Melanin Inhibitory Effect of Tuber himalayense Isolated in Incheon, Korea.

There has been a growing interest in skin beauty and antimelanogenic products. Melanogenesis is the process of melanin synthesis whereby melanocytes are activated by UV light or hormone stimulation to produce melanin. Melanogenesis is mediated by several enzymes, such as tyrosinase (TYR), microphthalmia-associated transcription factor (MITF), tyrosinase-related protein-1 (TRP-1), and TRP-2. In this study, we investigated the effect of Tuber himalayense extract on melanin synthesis in α-melanocyte-stimulating hormone (α-MSH)-treated B16F10 melanoma cells. We confirmed that T. himalayense extract was not toxic to α-MSH-treated B16F10 melanoma cells and exhibited a significant inhibitory effect on melanin synthesis at concentrations of 25, 50, and 100 µg/ml. Additionally, the T. himalayense extract inhibited melanin, TRP-1, TRP-2, tyrosinase, and MITF, which are enzymes involved in melanin synthesis, in a concentration-dependent manner. Furthermore, T. himalayense extract inhibited the mitogen-activated protein kinase (MAPK) pathways, such as extracellular signal-regulated kinase-1/2 (ERK), c-Jun N-terminal kinase (JNK), and p38. Therefore, we hypothesized that various components of T. himalayense extract affect multiple factors involved in melanogenesis in B16F10 cells. Our results indicate that T. himalayense extract could potentially be used as a new material for preparing whitening cosmetics.

The whitening effect of cuscutin responsible for traditional use of Bergenia purpurascens.

ETHNOPHARMACOLOGICAL RELEVANCE: The roots and rhizomes of Bergenia purpurascens (Hook. f. et Thomson) Engl., was used as a sunscreen to protect against ultraviolet rays in Tibet of China historically, but its skin whitening constituents and pharmacological effects of this plant remained unknown. AIM OF THE STUDY: To investigate the anti-melanogenesis effect of B. purpurascens in vitro and in vivo, and then explore the preliminary mechanism. MATERIALS AND METHODS: An ultraviolet B (UVB)-induced skin injury model of mice was used to verify the ameliorative effect of B. purpurascens extract (BPE) on ultraviolet damage. Then, alpha-melanocyte stimulating hormone (α-MSH)-induced murine melanoma cell line (B16F10) melanin generation model was further adopted to approval the effects of BPE and its bioactive compound, cuscutin, in vitro. Moreover, α-MSH stimulated melanogenesis model in zebrafish was employed to confirm the anti-pigmentation effect of cuscutin. Then, proteins expressions associated with melanin production were observed using western blotting assay to explore preliminary mechanism. RESULTS: BPE inhibited UVB-induced mice injury and restored skin barrier function observably in vivo. BPE and cuscutin suppressed the overproduction of melanin in α-MSH induced B16F10 significantly, in which cuscutin exhibited better effect than well-known whitening agent α-arbutin at same 10 µg/mL concentration. Moreover, the pigmentation of zebrafish embryo was decreased by cuscutin. Finally, cuscutin showed significant downregulation of expressions of tyrosinase (TYR) and tyrosinase related protein-1 (TRP-1), TRP-2 and microphthalmia-associated transcription factor (MITF) in the melanogenic signaling pathway. CONCLUSION: B. purpurascens extract and its major bioactive constituent, cuscutin, showed potent anti-melanogenesis and skin-whitening effect by targeting TYR and TRP-2 proteins for the first time, which supported its traditional use.

Inhibition of α-melanocyte-stimulating hormone-induced melanogenesis and molecular mechanisms by polyphenol-enriched fraction of Tagetes erecta L. flower.

