| Abstract |
Placental extract (PE) is employed as a skin-whitening agent in Japan, where this material is used in cosmetics and dietary supplements. However, the mechanism of PE's anti-melanogenic activity remains poorly understood. The goal of the present study was to elucidate the mechanism of PE's inhibitory effect on melanogenesis in B16 murine melanoma cells. Specifically, the activity of equine PE (EPE) against tyrosinase and melanogenic proteins was evaluated. The effects of EPE on tyrosinase activity and melanin content were assessed spectrophotometrically. This analysis showed that EPE inhibits melanogenesis in melanoma cells in a dose-dependent manner without affecting cell proliferation. EPE did not directly inhibit the enzymatic activity of tyrosinase. Western blot analysis demonstrated that EPE exposure also led to decreases in the protein levels of tyrosinase and tyrosinase-related protein 1 (TRP-1) in melanoma cells, without affecting the levels of the mRNAs encoding these proteins. This analysis further demonstrated that the EPE-induced depletion of tyrosinase and TRP-1 resulted from the induction, by EPE, of proteasome-mediated proteolytic degradation. Notably, the EPE-induced depletion of tyrosinase and TRP-1 was prevented by joint exposure to EPE and the proteasome inhibitor MG132. Similar experiments showed that the exposure of melanoma cells to MG132 abrogates the inhibition of melanogenesis by EPE. Immunoprecipitation analysis further revealed that EPE induces the ubiquitination of tyrosinase and TRP-1. Taken together, these results suggested that EPE induces proteasomal degradation of tyrosinase and TRP-1 by enhancing the ubiquitination of these targets, leading to the depletion of these proteins and the inhibition of melanogenesis.
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