The observation statistics suggested that the haemolymph melanization speed of larvae became fast and the growth inhibition of Escherichia coli was strong as the quantities of feeding on mulberry leaves increased. The RT-PCR result showed that the mRNA expressions of melanin biosynthesis enzyme BmTan, BmPo-1, BmYellow-f and BmDdc were high in the haemolyph of 5 L 3 d larvae. The qPCR analysis showed Bmtan, Bmddc, Bmyellow, Bmebony and Bmblack, especially Bmddc expression were significantly higher in black disease larvae than in normal larvae. Compared with control, Ddc inhibitors drastically inhibited the lipopolysaccharide-induced haemolymph melanization. In addition, the content of Dopa and Dopamine markedly rose after E. coli injection. These indicated that haemolymph melanization was linked to immune defenses and Bmddc may play a role in melanization response of haemolymph immune in silkworm.
Animals ; Bombyx ; enzymology ; genetics ; microbiology ; Escherichia coli ; Genes, Insect ; Hemolymph ; chemistry ; Larva ; Melanins ; biosynthesis

Eumelanin is a heteropolymer that is generally composed of hydroxylated indole residues and plays diverse protective functions in various species. Melanin is derived from the amino acid tyrosine and production of melanin is a highly complex oxidative process with a number of steps that can either proceed enzymatically or non-enzymatically. Although melanin plays important protective roles in many species, during melanization, particularly in steps that can proceed non-enzymatically, many toxic intermediates are produced, including semiquinones, dopaquinone, indole-quinones and moreover, the production of many reactive oxygen species. To mitigate the production of reactive species, a number of proteins that regulate the biochemical process of melanization have evolved in various living species, which is closely related to adaptation and physiological requirements. In this communication, we discuss differences between non-enzymatic and enzymatic processes of melanization and the enzymatic regulation of melanization in difference species with an emphasis on differences between mammals and insects. Comparison between melanization and insect sclerotization is also emphasized which raises some interesting questions about the current models of these pathways.
Animals ; Biological Evolution ; Humans ; Insecta ; metabolism ; Mammals ; metabolism ; Melanins ; biosynthesis ; chemistry ; Models, Biological ; Species Specificity

OBJECTIVETo investigate the co-culture of keratinocytes with Malassezia isolates which cause the pityriasis versicolor with different color and to analyze the changes of cytokines associated with melanogenesis.

METHODSThe effects of Malassezia species with different proportions on the growth rate of keratinocytes was assessed with 5 g/L methyl thiazolyl tetrazolium (MTT). Co-culture of keratinocytes and Malassezia species were performed with isolates from hyer- and hypo-pigmentation areas of pityriasis versicolor. The supernatants were collected at different time points, and the changes of basic fibroblast growth factor (b-FGF), endothelin-1 (ET-1), nerve growth factor-beta (NGF-beta), interleukin-1alpha (IL-1alpha), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), stem cell factor (SCF) were recorded. Three control groups were established accordingly.

RESULTSWhen the ratio between keratinocytes and Malassezia species was lower than 1: 10, the growth rate of keratinocytes was not affected by Malassezia (P > 0.05). When the ratio was increased above 1:20, the growth rate of keratinocytes was significantly inhibited by Malassezia (P < 0.01). The secretions of IL-1alpha, IL-6, TNF-alpha, and ET-1 was significantly increased after the co-culture of keratinocytes and Malassezia (P < 0.01), while those of b-FGF, NGF-beta, and SCF had no significant changes (P > 0.05). Compared with the isolates from the hypo-pigmentation area, ET-1 induced by isolate from hyperpigmentation area significantly increased (P < 0.01).

CONCLUSIONWhen Malassezia isolates are co-cultured with keratinocytes, the secretions of cytokines associated with melanogenesis may differ from each other. ET-1 may play certain role in the hyper-pigmentation of pityriasis versicolor.

Cell Proliferation ; Cells, Cultured ; Cytokines ; biosynthesis ; Humans ; Keratinocytes ; cytology ; metabolism ; microbiology ; Malassezia ; isolation & purification ; physiology ; Melanins ; biosynthesis ; Tinea Versicolor ; microbiology

OBJECTIVETo investigate the interference effect of nicotinamide on UVA-induced melanin genesis and melanin transport in human skin melanocyte.

METHODSThe optimum UVA dose expected to cause cell proliferation: 0.2 J/cm(2), nicotinamide was added immediately after the 0.2 J/cm(2) UVA exposure and the melanin content, cell cycles, cell apoptosis and mRNA express level were measured respectively.

RESULTSMelanin content in melanocytes was increased significantly after exposed to 0.2 J/cm(2) UVA. Melanin content in melanocytes was decreased after treatment with 10.0 mmol/ml nicotinamide following UVA exposure, but the cell cycles and the cell apoptosis rate were not significantly altered. mRNA express levels of TYR, TRP-1 were modulated by nicotinamide.