BACKGROUND: The pursuit for safe and efficacious skin-whitening agents has prompted a dedicated exploration of plant-derived compounds. Notably, Tagetes erecta L. flowers have been used as a medicinal extract and possessed in vitro mushroom tyrosinase activity. However, whether polyphenol-enriched fraction extracted from T. erecta L. flowers (TE) regulates melanogenesis within cellular and animal models has not yet been investigated. PURPOSE: This study aimed to investigate the effect of TE as a prospective inhibitor of melanogenesis. METHODS: Through advanced UPLC-QTof/MS analysis, the components of TE were analyzed. Anti-melanogenic effects of TE were evaluated in α-melanocyte-stimulating hormone (α-MSH)-stimulated B16F10 melanoma cells by measuring cell viability assay, extracellular and intracellular melanin biosynthesis, cyclic adenosine monophosphate (cAMP) production, and melanogenesis-related gene and protein expression. Zebrafish larvae were employed for in vivo studies, assessing both heart rate and melanogenesis. Furthermore, molecular docking analyses were employed to predict the interaction between TE components and the melanocortin 1 receptor (MC1R). Direct binding activity of TE components to MC1R was compared with [Nle4, d-Phe7]-MSH (NDP-MSH). RESULTS: TE was found to contain significant phenolic compounds such as patulitrin, quercetagetin, kaempferol, patuletin, and isorhamnetin. This study revealed that TE effectively inhibits melanin biosynthesis in both in vitro and in vivo models. This inhibition was attributed to interference of TE with the cAMP-cAMP response element-binding protein (CREB)-microphthalmia-associated transcription factor (MITF)-tyrosinase pathway, which plays a pivotal role in regulating melanogenesis. Importantly, TE exhibited the remarkable ability to curtail α-MSH-induced melanogenesis in zebrafish larvae without impacting heart rates. Molecular docking analyses predicted that the components of TE possibly interact with the melanocortin 1 receptor, suggesting their role as potential inhibitors of melanin biosynthesis. However, through the direct binding activity compared with NDP-MSH, any TE components did not directly bind to MC1R, suggesting that TE inhibits α-MSH-induced melanogenesis by inhibiting the cAMP-mediated intracellular signaling pathway. The assessment of anti-melanogenic activity, conducted both in vitro and in vivo, revealed that patulitrin and patuletin exhibited significant inhibitory effects on melanin formation, highlighting their potency as major contributors. DISCUSSION: This investigation demonstrated the considerable potential of TE as a natural remedy endowed with remarkable anti-melanogenic properties. The demonstrated capacity of TE to attenuate melanin production by modulating the cAMP-CREB-MITF-tyrosinase pathway underscores its central role in management of disorders associated with excessive pigmentation. Importantly, the implications of these findings extend to the cosmetics industry, where TE emerges as a prospective and valuable ingredient for the formulation of skin-whitening products. The elucidated interactions between TE components and MC1R not only provide insight into a potential mechanism of action but also elevate the significance of this study. In summary, this study not only contributes to our comprehension of pigmentation-related conditions but also firmly establishes TE as a secure and natural strategy for the regulation of melanin production. The innovative aspects of TE propel it into the forefront of potential interventions, marking a noteworthy advancement in the pursuit of effective and safe solutions for pigmentation disorders.

Integrative pharmacology reveals the mechanisms of Erzhi pills, A traditional Chinese formulation, stimulating melanogenesis.

ETHNOPHARMACOLOGICAL RELEVANCE: Erzhi pills (EZP), a traditional Chinese medicine formula prescribed for the treatment of vitiligo, has shown promising efficacy. However, the oral bioactive components and mechanisms underlying the promotion of melanogenesis by EZP remain unclear. AIM OF THE STUDY: This study aimed to investigate the pharmacological basis and mechanism of EZP in promoting melanogenesis. MATERIALS AND METHODS: UHPLC-TOF-MS analysis was used to identify absorbed phytochemicals in serum after oral administration of EZP. Network pharmacology methods were used to predict potential targets and pathways involved in the melanogenic activity of EZP, resulting in the construction of a "compound-target-pathway" network. Zebrafish and B16F10 cells were used to evaluate the effects of EZP on tyrosinase activity and melanin content. Western blot and ELISA analyses were used to validate the effects of EZP on melanogenesis-related proteins, including MITF, TYR, CREB, p-CREB, and cAMP. RESULTS: UHPLC-TOF-MS analysis identified 36 compounds derived from EZP in serum samples. Network pharmacology predictions revealed 89 target proteins associated with the identified compounds and closely related to vitiligo. GO and KEGG analyses indicated the involvement of the cAMP/PKA signaling pathway in the promotion of melanogenesis by EZP. Experimental results showed that EZP increased tyrosinase activity and melanin content in zebrafish and B16F10 cells without inducing toxicity. Western blot and ELISA results suggested that the melanogenic effect of EZP may be related to the activation of the cAMP/PKA signaling pathway. These results confirm the feasibility of combining serum pharmacological and network pharmacological approaches. CONCLUSIONS: EZP have the potential to increase tyrosinase activity and melanin content in zebrafish and cells possibly through activation of the cAMP/PKA pathway.

Tribuloside acts on the PDE/cAMP/PKA pathway to enhance melanogenesis, melanocyte dendricity and melanosome transport.

