Expression Of TRP-2 Research Articles

Inhibitory Efficacy of Smilax china L. on MITF, TRP-1, TRP-2, Tyrosinase Protein and mRNA Expression in Melanoma Cell (B16F10)

Inhibitory Efficacy of Smilax china L. on MITF, TRP-1, TRP-2, Tyrosinase Protein and mRNA Expression in Melanoma Cell (B16F10)

Protein Expression Analysis of Melanocyte Differentiation Antigen TRP-2

Melanocyte differentiation antigens, such as gp100, tyrosinase, and Melan-A and their corresponding antibodies HMB45, T311, and A103, are major diagnostic tools in surgical pathology. Little is known about tyrosinase-related protein 2 (TRP-2, or dopachrome tautomerase/DCT) another melanocyte differentiation antigen, which is an enzymatic component of melanogenesis. We identified a commercial reagent to TRP-2, monoclonal antibody (mAb) C-9 and undertook a comprehensive analysis to assess its specificity and usefulness for surgical pathology. Subsequently, we analyzed panels of normal tissues and tumors. We show that TRP-2 is regularly expressed in melanocytes of the normal skin. In cutaneous nevi, TRP-2 is present in junctional as well as in dermal nevocytes. In malignant tumors, C-9 reactivity is restricted to melanocytic and related lesions and present in 84% and 58% of primary and metastatic melanomas, respectively. Ten primary melanomas of the anorectal mucosa were all positive. Like the other melanocyte differentiation antigens, TRP-2 was absent in 6 desmoplastic melanomas. Also, only 2 of 9 angiomyolipomas were TRP-2 positive. We conclude that mAb C-9 is a valuable reagent for the analysis of TRP-2 expression in archival surgical pathology material. The expression pattern of TRP-2 in melanocytic and related lesions appears to parallel other melanocyte differentiation antigens, although the overall incidence is lower than other antigens, such as Melan-A or gp100.

Antimelanogenic effects of the novel melanogenic inhibitors daidzein and equol, derived from soymilk fermented with Lactobacillus plantarum strain TWK10, in B16F0 mouse melanoma cells

Melanin functions to protect the skin; however, excessive production of melanin can cause hyperpigmentation. The antimelanogenic effects of daidzein and its microbial metabolite, equol, and the mechanisms through which they inhibit melanogenesis in B16 melanoma cells were investigated. At concentrations of less than 50 and 40 µM, respectively, daidzein and equol had no cytotoxic effects on B16 melanoma cells stimulated with α-melanocyte-stimulating hormone (α-MSH). Daidzein and equol reduced melanin content in these cells by downregulating the transcription of genes encoding melanocortin 1 receptor. Additionally, these compounds interfered with the phosphorylation of p38 mitogen-activated protein kinase and induced the phosphorylation of extracellular signal-regulated kinase 1/2 and glycogen synthase kinase 3β. Moreover, daidzein and equol decreased the expression of tyrosinase, TRP-1, and TRP-2, thereby blocking melanin production in α-MSH-stimulated B16 melanoma cells. In conclusion, daidzein and equol were noncytotoxic in B16 melanoma cells and exerted antimelanogenic effects. The functions of these bioactive compounds in fermented soymilk were confirmed, and they may be useful for developing functional foods and cosmetics, extending the range of applications of crops containing them.

Pigmentation Effect of Rice Bran Extracted Minerals Comprising Soluble Silicic Acids.

Our investigation focused on identifying melanogenesis effect of soluble minerals in rice bran ash extract (RBE) which include orthosilicic acid (OSA). Melanocytes were apparently normal in terms of morphology. It was, however, shown that they were stressed a little in the RBE and OSA added media in aspect of LDH activity. Melanin synthesis and intracellular tyrosinase activity were increased by treatment of RBE which is similar to that of OSA. The Western blotting results showed that TRP-1, tyrosinase, and MITF expression levels were 2-3 times higher in the OSA and RBE groups compared to the control group which promoted melanin synthesis through CREB phosphorylation. Moreover, histology and immunohistochemistry were shown to have similar result to that of protein expression. As a result, minerals which comprise orthosilicic acid has the potential to promote melanogenesis and both RBE and OSA have similar cell viability, protein expression, and immunostaining results, suggesting that RBE comprises specific minerals which promote melanin synthesis through increasing of MITF and CREB phosphorylation. Therefore, RBE could be used as a novel therapeutic approach to combat melanin deficiency related diseases by stimulating melanocytes via its soluble Si and mineral components.

