12-O-tetradecanoylphorbol-13-acetate not only modulates proliferation rates, but also alters antigen expression and LAK-cell susceptibility of normal human melanocytes in vitro.
For serial cultivation of normal human melanocytes media supplemented with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) are largely employed. By using a culture medium that permits cultivation of melanocytes without TPA, the effects of TPA on melanocyte proliferation, phenotype, and susceptibility to lymphokine-activated killer cells were studied. Addition of 50 ng/ml TPA to the medium induced rapid dendrite formation and increased the cell proliferation rate by 16-63% in mitogen-rich media (four of seven cultures, p < 0.01), and by 237% in mitogen-reduced media (p < 0.001). Furthermore, several phenotypic changes indicating early stages of melanocyte transformation were induced by 50 ng/ml TPA. These included increased expression of melanoma progression-associated antigens such as A.1.43 and A.10.33, upregulation of nerve-growth factor receptor as well as of the melanocyte-activation marker HMB-45 and of histocompatibility class I antigens. In contrast, the expression of the differentiation marker K.1.2 and of intercellular adhesion molecule-1 was decreased in TPA-treated cultures. Most of these changes persisted even after removal of TPA from the culture medium (> or = 2 weeks). Staurosporine, a protein kinase C inhibitor, modulated melanocyte-antigen expression similar to TPA, suggesting that protein kinase C downmodulation rather than activation by TPA is involved. In addition to the antigenic alterations, the susceptibility of TPA-treated melanocytes to lymphokine-activated killer cell cytotoxicity decreased by 40% (p < 0.01), possibly due to their altered surface antigen expression. The presented data reveal that the tumor promoter TPA hitherto used as a supplement of melanocyte culture media induces profound phenotypic and functional changes of the cultured cells, indicating incipient transformation of normal human melanocytes in vitro.
Department of Dermatology, University Medical Center Steglitz, Free University of Berlin, Germany.
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Krasagakis
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Garbe
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Krüger-Krasagakes
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Orfanos
D000470:Alkaloids / Q000494:pharmacology; D000941:Antigens / Q000276:immunology*; D002455:Cell Division / Q000187:drug effects; D003602:Cytotoxicity, Immunologic; D006801:Humans; D015979:Killer Cells, Lymphokine-Activated / Q000502:physiology*; D007694:Killer Cells, Natural / Q000502:physiology; D008544:Melanocytes / Q000276:immunology* / Q000502:physiology*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D012016:Reference Values; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D000470:Alkaloids; D000941:Antigens; D011493:Protein Kinase C; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate