DIFFERENTIAL MODULATION OF EPIDERMAL KERATINIZATION IN IMMORTALIZED (HACAT) AND TUMORIGENIC HUMAN SKIN KERATINOCYTES (HACAT-RAS) BY RETINOIC ACID AND EXTRACELLULAR CA2+

The growth and differentiation response to retinoic acid (RA) was stud ied in the human keratinocyte line HaCaT and tumorigenic clones transf ected with c-Ha-ras oncogene (HaCaT-ras). Differentiation (mainly kera tin synthesis) was evaluated and correlated to cell proliferation in v itro but also growth behaviour in vivo (tumorigenicity). Comparable to normal keratinocytes, HaCaT cells and ras clones showed increased exp ression of the epidermal suprabasal keratins K1 and K10 upon RA deplet ion of the media (delipidized serum), while simple epithelial type ker atins K7, K8 and K18 as well as K19 and K13 (typical of internal strat ified epithelia) were almost completely suppressed. The cell density-d ependent increase of K1 and K10 at intermediate RA levels (as in regul ar media with untreated serum) was also observed at Ca2+ levels below 0.1 mM, thus being clearly unrelated to stratification, whereas K13 sy nthesis was Ca2+-dependent and initiated with stratification. The effe cts on keratins were fully reversed by increasing RA concentrations. T here was only mild stimulation of proliferation at RA doses (10(-10) t o 10(-8) M) not directly corresponding to suppression of keratinizatio n. Thus, the negative RA influence on K1 and K10, opposed to the effec t on simple keratins, substantiates the preserved regulatory capacity rendering these cells appropriate models for biological testing. Among the various tumorigenic HaCaT-ras clones highly and moderately differ entiating ones could be distinguished, accordingly induction in vitro led to a comparable spectrum of differentiation markers (K1 and K10 ap pearing early, and filaggrin late) as growth in vivo. These in vitro r esults demonstrate that, in spite of some differences in RA sensitivit y, virtually all clones possess the epidermal differentiation repertoi r which is regulated according to the same principles. Finally, this c onfirms our in vivo data that differentiation potential is not inverse ly related to the state of transformation or tumorigenicity.