ETHNOPHARMACOLOGICAL RELEVANCE: Tribuloside, a natural flavonoid extracted from Chinese medicine Tribulus terrestris L., has shown potent efficacy in treating various diseases. In China, the fruits of Tribulus terrestris L. have long been utilized for relieving headache, dizziness, itchiness, and vitiligo. Water-based extract derived from Tribulus terrestris L. can enhance melanogenesis in mouse hair follicle melanocytes by elevating the expression of α-melanocyte stimulating hormone (α-MSH) and melanocortin-1 recepter (MC-1R). Nevertheless, there is a lack of information regarding the impact of tribuloside on pigmentation in both laboratory settings and living organisms. AIM OF THE STUDY: The present research aimed to examine the impact of tribuloside on pigmentation, and delve into the underlying mechanism. MATERIALS AND METHODS: Following the administration of tribuloside in human epidermal melanocytes (HEMCs), we utilized microplate reader, Masson-Fontana ammoniacal silver stain, transmission electron microscopy (TEM) and scanning electron microscopy (SEM) to measure melanin contents, dendrite lengths, melanosome counts; L-DOPA oxidation assay to indicate tyrosinase activity, Western blotting to evaluate the expression of melanogenic and associated phosphodiesterase (PDE)/cyclic adenosine monophosphate (cAMP)/cyclic-AMP dependent protein kinase A (PKA) pathway proteins. A PDE-Glo assay to verify the inhibitory effect of tribuloside on PDE was also conducted. Additionally, we examined the impact of tribuloside on the pigmentation in both zebrafish model and human skin samples. RESULTS: Tribuloside had a notable impact on the production of melanin in melanocytes, zebrafish, and human skin samples. These functions might be attributed to the inhibitory effect of tribuloside on PDE, which could increase the intracellular level of cAMP to stimulate the phosphorylation of cAMP-response element binding (CREB). Once activated, it induced microphthalmia-associated transcription factor (MITF) expression and increased the expression of tyrosinase, Rab27a and cell division cycle protein 42 (Cdc42), ultimately facilitating melanogenesis, melanocyte dendricity, and melanin transport. CONCLUSION: Tribuloside acts on the PDE/cAMP/PKA pathway to enhance melanogenesis, melanocyte dendricity, and melanosome transport; meanwhile, tribuloside does not have any toxic effects on cells and may be introduced into clinical prescriptions to promote pigmentation.

Inhibitory Effects of Polyphenol- and Flavonoid-Enriched Rice Seed Extract on Melanogenesis in Melan-a Cells via MAPK Signaling-Mediated MITF Downregulation.

Melanin production is an important process that prevents the host skin from harmful ultraviolet radiation; however, an overproduction of melanin results in skin diseases. In the present study, we determined the antioxidative and anti-melanogenic activities of polyphenol- and flavonoid-enriched rice seed extracts in melan-a cells. The polyphenol and flavonoid content of Hopum (HP) and Sebok (SB) rice seed extracts was measured. The antioxidant capacity was determined using the ABTS radical scavenging method. SB contained high amounts of polyphenols and flavonoids, which significantly increased antioxidative activity compared with HP. Various concentrations of these extracts were evaluated in a cytotoxicity using melan-a cells. At 100 µg/mL, there was no significant difference for all treatments compared with untreated cells. Therefore, 100 µg/mL was selected as a concentration for the further experiments. SB significantly suppressed the phosphorylation/activation of p-38 MAPK, increased the expression of phosphorylated ERK 1/2 and Akt, and downregulated the microphthalmia-associated transcription factor (MITF). This resulted in decreased levels of tyrosinase and tyrosinase-related protein-1 and -2. These results indicate the potential of polyphenol- and flavonoid-enriched rice seed as a treatment for hyperpigmentation.

Pueraria protein extract inhibits melanogenesis and promotes melanoma cell apoptosis through the regulation of MITF and mitochondrial­related pathways.

Pueraria Lobata Radix (P. Lobata Radix) is an edible traditional Chinese medicine that contains various active compounds. Proteins from P. Lobata Radix have become the subject of increased interest in recent years. In evaluating the whitening effect on the skin, the present study found that the P. Lobata Radix water­soluble total protein extract (PLP) had the strongest inhibitory effect on tyrosinase activity. In the present study, the anti­melanogenic effect of PLP and the inhibitory effect on B16 melanoma cells were investigated. PLP significantly reduced the tyrosinase activity and melanin content in B16 melanoma cells. Mechanistically, PLP inhibited melanogenesis by decreasing the expression of tyrosinase, tyrosinase­related protein (TRP)­1 and TRP­2 through downregulation of the microphthalmia­associated transcription factor (MITF) gene, which was mediated by inhibition of p38 mitogen­activated protein kinase signaling. In addition, PLP inhibited cell viability and triggered apoptosis of B16 cells in a dose­dependent manner. Exposure to PLP reduced the mitochondrial membrane potential (MMP) and decreased ATP generation, leading to mitochondria­related apoptosis of B16 melanoma cells. The expression levels of succinate dehydrogenase (SDH) and its two related subunits (SDHA and SDHB) were downregulated significantly by PLP, which may be associated with the regulation of mitochondrial energy metabolism by PLP. These results may explain why MMP collapse and reduced ATP generation were observed in B16 melanoma cells treated with PLP. Finally, the present study demonstrated that the inhibition of melanin synthesis by PLP was correlated with the regulation of antioxidant enzymes to reduce reactive oxygen species levels. These results suggested that PLP inhibits melanogenesis by downregulating the expression of MITF­related melanogenic enzymes and triggering apoptosis through mitochondria­related pathways.

