Antigens, CD34 ; metabolism ; Biomarkers ; metabolism ; Cell Differentiation ; physiology ; Hair Follicle ; cytology ; Humans ; Intramolecular Oxidoreductases ; metabolism ; Keratinocytes ; metabolism ; Melanins ; metabolism ; Melanocytes ; metabolism ; PAX3 Transcription Factor ; metabolism ; Stem Cells ; metabolism

BACKGROUNDTopical tacrolimus has been used for vitiligo as a common treatment option for more than ten years while the underlying mechanism is still uncertain. The aim of this study was to investigate the direct effects of tacrolimus on the melanogenesis and migration on human A375 melanoma cells. The expression of c-KIT mRNA and protein of human A375 cells were also investigated.

METHODSThe cultured A375 human melanoma cells were randomly assigned to control and tacrolimus treatment groups (10, 10(2), 10(3) and 10(4) nmol/L). The cell proliferation was measured with Cell Counting Kit-8 assays. Melanin content was measured with NaOH method. Transwell migration assay was used to measure cell migration. The expression of c-KIT mRNA and protein were measured with real-time fluorescence quantitative polymerase chain reaction and immunohistochemistry respectively.

RESULTSThe cell proliferation of the 10(3) and 10(4) nmol/L tacrolimus groups were significantly lower (0.666 ± 0.062 and 0.496 ± 0.038) as compared with the control (0.841 ± 0.110, P < 0.05). The mean melanin content in all groups treated with different concentration of tacrolimus (10, 10(2), 10(3), 10(4) nmol/L) increased compared with the control group (P < 0.05). Dose-dependent increase in cell migration were seen in all tacrolimus-treated groups (P < 0.01). The expression of c-KIT mRNA level in A375 cells exposed to tacrolimus (10(3) and 10(4) nmol/L) had significantly increased by 3.03-fold and 3.19-fold respectively compared with the control (P < 0.05).

CONCLUSIONSAlthough tacrolimus had no effects on cell proliferation on A375 human melanoma cells, it could increase the melanin content and cell migration. The expression of c-KIT mRNA and protein increased dose-dependently in tacrolimus-treated groups as compared with the control. Our study demonstrated that tacrolimus could enhance the melanogenesis and cell migration on A375 cells.

Cell Line, Tumor ; Cell Movement ; drug effects ; genetics ; Cell Proliferation ; drug effects ; Humans ; Immunohistochemistry ; Melanins ; metabolism ; Melanocytes ; cytology ; drug effects ; Proto-Oncogene Proteins c-kit ; genetics ; metabolism ; RNA, Messenger ; genetics ; Tacrolimus ; pharmacology

Calcitonin ; Carcinoma, Papillary ; drug therapy ; pathology ; surgery ; Female ; Humans ; Melanins ; Melanocytes ; cytology ; pathology ; Middle Aged ; Prognosis ; S100 Proteins ; Thyroid Neoplasms ; drug therapy ; pathology

OBJECTIVETo explore the method of constructing tissue-engineered skin using melanocytes and bone marrow mesenchymal stem cells (BMSCs) in vivo.

METHODSMelanocytes were isolated from human foreskin. BMSCs were isolated from human bone marrow. Both of them were co-cultured at a ratio of 1:10, and then were implanted into the collagen membrane to construct the tissue-engineered skin, which was applied for wound repair in nude mice. The effectiveness of wound repair and the distribution of melanocytes were evaluated by morphological observation, in vivo 4,6-diamidino-2-phenylindole, dihydrochloride (DAPI) fluorescent staining tracing, HE staining, S-100 immunohistochemistry, and transmission electron microscopy.

RESULTSThe wounds were satisfactorily repaired among the nude mice. The melanocytes were distributed in the skin with normal structure, as confirmed by DAPI fluorescent staining tracing, HE staining, S-100 immunohistochemistry, and transmission electron microscopy.

CONCLUSIONMelanocytes and BMSCs, after proper in vitro culture at an appropriate ratio, can construct the tissue-engineered skin with I type collagen membrane.

Animals ; Bone Marrow Cells ; cytology ; Cells, Cultured ; Coculture Techniques ; Collagen Type I ; Humans ; Melanocytes ; cytology ; Mesenchymal Stromal Cells ; cytology ; Mice ; Mice, Nude ; Skin ; injuries ; Skin, Artificial ; Tissue Engineering

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

OBJECTIVETo investigate the effects of 8-methoxypsoralen on human melanocytes [Ca(2+)]i and cytoskeleton actin organization in vitro.

METHODSHuman melanocytes were obtained from normal foreskins. Laser confocal microscope was employed to measure [Ca(2+)]i and rhodamine-conjugated phalloidin was used to visualize the cytoskeleton actin.

RESULTS8-methoxypsoralen increased [Ca(2+)]i and induced organization of actin stress fiber cytoskeleton.

CONCLUSION8-methoxypsoralen might influence the migration of melanocytes by increasing the intracellular free Ca(2+) concentration and cytoskeleton actin reorganization.