마우스 흑색종세포에서 Purpurogallin의 멜라닌 생성 억제 효과

Melanin is one of the most important factors affecting skin color. Melanogenesis is the bioprocess of melanin production by melanocytes in the skin and hair follicles and is mediated by several enzymes, such as tyrosinase, tyrosinase related protein (TRP)-1, and TRP-2. Convenient enzymatic transformation of the simple phenol pyrogallol with polyphenol oxidase originating from pear to an oxidative product, purpurogallin, was efficient. The structure of the pyrogallol oxidation product was identified on the basis of spectroscopic methods. The biotransformation product purpurogallin showed significant inhibitory effects against both melanin synthesis and tyrosinase activity in a dose-dependent manner in B16 melanoma cells. In addition, purpurogallin significantly attenuated melanin production by inhibiting TRP-1, and TRP-2 expression through modulation of their corresponding transcription factors, and microphthalamia- associated transcription factor in B16 cells. Consequently, purpurogallin derived from convenient enzymatic transformation of pyrogallol might be a beneficial material for reducing skin hyperpigmentation.

은행 열매 추출물의 멜라닌 생성 저해효과

In this study, we investigated the inhibitory effect on melanin synthesis of Ginkgo biloba seed oil. The results showed 9.96% inhibitory effect scavenging activity on DPPH and showed a value of 1.33 mM of FeSO4 at a concentration of 0.06% in DMSO by using FRAP assay. G. biloba seed oil inhibited tyrosinase activity up tp 37.72% and suppressed the biosynthesis melanin up to 48.02% at 0.06% in B16/F10 mouse melanoma cell. In G. biloba seed oil treated group tyrosinase, TRP-1, TRP-2 and MITF gen expression levels significantly decreased compared to the contral group at a concentration of 0.04% and 0.06%. In conclusion, these results indicated that G. biloba seed oil extract have a good antimelanogenetic effects.

Inhibitory Effect of Dried Pomegranate Concentration Powder on Melanogenesis in B16F10 Melanoma Cells; Involvement of p38 and PKA Signaling Pathways.

Plants rich in antioxidant substances may be useful for preventing skin aging. Pomegranates, containing flavonoids and other polyphenolic compounds, are widely consumed due to their beneficial properties. We examined the underlying mechanisms of dried pomegranate concentrate powder (PCP) on melanin synthesis in B16F10 melanoma cells. The antioxidant effects of PCP were determined by measuring free radical scavenging capacity and transcript levels of antioxidant enzymes. To explore the inhibitory effects of PCP on melanin synthesis, we measured tyrosinase activity and melanin content in α-melanocyte stimulating hormone (α-MSH)-stimulated B16F10 cells. In addition, the levels of tyrosinase-related protein-1 (TRP-1), TRP-2, tyrosinase, and microphthalmia-associated transcription factor (MITF) expression were determined by Western blotting. Changes in the phosphorylation status of protein kinase A (PKA), cAMP response element-binding protein (CREB), mitogen-activated protein kinases (MAPKs), phosphatidylinositol 3-kinase (PI3K), serine/threonine kinase Akt, and glycogen kinase 3β (GSK3β) were also examined. The free radical scavenging activity of PCP increased in a dose-dependent manner. In PCP-treated B16F10 cells, transcript levels of glutathione peroxidase-1 (GPx-1) were increased compared with α-MSH-stimulated cells. In addition, PCP led to the down-regulation of phospho-p38, phospho-PKA, phospho-CREB, phospho-GSK3β, MITF, and TRP-1 compared with α-MSH-stimulated B16F10 cells. We believe this effect may be associated with PCP activity, which leads to the inhibition of melanin production and tyrosinase activity. These results suggest that PCP decreases tyrosinase activity and melanin production via inactivation of the p38 and PKA signaling pathways, and subsequently decreases phosphorylation of CREB, MITF, and melanogenic enzymes. These observations provided new insights on the molecular mechanisms of the skin-whitening property of PCP.