Morus alba L. root decreases melanin synthesis via sphingosine-1-phosphate signaling in B16F10 cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Morus alba L. has long been used for beauty in many Asian countries and regions, including anti-aging and hyperpigmentation. AIM OF THE STUDY: This study aimed at the inhibitory effect of Morus alba L. root on melanogenesis in B16F10 melanoma cells and the mechanism involved. MATERIALS AND METHODS: This study evaluated the anti-melanogenic effect of Morus alba L. root extract (MAR) on B16F10 melanoma cells by assessing cell viability, melanin accumulation, cellular tyrosinase activity, intra/inter-cellular S1P levels, cellular S1P-related metabolic enzyme activity, and western blot analysis. In addition, the potential S1P lyase (S1PL) inhibitory constituents in MAR were identified by LC-MS/MS. RESULTS: Without affecting the viability of B16F10 melanoma cells, MAR inhibited intracellular tyrosinase activity in a dose-dependent manner, thereby reducing the accumulation of melanin. MAR also downregulated the expression level of MITF via activating the ERK signaling pathway. Furthermore, MAR increased the intra/inter-cellular S1P by inhibiting S1PL. Several compounds with inhibitory S1PL activity have been identified in MAR, such as mulberroside A and oxyresveratrol. CONCLUSIONS: The anti-melanogenic effects of MAR mainly involve promoting MITF degradation mediated via S1P-S1PR3-ERK signaling through increasing cellular S1P levels by inhibiting S1PL activity.

Ligularia fischeri ethanol extract: An inhibitor of alpha-melanocyte-stimulating hormone-stimulated melanogenesis in B16F10 melanoma cells.

BACKGROUND: Ligularia fischeri is a perennial herb isolated from plants of the Asteraceae family. Ligularia fischeri is distributed throughout Korea, Japan, eastern Siberia, and China. AIMS: The aim of this study is to examine the intracellular inhibitory effect of Ligularia fischeri ethanol extract on melanin synthesis and expression of tyrosinase and tyrosinase-related protein 1 and 2. In addition, we analyzed the mitogen-activated protein kinase signaling pathway and microphthalmia-associated transcription factor in alpha-melanocyte-stimulating hormone-stimulated B16F10 melanoma cells. METHODS: To assess the inhibition of melanogenesis in alpha-melanocyte-stimulating hormone-stimulated B16F10 melanoma cells, the expression of melanogenesis-related genes was investigated by quantitative real-time polymerase chain reaction, while western blotting was performed to determine protein expression levels. RESULTS: We confirmed that the ethanol extract of Ligularia fischeri inhibited melanin synthesis in vitro by decreasing tyrosinase and tyrosinase-related protein 1 and 2 expression. Furthermore, we revealed that tyrosinase expression was regulated by the suppression of microphthalmia-associated transcription factor expression and activation of extracellular signal-regulated kinase phosphorylation. The ethanol extract of Ligularia fischeri inhibited melanogenesis by activating extracellular signal-regulated kinase phosphorylation and suppressing microphthalmia-associated transcription factor and tyrosinase expression. CONCLUSIONS: Ligularia fischeri ethanol extract may be used as an effective skin whitening agent in functional cosmetics.

SNA077, an Extract of Marine Streptomyces sp., Inhibits Melanogenesis by Downregulating Melanogenic Proteins via Inactivation of cAMP/PKA/CREB Signaling.

Excess melanin in skin is known to be the main cause of hyper-pigmentary skin diseases such as freckles and lentigo. This study aimed to evaluate the depigmenting efficacy of an extract from the marine microorganism strain, Streptomyces sp. SNA077. To determine the anti-melanogenic efficacy of SNA077, we assessed the melanin contents of SNA077-treated B16, Melan-a, and MNT-1 cells. We observed the expression of key enzymes in melanogenesis via qRT-PCR and Western blot analyses. We further estimated the skin-whitening effect of SNA077 using a skin-equivalent model. SNA077 dramatically decreased the melanin production of B16 cells, Melan-a, and MNT-1 cells. In B16 cells treated with SNA077, the activity of cellular tyrosinase was clearly inhibited. In addition, the mRNA and protein expression levels of melanogenic genes were suppressed by SNA077 treatment in B16 and MNT-1 cells. Upstream of tyrosinase, the expression levels of phospho-CREB, phospho-p38, PKA activity, cyclic AMP production, and MC1R gene expression were inhibited by SNA077. Finally, SNA077 clearly showed a skin-brightening effect with a reduced melanin content in the skin tissue model. Collectively, our results suggest for the first time that an extract of marine Streptomyces sp. SNA077 could be a novel anti-melanogenic material for skin whitening.

Molecular Analysis of the Melanogenesis Inhibitory Effect of Saponins-Rich Fraction of Argania spinosa Leaves Extract.

