Stimulated B16 Cells Research Articles

5G Electromagnetic Radiation Attenuates Skin Melanogenesis In Vitro by Suppressing ROS Generation.

Recently, the impacts of 5G electromagnetic radiation (EMR) with 28 GHz on human health have been attracting public attention with the advent of 5G wireless communication. Here, we report that 5G (28 GHz) EMR can attenuate the skin pigmentation in murine melanoma cells (B16F10) and a 3D pigmented human epidermis model (Melanoderm™). B16 cells were exposed to 5G (28 GHz) with or without α-MSH for 4 h per day. Interestingly, 5G attenuated α-MSH-induced melanin synthesis. Fontana–Masson staining confirmed that the dendritic formation of α-MSH stimulated B16 cells was diminished by 5G exposure. To confirm the anti-melanogenic effect of 5G EMR, MelanoDerm™ was irradiated with 5G at a power intensity of 10 W/m2 for 4 h a day for 16 days and melanin distribution was detected with Fontana–Masson staining, which supported the anti-melanogenic effect of 5G EMR. Consistently, 5G EMR suppressed α-MSH induced upregulation of melanogenic enzymes; tyrosinase, TRP-1, and TRP-2. Of note, 5G EMR attenuated ROS production stimulated by α-MSH and H2O2, suggesting that 5G EMR may dissipate ROS generation, which is pivotal for the melanin synthesis. Collectively, we demonstrated that 5G EMR can attenuate skin pigmentation by attenuating ROS generation.

Glucose Exerts an Anti-Melanogenic Effect by Indirect Inactivation of Tyrosinase in Melanocytes and a Human Skin Equivalent.

Sugars are ubiquitous in organisms and well-known cosmetic ingredients for moisturizing skin with minimal side-effects. Glucose, a simple sugar used as an energy source by living cells, is often used in skin care products. Several reports have demonstrated that sugar and sugar-related compounds have anti-melanogenic effects on melanocytes. However, the underlying molecular mechanism by which glucose inhibits melanin synthesis is unknown, even though glucose is used as a whitening as well as moisturizing ingredient in cosmetics. Herein, we found that glucose significantly reduced the melanin content of α-melanocyte-stimulating hormone (MSH)-stimulated B16 cells and darkly pigmented normal human melanocytes with no signs of cytotoxicity. Furthermore, topical treatment of glucose clearly demonstrated its whitening efficacy through photography, Fontana-Masson (F&M) staining, and multi-photon microscopy in a pigmented 3D human skin model, MelanoDerm. However, glucose did not alter the gene expression or protein levels of major melanogenic proteins in melanocytes. While glucose potently decreased intracellular tyrosinase activity in melanocytes, it did not reduce mushroom tyrosinase activity in a cell-free experimental system. However, glucose was metabolized into lactic acid, which can powerfully suppress tyrosinase activity. Thus, we concluded that glucose indirectly inhibits tyrosinase activity through conversion into lactic acid, explaining its anti-melanogenic effects in melanocytes.

Mannosylerythritol lipids inhibit melanogenesis via suppressing ERK-CREB-MiTF-tyrosinase signalling in normal human melanocytes and a three-dimensional human skin equivalent.

Hyperpigmentation is caused by excessive production of melanin in melanocytes. Mannosylerythritol lipids (MELs) are glycolipid biosurfactants that are abundantly produced by yeasts and used commercially in cosmetics. However, the potential depigmenting efficacy of MELs has not been evaluated. In this study, the depigmentary effect of MELs was tested in primary normal human melanocytes (NHMs), α-melanocyte-stimulating hormone (MSH)-stimulated B16 cells (murine melanoma cells) and a human skin equivalent (MelanoDerm) using photography, Fontana-Masson (F&M) staining and two-photon microscopy. Mannosylerythritol lipids significantly decreased the melanin contents in NHMs and α-MSH-stimulated B16 cells. Consistent with these findings, MELs treatment had a clear whitening effect in a human skin equivalent, brightening the tissue colour and reducing the melanin content. The molecular mechanism underlying the anti-melanogenic effect of MELs treatment was examined by real-time PCR and Western blotting. Mechanistically, MELs clearly suppressed the gene expression levels of representative melanogenic enzymes, including tyrosinase, Tyrp-1 and Tyrp-2, by inhibiting the ERK/CREB/MiTF signalling pathway in NHMs. This work demonstrates for the first time that MELs exert whitening effects on human melanocytes and skin equivalent. Thus, we suggest that MELs could be developed as a potent anti-melanogenic agent for effective whitening, beyond their use as a biosurfactant in cosmetics.