Dioscin Derived from Solanum melongena L. "Usukawamarunasu" Attenuates α-MSH-Induced Melanogenesis in B16 Murine Melanoma Cells via Downregulation of Phospho-CREB and MITF.

This study aimed to chemically isolate and explore an antimelanogenesis inducer in extracts of Solanum melongena L. "Usukawamarunasu" eggplant. We successfully identified dioscin ([25R]-Spirost-5-en-3β-yl) 2-O-(6-deoxy-α-L-mannopyranosyl) - 4-O-(6-deoxy-α-L-mannopyranosyl)-β-D-glucopyranoside] in the plant, and examined the effects of α-melanocyte-stimulating hormone (MSH)-induced melanogenesis in B16 murine melanoma cells by this plant-derived dioscin. Immunoblot analysis suggested that dioscin reduced the expression of tyrosinase, tyrosinase-related protein-1 (TRP-1), and TRP-2, resulting in inhibition of intracellular production of melanin. In addition, dioscin caused reduction of phosphorylated cAMP-responsive element binding protein 1 transcription factors (CREB), which led to a reduction of microphthalmia-related transcription factor (MITF) in α-MSH-stimulated cells, but did not affect phosphorylation of extracellular signal-regulated kinase. Furthermore, dioscin significantly downregulated the expression of tyrosinase, TRP-1, and TRP-2, which led to the reduction of α-MSH-induced melanogenesis in B16 cells. These results suggest that dioscin may decrease the level of MITF via inhibition of phosphorylation of CREB in α-MSH-induced melanogenesis in B16 cells.

Proton pump inhibitors decrease melanogenesis in melanocytes.

Proton pump inhibitors (PPIs) are widely used as inhibitors of gastric juice secretion for treatment of gastroesophageal reflux disease. However, there are no previous studies of the effects on melanogenesis resulting from PPI treatments. Therefore, the aim of the present study was to investigate the effects of PPIs on melanogenesis in melan-a cells derived from immortalized mouse melanocytes. Tyrosinase activity and copper-chelating activity were measured spectrophotometrically. In addition, the melanin content and viability of melan-a cells treated with PPIs were assessed and the mRNA levels of melanogenesis-associated genes were measured by reverse transcription-polymerase chain reaction. Treatment with rabeprazole, but not the other PPIs tested, resulted in strong, dose-dependent inhibition of mushroom tyrosinase (TYR). By contrast, each of the PPIs tested exhibited copper-chelating activity. Treatment of melan-a cells with 100 µM concentrations of the PPIs resulted in significantly reduced melanin synthesis and reduced expression of several melanogenesis-associated genes, including TYR, TYR-related protein-1 (TRP-1) and TRP-2, and microphthalmia-associated transcription factor, but did not result in cytotoxic effects. These results suggest that PPIs inhibit melanin biosynthesis in melan-a cells via the downregulation of melanogenesis-associated genes. Furthermore, the findings indicate that PPIs in general could be utilized as skin-whitening agents and/or as biomaterial for treating hyperpigmentation disorders.

Inhibition of melanogenesis by β-caryophyllene from lime mint essential oil in mouse B16 melanoma cells.

Volatile essential oils of mint species are used for cosmetics and in skin care products. In this study, we evaluated the main chemical components of the lime mint and the anti-melanogenic properties of its main components. The essential oil was analysed by gas chromatography-mass spectrometry (GC/MS). The anti-melanogenic effects of mint essential oil and β-caryophyllene were investigated in B16F10 murine melanoma cells. The main components of lime mint essential oil were found to be D-limonene (41.10%), D-carvone (8.58%), δ-selinene (6.73%) and β-caryophyllene (6.24%). The lime mint essential oil reduced melanin production in a dose-dependent manner in murine B16F10 cells. β-Caryophyllene, one of the main compounds in lime mint essential oil, could reduce melanogenesis by down-regulating the expression of MITF, TRP-1, TRP-2 and tyrosinase, resulting in a decrease in melanin content decrease. These results reveal that lime mint essential oil and β-caryophyllene are considered to be valuable as potential skin-whitening agents.

