Skin organ culture model for examining epidermal melanization

To clarify the regulatory mechanism of skin pigmentation through epidermal proliferation and differentiation, organ cultures of pigmented guinea pig skins have been studied for their melanogenic responses to exogenous stimuli. Melanogenic activity was measured by both tyrosinase activity assessed by observing release of 3H2O from tissue after incubation with 3H-tyrosine and melanin synthesis, indicated by the incorporation of 14C-2-thiouracil into tissue. Those skin explants that were maintained in serum-free media under air conditions equilibrated with 5% CO2, 50% O2, and 45% N2 formed new, chemically analyzable pigment in the tissue within 4 d of culture. This melanization was accompanied by an increased number of melanocytes as well as by enhanced tyrosinase activity and elevated melanin synthesis. This organ culture system responded well to known tyrosinase inhibitors such as phenylthiourea, which decreased melanogenic activity. Of the arachidonic acid metabolites tested, PGE2, LTC4, LTB4, and 5-HETE were found to significantly stimulate melanogenic activity as indicated by tyrosinase activity, whereas PGF2-alpha, 12-HETE, and 15-HETE did not. Among several known growth factors, only bFGF significantly stimulated melanogenesis in the organ cultured melanocytes. TGF-alpha, which increased DNA synthesis, had a slightly stimulating effect on melanogenesis, whereas TGF beta, which inhibited DNA synthesis, did not stimulate melanogenesis, but rather slightly decreased it. 8-methoxypsoralen+ultraviolet A-treated skin that underwent a marked pigmentation within 14 d in vivo demonstrated enhanced melanogenesis in the organ culture system in proportion to its in vivo pigmentation. Our organ culture system might provide an opportunity to examine the mechanism of action of epidermal melanization.