• Journal of Ginseng Research
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• 제45권5호
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• pp.555-564
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• 2021

Background: Ginsenosides of Panax ginseng are used to enhance skin health and beauty. The present study aimed to investigate the potential use of ginsenoside Rf (Rf) from Panax ginseng as a new anti-pigmentation agent. Methods: The anti-melanogenic effects of Rf were explored. The transcriptional activity of the cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) and the expression levels of tyrosinase, microphthalmia-associated transcription factor (MITF), and tyrosinase-related proteins (Tyrps) were evaluated in melanocytes and UV-irradiated ex vivo human skin. Results: Rf significantly inhibited Forskolin (FSK) or UV-stimulated melanogenesis. Consistently, cellular tyrosinase activity and levels of MITF, tyrosinase, and Tyrps were downregulated. Furthermore, Rf suppressed MITF promoter activity, which was stimulated by FSK or CREB-regulated transcription coactivator 3 (CRTC3) overexpression. Increased CREB phosphorylation and protein kinase A (PKA) activity induced by FSK were also mitigated in the presence of Rf. Conclusion: Rf can be used as a reliable anti-pigmentation agent, which has a scientifically confirmed and reproducible action mechanism, via inhibition of CREB/MITF pathway.

• The Korean Journal of Physiology and Pharmacology
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• 제26권2호
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• pp.113-123
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• 2022

Diarylpropionitrile (DPN), a selective agonist for estrogen receptor β (ERβ), has been reported to regulate various hormonal responses through activation of ERβ in tissues including the mammary gland and brain. However, the effect of DPN on melanogenesis independent of ERβ has not been studied. The aim of this study is to examine the possibility of anti-melanogenic effect of DPN and its underlying mechanism. Melanin contents and cellular tyrosinase activity assay indicated that DPN inhibited melanin biosynthesis in alpha-melanocyte stimulating hormone-stimulated B16F10 melanoma cell line. However, DPN had no direct influence on in vitro tyrosinase catalytic activity. On the other hand, 17β-estradiol had no effect on inhibition of melanogenesis, suggesting that the DPN-mediated suppression of melanin production was not related with estrogen signaling pathway. Immunoblotting analysis showed that DPN down-regulated the expression of microphthalmia-associated transcription factor (MITF), a central transcription factor of melanogenesis and its down-stream genes including tyrosinase, tyrosinase-related protein (TRP)-1, and TRP-2. Also, DPN attenuated the phosphorylation of protein kinase A (PKA) and cAMP-response element-binding protein (CREB). Additionally, DPN suppressed the melanin synthesis in UVB-irradiated HaCaT conditioned media culture system suggesting that DPN has potential as an anti-melanogenic activity in physiological conditions. Collectively, our data show that DPN inhibits melanogenesis via downregulation of PKA/CREB/MITF signaling pathway.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2021년도 춘계학술대회
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• pp.54-54
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• 2021

The Melanoma Research Coalition reported melanoma affects humans of various races. This study was conducted to confirm the inhibitory effect of melanogenesis in B16 F10 cells of Nypa fruticans Wurmb of ethyl acetate fraction (NEF). Nypa fruticans Wurmb is an important component of the East Asian mangrove vegetation. It belongs to Araceae family. Traditionally, N. fruticans was used to treat various diseases such as asthma, sore throat, liver disease, a pain reliever, and can also be used as sedative and carminative. The present study, the inhibitory effect on melanogenesis was determined by Western blotting and RT-qPCR. The level of expression of tyrosinase, TRP-1, and TRP-2 is regulated by microphthalmia-associated transcription factor (MITF) and cAMP, and cAMP affects the activity of protein kinase A (PKA). Activated PKA stimulates the phosphorylation of cAMP-reactive element-binding protein (CREB) in the nucleus, thereby increasing the amount of MITF expression and enhancing melanogenesis. Western blotting and RT-qPCR analysis showed that NEF treatment decreased the expression of tyrosinase. Similarly, TRP-1 and TRP-2 levels were decreased, which were decreased significantly at compared with the untreated control. Also, NEF attenuated the IBMX mediated increase in the intracellular cAMP level and the phosphorylation of PKA. In conclusion, NEF significantly inhibited the expressions of melanogenesis through cAMP/PKA/CREB signaling pathways.

