Inhibitory effect of the water-soluble polymer-wrapped derivative of fullerene on UVA-induced melanogenesis via downregulation of tyrosinase expression in human melanocytes and skin tissues

The C60-fullerene derivatives are expected, as novel and potent anti-oxidants, to more effectively protect skin cells against oxidative stress. UVA-induced oxidative stress is considered to promote melanogenesis and serious skin damage. The effect of any fullerene derivatives on UVA-induced melanogenesis is still unknown. Here, we evaluated effects of a water-soluble polyvinylpyrrolidone (PVP)-wrapped fullerene derivative (named “Radical Sponge®” because of its anti-oxidant ability) on melanogenesis, which was promoted by UVA-irradiation to human melanocytes and skin tissues. Radical Sponge® markedly scavenged UVA-induced reactive oxygen species (ROS) inside human melanocytes as shown by fluorometry using the redox indicator CDCFH-DA. After treatment with Radical Sponge® or other agents, human melanocytes and skin tissues were irradiated by UVA. Then, cellular melanin content, tyrosinase activity and the ultrastructural change of skin melanosomes were examined. Radical Sponge® showed to significantly inhibit UVA-promoted melanogenesis in normal human epidermis melanocytes (NHEM) and human melanoma HMV-II cells within a non-cytotoxicity dose range. As compared with two whitening agents, arbutin and l-ascorbic acid, Radical Sponge® demonstrated the stronger anti-melanogenic potential according to spectrophotometric quantification for extracted melanin. In human skin cultures also, UVA-promoted melanin contents were repressed by Radical Sponge® according to Fontana–Masson stain, suggesting its ability to repress UVA-induced tanning. Transmission electron microscopic ultrastructural images also proved that UVA-increased melanosomes in human skin tissue were obviously reduced by Radical Sponge®. The UVA-enhanced tyrosinase enzymatic activity in NHEM melanocytes was inhibited by Radical Sponge® more markedly than by arbutin and l-ascorbic acid. The UVA-enhanced tyrosinase protein expression, together with cell-size fatness and dendrite-formation, was also inhibited more markedly by Radical Sponge® according to immunostain and flow cytometry using anti-tyrosinase antibody. Thus the depigmentating action of Radical Sponge® might be due to its down-regulating effect on the tyrosinase expression, which is initiated by UVA-caused ROS generation.