Plant saponins are abundant and diverse natural products with a great potential for use in drug-discovery research. Here, we evaluated extracts of saponins-rich fractions of argan leaves and argan oil extraction byproducts (shell, pulp, press cake) for their effect on melanogenesis. Results show that from among the samples tested, only the saponins-rich fraction from leaves (ALS) inhibited melanin production in B16 murine melanoma (B16) cells. The mechanism of the melanogenesis inhibition was elucidated by determining the protein and mRNA expression of melanogenesis-associated enzymes tyrosinase (TYR), tyrosinase-related protein 1 (TRP1), and dopachrome tautomerase (DCT), and microphthalmia-associated transcription factor (MITF), and performing DNA microarray analysis. Results showed that 10 µg/mL ALS significantly inhibited melanogenesis in B16 cells and human epidermal melanocytes by 59% and 48%, respectively, without cytotoxicity. The effect of ALS on melanogenesis can be attributed to the decrease in TYR, TRP1, and MITF expression at the protein and mRNA levels. MITF inhibition naturally led to the downregulation of the expression of Tyr and Trp1 genes. Results of the DNA microarray analysis revealed the effect on melanogenesis-associated cAMP and Wnt signaling pathways' genes. The results of this study suggest that ALS may be used in cosmeceuticals preparations for hyperpigmentation treatment.

The melanin inhibitory effect of plants and phytochemicals: A systematic review.

BACKGROUND: Melanin plays an important role in protecting human skin, while excessive synthesis of melanin can cause abnormal pigmentation and induce skin diseases. Long-term use of commercial whitening agents in managing skin melanin such as kojic acid and arbutin can lead to some negative effects such as dermatitis and liver cancer. Although past studies have researched the melanin inhibitory effect of plant extracts, the effective dose and mechanisms are not well summarized and discussed. This study aims to explore the melanin inhibitory property of phytochemicals and tries to answer the following research questions: (1) Which plant extracts and phytochemicals could inhibit melanin biosynthesis in the skin? what is the mechanism of action? (2) Have human trials been conducted to confirm their melanin inhibitory effect? (3) If not, which phytochemicals are recommended for further human trials? This article would provide information for future research to develop natural and safe skin whitening products. METHODS: A preferred reporting items for systematic reviews and meta-analyses (PRISMA) systematic review method and OHAT risk-of-bias tool were applied to screen literature from 2000 to 2021 and 50 research articles met the selection criteria. RESULTS: Flavonoids, phenolic acids, stilbenes and terpenes are main classes of phytochemicals responsible for the melanin inhibitory effects. The in vitro/in vivo melanin inhibitory effects of these plant extracts/phytochemicals are achieved via three main mechanisms: (1) the ethyl acetate extract of Oryza sativa Indica cv., and phytochemicals such as galangin and origanoside could manage melanin biosynthesis through competitive inhibition, non-competitive inhibition or mixed-type inhibition of tyrosinase; (2) phytochemicals such as ginsenoside F1, ginsenoside Rb1 and 4­hydroxy-3-methoxycinnamaldehyde could inhibit melanogenesis through down-regulating microphthalmia-related transcription factor (MITF) gene expression via different signalling pathways; (3) the ethanolic extracts of Dimorphandra gardneriana, Dimorphandra gardneriana, Lippia microphylla and Schinus terebinthifolius have a good ultraviolet absorption ability and high sun protective factor (SPF) values, thereby inhibiting UV induced melanogenesis in the skin. CONCLUSION: Although many plant extracts and phytochemicals have been found to inhibit melanin production, most of the results were only proved in cellular and/or animal models. Only the ethyl acetate extract of Oryza sativa Indica cv. panicle, and ginsenoside F1 were proved effective in human trials. Animal studies proved the effectiveness of galangin, origanoside, ginsenoside Rb1 and 4­hydroxy-3-methoxycinnamaldehyde with effective dose below 3 mM, and therefore recommended for future human trial. In addition, cellular studies have demonstrated the effectiveness of oxyresveratrol, mulberroside A, kurarinol, kuraridinol, plumbagin, (6aR,11aR)-3,8-dihydroxy-9­methoxy pterocarpan, ginsenoside Rh4, cardamonin, nobiletin, curcumin, ß-mangostin and emodin in inhibiting melanin synthesis at low concentrations of 20 µM and proved the low SPF values of Dimorphandra gardneriana, Dimorphandra gardneriana, Lippia microphylla and Schinus terebinthifolius extracts, and therefore recommended for further animal and human trials.

Vegetable-derived indole enhances the melanoma-treating efficacy of chemotherapeutics.

