Studies on guinea pig skin cell cultures. VI. Growth kinetics of epidermal keratinocytes and dermal fibroblasts

Abstract

The growth kinetics of epidermal keratinocytes (EK) and dermal fibroblasts (DF) have been determined by four different methods: incorporation of 3H-thymidine into DNA (3H-microgram DNA ratio); 3H-thymidine/14C amino acid incorporation ratio (3H:14C ratio); 3H-thymidine labelled nuclei, and colchicine-blocked metaphase counts. The growth curve of EK was no different when plotted with the 3H:14C ratio than with the 3H-microgram DNA ratio. However, this was not true for DF. The replacement of sodium bicarbonate with Hepes buffer in the culture medium did not greatly affect the shape of the EK growth curve, whereas the DF growth curve became diphasic instead of monophasic. The elimination of mature (differentiated) keratinocytes from the very onset of EK culture had a profound effect on the EK growth curve. DNA synthesis peaked at day 1 in cultures without, instead of day 9 in cultures with differentiated cells. Furthermore, mitotic activity did not show up before day 6. This suggests that (i) EK in culture are sensitive to the G1 inhibitor released by differentiated epidermal cells, and (ii) they remain in G2 for about 5 days. Thus, EK in culture seem to continue to be susceptible, as in vivo, to homeostatic regulation through the action of G1-G2 inhibitors.