Effect of quercetin derivatives on melanogenesis stimulation of melanoma cells

Controlling melanogenesis is important for maintaining the good health and cosmetic appearance of a human body. This study aims to search the active compounds from natural products exhibiting the melanogenesis modulating activity and elucidate the mechanism underlying the observed activity. Two novel quercetin glycosides namely 4′-O-β-d-glucopyranosyl-quercetin-3-O-β-d-glucopyranosyl-(1 → 4)-β-d-glucopyranoside 1 and 4′-O-β-d–glucopyranosyl-(1 → 2)-β-d-glucopyranosyl-quercetin-3-O-β-d-glucopyranosyl-(1 → 4)-β-d-glucopyranoside 2 were isolated and identified from Helminthostachys zeylanica roots 50 % ethanol extract. Compound 1 exhibited intracellular melanogenesis stimulatory activity, while 2 showed no effect even the structural similarity. To understand the structure–activity relationships, twelve quercetin glycosides and seven methylquercetins were synthesized from rutin as starting material. As the result of bioassay using synthesized nineteen quercetin derivatives in B16 melanoma cells, some quercetin-3-O-β-d-glucopyranosides stimulated the intracellular melanogenesis. On the other hand, synthesized 3-O-methylquercetin 12 and 3,4′,7-O-trimethylquercetin 15 increased both intra and extracellular melanin contents with no cytotoxicity. Compoud 15 increased the phosphorylated p38 mitogen-activated protein kinase (MAPK) and microphthalmia-associated transcription factor (MITF) which regulates the expression of tyrosinase, TRP-1 and TRP-2. While 12 enhanced the expression of the melanogenic enzymes without involving the MITF, as evidenced by its lack of any stimulation of the expression of MITF and p-p38 MAPK. This result indicates that 12 may stimulate the expression of tyrosinase, TRP-1, and TRP-2 by stimulating currently unidentified transcriptional factors and/or by regulating the degradation of melanogenic enzymes.

ADAM protease inhibitors reduce melanogenesis by regulating PMEL17 processing in human melanocytes

BackgroundADAMs (a disintegrin and metalloprotease) are a family of proteases involved in ectodomain shedding that play a role in various biological processes such as cell adhesion and migration. ADAM10 and ADAM17 are suggested to be involved in pigmentary disorders. ObjectiveWe examined the effect of ADAM protease inhibitors on the modulation of melanogenesis in normal human epidermal melanocytes (NHEM). MethodsNHEMs and B16F10 treated with ADAM protease inhibitors were analyzed. AlamarBlue cell proliferation assay, melanin content assay, tyrosinase activity assay, Western blotting analysis, electron microscopic analysis, and RNA interference were employed. ResultsIn NHEMs, melanin content was reduced by treatment with ADAM protease inhibitors. The inhibitors did not change the protein expression of tyrosinase, TRP-1, and MITF. In B16F10 cells, treatment of the cells with ADAM protease inhibitor diminished the α-MSH-induced increase in melanin content. Electron microscopy showed that the number of fibrillar and mature melanosomes was significantly reduced and that the vacuolar compartments were filled with dense unstructured aggregates after treatment with ADAM protease inhibitors. We therefore focused on the processing of PMEL17, a melanosomal glycoprotein that forms a fibrillar matrix on which melanin gets deposited. Proteolytic processing of PMEL17 is required to form functional fibrils during melanogenesis. Recently, γ-secretase and β-site amyloid precursor protein-cleaving enzyme 2 (BACE2) were found to cleave PMEL17. We found that ADAM protease inhibitors exerted effects on the processing of C-terminal and N-terminal fragments of PMEL17. Using BACE2 siRNA and γ-secretase inhibitor, we showed that ADAM protease inhibitor affected PMEL17 processing in a γ-secretase and BACE2-independent mechanism. ConclusionSeveral proteases, including ADAM proteases, can contribute to the formation of fibrils and their assembly into sheets in melanosomes.

Effect of Dendrobium moniliforme on Melanogenic Protein Expression in B16F10 Melanoma Cells

The Dendrobium moniliforme extract (DME) was successively partitioned using n-hexane, CH2Cl2, EtOAc, BuOH, and water. The results indicate that the Dendrobium moniliforme fraction extracted using CH2Cl2 (DMC) was an effective inhibitor of melanogenesis in murine melanoma cells (B16F10). To elucidate the mechanism of the effect of DME on melanogenesis, we performed western blotting of the melanogenic proteins. DME inhibited tyrosinase and, tyrosinase-related protein-1 (TRP-1) and TRP-2 expressions. Futher, we confirmed that DME decreased the protein level of melanocyte-specific isoform of microphthalmia-associated transcription factor (MITF) proteins, which decreased tyrosinase and related genes in B16F10 melanoma cells. On the basis of the results, we suggest that D. moniliforme is effective against hyperpigmentation disorders and that it be considered a possible anti-melanogenic agent in topical application.