CONCLUSIONNicotinamide has more effect on decreasing melanin genesis after UVA exposure, nicotinamide also plays a role in modulating the mRNA express of TYR, TRP-1 gene. It is possible to consider nicotinamide as an efficient and safe sun screen to provide a certain level of protection for UVA exposed skin.

Cells, Cultured ; Humans ; Melanins ; biosynthesis ; Melanocytes ; drug effects ; metabolism ; radiation effects ; Niacinamide ; pharmacology ; Ultraviolet Rays ; adverse effects

OBJECTIVETo investigate the effects of Liuweidihuang Decoction on the proliferation and melanin synthesis of cultured human melanocytes in vitro.

METHODSMTT assay was used to assess the proliferation of cultured human melanocytes after treatment with Liuweidihuang Decoction, and NaOH assay was employed to determine melanin synthesis.

RESULTSLiuweidihuang Decoction at the concentration of 5%-15% did not affect the proliferation of the melanocytes (P>0.05), and the decoction at 20%-30% significantly inhibited the cell proliferation, especially at the optimal concentration of 30% (P<0.01). The decoction promoted melanin synthesis in the melanocytes at the concentration of 10%-15% with the optimal concentration of 15% (P<0.01) but failed to produce such an effect at 20%-30%.

CONCLUSIONLiuweidihuang Decoction exert a two-way regulation on the proliferation and melanin synthesis of cultured human melanocytes in vitro.

Cell Proliferation ; drug effects ; Cells, Cultured ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Melanins ; biosynthesis ; Melanocytes ; cytology ; drug effects ; metabolism

OBJECTIVE: To explore the relationship between melanin synthesis and the congenital high myopia of albino guinea-pigs. METHODS: Twelve albino guinea-pigs and 12 pigmented guinea-pigs of 220~250 grams (aged 5~6 weeks) were chosen at random. The eyes were examined with retinoscopy, A-scan ultrasonography and vernier caliper. The retinal structures were examined with light and electronic microscope. RESULTS: The diopter was -19.17 D in albino guinea-pigs and +0.63 D in pigmented guinea-pigs on average. Compared with the pigmented guinea-pigs, the axial dimensions of the albino guinea-pigs were elongated. There was significant difference between the albino guinea-pigs and the pigmented guinea-pigs. The retinal thickness, pigment granules and membrane disc of the outer segment of the visual cells decreased in the albino guinea-pigs, and the membrane disc space became narrow. The normal retinal thickness, plenty of pigment granules , membrane disc and wide membrane disc space could be observed in the pigmented guinea-pigs. CONCLUSION Albino guinea-pigs have high myopia, and pigmented guinea pigs have light hyperopia. There are structural differences in the retina between albino guinea-pigs and pigmented guinea-pigs. The abnormity of albino guinea-pigs provides optical foundation for its high myopia.
Albinism ; complications ; Animals ; Guinea Pigs ; Melanins ; biosynthesis ; Microscopy, Electron, Scanning ; Myopia ; congenital ; etiology ; metabolism ; Random Allocation ; Retina ; metabolism ; pathology ; ultrastructure ; Skin Pigmentation

OBJECTIVETo evaluate the transfect results of recombinant adenovirus vector carrying tyrosinase gene (Ad-tyr) in vitro by magnetic resonance imaging (MRI) after the Ad-tyr was transfected into HepG2 cell.

METHODSThe Ad-tyr which carried the full-length cDNA of tyrosinase gene was transfected into HepG2 cell. The transfected cells were scan by MRI sequences of T1 weighted image (T1WI) , T2 weighted image (T2WI) , and short time inversion recovery (STIR) to observe the MRI signals of expressed melanin. Masson-Fontana staining was performed to search for melanin granules in transfected cells. Real-time PCR method was used to search for cDNA of tyrosinase gene.

RESULTSAd-tyr was transfected into HepG2 cells and synthesized a large amount of melanin inside. The synthesized melanin of 1 x 10(6) cells which had been transfected by Ad-tyr with the 50, 150, and 300 multiplicity of infection separately were all sufficient to be detected by MRI and showed high signals in MRI T1WI, T2WI, and STIR sequences. The signal intensities of MRI were positively correlated to the amounts of transfected Ad-tyr. The melanin granules were found in HepG2 cells in Masson-Fontana staining. The cDNA amount of tyrosinase gene in transfected HepG2 cells, which was detected by real-time PCR, was remarkably higher than that in nontransfected cells.

CONCLUSIONThe synthesized melanin of HepG2 cells, which controlled by expression of exogenous gene, can be detected by MRI, indicating that the adenovirus vector can efficiently carry the tyrosinase gene into HepG2 cells.