• Journal of Ginseng Research
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• 제47권6호
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• pp.714-725
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• 2023

Background: Our previous investigation indicated that the preparation of Panax ginseng Meyer (P. ginseng) inhibited melanogenesis. It comprised salicylic acid (SA), protocatechuic acid (PA), p-coumaric acid (p-CA), vanillic acid (VA), and caffeic acid (CA). In this investigation, the regulatory effects of P. ginseng phenolic acid monomers on melanin production were assessed. Methods: In vitro and in vivo impact of phenolic acid monomers were assessed. Results: SA, PA, p-CA and VA inhibited tyrosinase (TYR) to reduce melanin production, whereas CA had the opposite effects. SA, PA, p-CA and VA significantly downregulated the melanocortin 1 receptor (MC1R), cycle AMP (cAMP), protein kinase A (PKA), cycle AMP-response element-binding protein (CREB), microphthalmia-associated transcription factor (MITF) pathway, reducing mRNA and protein levels of TYR, tyrosinase-related protein 1 (TYRP1), and TYRP2. Moreover, CA treatment enhanced the cAMP, PKA, and CREB pathways to promote MITF mRNA level and phosphorylation. It also alleviated MITF protein level in α-MSH-stimulated B16F10 cells, comparable to untreated B16F10, increasing the expression of phosphorylation glycogen synthase kinase 3β (p-GSK3β), β-catenin, p-ERK/ERK, and p-p38/p38. Furthermore, the GSK3β inhibitor promoted p-GSK3β and p-MITF expression, as observed in CA-treated cells. Moreover, p38 and ERK inhibitors inhibited CA-stimulated p-p38/p38, p-ERK/ERK, and p-MITF increase, which had negative binding energies with MC1R, as depicted by molecular docking. Conclusion: P. ginseng roots' phenolic acid monomers can safely inhibit melanin production by bidirectionally regulating melanin synthase transcription. Furthermore, they reduced MITF expression via MC1R/cAMP/PKA signaling pathway and enhanced MITF post-translational modification via Wnt/mitogen-activated protein kinase signaling pathway.

• Journal of Microbiology and Biotechnology
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• 제32권8호
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• pp.982-988
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• 2022

Licorice (Glycyrrhiza) has been used as preventive and therapeutic material for hyperpigmentation disorders. Previously, we isolated noble compounds including dehydroglyasperin C (DGC), dehydroglyasperin D (DGD) and isoangustone A (IAA) from licorice hexane/ethanol extracts. However, their anti-melanogenic effects and underlying molecular mechanisms are unknown. The present study compared effects of DGC, DGD and IAA on pigmentation in melan-a melanocytes and human epidermal melanocytes (HEMn). DGD exerted the most excellent anti-melanogenic effect, followed by DGC and IAA at non-cytotoxic concentrations. In addition, DGD significantly inhibited tyrosinase activity in vitro cell-free system and cell system. Western blot result showed that DGD decreased expression of microphthalmia-associated transcription factor (MITF), tyrosinase and tyrosinase-related protein-1 (TRP-1) in melan-a cells and HEMn cells. DGD induced phosphorylation of MITF, ERK and Akt signal pathway promoting MITF degradation system. However, DGD did not influence p38 and cAMP-dependent protein kinase (PKA)/CREB signal pathway in melan-a cells. These result indicated that DGD inhibited melanogenesis not only direct regulation of tyrosinase but also modulating intracellular signaling related with MITF level. Collectively, these results suggested a protective role for DGD against melanogenesis.