Chemical composition, antioxidant, anti-melanogenic and anti-inflammatory activities of Glechoma hederacea (Lamiaceae) essential oil

Essential oils show important biological activities, which account for the development of aromatherapy used in complementary and alternative medicine. The aim of this study was to evaluate the chemical composition and the antioxidant, anti-melanogenic, and anti-inflammatory properties of Glechoma hederacea essential oil (GH-EO). The essential oil composition was analyzed by gas chromatography and mass spectrometry. The most abundant compound in GH-EO was trans-3-pinanone (41.4%), and other major compounds were 4,5,6,7-Tetrahydro-5-isopropenyl-3,6-beta-dimethyl-6-alpha-vinylbenzofuran (10.8%), β-caryophyllene (10.2%), and spathulenol (4.3%). Our results indicated that GH-EO shows strong lipid peroxidation inhibitory activity for the moderation of β-carotene bleaching. GH-EO can markedly decrease melanin production and tyrosinase activity in α-melanin-stimulating-hormone (α-MSH)-stimulated B16 cells. The results also demonstrated that the activity of GH-EO on melanogenesis might be the result of its potent antioxidative ability (P < 0.05), which was reflected in the decreased cellular oxidant and malondialdehyde (MDA) levels and in the recovered activities of glutathione peroxidase (GPX) and superoxide dismutase (SOD) in α-MSH stimulated B16 cells. GH-EO may suppress the inflammatory responses of LPS-stimulated RAW 264.7 macrophages, including nitric oxide (NO) production, by regulating the expression of the inflammation-related enzymes inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and heme oxygenase-1 (HO-1), and the inflammatory cytokine tumor necrosis factor-α (TNF-α), which decreases LPS-induced MDA levels and DNA damage.

Melaleuca quinquenervia essential oil inhibits α-melanocyte-stimulating hormone-induced melanin production and oxidative stress in B16 melanoma cells

BackgroundEssential oils are odorous, volatile products of plant secondary metabolism, which are found in many leaves and stems. They show important biological activities, which account for the development of aromatherapy used in complementary and alternative medicine. The essential oil extracted from Melaleuca quinquenervia (Cav.) S.T. Blake (paperbark) (MQ-EO) has various functional properties. PurposeThe aim of this study is to investigate the chemical composition of MQ-EO by using gas chromatography-mass spectrometry (GC-MS) and evaluate its tyrosinase inhibitory activity. MethodsGas chromatography-mass spectrometry (GC-MS)-based metabolomics was used to identify 18 components in MQ-EO. The main components identified were 1,8-cineole (21.60%), α-pinene (15.93%), viridiflorol (14.55%), and α-terpineol (13.73%). B16 melanoma cells were treated with α-melanocyte-stimulating hormone (α-MSH) in the presence of various concentrations of MQ-EO or its major compounds. Cell viability was accessed by MTT assay and cellular tyrosinase activity and melanin content were determined by using spectrophotographic methods. The antioxidant mechanism of MQ-EO in α-MSH stimulated B16 cells was also investigated. ResultsIn α-melanocyte-stimulating hormone (α-MSH)-stimulated murine B16 melanoma cells, MQ-EO, 1,8-cineole, α-pinene, and α-terpineol significantly reduced melanin content and tyrosinase activity. Moreover, MQ-EO, 1,8-cineole, α-pinene, and α-terpineol decreased malondialdehyde (MDA) levels. In addition, restored glutathione (GSH) levels, glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase activities were increased in α-MSH-stimulated B16 cells. MQ-EO not only decreased apoptosis but also reduced DNA damage in α-MSH stimulated B16 cells. These results showed that MQ-EO and its main components, 1,8-cineole, α-pinene, and α-terpineol, possessed potent anti-tyrosinase and anti-melanogenic activities besides the antioxidant properties. ConclusionsThe active functional components of MQ-EO were found to be 1,8-cineole, α-pinene, and α-terpineol. Consequently, the results of present study suggest that MQ-EO is non-cytotoxic and can be used as a skin-whitening agent, both medically and cosmetically.

