In Vitro Transformation of Epidermal Cells from Newborn Mice

Cultures of epidermal cells obtained from newborn BALB/c mice were used to study in vitro transformation of epithelial cells. One-day-old primary cultures plated at 10⁶ cells/ml were treated for 2 days with various concentrations of N-methyl-N'-nitro-N-nitrosoguanidine, 3-methylcholanthrene, and 3-methylcholanthrene-11,12-epoxide. Untreated newborn epidermal cells both divide and keratinize in vitro in Medium 199 supplemented with 10% fetal calf serum. After a few weeks the cells enlarge, show signs of senescence, and then die after about 3 months in vitro. The epidermal cells treated with N-methyl-N'-nitro-N-nitrosoguanidine, 3-methylcholanthrene, or 3-methylcholanthrene-11,12-epoxide went through a similar crisis, appearing as though they too were going to die at 3 months. However, small populations of the remaining cells began to proliferate into colonies that soon grew to confluence. These epithelial cells were characterized by rapid growth, loss of visible keratinization, and subculturability, having been passaged 12 times, in contrast to untreated cells, which are not subculturable. Electron microscope studies did not reveal any true desmosomes, but junctional complexes were present in all of the cell strains examined. The injection of 10⁶ cells from the various cell strains into athymic “nude” or syngeneic mice resulted in rapidly growing solid tumors, which were characterized as highly anaplastic “undifferentiated” tumors.