• 한방안이비인후피부과학회지
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• 제28권1호
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• pp.41-52
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• 2015

Objective : This study investigated the inhibitory mechanism of the hypopigmentating effects on ethanol extract of Rosae rugosae Flos (ERR) that has not yet been examined. Methods : We analyzed the anti-melanogenic effects of ethanol extracts from Rosae rugosae Flos by tyrosinase activity, melanin contents. We also examined protein expression levels of tyrosinase, TRP-1, TRP-2, MITF and ERK by western blot analysis in melanoma cells. Results : In this investigation, ERR effectively reduced ${\alpha}$-MSH-stimulated melanin synthesis by suppressing expression of tyrosinase and tyrosinase-related protein-1 (TRP-1). On the other hand, the expression of tyrosinase-related protein-2 (TRP-2) were not affected by treatment with ERR. ERR inhibited the expression of microphthalmia-associated transcription factor (MITF) as a key transcription factor for tyrosinase expression regulating melanogenesis. The upstream signaling pathway including cAMP response element-binding protein (CREB) and MAPKs were also inhibited by ERR. Pretreatment with PD98059, ERK inhibitor, attenuated the inhibitory effect of ERR on ${\alpha}$-MSH-induced tyrosinase activity. Conclusions : Our study suggested that the anti-melanogenic activity of ERR is correlated with the suppression of tyrosinase gene through CREB/MITF/ERK pathway.

• 한국응용과학기술학회지
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• 제38권3호
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• pp.883-890
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• 2021

본 연구는 B16F10 melanoma 세포에서 산딸기 잎 추출물(Rubus crataefifolius Leaf Extract, RCLE)의 멜라닌 생성 억제 효과를 확인하고자 수행되었다. α-MSH로 자극한 B16F10 melanoma 세포에서 멜라닌 함량과 tyrosinase 활성, 멜라닌 생성관련 효소들인 TRP-1, TRP-2 및 MITF의 단백질 발현 수준을 조사하였고 이에 대한 RCLE의 효과를 검증하였다. RCLE는 tyrosinase 활성과 멜라닌 생성을 효과적으로 억제하였고 멜라닌 생성 경로에 관여하는 PKA와 CREB의 인산화와 MITF의 발현을 억제하였으며 멜라닌 생성 관련 단백질의 발현을 하향 조절하였다. 이러한 결과는 RCLE가 MITF 발현을 억제하여 α-MSH로 자극된 멜라닌 합성을 억제한다는 것을 보여준다. 따라서 이러한 연구결과는 RCLE가 과도한 멜라닌 생성으로 인한 색소 침착을 완화시킬 수 있는 기능성 화장품 소재로 활용될 수 있음을 시사한다.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2023년도 임시총회 및 춘계학술대회
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• pp.56-56
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• 2023

The cone of Pinus rigida × Pinus taeda (PRT), a plant in the Pinaceae family, has long been used in traditional medicine to treat hemostasis, bruises, and burns. Previous research has shown that regulating oxidation-reduction reactions in reactive oxygen species can help inhibit melanogenesis, the process of melanin synthesis, which is a common target for addressing hyperpigmentation. Inhibiting tyrosinase is also known to be effective in this regard. Based on these findings, we conducted an investigation into the inhibitory effect of the ethyl acetate fraction of PRT (ERT) on melanogenesis in B16 F10 cells. We know that the expression levels of melanin biosynthesis-related proteins, including tyrosinase, TRP-1, and TRP-2, are regulated by MITF (microphthalmia-associated transcription factor) and cAMP, with cAMP affecting the activity of protein kinase A (PKA). PKA can reduce melanogenesis, and CREB reduces the phosphorylation of melanin-producing enzymes. In addition, the MAPK signaling pathway, composed of ERK, JNK, p38, and other factors, is also known to play a role in the inhibition of melanogenesis in melanocytes. Our immunoblotting results showed that ERT inhibited the expression of melanin production-related proteins (tyrosinase, TRP-1, TRP-2, and MITF) that were significantly increased by a-MSH treatment to promote melanin production. Furthermore, the phosphorylation levels of factors related to cAMP/PKA/CREB and MAPK signaling pathways were significantly reduced without affecting the total form. In conclusion, we believe that treatment with ERT can inhibit melanin synthesis by modulating the phosphorylation of cAMP/PKA/CREB and MAPK signaling pathways at the cellular level. These findings suggest the potential of ERT as a raw material for functional cosmetics and pharmaceuticals, thanks to its antioxidant activity and ability to inhibit melanogenesis. We thought that these findings of ERT as a natural plant resource will inspire further research and development in this area.

