Summary
A procedure for the preparation and cultivation of rat epidermal basal cells from full thickness skin resulted in greater than 99% viability and 90% plating efficiency. However, attempts to subculture monolayers of these epithelial cells grown in medium with serum as the only supplement were totally unsuccessful. When hydrocortisone and insulin were added to the medium, subcultivation of primary growth was obtained. It was demonstrated that hydrocortisone at concentrations as low as 0.1 μg/ml was necessary for at least the initial attachment of the cells to the substrate—an essential step in subcultivation. Increasing concentrations of insulin (0.1 to 50 μg/ml) caused the rate of proliferation and the cell density to increase, but insulin alone did not support subcultivation.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Earle, W. R. Long term cultivation of animal tissue cells in large cultures. Fed. Proc. 17: 967–974; 1958.
Karasek, M.; Moore, T. Isolation and properties of a germinative and nongerminative cell population from skin. J. Invest. Derm. 54:430; 1970.
Vaughan, F. L.; Bernstein, I. A. Studies of proliferative capabilities in isolated epidermal basal and differentiated cells. J. Invest. Derm. 56: 454–466; 1971.
Karasek, M.; Charlton, M. F. Growth of postembryonic skin epithelial cells on collagen gels. J. Invest. Dermatol. 56:205–210; 1971.
Freeman, A. E.; Ingel, H. J.; Herrman, B. J.; Kleinfeld, K. N. Growth and characterization of human skin epithelial cell cultures. In Vitro 12: 352–262; 1976.
Rheinwald, J. G.; Green, H. Serial cultivation of strains of human epidermal keratinocytes: The formation of keratinizing colonies from single cells. Cell 6:331–341; 1975.
Peehl, D. M.; Ham, R. G. Growth and differentiation of human keratinocytes without a feeder layer or conditioned medium. In Vitro 16: 516–525; 1980.
Eisinger, E.; Lee, J. S.; Hefton, J. M.; Darzynkiewicz, Z.; Chiao, J. W.; DeHarven, E. Human epidermal cell cultures: Growth and differentiation in the absence of dermal components or medium supplements. Proc. Natl. Acad. Sci. USA 76:5340–5344; 1979.
Peehl, D. M.; Ham, R. G. Clonal growth of human keratinocytes with small amounts of dialyzed serum. In Vitro 16:526–538; 1980.
cohen, S.; Savage, C. R. Recent studies on the chemistry and biology of epidermal growth factor. Recent Prog. Horm. Res. 31:551–572; 1974.
Rheinwald, J. G.; Green, H. Epidermal growth factor and the multiplication of cultured human epidermal keratinocytes. Nature 265:421–424; 1977.
Hayashi, I.; Larher, J.; Sato, G. Hormonal growth control of cells in culture. In Vitro 14: 23–30; 1978.
Vaughan, F. L.; Mitra, R. S.; Bernstein, I. A. Synthesis of DNA in isolated nuclei from differentiated mammalian epidermal cells. J. Invest. Dermatol. 66:355–359; 1976.
Phillips, J. H.; Terryberry, J. E. Counting actively metabolizing tissue culture cells. Exp. Cell Res. 13:341–347; 1957.
Picton, W.; Devitt, H.; Forgie, M. A. Practical applications of the Quantimet 720, an image analyzing computer, in the field of investigative dermatology. Br. J. Dermatol. 95:341–348; 1976.
Marchok, A. C.; Rhoton, J.; Nettesheim, P. Long term culture of epithelial cells from normal rat tracheas. In Vitro 12:320; 1976.
Van Scott, E. J. Mechanical separation of the epidermis from the corneum. J. Invest. Dermatol. 18:377–379; 1955.
Szabo, G. A modification of the technique of “skin splitting” with trypsin. J. Pathol. Bacteriol. 70: 545; 1955.
Price, F. M.; Camalier, R. F.; Gantt, R.; Taylor, W. G.; Smith, G. H.; Sanford, K. K. A new culture medium for human skin epithelial cells. In Vitro 16:147–158; 1980.
Hayashi, I.; Larner, J.; Sato, G. Hormonal growth control of cells in culture. In Vitro 14:23–30; 1978.
Poste, G. Tissue dissociation with proteolytic enzymes. Exp. Cell Res. 65:359–367; 1971.
Schwartz, A. G.; Amos, H. Insulin dependence of cells in primary culture: Influence of ribosome integrity. Nature 219:1366–1367; 1968.
Author information
Authors and Affiliations
Additional information
This work was supported by Grants 1-R01-CA-19988 and 5-R01-AM-15206 from the National Institutes of Health, U.S. Public Health Service.
Rights and permissions
About this article
Cite this article
Vaughan, F.L., Kass, L.L. & Uzman, J.A. Requirement of hydrocortisone and insulin for extended proliferation and passage of rat keratinocytes. In Vitro 17, 941–946 (1981). https://doi.org/10.1007/BF02618418
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02618418