Food-drug interaction is an important but overlooked issue. For example, little is known concerning whether or not the chemotherapy of cancers is affected by the well-defined dietary chemicals such as 2-(indol-3-ylmethyl)-3,3'-diindolylmethane (LTr1) derived from daily consumed cruciferous vegetables. This work, inspired by the described melanogenesis reduction by certain indoles, presents that LTr1 mitigates the melanogenesis and thus potentiates the in vitro and in vivo anti-melanoma effectiveness of different chemotherapeutic agents including dacarbazine, vemurafenib, and sorafenib. In B16 melanoma cells, LTr1 was shown to inhibit the melanogenesis by acting towards the regulatory (R) subunit of protein kinase A (PRKAR1a) associated with the phosphorylation of cAMP-response element binding protein (CREB). This allows LTr1 to reduce the expression of melanogenesis-related enzymes such as tyrosinase (TYR), tyrosinase-related protein 1 (TYRP1), and tyrosinase-related protein 2 (TYRP2). Furthermore, LTr1 was addressed to bind to the aryl hydrocarbon receptor (AhR) and up-regulate the expression of CYP1A1 encoding cytochrome P450 1A1, leading to the escalation of reactive oxygen species (ROS) level. The increased ROS generation promotes the cysteine-to-cystine transformation to inhibit the pheomelanogenesis in melanomas. Collectively, the work identifies LTr1 as a new melanogenesis inhibitor that modulates the PKA/CREB/MITF and AhR/CYP1A1/ROS pathways, thereby providing a new option for (re)sensitizing melanomas to chemotherapeutics.

Inhibitory activity of soybean (Glycine max L. Merr.) Cell Culture Extract on tyrosinase activity and melanin formation in alpha-melanocyte stimulating Hormone-Induced B16-F10 melanoma cells.

BACKGROUND: Hyperpigmentation, which causes excessive melanin synthesis and accumulation, is an important issue in the cosmetic industry. Since compounds developed against hyperpigmentation often come with side effects such as skin irritation and contact dermatitis, new studies focus on the use of natural agents that have no side effects. METHODS AND RESULTS: In this study, it was found that the effects of soybean cell culture extract (SCE) on alpha-melanocyte-stimulating hormone (α-MSH) induced melanogenesis in B16-F10 murine melanoma cells. The cells were incubated with SCE for 48 h after treatment with α­MSH for 24 h to analysis the melanin content, cellular tyrosinase activity, and gene and protein expression. SCE at 1 mg/mL decreased melanin content and tyrosinase activity by 34% and 24%, respectively, compared to the α-MSH-treated group, which did not decrease cell viability. In addition, SCE (1 mg/mL) downregulated the expression of tyrosinase (TYR), tyrosinase-related protein (TRP)-1, tyrosinase-related protein (TRP)-2, and microphthalmia-associated transcription factor (MITF) genes 1.5-, 1.5-, 2-, and 2-fold, respectively. Furthermore, SCE inhibited the expression of TYR, TRP1, and TRP2 by decreasing the expression of MITF, as shown in a western blot assay. CONCLUSIONS: This study suggests that SCE reveals dose-dependent inhibition of melanin synthesis through the suppression of tyrosinase activity as well as molecular levels of TYR, TRP1, TRP2, and MITF in B16-F10 murine melanoma cells. Therefore, SCE has the potential to be an effective and natural skin-whitening agent for application in the cosmetic industry.

Anti-melanogenesis effect from Wampee fruit pectin via α-MSH/TRY pathway in A375 cells.

BACKGROUND: Polysaccharides from wampee have been reported to process various biological activities, while the relationship between structure and bioactivities has been barely addressed. Pectin, an abundant water-soluble polysaccharide in wampee, showed significant antioxidant activity, which was associated with the anti-melanogenic activity. Therefore, this study investigated the physicochemical characteristics and the anti-melanogenesis effect of pectin extracted from wampee fruit in A375 cells. METHODS: The physicochemical characterization of pectin from wampee fruit was investigated by gel chromatography (GCP), FT-IR spectroscopy, and NMR spectroscopy methods. The anti-melanogenesis effects and mechanism were evaluated by mushroom tyrosine enzyme and human melanin cell model in vitro. RESULTS: The results showed that a molecular weight of 5.271 × 105 Da wampee fruit pectin (WFP) were mainly composed of mannose (Man), ribose (Rib), rhamnose (Rha), glucuronic acid (Glc A), glucose (Glc), galacturonic acid (Gal A), galactose (Gal), and arabinose (Ara), which linked with →4)-ß-D-Galp-(1 → units. The current study revealed that WFP could significantly suppress mushroom TRY activity in vitro. Furtherly, WFP significantly reduced intracellular and extracellular melanin formation in A375 melanoma cells depending on the presence of alpha-melanocyte stimulating hormone (α-MSH). TRY activity was only inhibited in α-MSH treated A375 cells. Western blot analysis demonstrated that WFP reverse α-MSH induced melanogenesis in A375 melanoma cells, including in down-regulated TRY, TYRP-1, TYRP-2, MITF and CREB expressions. CONCLUSION: These results indicated that WFP could inhibit α-MSH induced melanogenesis in A375 melanoma cells via α-MSH/TRY pathway. In conclusion, these data provided a new perspective to annotate WFP anti-melanogenesis activity mechanism.

