CLONAL GROWTH IN VITRO OF EPITHELIAL CELLS FROM NORMAL HUMAN TISSUES

Tissue culture strains of cells from four different normal human tissues—liver, conjunctiva, kidney, and appendix—have been grown by the plating procedure previously developed for the HeLa strain of cervical carcinoma cells. This technique results in colony formation from isolated single cells, in a manner completely analogous to the plating of bacteria in semisolid nutrient media. Clonal cell strains have been isolated from each cell type. All behaved exactly alike in all properties studied except that some differences in plating efficiency were displayed in some of the growth media employed. The cells from normal human tissues resembled the HeLa S3 carcinomatous cell in the following properties:— (a) Single cells displayed a plating efficiency close to 100 per cent in an appropriate medium. (b) They all grew as an epithelial sheet on glass, the cells being closely packed and polygonal in shape. (c) They had mean generation times of 20 to 23 hours in the nutrient media employed, (d) The mitotic frequency was constant, and therefore the duration of mitosis was the same for all the strains studied, (e) The incidence of multinuclearity and giant formation was very low and similar in both types of cells. (f) Both classes of cells had the same total volume, and the same nuclear cross-sectional area. (g) Both also showed a tendency to spread more in the presence of human serum (concentration of 20 per cent or more) than in porcine serum. However, this differential morphological response was much more marked in the HeLa cell than in those from normal tissues. The only difference noted in the behavior of these two groups of cells lay in the tendency of the cells from normal tissues always to exhibit a greater cross-sectional area when spread on glass than the HeLa cell in the same medium. The frequency of occurrence of different types of multinuclearity in the HeLa cell and cells from normal tissues has been measured. The data suggest that multinuclearity depends on two factors: a necessary, predisposing state in the cell, and a random, independent event causing the appearance of an additional nucleus in such a prepared cell.