Requirement of hydrocortisone and insulin for extended proliferation and passage of rat keratinocytes.

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 micrograms/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 micrograms/ml) caused the rate of proliferation and the cell density to increase, but insulin alone did not support subcultivation.