(+)-Magnolin Enhances Melanogenesis in Melanoma Cells and Three-Dimensional Human Skin Equivalent; Involvement of PKA and p38 MAPK Signaling Pathways.

Magnoliae Flos is a traditional herbal medicine used to treat nasal congestion associated with headache, empyema, and allergic rhinitis. In our preliminary screening of crude drugs used in Japanese Kampo formulas for melanin synthesis, the methanol extract of Magnoliae Flos was found to exhibit strong melanin synthesis activity. However, there have been no studies evaluating the effects of Magnoliae Flos or its constituents on melanogenesis. The present study aimed to isolate the active compounds from Magnoliae Flos that activate melanin synthesis in melanoma cells and three-dimensional human skin equivalent, and to investigate the molecular mechanism underlying melanin induction. The methanol extract of Magnoliae Flos induced an increase of melanin content in both B16-F1 and HMV-II cells. A comparison of melanin induction by three fractions prepared from the extract showed that the ethyl acetate fraction markedly induced melanin synthesis. Bioassay-guided separation of the ethyl acetate fraction resulted in the isolation of seven lignans (1:  - 7: ). Among them, (+)-magnolin (5: ) strongly induced melanin synthesis and intracellular tyrosinase activity. Furthermore, the ethyl acetate fraction and 5: clearly induced melanin content in a three-dimensional human skin equivalent. Molecular analysis revealed that 5: triggered the protein expression of tyrosinase, tyrosinase-related protein-1, and tyrosinase-related protein-2. Further analysis of transcriptional factors and signaling pathways demonstrated that 5: induces the protein expression of tyrosinase, tyrosinase-related protein-1, and tyrosinase-related protein-2 activated by the protein kinase A- and p38 mitogen-activated protein kinase-dependent pathways, leading to cAMP-responsive element-binding protein phosphorylation and microphthalmia-associated transcription factor expression. These findings demonstrate the potential of 5: as a potent therapeutic agent for hypopigmentation.

Inhibitory effects and underlying mechanisms of Artemisia capillaris essential oil on melanogenesis in the B16F10 cell line.

The present study investigated the anti­melanogenic activity of 10 essential oils using the B16F10 cell model. Initially, a wide range of concentrations of these essential oils were screened in order to determine their toxicity levels. The assigned non­toxic concentrations of the tested essential oils were then used to evaluate their effects on melanogenesis. The effects of the essential oils with potent anti­melanogenic activity on cell proliferation, protection against H2O2­induced cell death and the expression of certain melanogenesis­related genes, including MITF, tyrosinase, tyrosinase related protein (TRP)­1 and TRP­2 were also evaluated. The results revealed that the essential oils extracted from Citrus unshiu, Juniperus chinensis L., Zanthoxylum piperitum and Artemisia capillaris (A. capillaris) inhibited melanogenesis. However, among these four extracts, only A. capillaris extract enhanced cell proliferation, exhibited anti­H2O2 activities and decreased the expression level of TRP­1. It was demonstrated that A. capillaris extract inhibited melanin synthesis via the downregulation of the TRP­1 translational level. These essential oil extracts, particularly that of A. capillaris, may thus be used as natural anti­melanogenic agents for therapeutic purposes and in the cosmetic industry for skin whitening effects with beneficial proliferative properties. However, further studies using in vivo models are required to validate these findings and to examine the effects of these extracts on various molecular pathways.

Calycosin, a Common Dietary Isoflavonoid, Suppresses Melanogenesis through the Downregulation of PKA/CREB and p38 MAPK Signaling Pathways.

Calycosin, a bioactive isoflavonoid isolated from root extracts of Astragalus membranaceus, has been reported to inhibit melanogenesis, the mechanism of which remains undefined. In this study, we interrogated the mechanistic basis by which calycosin inhibits melanin production in two model systems, i.e., B16F10 melanoma cells and zebrafish embryos. Calycosin was effective in protecting B16F10 cells from α-melanocyte-stimulating hormone (α-MSH)-induced melanogenesis and tyrosinase activity. This anti-melanogenic effect was accompanied by decreased expression levels of microphthalmia-associated transcription factor (MITF), a key protein controlling melanin synthesis, and its target genes tyrosinase and tyrosinase-related protein-2 (TRP-2) in calycosin-treated cells. Mechanistically, we obtained the first evidence that calycosin-mediated MITF downregulation was attributable to its ability to block signaling pathways mediated by cAMP response element-binding protein (CREB) and p38 MAP kinase. The protein kinase A (PKA) inhibitor H-89 and p38 inhibitor SB203580 validated the premise that calycosin inhibits melanin synthesis and tyrosinase activity by regulating the PKA/CREB and p38 MAPK signaling pathways. Moreover, the in vivo anti-melanogenic efficacy of calycosin was manifested by its ability to suppress body pigmentation and tyrosinase activity in zebrafish embryos. Together, these data suggested the translational potential of calycosin to be developed as skin-lightening cosmeceuticals.

