Nicotinic Acid Hydroxamate Downregulated the Melanin Synthesis and Tyrosinase Activity through Activating the MEK/ERK and AKT/GSK3β Signaling Pathways

Abstract

In this study, nicotinic acid hydroxamate (NAH), a nicotinic acid derivative, was found to show dose-dependent inhibition of melanin content and tyrosinase activity of murine melanoma B16F10 cells with or without being cotreated with cAMP stimulators. In the studies on signaling pathways for skin whitening, NAH-treated B16F10 cells resulted in a decrease in the expression of tyrosinase, tyrosinase-related protein-1, and microphthalmia-associated transcription factor (MITF). PD98059 and LY294002 additions were obviously to increase melanin contents in B16F10 cells; however, they were reversed by NAH cotreatments. NAH-mediated increases in the phosphorylation of mitogen-activated protein kinase kinase (MEK)/ERK and AKT/glycogen synthase kinase-3β (GSK3β) were also found, which in turn led to the inhibition of MITF expression and then downregulated tyrosinase and tyrosinase-related protein (TRP)-1 expressions. These results suggest that NAH may be an active component in the inhibition of melanogenesis, which will have potential uses as cosmetics for whitening and need further investigation.

Glycine hydroxamate inhibits tyrosinase activity and melanin contents through downregulating cAMP/PKA signaling pathways

“…The B16F10 melanoma cells were cultured following the methods of Lin et al (2012b). Different concentrations of GH, Gly and arbutin (0.5-3.5 mM) were added to a seeded 24-well microtiter plate and incubated in a humidified incubator at 5 % CO 2 and 37 °C for 24 h. Five microliter of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT, 5 mg/mL) was added under light protection and after 4 h, 100 μL of DMSO was added to dissolve the formed formazans.…”

“…The absorbance values at 475 nm were compared against a standard curve of mushroom TYR. To determine melanin content in B16F10 cells, the extraction and analysis were performed as per the previous report (Lin et al 2012b). …”

“…After electrophoresis, the Western blot analysis was performed as previously reported (Lin et al 2012b) using the primary antibodies against β-actin, MITF, CREB, and p-CREB (1000-fold dilution by 0.25 % gelatin in NaCl/EDTA/Tris (NET) solution) and TYR, TRP-1, TRP-2 (500-fold dilution by 0.25 % gelatin in NET solution). The inhibitory effects of GH treatment on protein expression were quantified and expressed as a relative density (% area) using the Syngene G:bBOX imaging system (Syngene, UK) equipped with the Gene Snap software (Syngene, UK).…”

Down-regulation of tyrosinase, TRP-1, TRP-2 and MITF expressions by citrus press-cakes in murine B16 F10 melanoma

“…In mammalian melanocytes, melanogenesis is controlled at least by three regulatory proteins, tyrosinase, and TRP1 and TRP2. In contrast, hyperpigmentation is by overactivity of TRPs, but not all skin-whitening agents can simultaneously inhibit TYR, TRP-1, and TRP-2, such as Arthrophytum scoparium, Glechoma hederacea and nicotinic acid hydroxamat [18][19][20] . However, our Western blotting assay showed that CCE treatment reduced the expression of all rate-limiting enzymes, including tyrosinase, TRP-1, and TRP-2 protein, and prevented abnormal accumulation of melanin in the process of melanogenesis.…”

“…Measurement of tyrosinase, TRP-1, TRP-2, and MITF in B16F10 cells by Western blot was undertaken as described previously by Yoon 20. Tyrosinase, TRP-1, TRP-2 and MITF bands were detected with the rabbit polyclonal anti-tyrosinase antibody (dilution 1:1 000), anti-TRP-1 antibody (dilution 1:1 000), anti-TRP-2 antibody and anti-MITF antibody (dilution 1:500), respectively, which were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA), and then further incubated with horseradish peroxidase-conjugated antirabbit IgG antibody at a 1:5 000 dilution.…”