Actins ; biosynthesis ; genetics ; Calcium ; metabolism ; Cell Movement ; drug effects ; Cells, Cultured ; Cytoskeletal Proteins ; biosynthesis ; genetics ; Humans ; Melanocytes ; cytology ; drug effects ; metabolism ; Methoxsalen ; pharmacology ; Skin ; cytology

In this study, the essential oil from lotus flower extract, including petals and stamens, was assessed with regard to its effects on melanogenesis in human melanocytes. The lotus flower essential oil was shown to stimulate melanin synthesis and tyrosinase activity in a dose-dependent manner. The lotus flower essential oil induced the expression of tyrosinase, microphthalmia-associated transcription factor M (MITF-M), and tyrosinase-related proten-2 (TRP-2) proteins, but not tyrosinase mRNA. Moreover, it increased the phosphorylation of ERK and cAMP response element binding protein (CREB). In order to verify the effective components of the lotus flower oil, its lipid composition was assessed. It was found to be comprised of palmitic acid methyl ester (22.66%), linoleic acid methyl ester (11.16%), palmitoleic acid methyl ester (7.55%) and linolenic acid methyl ester (5.16%). Among these components, palmitic acid methyl ester clearly induced melanogenesis as the result of increased tyrosinase expression, thereby indicating that it may play a role in the regulation of melanin content. Thus, our results indicate that lotus flower oil may prove useful in the development of gray hair prevention agents or tanning reagents.
Blotting, Western ; Cell Proliferation ; Cyclic AMP/metabolism ; Cyclic AMP Response Element-Binding Protein/genetics/metabolism ; Flowers/*chemistry ; Gas Chromatography-Mass Spectrometry ; Humans ; Intramolecular Oxidoreductases/genetics/metabolism ; Lotus/*chemistry ; Melanins/*biosynthesis ; Melanocytes/*drug effects/metabolism ; Microphthalmia-Associated Transcription Factor/genetics/metabolism ; Monophenol Monooxygenase/genetics/metabolism ; Phosphorylation ; Plant Oils/*pharmacology ; RNA, Messenger/genetics/metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Skin/cytology/drug effects/metabolism

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

BACKGROUNDLatanoprost, a prostaglandin F2alpha analog, has been shown to be an effective intraocular pressure lowering agent which acts by inducing ciliary muscle cells to synthesise matrix metalloproteinases. However, the response of ciliary melanocytes to latanoprost has never been reported. This research has investigated the ability of latanoprost to induce matrix metalloproteinase-1 expression in human ciliary melanocytes, and thereby advance the understanding of the mechanism of PGF(2alpha) in decreasing intraocular pressure.

METHODSIn vitro human ciliary melanocytes were treated for 48 hours with five different concentrations of latanoprost (100, 150, 200, 500, and 1000 nmol/L). Ciliary melanocytes treated with 0.01% ethanol (vehicle) were used as a control. The expression of matrix metalloproteinase-1 in ciliary melanocytes was determined by Western blotting and immunofluorescent staining.

RESULTSWestern blotting showed that the expression of matrix metalloproteinase-1 in ciliary melanocytes was induced by latanoprost, and the level of expression was dependent on the concentration of latanoprost in the culture medium. Immunofluorescent staining showed that matrix metalloproteinase-1 was confined to the ciliary melanocyte cytoplasm.

CONCLUSIONSLatanoprost induced the expression of matrix metalloproteinase-1 in human ciliary melanocytes in a dose-dependent manner. Ciliary melanocytes, as well as ciliary muscle cells, may also play an important role in uveoscleral outflow modulation.

Cells, Cultured ; Ciliary Body ; cytology ; drug effects ; enzymology ; Female ; Fluorescent Antibody Technique ; Humans ; Immunoblotting ; Male ; Matrix Metalloproteinase 1 ; analysis ; Melanocytes ; drug effects ; enzymology ; Prostaglandins F, Synthetic ; pharmacology

OBJECTIVETo construct an in vitro equivalent of the pigmented skin using tissue engineering methods.

METHODSSurgically removed foreskins was used as the source of keratinocytes and melanocytes harvested by routine tissue digestion. The fibroblasts were enriched by tissue block culture and seeded into the scaffold constructed using mouse tail collagens to construct the pigmented skin equivalent model. The general structure and the melanocyte distribution and growth status in this model were observed with HE staining and Fontana Masson staining. The ultrastructure of the constructed pigmented skin equivalent was observed by transmission electron microscope.

RESULTS AND CONCLUSIONThe pigmented skin equivalent model was structurally intact, and allowed optimal cell growth. Fontana Masson staining identified in the basal layer numerous melanocytes in normal growth, and the constructed model was structurally similar to normal skin tissue, suggesting successful construction of the pigmented skin equivalent model.

Animals ; Foreskin ; cytology ; Humans ; Keratinocytes ; cytology ; Male ; Melanocytes ; cytology ; Mice ; Skin Pigmentation ; Skin, Artificial ; Tissue Engineering ; methods