Down-regulation of MITF, TRP-1, TRP-2, and tyrosinase expressions by compounds isolated from Pruni persicae Flos in murine B16F10 melanoma

Three single compounds were isolated by the fractionation from Pruni persicae Flos, and its chemical structure was analyzed using 1H-, 13C-nuclear magnetic resonance, fast atom bombardment -Mass, and fourier transform infrared spectrometer analyses. The compounds were determined as quercetin 3-O-rhamnoside, kaempferol 3-O-β-neohesperidoside, and kaempferol 3-O-β-d-glucoside. To prove the whitening effect of these compounds, B16F10 melanoma was treated with quercetin-3-O-rhamnoside, kamferol-3-O-β-neohesperidoside or kaemferol-3-O-β-d-glucoside to confirm cell cytotoxicity using 3-[4,5-dimethylthiazol-2yl]-2,5-diphenyl-tetrazolium-bromide assay. As a result, cell cytotoxicity was examined at more than 20% in 20 μg/mL of compounds isolated from Pruni persicae Flos. Tyrosinase activity and melanin production were examined at 10 μg/mL of each compounds. Results showed quercetin-3-O-rhamnoside, kamferol-3-O-β-neohesperidoside or kaemferol-3-O-β-d-glucoside reduced tyrosinase activity in a dose-dependent manner. Moreover, quercetin-3-O-rhamnoside showed reduction of tyrosinase activity by 37.6% at 10 μg/mL of quercetin-3-O-rhamnoside as well as the reduction of melanin contents by 58.8%. In addition, quercetin-3-O-rhamnoside, kamferol-3-O-β-neohesperidoside, and kaemferol-3-O-β-d-glucoside reduced tyrosinase, tyrosinase-related proteins 1, and 2, microphthalmia associated transcription factor expressions, which were related to whitening effect, as well as both of mRNA and protein level of cyclic adenosine monophosphate as upstream signals. More interestingly, quercetin-3-O-rhamnoside strongly reduced both mRNA and protein expressions than other compounds. Therefore, we determined the whitening effect from isolated compound fractionated from Pruni persicae Flos. Moreover, strong whitening activity of quercetin-3-O-rhamnoside was examined, which suggested the potential application of this compound in functional cosmetic development.

Anti-melanogenic properties of honeybee (Apis mellifera L.) venom in α-MSH-stimulated B16F1 cells

Since ancient times, honeybee (Apis mellifera L.) venom (BV) has been applied to the skin as an anti-ageing remedy. To further access BV as an effective whitening agent for cosmetics and potential external treatment for topical use, we investigated its ability to inhibit tyrosinase activity and subsequent melanin biosynthesis. B16F1 melanoma cells were treated with 10 nM α-melanocyte stimulating hormone (α-MSH) and then with various doses of BV. BV inhibited direct tyrosinase activity and cellular tyrosinase activity, which decreased melanin synthesis in α-MSH-stimulated B16F1 melanoma cells. In addition, we used reverse transcription-polymerase chain reaction and Western blotting for melanogenesis-related genes such as tyrosinase-related protein (TRP)-1 and TRP-2 to examine the mechanisms underlying the inhibitory effects of BV on melanogensis. BV decreased the mRNA and protein expression of TRP-1 and TRP-2. These findings suggest that BV induced the downregulation of melanogenesis by inhibiting TRP-1 and TRP-2 activation.