The melanogenesis-inhibitory effect and the percutaneous formulation of ginsenoside Rb1.

Ginsenoside Rb1 (Rb1) is the most predominant ginsenoside isolated from the roots of ginseng (Panax ginseng C. A. Meyer). This compound is active in various human biological pathways that are involved in human collagen synthesis and inhibition of cell apoptosis. In this study, the skin-whitening effects of Rb1 were investigated in B16 melanoma cells. Our results showed that Rb1 inhibited melanogenesis in α-melanocyte-stimulating hormone (α-MSH)-stimulated B16 cells in a dose-dependent manner, which collectively indicated that Rb1 may have skin-whitening effects and may be formulated into skin-whitening products for skin care. Accordingly, a ginsenoside collagen transdermal patch was developed as a vehicle to topically deliver Rb1 into pig skin. The percutaneous permeation, retention within skin, and release in vitro of Rb1 from seven transdermal patch formulas were studied. It was determined that the best formula for ginsenoside collagen transdermal patch is made of protein collagen hydrolysate powder (PCHP) 2.0% (w/w), methyl cellulose (MC) 0.5% (w/w), polyethyleneglycol 6000 (PEG6000) 0.5% (w/w), ginsenoside 0.036% (w/w), azone 0.4% (v/w), menthol 0.20% (w/w), and water.

Effect of Vetiveria zizanioides essential oil on melanogenesis in melanoma cells: downregulation of tyrosinase expression and suppression of oxidative stress.

The major objective of this study was to estimate the hypopigmentation function of the essential oil from Vetiveria zizanioides (VZ-EO). Our results indicated that VZ-EO exhibits potent lipid peroxidation inhibitory activity to moderate the bleaching of β-carotene and to maintain the cellular glutathione (GSH) levels. VZ-EO can markedly decrease melanin production and tyrosinase activity in α-melanin-stimulating-hormone- (α-MSH-) stimulated B16 cells. The effect of VZ-EO on melanogenesis is achieved by the suppression of cellular tyrosinase expression. The results demonstrated that the activity of VZ-EO on melanogenesis might be the result of its potent antioxidative ability, which was reflected in the decreased cellular oxidant and malondialdehyde (MDA) levels and the recovered activities of superoxide dismutase (SOD), glutathione peroxidase (GPX), and catalase (CAT) in α-MSH-stimulated B16 cells. The most abundant compound in VZ-EO is cedr-8-en-13-ol (12.4%), which has a strong capability to inhibit lipid peroxidation. Therefore, VZ-EO has the potential to become an ingredient in future hypopigmentation drugs, foods, and cosmetics.

Effect of epigallocatechin-3-gallate on epitheliai-mesenchymal transition in melanoma via the trans-forming growth factor-151/signal transducer and activators of transcription 3 pathway

Objective To investigate the effects of epigallocatechin-3-gallate （EGCG） on epithe- lial-mesenchymal transition （EMT） through transforming growth factor （TGF）-l/signal transducer and activators of transcription 3 （ STAT3 ） signal pathway in malignant melanoma. Methods The cell morphol- ogy, migration and invasion ability of TGFq31-stimulated B16 cells were studied under a phase-contrast mi- croscope 24 h after co-incubation with different concentrations （5, 25, and 50 rag/L） of EGCG. The mi- gration ability of the cells was detected by Scrape-wound migration assay, and the invasion ability was stud- ied by Transwell. Western blotting was employed to study the expression of p-stat3 and E-cadherin. The animal model of B16 tumour was established, and E-cadhein mRNA expression was detected by using re- verse transcription-polymerase chain reaction （RT-PCR）. Results （ 1 ） TGF-I （ 5 pg/L）-stimulated B16 cells were long-spindle shaped with loose cell-cell connection. Twenty-four h later after co-incubation with different concentrations of EGCG, B16 cells became more connected, with the degree of change pro- portional to EGCG concentrations. There were increased distance in Scrape-wound migration assay, and Tr- answell number was 97. 00 ＋9. 97, 60. 30 ±4. 00 and 38.75 ±7. 18 respectively. The invasion ability was negatively correlated to the EGCG concentration （ r = - 0. 92, P 〈 0. 05 ） ; （ 2 ） The effects of reduced p- STAT3 expression and increased E-cadherin expression under the effects of different concentrations of EGCG were concentration-dependent. The expression levels of p-STATS and E-Cadherin were also nega- tively correlated （ r = - 0. 805, P 〈 0. 05 ） ; （ 3 ） The obviously increased level of E-cadhein mRNA inEGCG intervened tumour tissue had statistically significant difference （ P 〈 O. O1 ）. Conclusion EGCG in- hibits EMT of B16 cells clearly through the TGF-[31/STAT 3 signal pathway. Key words: Epigallocatechin-3-gallate ; Epithelial-mesenehymal transition ; Transforminggrowth factor-131 ; Melanoma

