• Korean Journal of Plant Resources
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• v.31 no.5
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• pp.547-553
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• 2018

This study was conducted to investigate the effect of branch extracts of Vaccinium oldhamii (VOB) on melanin synthesis in B16F10 cells. VOB promoted melanin production in absence or presence of ${\alpha}$-melanocyte-stimulating hormone (${\alpha}-MSH$) in B16F10 cells. However, VOB did not affect the expression of tyrosinase and TRP-1 associated with melanin synthesis at the mRNA and protein levels in B16F10. But, VOB decreased TRP-2 protein level and induced tyrosinase activation in B16F10 cells. Inhibition of tyrosinase activity and tyrosinase knockdown attenuated VOB-mediated melanin synthesis. In conclusion, VOB may stimulate melanin synthesis through activating tyrosinase activity.

• Journal of Acupuncture Research
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• v.34 no.1
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• pp.1-9
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• 2017

Objectives : We investigated the synergistic effects of bee venom (BV) and natural killer (NK) cells on B16F10 melanoma cell apoptosis mediated by IL-4. Methods : We performed a cell viability assay to determine whether BV can enhance the inhibitory effect of NK-92MI cells on the growth of B16F10 melanoma cells, and western blot analysis to detect changes in the expression of IL-4, $IL-4R{\alpha}$, and other apoptosis-related proteins. EMSA was performed to observe the activity of STAT6. To confirm that the inhibitory effect of BV and NK cells was mediated by IL-4, the above tests were repeated after IL-4 silencing by siRNA (50 nM). Results : B16F10 melanoma cells co-cultured with NK-92MI cells and simultaneously treated by BV ($5{\mu}g/ml$) showed a higher degree of proliferation inhibition than when treated by BV ($5{\mu}g/ml$) alone or co-cultured with NK-92MI cells alone. Expression of IL-4, $IL-4R{\alpha}$, and that of other pro-apoptotic proteins was also enhanced after co-culture with NK-92MI cells and simultaneous treatment with BV ($5{\mu}g/ml$). Furthermore, the expression of anti-apoptotic bcl-2 decreased, and the activity of STAT6, as well as the expression of STAT6 and p-STAT6 were enhanced. IL-4 silencing siRNA (50 nM) in B16F10 cells, the effects of BV treatment and NK-92MI co-culture were reversed. Conclusion : These results suggest that BV could be an effective alternative therapy for malignant melanoma by enhancing the cytotoxic and apoptotic effect of NK cells through an IL-4-mediated pathway.

• The Journal of Pediatrics of Korean Medicine
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• v.20 no.1
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• pp.257-272
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• 2006

Objective : In this study, the ability of Oriental medicine Samultanggamibang(SMTG) to induce apoptosis was investigated in B16F10 melanoma cells. Method : Tetrazolium-based colorimetric assay was performed for cytotoxicity test. Several new assays for the basis of biochemical events associated with apoptosis such as DNA fragmentation by a flow cytometry, caspase-3 activation and PARP cleavage by Western blotting should be carried out potentially useful for the basis of biochemical events associated with apoptosis such as a flow cytometry and caspase-3 activation. Results : (1) The number of B16F10 melanoma cells was less than 30 % after exposure to 1 mg/ml SMTG for 48 h. SMTG increased cytotoxicity of B16F10 melanoma cells in a dose- and time-dependent manner. (2) The percentage of apoptotic cells by flow cytometric analysis of the DNA-stained cells increased to 21 % at 24 h and 25 % at 48 h after treatment with 1 mg/ml SMTG. (3) SMTG-induced apoptosis was accompained by the activation of caspase-3 and the specific proteolytic cleavage of poly-ADP-ribose polymerase. (4) SMTG induces the activation of caspase-3 and the specific proteolytic cleavage of poly-ADP-ribose polymearse and eventually leads to apoptosis through c-Jun NH2-terminal protein kinase (JNK)-dependent manner in B16F10 melanoma cells. Conclusion : SMTG had a strong cytotoxic effect of B16F10 melanoma cells.

• Fisheries and Aquatic Sciences
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• v.21 no.9
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• pp.21.1-21.8
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• 2018

In the present study, we first evaluated the melanin inhibitory effect of four crude 70% ethanol extracts separated from soft corals abundantly growing along the seawaters of Jeju Island, South Korea, including Dendronephthya castanea (DC), Dendronephthya gigantea (DG), Dendronephthya puetteri (DP), and Dendronephthya spinulosa (DS). Among the four ethanol extracts, the ethanol extract of DP (DPE) did not possess any cytotoxic effect on B16F10 cells. However, all other three extracts showed a cytotoxic effect. Also, DPE reduced the melanin content and the cellular tyrosinase activity without cytotoxicity, compared to the ${\alpha}-MSH$-stimulated B16F10 cells. Specifically, DPE downregulated the expression levels of tyrosinase and microphthalmia-associated transcription factor by activating the ERK signaling cascade in ${\alpha}-MSH$-stimulated B16F10 cells. Interestingly, the melanin inhibitory effect of DPE was abolished by the co-treatment of PD98059, an ERK inhibitor. According to these results, we suggest that DPE has whitening capacity with the melanin inhibitory effects by activating ERK signaling and could be used as a potential natural melanin inhibitor for cosmeceutical products.

