:632–644; 1981.
Friedman, E.; Thor, A.; Horan Hand, P., et al. Surface expression of tumor-associated antigens in primary cultured human colonic epithelial cells from carcinomas, benign tumors and normal tissues. Cancer Res. 45:5648–5655; 1985.
Hammond, S. L.; Ham, R. G.; Stampfer, M. R. Serum-free growth of human mammary epithelial cells: rapid clonal growth in defined medium and extended serial passage with pituitary extract. Proc. Natl. Acad. Sci. USA 81:5435–5439; 1984.
Higgins, P. J.; Staiano-Coico, L.; Evenson, D. P., et al. Characterization and carcinogen sensitivity of an established endothelial-like cell like derived from adult rat liver tissue. Oncology 41:331–337; 1984.
Jauregui, H. O.; McMillan, P. N.; Driscoll, J., et al. Attachment and long-term survival of adult rat hepatocytes in primary monolayer cultures: comparison of different substrata and tissue culture media formulations. In Vitro 22:13–22; 1986.
Lechner, J. F.; Haugen, A.; McClendon, I. A., et al. Clonal growth of normal adult human bronchial epithelial cells in a serum-free medium. In Vitro 18:633–642; 1982.
Masui, T.; Wakefield, L. M.; Lechner, J. F., et al. Type beta transforming growth factor is the primary differentiation-inducing serum factor for normal human bronchial epithelial cells. Proc. Natl. Acad. Sci. USA 83:2438–2442; 1986.
Moses, H. L.; Tucker, R. G.; Leof, E. B., et al. Type beta transforming growth factor is a growth stimulator and a growth inhibitor. In: Feramisco, J.; Ozanne, B.; Stiles, G., eds. Cancer cells, vol. 3. Cold Spring Harbor, NY: Cold Spring Harbor Laboartory 1985:65–71.
Moyer, M. P.; Aust, J. B. Human colon cells: culture and in vitro transformation. Science 224:1445–1447; 1984.
Siddiqui, K. M.; Chopra, D. P. Primary and long-term epithelial cell cultures from human fetal normal colonic mucosa. In Vitro 20:859–868; 1984.
Sun, T.-T.; Shih, C.; Green, H. Keratin cytoskeleton in epithelial cells of internal organs. Proc. Natl. Acad. Sci. USA 76:2813–2817; 1979.
Thompson, A. A.; Dilworth, S.; Hay, R. J. Isolation and culture of colonic epithelial cells in serum-free medium. J. Tissue Cult. Methods 9:117–122; 1985.
Tsao, M. C.; Walthall, B. J.; Ham, R. C. Clonal growth of normal human epidermal keratinocytes in a defined medium. J. Cell. Physiol. 110:219–229; 1982.
Yang, J.; Richards, J.; Guzman, R., et al. Sustained growth in primary cultures of normal mammary epithelial cells embedded in collagen gels. Proc. Natl. Acad. Sci. USA 77:2088–2092; 1980.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Buset, M., Winawer, S. & Friedman, E. Defining conditions to promote the attachment of adult human colonic epithelial cells. In Vitro Cell Dev Biol 23, 403–412 (1987). https://doi.org/10.1007/BF02623855
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02623855