Effect of cultured medium of human umbilical cord blood-derived mesenchymal stem cells on melanogenic enzyme activity in mouse B16 melanoma cells

The cultured medium of mesenchymal stem cell is a valuable resource for a variety of stem cell-derived cytokines and growth factors with a potential therapeutic effect on skin diseases. Here, we investigated the effect of the cultured medium of human umbilical cord blood-derived mesenchymal stem cell (CM-hCBSC) on melanogenesis in mouse B16 melanoma cells stimulated by α-melanocyte stimulating hormone (α-MSH). Our results show that CM-hCBSC inhibits melanin synthesis due to the decrease in cellular tyrosinase activity in B16 melanoma cells without cytotoxicity. The activity of melanogenic enzymes such as tyrosinase, tyrosinase-related protein-1 (TRP-1), and TRP-2 were evaluated by western blot and semi-quantitative RT-PCR. The CM-hCBSC down-regulated the protein and mRNA expression levels of tyrosinase and TRP-1 in a dose-dependent manner. In addition, TRP-2 protein and mRNA expression levels were found to be down-regulated to a lesser extent. We also measured the level of cytokines in CM-hCBSC using the Luminex total system. It was found that the top enriched cytokine in the CMhCBSC was interukine-6 (IL-6), known as a potent regulator of melanin synthesis together with IL-1α and tumor necrosis factor α (TNF- α). These results indicate that secretary factors including IL-6 in the CM-hCBSC may inhibit melanin synthesis by down-regulating the expression of tyrosinase and TRP1. This study suggests that CMhCBSC has a therapeutic effect as an anti-melanogenic agent and may be effective against hyperpigmentation disorders.

Inhibitory effects of Taraxacum mongolicum with phreatic water on melanin synthesis

BackgroundRecently, people have begun showing heightened interest in skin whitening. Melanin is an important factor that determines skin color. The purpose of this study is to investigate the inhibitory effect of Taraxacum mongolicum (TAM) with phreatic water (PW) from Dogo Hot Springs on melanin synthesis. MethodsWe assessed the inhibitory effects of TAM on melanin synthesis in B16F10 mouse melanoma cells. The mRNA levels of tyrosinase related protein (TRP)-1, TRP-2, tyrosinase, MITF, ERK, and PKA protein were analyzed with reverse transcription polymerase chain reaction and Western blot analysis. We also assessed the inhibitory effects of TAM with PW on melanin synthesis in HRM-2 melanin-possessing hairless mice. After UVB irradiation, differences in melanin were analyzed with an image analysis software between the left dorsal skin (untreated part) and the right dorsal skin (treated part). The mRNA levels of TRP-1, TRP-2, and matrix metalloproteinase (MMP)-9 were analyzed with real-time quantitative polymerase chain reaction. The dorsal skins were analyzed with histological test by hematoxylin and eosin staining. ResultsTAM inhibited the TRP-1, TRP-2, tyrosinase, MITF mRNA gene expression, and PKA protein expression on the concentration-dependent B16F10 cell. Moreover, TAM increased the ERK mRNA gene expression in the B16F10 cell. After UVB irradiation, TAM with PW increased the differences in melanin between the left dorsal skin (untreated part) and the right dorsal skin (treated part) in HRM-2 mice. TAM with PW inhibited the TRP-1, TRP-2, and MMP-9 mRNA gene expression in HRM-2 mice. TAM with PW decreased the epidermal thickness, around the cell deformation, keratinization, and infiltration in HRM-2 mice. ConclusionThese results indicate that TAM with PW has the inhibitory effect of decreasing the melanin synthesis.

S-(-)-10,11-dihydroxyfarnesoic acid methyl ester inhibits melanin synthesis in murine melanocyte cells.

The development of antimelanogenic agents is important for the prevention of serious aesthetic problems such as melasmas, freckles, age spots, and chloasmas. In the course of screening for melanin synthesis inhibitors, we found that the culture broth from an insect morphopathogenic fungus, Beauveria bassiana CS1029, exhibits potent antimelanogenic activity. We isolated and purified an active metabolite and identified it as S-(−)-10,11-dihydroxyfarnesoic acid methyl ester (dhFAME), an insect juvenile hormone. To address whether dhFAME inhibits melanin synthesis, we first measured the size of the melanin biosynthesis inhibition zone caused by dhFAME. dhFAME also showed inhibitory activity against mushroom tyrosinase in Melan-a cells. Intracellular, dose-dependent tyrosinase inhibition activity was also confirmed by zymography. In addition, we showed that dhFAME strongly inhibits melanin synthesis in Melan-a cells. Furthermore, we compared levels of TYR, TRP-1, TRP-2, MITF, and MC1R mRNA expression by reverse-transcription polymerase chain reaction and showed that treatment of Melan-a cells with 35 μM dhFAME led to an 11-fold decrease in TYR expression, a 6-fold decrease in TRP-2 expression, and a 5-fold decrease in MITF expression. Together, these results indicate that dhFAME is a potent inhibitor of melanin synthesis that can potentially be used for cosmetic biomaterial(s).