Adenoviridae ; genetics ; metabolism ; Gene Transfer Techniques ; Genetic Vectors ; genetics ; Hep G2 Cells ; Humans ; Magnetic Resonance Imaging ; methods ; Melanins ; analysis ; genetics ; Monophenol Monooxygenase ; biosynthesis ; genetics ; Transfection

Glycyrrhizin (GR), triterpenoid saponin composed of one glycyrrhetinic acid (GA) and two glucuronic acids, is a main constituent of the hydrophilic fraction of licorice (Glycyrrhiza glabra) extracts and is known to have a wide range of pharmacological actions. In this study, we investigated the mechanism of GR effect on melanogenesis in B16 murine melanoma cells. The cellular levels of tyrosinase mRNA, protein, enzyme activities and melanin contents were increased by GR in a dose dependent manner. Expression of tyrosinase-related protein-2 (TRP-2) mRNA was also increased by GR, however, no significant change was observed on TRP-1. No cytotoxicity was observed at the effective concentration range of GR. GA showed no effect on melanogenesis at the equivalent nontoxic concentrations, indicating that glycoside structure is important in the stimulatory effect of GR on melanogenesis. These results indicate that GR-induced stimulation of melanogenesis is likely to occur through the transcriptional activation.
Animal ; Blotting, Western ; Glycyrrhetinic Acid/pharmacology ; Glycyrrhizic Acid/*pharmacology ; Intramolecular Oxidoreductases/genetics/metabolism ; Melanins/*biosynthesis ; Melanoma, Experimental/enzymology/*metabolism ; Mice ; Monophenol Monooxygenase/genetics/metabolism ; Proteins/genetics/metabolism ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo observe the effect of aloesin, tea polyphenols, arbutin on melanocytes in the pigmented skin equivalent model.

METHODSFirst, we constructed the pigmented skin equivalent model in vitro. And then we detected the effect of aloesin, tea polyphenols and arbutin on the cells' shape, tyrosinase activity and formation of melanin in the constructed pigmented skin equivalent.

RESULTSThree depigmenting agents showed an inhibition effect on the tyrosinase activity of melanocytes and reduced significantly melanin content in the pigmented skin equivalent model, in which the tea polyphenols had the strongest effect, and then was the aloesin. But the tea polyphenols showed the strongest toxicity, while the aloesin and arbutin had a much lower toxicity.

CONCLUSIONSAll the three depigmenting agents showed a concentration dependent suppression effect on the tyrosinase activity and formation of melanin, in which the tea polyphenols was the strongest effect( P <0.05). Aoesin has a good suppression effect on the tyrosinase activity and formation of melanin, but has a much lower toxicity, which could be used as a safe depigmenting agent.

Arbutin ; pharmacology ; Cells, Cultured ; Chromones ; pharmacology ; Flavonoids ; pharmacology ; Foreskin ; cytology ; Glucosides ; pharmacology ; Humans ; Male ; Melanins ; biosynthesis ; Melanocytes ; drug effects ; Phenols ; pharmacology ; Pigmentation ; Polyphenols ; Skin ; drug effects ; Skin Aging ; drug effects

OBJECTIVEDespite the causes for melanin increase, the increased gene expression of TYR is a common pathological process. Based on this viewpoint, antisense-S-Oligo of TYR was designed and synthesized to regulate synthesis of melanin in order to explore the treatment for skin pigmentation.

METHODSThe cultured melanocytes were divided into 3 groups. The group 1 was treated with endothelin, group 2 treated with ultraviolet ray and group 3 was used as the control. In each group, the 5' antisense-S-Oligo, the 3' antisense-S-Oligo, the mixed antisense-S-Oligo of TYR or Dotap only was added. The melanin content and TYR gene expressions were examined.

RESULTSThe 5' antisense-S-Oligo, the 3' antisense-S-Oligo and the mixed antisense-S-Oligo significantly inhibited the increase of melanin content and TYR gene expression, which were caused by endothelin or ultraviolet ray treatment. Of the three treatments, the 3' antisense-S-Oligo showed the strongest effect.

CONCLUSIONAntisense-S-Oligo has significant regulating effects on TYR gene expression and melanin content. The 3' antisense-S-Oligo is more effective than the 5' antisense-S-Oligo.

3' Flanking Region ; genetics ; 5' Flanking Region ; genetics ; Endothelins ; pharmacology ; Gene Expression ; Melanins ; biosynthesis ; Melanocytes ; drug effects ; metabolism ; radiation effects ; Oligodeoxyribonucleotides, Antisense ; genetics ; pharmacology ; Tyrosine ; genetics ; metabolism ; Ultraviolet Rays