Melanin Production In Melanoma Cells Research Articles

Milk Exosome-Derived MicroRNA-2478 Suppresses Melanogenesis through the Akt-GSK3β Pathway

Exosomes participate in intercellular communication by transferring molecules from donor to recipient cells. Exosomes are found in various body fluids, including blood, urine, cerebrospinal fluid and milk. Milk exosomes contain many endogenous microRNA molecules. MicroRNAs are small noncoding RNAs and have important roles in biological processes. The specific biological functions of milk exosomes are not well understood. In this study, we investigated the effects of milk exosomes on melanin production in melanoma cells and melanocytes. We found that milk exosomes decreased melanin contents, tyrosinase activity and the expression of melanogenesis-related genes in melanoma cells and melanocytes. Bovine-specific miR-2478 in exosomes inhibited melanin production. We found that Rap1a is a direct target gene of miR-2478 in melanoma cells and melanocytes. MiR-2478 overexpression decreased Rap1a expression, which led to downregulated melanin production and expression of melanogenesis-related genes. Inhibition of Rap1a expression decreased melanogenesis through the Akt-GSK3β signal pathway. These results support the role of miR-2478 derived from milk exosomes as a regulator of melanogenesis through direct targeting of Rap1a. These results show that milk exosomes could be useful cosmeceutical ingredients to improve whitening.

SIRT7 gene knockout using CRISPR/Cas9 system enhances melanin production in the melanoma cells

Melanin is a prominent pigment of skin and hair, and its deficiency can cause various disorders such as hair graying and albinism. The improvement of melanin production at a genetic level could offer an effective and permanent solution. Recently, SIRT7 has evoked an interest in the study of hair follicle stem cells, but its role in melanin synthesis remains unclear. In the present study, we have first successfully developed SIRT7 gene KO melanoma cells using the CRISPR/Cas9 system. It was found that the SIRT7 gene KO enhanced melanin production in melanoma cells. To validate the role of SIRT7 in melanin production, RT-PCR, western blot, and immunofluorescence staining assays were performed. The expression levels of melanin-producing genes and proteins (MITF, TRP1, TRP-2, TYR, TH) were significantly increased in SIRT7 gene KO cells compared to normal cells. In addition, melanin production was increased in KO cells higher than in normal cells through the image analysis. All these results suggest that SIRT7 could play an essential role in regulating melanin production, providing an alternative drug target to treat pigmentary disorders.

Synthesis and Cytotoxic Evaluation of Kojic Acid Derivatives with Inhibitory Activity on Melanogenesis in Human Melanoma Cells.

Malignant melanoma is an agressive tumour related to the overproduction of melanin, which provides colors of skin, eyes and hair. In addition contributing to the risk of malignant melanoma, abnormal production of melanin has many drawbacks, including hyperpigmentation, post-inflammatory pigmentation, melasma and skin aging. Kojic acid is currently employed in order to lighten skin pigmentation and provide depigmentation. Mannich bases of kojic acid with the structure of 2-substituted-3-hydroxy-6-hyroxymethyl/chloromethyl/ methyl/morpholinomethyl/piperidinylmethyl/pyrrolidinylmethyl-4H-pyran-4-one (compounds 1-23) were synthesized by the reaction of kojic acid/chlorokojic acid/allomaltol and substituted benzylpiperazine derivatives in the presence of formaline. To obtain the cyclic amine (morpholine, piperidine and pyrrolidine) derivatives, nucleophilic substitutions were carried out. Cytotoxic effects on A375 human malignant melanoma, HGF-1 human gingival fibroblasts, and MRC-5 human lung cell lines were investigated by sulphorhodamine B assay. Control agents were vemurafenib, dacarbazine, temozolomide, and lenalidomide, which are the commercially available drugs for the treatment of malignant melanoma. Cytotoxic action against melanoma cells was significantly more efficacious (IC50: 11.26-68.58 μM) than the FDA-approved drugs except for vemurafenib. Fourteen of the compounds were proven to have higher IC50 values for the non-cancerous cell lines, HGF-1, and MRC-5 cells. Melanogenesis inhibition assay was performed to observe the ability of the drugs to inhibit melanin production and certain compounds were shown to be capable of actively inhibiting melanin production in melanoma cells. Mannich bases of kojic acid derivatives may be promising therapeutic agents, since some have more potent effects on melanoma cells than previously FDA-approved drugs for the treatment of malignant melanoma.

Inhibitory effects of Dunaliella salina on tyrosinease activity and melanin production in melanoma cells

Inhibitory effects of Dunaliella salina on tyrosinease activity and melanin production in melanoma cells

Inhibitory effect of ephedrannins A and B from roots of Ephedra sinica STAPF on melanogenesis

BackgroundMelanogenesis, a process producing the pigment melanin in human skin, eyes and hair, is a major physiological response against various environmental stresses, in particular exposure to ultraviolet radiation, and its pathway is regulated by a key enzyme, tyrosinase. In this study, we evaluated the effects of ephedrannins A and B, which are polyphenols from the roots of Ephedra sinica, commonly used in herbalism in oriental countries, on mushroom tyrosinase and melanogenesis in B16F10 melanoma cells. MethodsTheir effects on mushroom tyrosinase were determined via kinetic studies using a spectrophotometric analysis and those on melanin and tyrosinase production in melanoma cells treated with α-MSH (melanin stimulating hormone) were examined using PCR and ELISA. ResultsBoth ephedrannins A and B exhibited concentration-dependent inhibitory effects on l-tyrosine oxidation by mushroom tyrosinase, and the inhibition mechanism was competitive and reversible with l-tyrosine as the substrate. In addition, melanin production in melanoma cells was also suppressed in a concentration-dependent manner by ephedrannins A and B without significant effects on cell proliferation at the concentrations tested. Both compounds showed inhibitory effects on melanin production by suppressing the transcription of tyrosinase in the cells. ConclusionBoth compounds exhibited significant inhibitory effects, but the inhibition by ephedrannin B was much more effective than that by ephedrannin A. Both ephedrannins A and B may be good candidates for a whitening agent for skin. General significanceThis is the first report that describes effective inhibition of melanin production by ephedrannins A and B isolated from Ephedra roots.