Kaliziri extract upregulates tyrosinase, TRP-1, TRP-2 and MITF expression in murine B16 melanoma cells.

BackgroundKaliziri extract (KZE) is a traditional Uyghur medicine (TUM), used by traditional hospitals in China as an injection for treatment of vitiligo for more than 30 years. Clinical application has shown that this medicine has obvious therapeutic effects. However, its phytochemical analysis and mechanism have not been examined.MethodsKZE was extracted from seeds of Kaliziri [Vernonia anthelmintica (L.) Willd.] in ethanol-water (80:20, v/v), its components were identified by LC-MS/MS, and the signaling pathway of melanin synthesis in KZE treated murine B16 melanoma cells was examined by western blotting.ResultsLiquid chromatography-mass spectrometry analysis confirmed that the main components of KZE are flavonoids. KZE increased the tyrosinase activity and melanin content in a dose-dependent manner at concentrations of 5-40 μg/ml, and treatment with 20 μg/ml of KZE enhanced the expression of tyrosinase in B16 cells in a time-dependent manner.ConclusionsKZE induced melanogenesis by increasing the expression of TYR, TRP-1, TRP-2 and MITF in B16 cells.

Limonoids from Azadirachta indica var. siamensis Extracts and Their Cytotoxic and Melanogenesis‐Inhibitory Activities

Six new limonoids, 7-benzoyl-17-epinimbocinol (5), 3-acetyl-7-tigloylnimbidinin (8), 1-isovaleroyl-1-detigloylsalanninolide (15), 2,3-dihydro-3α-methoxynimbolide (16), deacetyl-20,21-epoxy-20,22-dihydro-21-deoxyisonimbinolide (26), and deacetyl-20,21,22,23-tetrahydro-20,22-dihydroxy-21,23-dimethoxynimbin (27), along with 28 known limonoids, 1-4, 6, 7, 9-14, 17-25, and 28-34, and two known flavonoids, 35 and 36, have been isolated from the extracts of bark, leaves, roots, and seeds of Azadirachta indica A. Juss. var. siamensis Valeton (Siamese neem tree; Meliaceae). The structures of the new compounds were elucidated on the basis of extensive spectroscopic analysis and comparison with literature data. All of these compounds were evaluated for their cytotoxic activities against leukemia (HL60), lung (A549), stomach (AZ521), and breast (SK-BR-3) cancer cell lines. Eleven compounds, 1, 2, 4-7, 13, 16, 17, 29, and 30, exhibited potent cytotoxicities against one or more cell lines with IC50 values in the range of 0.1-9.3 μM. Compound 16 induced apoptotic cell death in AZ521 cells upon evaluation of the apoptosis-inducing activity by flow cytometric analysis. Western blot analysis on AZ521 cells revealed that compound 16 activated caspases-3, -8, and -9, while increasing the ratio of Bax/Bcl-2. This suggested that 16 induced apoptosis via both mitochondrial and death receptor pathways in AZ521. In addition, upon evaluation of all compounds against the melanogenesis in B16 melanoma cells induced with α-melanocyte-stimulating hormone (α-MSH), 20 limonoids, i.e., 1-3, 6, 9-11, 18, 19, 21-29, 32, and 34, and two flavonoids, 35 and 36, exhibited melanogenesis-inhibitory activities, with no, or almost no, toxicities to the cells at lower and/or higher concentrations, which were more potent than the reference arbutin, a known melanogenesis inhibitor. Western blot analysis showed that nimbin (18) reduced the protein levels of microphtalmia-associated transcription factor (MITF), tyrosinase, tyrosine-related protein 1 (TRP-1), and TRP-2 mostly in a concentration-dependent manner, indicating that 18 inhibits melanogenesis on a α-MSH-stimulated B16 melanoma cells by, at least in part, inhibiting the expression of MITF, followed by decreasing the expression of tyrosinase, TRP-1, and TRP-2.