• 생명과학회지
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• 제32권5호
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• pp.355-361
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• 2022

멜라닌 색소는 피부색의 주요 원인이다. 멜라닌 색소는 멜라닌 세포에서 생성된 다음 각질 세포로 전달되어 결국 피부 표면에 다양한 색상을 부여한다. 많은 탈색제 및 피부 미백제가 개발되었지만, 색소 침착을 감소시키기 위한 재료에 대한 수요는 여전히 증가하고 있다. 본 연구에서 천연 화합물을 사용하여 탈색 및 피부 미백에 대한 재료를 찾으려고 시도한 결과 겨우살이(Viscum album var. coloratum) 추출물이 색소 침착을 억제할 수 있음을 발견하였다. 인간 멜라닌 세포에 겨우살이 추출물(mistletoe extracts, ME)을 처리했을 때 색소 침착이 극적으로 감소하였다. 프로모터 리포터 분석은 ME 처리가 HM3KO 흑색종 세포에서 microphthalmia-associated transcription factor (MITF), melanophilin (MLPH), tyrosinase related protein 2 (TRP-2), and tyrosinase (TYR) 유전자의 전사를 억제한다는 것을 보여주었다. 일관되게 ME는 MITF, TRP-1 및 TYR과 같은 색소 침착 관련 분자의 단백질 수준을 감소시켰다. 또한 ME는 cAMP Responsive Element Binding Protein (CREB), AKT 및 ERK의 인산화를 감소시켰다. 이러한 결과는 ME가 색소 침착과 관련된 세포 내 신호 전달의 조절을 통해 멜라닌 생성을 억제한다는 것을 시사한다. 끝으로 ME는 색소 침착에 대한 생체 내 평가 모델인 제브라피쉬 배아의 멜라닌 생성을 현저하게 억제하였다.

• 한방안이비인후피부과학회지
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• 제24권1호
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• pp.25-35
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• 2011

Objective : The present study was designed to assess the potential inhibitory activity of an ethanol extract of Belamcandae Rhizoma (EBR) on the alpha-melanocyte stimulating hormone (${\alpha}$-MSH)-induced melanogenesis signal pathway in B16F10 melanoma cells. Methods : Several experiments were performed in B16F10 melanoma cells. We studied tyrosinase activity, melanin content, cell-free tyrosinase activity and DOPA stain, and performed Western blots and RT-PCR for proteins and mRNA involved in melanogenesis. Results : ${\alpha}$-MSH-induced tyrosinase activity and melanin content were inhibited significantly by EBR. EBR markedly suppressed the protein expression level of tyrosinase in B16F10 melanoma cells. On the other hand, the expression of tyrosinase-related protein-1 (TRP-1) and -2 (TRP-2; DCT) were not affected by EBR. To elucidate the mechanism of the depigmenting property of EBR, we examined the involvement EBR in cAMP response element binding (CREB) protein phosphorylation and microphthalmia-associated transcription factor (MITF) signalling induced by ${\alpha}$-MSH. EBR did not regulate CREB phosphorylation and MITF expression by ${\alpha}$-MSH. Nevertheless, the mRNA expression of tyrosinase was significantly attenuated by EBR treatment without changes in the expression of TRP-1 and -2 mRNA. Conclusion : Our study suggested that EBR inhibits ${\alpha}$-MSH-induced melanogenesis by suppressing tyrosinase mRNA.