Inhibitory effect of homochlorcyclizine on melanogenesis in α-melanocyte stimulating hormone-stimulated mouse B16 melanoma cells

The histamine receptor H1 antagonist homochlorcyclizine (HC) has been widely used as an antihistamine agent for the treatment of allergies. However, the effect of HC on skin pigmentation is not known. In the present study, we investigated the inhibitory effect of HC on melanogenesis in mouse B16 melanoma cells. Our results showed that HC inhibited melanogenesis in either α-melanocyte stimulating hormone (α-MSH)- or 3-isobutyl-1-methylxanthin (IBMX)-stimulated B16 cells in a dose-dependent manner. Despite the strong inhibition of melanogenesis by HC, it was surprisingly found that HC did not reduce either cellular or melanosomal tyrosinase activity in α-MSH-stimulated B16 cells. In addition, HC also did not directly inhibit either murine or mushroom tyrosinase activity in the cell-free system. Moreover, western blotting and reverse-transcription polymerase chain reaction (RT-PCR) analyses respectively confirmed that HC did not downregulate levels of tyrosinase protein and its mRNA in α-MSH-stimulated B16 cells. These results clearly demonstrated that HC inhibits melanogenesis of B16 cells by a mechanism other than reduction of the cellular tyrosinase activity. From the present study, HC was proven to be a good candidate as a skin-whitening agent for treatment of skin hyperpigmentation, and this generic drug might be suitable for use in combination with other depigmenting agents due to its unique inhibition mechanism.

Melanogenesis inhibition by a crude extract of Magnolia officinalis

In the present study, we investigated the inhibitory effect of a crude extract from Magnolia officinalis (MOE) on melanogenesis in both mouse B16 melanoma cells and zebrafish. Our results showed that MOE inhibited melanogenesis in either a-melanocyte stimulating hormone (a-MSH)- or 3-isobutyl-1-methylxanthin (IBMX)-stimulated B16 cells in a dose-dependent manner with an IC50 value of 9.3 µg/ml. In addition, MOE also inhibited cellular tyrosinase activity with an IC50 value of 13.4 µg/ml while no inhibitory activity was found by MOE against cell-free tyrosinase activity. Moreover, western blotting and real time reverse-transcription polymerase chain reaction (qRT-PCR) analyses, respectively confirmed that MOE downregulated levels of tyrosinase protein but not that of its mRNA in a-MSH-stimulated B16 cells. These results demonstrated that MOE inhibits melanogenesis of B16 cells by a pre-translational regulation on tyrosinase gene expression. On the other hand, when using zebrafish as a depigmenting assay system, MOE could inhibit both melanogenesis and tyrosinase activity in the in vivo model. From the present study, MOE was proven to be a good candidate as a skin-whitening agent for treatment of skin hyperpigmentation. Key words: Magnolia officinalis, melanogenesis, tyrosinase, melanin, inhibition.

Stimulation of melanogenesis in murine melanoma cells by 2-mercapto-1-(β-4-pyridethyl) benzimidazole (MPB)

The effects of 2-mercapto-1-(β-4-pyridethyl) benzimidazole (MPB), one of the benzimidazole derivatives designed for a nucleic acid analogue, on melanogenesis of murine B16-F10 melanoma cell lines were investigated. MPB (40 μM) induced a striking dendricity in B16 melanoma cells within 12 h and maximal dendricity between 48 and 72 h. The stimulation of melanin synthesis was observed after only 2 days of treatment together with a dose-dependent growth inhibition. Moreover, MPB increased the activity of tyrosinase through the expression of tyrosinase mRNA without increasing the intracellular cyclic AMP content. MPB-induced melanogenesis was inhibited by novel protein kinase A inhibitors, KT-5720 and H-85. These findings indicate that MPB stimulated B16 cells to terminally differentiate and may be a useful drug in studying the regulation of melanogenesis.