Green tea polyphenols inhibit malignant melanoma progression via regulating circ_MITF/miR-30e-3p/HDAC2 axis.

Green tea polyphenols (GTPs) are regarded as anticancer substances and have been revealed to play significant roles in the development of malignant melanoma. However, the mechanisms by which GTPs perform anticarcinogenic activity are not well elucidated. Cellular function assays revealed that GTPs inhibited melanoma cell proliferation, migration, invasion, epithelial-mesenchymal transition (EMT), and promoted apoptosis in vitro. Circ_MITF expression was elevated in melanoma tissues and cells but was decreased by GTPs in cells. Functional experiments indicated circ_MITF overexpression reversed the anticancer effects of GTPs on melanoma cells. Then the underlying mechanism analysis suggested that circ_MITF served as a sponge for miR-30e-3p to upregulate the level of HDAC2. MiR-30e-3p reexpression attenuated the regulatory effects of circ_MITF on GTPs-treated melanoma cells. Silencing of miR-30e-3p promoted the malignant phenotypes in GTPs-treated melanoma cells, which were reversed by HDAC2 knockdown. Preclinically, administration of GTPs suppressed the expression of downstream target genes and repressed tumorigenesis of xenografts in nude mice. In all, GTPs suppressed melanoma progression by regulating circ_MITF/miR-30e-3p/HDAC2 axis, providing a potential therapeutic strategy for human malignant melanoma intervention.

Assessment of Risk of Hereditary Predisposition in Patients With Melanoma and/or Mesothelioma and Renal Neoplasia.

Importance: In BAP1 tumor predisposition syndrome, clear cell renal cell carcinoma (RCC) is frequently associated with melanoma and/or mesothelioma, while germline MITF p.E318K alterations are being increasingly reported in melanoma/RCC. Limited data exist on the co-occurrence of melanoma and/or mesothelioma with renal neoplasia and the prevalence of associated germline alterations. Objective: To assess the frequency of melanoma and/or mesothelioma co-occurring with renal neoplasia using our institutional nephrectomy registry and to determine the prevalence of BAP1 and MITF alterations within this cohort. Design, Setting, and Participants: In this genetic association study, medical records from 8295 patients from 1970 to 2018, renal neoplasia co-occurring with melanoma and/or mesothelioma within a single institutional nephrectomy registry was reevaluated based on contemporary histopathologic criteria and the medical records were reviewed. Data were analyzed from September 2019 to May 2021. Main Outcomes and Measures: Identified cases were screened for BAP1 loss using immunohistochemistry; while patients with melanoma and clear cell RCC were screened for MITF p.E318K alterations. Tumors from patients with potential germline alterations were analyzed with comprehensive molecular profiling using a 514-gene next generation sequencing panel. Results: Of a total of 8295 patients, 93 (1.1%; 95% CI, 0.9%-1.4%) had melanoma and/or mesothelioma co-occurring with renal neoplasia (cutaneous melanoma, n = 76; uveal melanoma, n = 11; mesothelioma, n = 6). A total of 69 (74.2%) were male; 24 (25.8%) were female; median age at diagnosis of renal neoplasia was 63 years (IQR, 58-70 years) and the median duration of follow-up was 8.5 years (IQR, 5.0-14.6 years). Two patients with clear cell RCC had germline BAP1 alterations in the setting of cutaneous melanoma and mesothelioma. Two patients with hybrid oncocytic tumors had biallelic inactivation of FLCN in a setting of Birt-Hogg-Dubé (BHD) syndrome associated with uveal melanoma and mesothelioma. Tumor-only screening of clear cell RCC associated with cutaneous (n = 53) and uveal melanoma (n = 6) led to the identification of 1 patient with a likely germline MITF p.E318K alteration. After excluding benign renal neoplasia (such as oncocytoma and angiomyolipoma), alterations of BAP1, FLCN, and MITF were identified in 5 of 81 patients (6.2%) with melanoma and/or mesothelioma and renal neoplasia. In contrast to hybrid oncocytic tumors in BHD, no unique genotype-phenotype correlations were seen for clear cell RCC with pathogenic BAP1/ MITF alterations and VHL loss of function variants. Four of 5 cases (80%) met current National Comprehensive Cancer Network criteria for germline testing based on a combination of age, multifocality, histologic findings, and family history. Conclusions and Relevance: In this genetic association study, findings support the continued use of these National Comprehensive Cancer Network criteria and suggest more stringent screening may be warranted in this patient population.