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Abstract: In the present study, we investigated the effects of heat treatment on melanogenesis in a mouse melanocyte cell line (Mel-Ab). It has been reported that activated extracellular signal-regulated kinase (ERK) is responsible for microphthalmia-associated transcription factor (MITF) degradation, which leads to a reduction in tyrosinase protein production and melanin synthesis. Here we demonstrate that heat treatment induces sustained ERK activation, which may inhibit melanogenesis. However, the specific ERK pathway inhibitors, PD98059 or U0126 did not restore heat-induced hypopigmentation. Furthermore, PD98059 or U0126 hardly blocked the heat-induced activation of ERK. These results suggest that heat treatment may inactivate protein phosphatase, and thus ERK activation is maintained. To support this hypothesis, we examined the effects of heat treatment on protein phosphatase 2A (PP2A) activity. The results obtained show that heat treatment inactivates PP2A, which may subsequently cause ERK activation and that heat treatment inhibits MITF promoter activity. Overall, our results demonstrate that heat treatment reduces melanin production in a temperature-dependent manner. [Copyright &y& Elsevier]

Transforming growth factor-β1 (TGF-β1) plays a pivotal role in cell proliferation, differentiation, and apoptosis. In this study, we investigated the effects of TGF-β1 on melanogenesis using a spontaneously immortalized mouse melanocyte cell line, Mel-Ab. Our results show that TGF-β1 significantly inhibits melanin synthesis in a concentration-dependent manner and that it reduces the activity of tyrosinase, the rate-limiting melanogenic enzyme. We also found that TGF-β1 reduces microphthalmia-associated transcription factor (MITF) promoter activity and decreased MITF, tyrosinase, tyrosinase-related protein-1 (TRP-1), and TRP-2 protein production. In addition, TGF-β1 was found to induce a delay in the activation of extracellular signal-regulated kinase (ERK) at 6 h, whereas many growth factors activate ERK transiently in minutes. Moreover, the specific ERK pathway inhibitor, PD98059 blocked the hypopigmenting effects induced by TGF-β1. PD98059 was also found to abrogate the TGF-β1-mediated down-regulation of MITF, tyrosinase, TRP-1, and TRP-2 production. These results suggest that the ERK pathway may be involved in the melanogenic signaling cascade, and that delayed ERK activation by TGF-β1 contributes to reduced melanin synthesis via MITF down-regulation. [Copyright &y& Elsevier]

In this study, the effects of (−)-epigallocatechin-3-gallate (EGCG) and/or hinokitiol (β-thujaplicin) on melanogenesis were investigated. Our results showed that both EGCG and hinokitiol significantly inhibited melanin synthesis in a concentration-dependent manner, and that their hypopigmenting effects were stronger than that of kojic acid, which is known to inhibit melanin formation in melanocytes and melanoma cells. Interestingly, EGCG did not show any additive hypopigmenting effect in combination with kojic acid, though EGCG did show a synergistic effect in combination with hinokitiol. Several reports indicate that the activation of extracellular signal-regulated kinase (ERK) induces microphthalmia-associated transcription factor (MITF) degradation. Accordingly, the effects of EGCG and hinokitiol on the ERK signaling pathway were examined. EGCG and hinokitiol induced neither ERK activation nor MITF degradation. On the other hand, both EGCG and hinokitiol reduced the protein levels of MITF and of tyrosinase, the rate limiting melanogenic enzyme, whereas kojic acid had no effect. In addition, hinokitiol strongly downregulated the activity of tyrosinase, whereas EGCG or kojic acid had only a little effect. These results show that both EGCG and hinokitiol reduce MITF production, and suggest that reduced tyrosinase activity by hinokitiol explains their synergistic effect on melanogenesis. [ABSTRACT FROM AUTHOR]

In this study, we investigated the effects of lysophosphatidic acid (LPA) on melanogenesis in Mel-Ab cells. We found that LPA significantly attenuates melanin synthesis, and reduces the activity of tyrosinase, the rate-limiting melanogenic enzyme. Interestingly, LPA was also found to induce the activation of a 90 kDa ribosomal S6 kinase (RSK-1), which is known to phosphorylate microphthalmia-associated transcription factor (MITF) at serine 409. Though it has been previously reported that the phosphorylation of MITF is followed by the degradation of MITF, we found that LPA significantly inhibited MITF promoter activity, and that this reduced MITF and tyrosinase protein production. Our results indicate that LPA contributes to reduced melanin synthesis via the down-regulation of MITF. [Copyright &y& Elsevier]

Summary: Background: Melanogenesis is regulated by a series of enzymes under the control of microphthalmia-associated transcription factor (MITF). Objective: The aim of this study was to examine melanosome-associated protein levels in Mel-Ab cells after hinokitiol treatment. Methods: We measured melanin contents and analyzed melanosome-associated protein levels using Western blot and RT-PCR analysis. Results: Hinokitiol markedly inhibited melanin synthesis and also reduced the protein levels of tyrosinase (TYR), tyrosinase-related protein 1 (TYRP-1), tyrosinase-related protein 2 (TYRP-2) and MITF in Mel-Ab cells. In addition, hinokitiol significantly increased the phosphorylations of extracellular signal-regulated kinases 1 and 2 (ERK1/2). Furthermore, reverse transcription-polymerase chain reaction (RT-PCR) analysis revealed that TYR and MITF mRNA levels were significantly decreased but that levels of TYRP-1 and TYRP-2 mRNA were unaffected by hinokitiol treatment. These results suggest that hinokitiol-induced ERK phosphorylation reduces MITF and TYR transcription, and mediates the action of hinokitiol on melanogenesis. Interestingly, the mRNAs of TYRP-1 and TYRP-2 were unaffected, although the protein levels of TYRP-1 and TYRP-2 were down-regulated. Thus, the effects of hinokitiol on the transcription of TYR may differ from its effects on TYRP-1 and TYRP-2. Conclusion: Therefore, we suggest that TYRP-1 and TYRP-2 may be regulated by post-translational degradation after hinokitiol treatment. [Copyright &y& Elsevier]