Keratinocyte-derived Laminin-332 Protein Promotes Melanin Synthesis via Regulation of Tyrosine Uptake

Melanocytes, which produce the pigment melanin, are known to be closely regulated by neighboring keratinocytes. However, how keratinocytes regulate melanin production is unclear. Here we report that melanin production in melanoma cells (B16F10 and MNT-1) was increased markedly on a keratinocyte-derived extracellular matrix compared with a melanoma cell-derived extracellular matrix. siRNA-mediated reduction of keratinocyte-derived laminin-332 expression decreased melanin synthesis in melanoma cells, and laminin-332, but not fibronectin, enhanced melanin content and α-melanocyte-stimulating hormone-regulated melanin production in melanoma cells. Similar effects were observed in human melanocytes. Interestingly, however, laminin-332 did not affect the expression or activity of tyrosinase. Instead, laminin-332 promoted the uptake of extracellular tyrosine and, subsequently, increased intracellular levels of tyrosine in both melanocytes and melanoma cells. Taken together, these data strongly suggest that keratinocyte-derived laminin-332 contributes to melanin production by regulating tyrosine uptake.

Skeletal Muscle Phenotypically Converts and Selectively Inhibits Metastatic Cells in Mice

Skeletal muscle is rarely a site of malignant metastasis; the molecular and cellular basis for this rarity is not understood. We report that myogenic cells exert pronounced effects upon co-culture with metastatic melanoma (B16-F10) or carcinoma (LLC1) cells including conversion to the myogenic lineage in vitro and in vivo, as well as inhibition of melanin production in melanoma cells coupled with cytotoxic and cytostatic effects. No effect is seen with non-tumorigenic cells. Tumor suppression assays reveal that the muscle-mediated tumor suppressor effects do not generate resistant clones but function through the down-regulation of the transcription factor MiTF, a master regulator of melanocyte development and a melanoma oncogene. Our findings point to skeletal muscle as a source of therapeutic agents in the treatment of metastatic cancers.

Inhibition of melanogenesis as a radiation sensitizer for melanoma therapy

Melanin pigment displays strong photo- and radioprotective properties, suggesting that inhibition melanogenesis could increase sensitivity of melanoma to ionizing radiation. We tested this concept in human melanoma cells cultured in either Ham's F10 medium to maintain amelanotic phenotype or DMEM to induce/stimulate melanin production, respectively; N-phenylthiourea (PTU) and D-penicillamine were used as an inhibitor of melanogenesis. Melanogenic activity was evaluated by visual inspection (color of cell pellets) or by measurement of tyrosinase (dopa oxidase) activity assay. Amelanotic cells or cells with various melanin content were exposed to gamma radiation and tested for viability and colony forming capability. Gamma radiation at doses of 2-15 Gy inhibited cell viability and colony forming efficiency in dose- and time-dependent manner, but pigmented melanoma cells were significantly more resistant to gamma radiation than nonpigmented cells (p < 0.05-0.001). Both PTU and D-penicillamine inhibited strongly tyrosinase activity and melanin production in melanoma cells (p < 0.05-0.001). Furthermore, inhibition of melanogenesis by PTU or D-penicillamine resulted in enhancement of melanoma cells sensitivity to killing by gamma rays. In conclusion, the results of these cell culture experiments give support to a clinical trial of pharmacologically induced decrease in melanin synthesis to enhance the efficacy of radiotherapy in advanced melanomas.

Biological actions of melanocyte-stimulating hormone.

Melanocyte-stimulating hormone (alpha-melanotropin, MSH) may function in a number of diverse physiological roles. MSH stimulates (1) rapid translocation of melanosomes (melanin granules) in dermal melanophores to effect rapid colour change and (2) melanogenesis in normal and abnormal (melanoma) epidermal melanocytes. Both actions involve (1) initial binding of the peptide on the melanocyte membrane, (2) transduction of signal to adenylate cyclase, and (3) increased cytosolic levels of cyclic AMP. Efforts to prepare radioiodinated MSH and analogues for radioreceptor studies using melanoma membranes and intact cells reveal that conventional iodination procedures inactivate the hormone because of oxidative and iodination effects on specific structural components of the peptide. These effects can be circumvented by the use of synthetically tailored MSH analogues. Transduction of signal from receptor to adenylate cyclase requires calcium, but prostaglandin or beta-adrenoceptor stimulation of melanophores does not. The nucleotide and metal ion requirements for mouse melanoma adenylate cyclase activity have been characterized. There is both a transcriptional and translational requirement for MSH stimulation of tyrosinase activity and melanin production in melanoma cells. Melanosome translocation within melanophores is enhanced in the absence of extracellular calcium. A model for the MSH control of melanosome movements suggests a bifunctional, but compartmentalized, role for calcium in the action of MSH.