Differentiation-related changes of cytokeratin expression in cultured keratinocytes and in fetal, newborn, and adult epidermis

Cytokeratin expression in differentiating cultured foreskin keratinocytes was studied using chain-specific anti-cytokeratin monoclonal antibodies directed against cytokeratins 4, 8, 10, 13, 18, and 19, respectively. Keratinocytes were cultured at low Ca2+ concentration (0.06 mM) to repress differentiation. At confluency, the cells were switched to high Ca2+ concentration (1.6 mM) to induce differentiation. Cells were harvested 0, 3, 8, 16, 24, 48, and 72 h after the switch. Keratinocytes cultured throughout at high Ca2+ concentration were also harvested. Immunoblots of cytokeratin preparations isolated from these cultures showed that cytokeratins 4, 13, and 19 were not present in nondifferentiating keratinocytes but could be detected from about 16 h after the Ca2+ switch. Immunohistochemical studies were performed on frozen sections of cell sheets incubated with anti-cytokeratin and anti-vimentin. Expression of cytokeratins 4, 13, and 19 was seen in superficial cells. Cytokeratin 10 was locally present in suprabasal and superficial cells. Vimentin was present in 40-70% of the basal cells and in only a few differentiating keratinocytes. Expression of cytokeratins 8 and 18 could not be detected. The same antibodies were also used to stain sections from fetal (15, 20, and 29 weeks), newborn (40 weeks), and mature (5 and 75 years) epidermis. In the 15-week-old epidermis, basal cells were positive for cytokeratins 8 and 19 and locally for cytokeratin 4; intermediate cells expressed cytokeratins 4, 10, 13, and 19; and the periderm contained cytokeratins 4, 8, 13, 18, and 19. In the 20-week-old epidermis, cytokeratin 4 had disappeared from the basal cell layer and cytokeratin 19 was present only locally; in the intermediate cell layer, cytokeratins 4 and 19 had disappeared; and in the periderm, the expression of the cytokeratins studied was the same as that in the 15-week-old epidermis. The basal cells of the 29-week-old fetal epidermis, the newborn epidermis, and the mature epidermis are negative with all antibodies tested, except for some scattered cells in the fetal and newborn skin, presumably Merkel cells, that were positive for cytokeratins 8, 18, and 19. Suprabasal cells in all specimens were positive only for cytokeratin 10. With respect to the cytokeratins studied, our results show that cultured differentiating keratinocytes resemble the suprabasal cells of early fetal epidermis. Basal cells of cultured keratinocytes resemble the basal cells of late fetal, newborn, and adult epidermis and therefore support previous observations.