• Journal of Physiology & Pathology in Korean Medicine
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• v.19 no.3
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• pp.662-670
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• 2005

Recently many efforts were focused to understand the mechanical insights of melanogenesis to develop the agents for hyper-pigmentation and hypo-pigmentation. In the melanin bio-synthetic pathway, tyrosinase is the rate limiting enzyme, and ${\alpha}$-melanocyte stimulating hormone(MSH) stimulates melanogenesis and enhances the melanin synthesis and the tyrosinase activity. The author has analyzed the effects of Biota Orientalis Folium on the basal melanogenic activities of B16 mouse melanoma cells, and on the ${\alpha}$-MSH or tyrosinase-induced melanogenesis. Biota Orientalis Folium alone markedly suppressed melanin content and tyrosinase activity in a dose-dependent manner. The decrease of cell propagation was observed in B16 cells treated with 200${\mu}$g/ml dose of Biota Orientalis Folium, indicating that Biota Orientalis Folium-induced depigmenting effect was caused by inhibition of melanin synthesis, not due to destruction of B16 cells. Pretreatment of the cells with Biota Orientalis Folium also suppressed the increase of ${\alpha}$-MSH (10 nM) induced melanin content and tyrosinase activity. Biota Orientalis Folium inhibited the revelation of ${\alpha}$-MSH induced tyrosinase protein and tyrosinase related protein and mRNA of tyrosinase in B16 melanoma cell. These results suggest that Biota Orientalis Folium inhibits melanogenesis and abrogates ${\alpha}$-MSH and tyrosinase-induced melanogenesis in B16 melanoma cells.

• Archives of Pharmacal Research
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• v.26 no.10
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• pp.861-867
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• 2003

Inhibitory effect of the lectins (KML-C) isolated from Korean mistletoe (KM; Viscum album coloratum) on tumor metastases produced by murine tumor cells (B16-BL6 melanoma, colon 26M3.1 carcinoma and L5178Y-ML25 lymphoma cells) was investigated in syngeneic mice. An intravenous (i.v.) administration of KML-C (20-50 ng/mouse) 2 days before tumor inoculation significantly inhibited lung metastases of both B16-BL6 and colon 26-M3.1 cells. The prophylactic effect of 50 ng/mouse of KML-C on lung metastasis was almost the same with that of 100 $\mu$ g/mouse of KM. Treatment with KML-C 1 day after tumor inoculation induced a significant inhibition of not only the experimental lung metastasis induced by B16-BL6 and colon 26M3.1 cells but also the liver and spleen metastasis of L5178Y-ML25 cells. Furthermore, multiple administration of KML-C given at 3 day-intervals after tumor inoculation led to a significant reduction of lung metastasis and suppression of the growth of B16-BL6 melanoma cells in a spontaneous metastasis model. In an assay for natural killer (NK) cell activity. i.v. administration of KML-C (50 ng/mouse) significantly augmented NK cytotoxicity against Yac-1 tumor cells 2 days after KML-C treatment. In addition, treatment with KML-C (50 ng/mouse) induced tumoricidal activity of peritoneal macrophages against B16-BL6 and 3LL cells. These results suggest that KML-C has an immunomodulating activity to enhance the host defense system against tumors, and that its prophylactic and therapeutic effect on tumor metastasis is associated with the activation of NK cells and macrophages.

• Journal of Acupuncture Research
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• v.35 no.4
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• pp.182-186
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• 2018

Background: The purpose of this study was to investigate the effects of wild ginseng pharmacopuncture on melanin production in B16/F10 murine melanoma cells. Methods: To determine the effect of wild ginseng pharmacopuncture solution on B16/F10 cells, cytotoxicity was evaluated using the 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyl-tetrazolium bromide (MTT) method. To observe B16/F10 cell growth, death, and morphological changes, Trypan blue solution was used. The Hosoi method was used to investigate the effect of wild ginseng pharmacopuncture solution on melanin production. The Martinez-Esparza method was used to investigate the effect of wild ginseng pharmacopuncture solution on tyrosinase activity. To determine the pathway involved in the melanogenesis in cells exposed to wild ginseng pharmacopuncture solution, a cell-free tyrosinase was used. Results: Following treatment with $200{\mu}L$ of wild ginseng solution, the cell survival rate was $76.32{\pm}2.45%$ which significantly decreased with higher concentrations (${\mu}L$) of wild ginseng (up to $200{\mu}L$). When $100{\mu}L$ of wild ginseng was used, the cell survival rate was $89.95{\pm}2.07%$. No morphological changes or abnormalities were observed in the B16/F10 murine melanoma cells as observed in the Trypan blue test. Melanin production was significantly reduced to $72.17{\pm}3.74%$ at $100{\mu}L$. Using $100{\mu}L$ of wild ginseng solution, tyrosinase activity was significantly decreased to $80.15{\pm}1.05%$. Wild ginseng pharmacopuncture solution reduced melanin production both directly and indirectly. Conclusion: This study suggests that wild ginseng pharmacopuncture solution may be effective in inhibiting melanin production. Further studies are needed to determine safe and effective clinical applications.

