Stimulation of Tyrosinase Activity and Melanin Formation of Cultured Melanoma Cells by Serum Deprivation Alone or in Combination with Dibutyryl Cyclic AMP and Theophylline

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Abstract
Serum removal from the media of serial monolayer cultures of Harding-Passey mouse melanoma during an incubation period of 3 days resulted in an exponentially declining DNA synthesis rate (measured by incorporation of [14C]thymidine) and inhibition of cell proliferation. Protein synthesis, as measured by the incorporation of radioactive leucine, was less affected then DNA synthesis. Incubation in serum-free culture medium resulted in significant rises of tyrosinase activity and cellular melanin content. Addition of dibutyryl cyclic AMP (Bu2cAMP, 5 .times. 10-4 M) and theophylline (5 .times. 10-4 M) to serum-free cultures caused a further striking increase of tyrosinae activity and melanin formation, while treatment of serum containing cultures with Bu2cAMP and theophylline showed only a slight rise in melanogenesis. These stimulatory effects are mediated by an increased intracellular cAMP level, since a correlation between the degree of melanogenesis and cellular cAMP content was indicated. Treatment of serum-free or serum-containing cultures with the phosphodiesterase inhibitor theophylline (5 .times. 10-4-1 .times. 10-3 M) alone revealed only a slight enhancement (about 20%) of melanogenesis. Because augmentation of melanogenesis by serum-free medium alone or together with Bu2cAMP and theophylline was prevented by cycloheximide (or actinomycin D), de novo protein synthesis seems required for these stimulatory effects.

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