Masae Okura1, Toshiharu Yamashita2, Yasue Ishii-Osai1, Momoko Yoshikawa1, Yasuyuki Sumikawa1, Kazumasa Wakamatsu3, Shosuke Ito3. 1. Department of Dermatology, Sapporo Medical University School of Medicine, Sapporo 060-8543, Japan. 2. Department of Dermatology, Sapporo Medical University School of Medicine, Sapporo 060-8543, Japan. Electronic address: yamasita@sapmed.ac.jp. 3. Department of Chemistry, Fujita Health University School of Health Sciences, Toyoake 470-1192, Japan.
Abstract
BACKGROUND: Rhododendrol (RD), a skin-whitening agent, is believed to be associated with cases of cosmetics-related leukoderma that have been reported in Japan. Recently, we have shown that RD is catalyzed by tyrosinase to produce putative toxic metabolites RD-catechol and RD-cyclic catechol. OBJECTIVE: To examine the cytotoxicity and production of reactive oxygen species (ROS) in melanocytic cells by RD and its metabolic products. METHODS: The growth inhibitory effect of RD or its metabolite on the normal human epidermal melanocyte (NHEM) and B16F1 cells was assessed by cell counting or WST assay. ROS production was detected by flow cytometry and confocal microscopy after cells were treated with 2',7'-dichlorofluorescein and RD or its metabolite. RESULTS: Growth of NHEM derived from African American (NHEMb) and B16F1 cells was suppressed by 300μM or more RD. Growth inhibitory activity of RD (IC50 of B16F1: 671μM) was weaker than hydroquinone (IC50 of B16F1: 28.3μM) or resveratrol (IC50 of B16F1: 27.1μM). Flow cytometric analysis detected ROS production in the NHEMb and B16F1 cells exposed to RD. However, neither RD nor H2O2 increased the subG1 fraction of these melanocytic cells. RD-catechol and RD-cyclic catechol inhibited growth of NHEMb and B16F1 cells much more strongly than did RD. RD-catechol, as well as RD, produced ROS detected by both flow cytometry and immunostaining, while RD-cyclic catechol produced a hardly detectable amount of ROS in B16F1 cells. CONCLUSIONS: These results suggest that RD exerts the cytotoxicity in melanocytic cells through its oxidative metabolites and that ROS plays a role in RD-mediated cytotoxicity.
BACKGROUND:Rhododendrol (RD), a skin-whitening agent, is believed to be associated with cases of cosmetics-related leukoderma that have been reported in Japan. Recently, we have shown that RD is catalyzed by tyrosinase to produce putative toxic metabolites RD-catechol and RD-cyclic catechol. OBJECTIVE: To examine the cytotoxicity and production of reactive oxygen species (ROS) in melanocytic cells by RD and its metabolic products. METHODS: The growth inhibitory effect of RD or its metabolite on the normal human epidermal melanocyte (NHEM) and B16F1 cells was assessed by cell counting or WST assay. ROS production was detected by flow cytometry and confocal microscopy after cells were treated with 2',7'-dichlorofluorescein and RD or its metabolite. RESULTS: Growth of NHEM derived from African American (NHEMb) and B16F1 cells was suppressed by 300μM or more RD. Growth inhibitory activity of RD (IC50 of B16F1: 671μM) was weaker than hydroquinone (IC50 of B16F1: 28.3μM) or resveratrol (IC50 of B16F1: 27.1μM). Flow cytometric analysis detected ROS production in the NHEMb and B16F1 cells exposed to RD. However, neither RD nor H2O2 increased the subG1 fraction of these melanocytic cells. RD-catechol and RD-cyclic catechol inhibited growth of NHEMb and B16F1 cells much more strongly than did RD. RD-catechol, as well as RD, produced ROS detected by both flow cytometry and immunostaining, while RD-cyclic catechol produced a hardly detectable amount of ROS in B16F1 cells. CONCLUSIONS: These results suggest that RD exerts the cytotoxicity in melanocytic cells through its oxidative metabolites and that ROS plays a role in RD-mediated cytotoxicity.