• Preventive Nutrition and Food Science
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• v.15 no.3
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• pp.213-220
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• 2010

To examine the potential of Terminalia chebula as a whitening agent, we measured antioxidant activity using DPPH$\cdot$, ABTS${\cdot}^+$ assays and ferric-reducing antioxidant power (FRAP) assays, and depigmenting activity using B16F10 melanoma cells. The intracellular reactive oxygen species (ROS) level was monitored by $H_2DCFDA$ fluorescence labeling, and melanin contents in B16F10 melanoma cells by 960 $J/m^2$ dose of UVA-induced oxidative stress. The radical-scavenging activities of T. chebula extract (TCE) were measured in terms of $EC_{50}$ values using DPPH$\cdot$, ABTS${\cdot}^+$ assays and FRAP value were 280.0 ${\mu}g/mL$, 42.2 ${\mu}g/mL$ and 113.1 ${\mu}mol$ $FeSO_4{\cdot}7H_2O/g$, respectively. We found that ROS and melanin concentrations were reduced by TCE treatments of 25 ${\mu}g/mL$ under UVA-induced oxidative stress. Tyrosinase activity and melanin contents in $\alpha$-melanocyte stimulating hormone (MSH)-induced melanoma cells both decreased dose-dependently in the treatment groups. TCE similarly reduced melanogenesis in B16F10 melanoma cells stimulated by $\alpha$-MSH as compared to arbutin as a positive control. T. chebula may prove to be a useful therapeutic agent for hyperpigmentation and an effective component in skin whitening and.or lightening cosmetics.

• Journal of Life Science
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• v.22 no.3
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• pp.318-323
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• 2012

This study involved observation of the inhibitory effect of 70% EtOH and water extracts from Doinseunggitang on melanin synthesis, tyrosinase activity, and western blotting using B16F10 melanoma cells. Doinseunggitang extracts inhibited melanin synthesis and tyrosinase activity in a dependent manner. As a result, it was found that Doinseunggitang 70% EtOH extracts inhibit melanin synthesis and tyrosinase activity, respectively, by 40% and 51%. In addition, western blotting analysis showed that 70% EtOH extracts inhibited tyrosinase, MITF, TRP-1, and TRP-2 expression. These results show that 70% ethanol extracts of Doinseunggitang could be developed as a skin whitening material in cosmetics.

• YAKHAK HOEJI
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• v.57 no.1
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• pp.24-31
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• 2013

Although statins, inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, have been shown to increase melanin synthesis, the exact mechanism of this action is not fully understood. In this study we investigated the possible involvement of intracellular $Ca^{2+}$ signal in the mechanism of stimulation of melanin synthesis induced by lovastatin in B16 cells. Lovastatin stimulated the production of melanin in a dose-dependent manner in the cells. Treatment with mevalonate, FPP and GGPP, precursors of cholesterol, did not significantly suppress the lovastatin-induced melanin production, suggesting that inhibition of cholesterol synthesis may not be involved in the mechanism of the action of lovastatin. In addition, lovastatin did not significantly alter the cAMP concentration and the stimulated production of melanin by lovastatin was not significantly changed by treatment with H89, a potent inhibitor of protein kinase A, which demonstrates that cAMP pathway may not be involved. However, lovastatin increased intracellular $Ca^{2+}$ concentration in a dose-related fashion. Treatment with EGTA, an extracellular $Ca^{2+}$ chelator did not significantly alter the lovastatin-induced intracellular $Ca^{2+}$ increase and melanin synthesis, whereas intracellular $Ca^{2+}$ reduction with BAPTA/AM and intracellular $Ca^{2+}$ release blockers (dantrolene and TMB-8) completely blunted these actions of lovastatin. Taken together, these results suggest that the intracellular $Ca^{2+}$ release may play an important role in the lovastatin-induced stimulation of melanin synthesis in B16 cells. These results further suggest that lovastatin may be useful for the treatment of hypopigmentation disorders, such as vitiligo.