Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:1.10.3.1 (tyrosinase)
 9,065 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cultures of differentiated melanocytes can readily be grown from the dissociated epidermis of neonatal mice, and immortal cell lines often develop from these. However, the first cells that grow and transiently dominate the cultures, while similar to melanocytes, are unpigmented. These have been shown to be precursors of melanocytes and may be termed melanoblasts. Under our previous standard culture conditions, involving the use of keratinocyte feeder cells, foetal calf serum, the phorbol ester 12-O-tetradecanoyl phorbol acetate (TPA) and cholera toxin, all the melanoblasts spontaneously differentiated to pigmented melanocytes within about 3 weeks. We now describe some factors affecting the proliferation and differentiation of diploid murine melanoblasts in the presence of serum. Murine stem cell factor/steel factor (SCF), basic fibroblast growth factor (bFGF) and murine leukaemia inhibitory factor/differentiation-inhibiting activity (LIF/DIA) all increased melanoblast numbers. SCF and LIF also slightly inhibited melanoblast differentiation, while cholera toxin and TPA promoted differentiation. Using some of these findings, and by regular replacement of keratinocyte or fibroblastoid feeder cells, we have established a clonal line of immortal murine melanoblasts, 'melb-a'. These cells express tyrosinase-related protein-2 but not, in general, tyrosinase. They can be induced to differentiate irreversibly to functional melanocytes (also proliferative and immortal) by plating in the absence of feeder cells. Thus a new immortal melanocyte line, 'melan-a2', has also been produced.
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PMID:A cloned, immortal line of murine melanoblasts inducible to differentiate to melanocytes. 754 May 32

The proto-oncogene c-Kit encodes a membrane receptor protein with intrinsic tyrosine kinase activity. Activation of c-Kit induces cell proliferation, differentiation or migration among different cell types. The present study provides evidence that c-Kit plays an important role in the cell differentiation rather than in cell proliferation in pigment cells. We found that normal human melanocytes and a limited number of melanoma cells, e.g. WM35, WM39 and G361 cell lines, expressed the c-Kit gene together with tyrosinase and TRP-1 genes. When exposed to alpha-melanocyte stimulating hormone, these three cell lines also showed an increased tyrosinase (dopa-oxidase) activity. By incubating these cells with 20 ng/ml of stem cell factor (SCF) which is a ligand of c-Kit receptor, we found a transient increase of tyrosinase activity 2-4 h post-incubation, indicating an early response of tyrosinase activation, either by elevating tyrosinase protein expression or by tyrosinase protein modification (e.g. phosphorylation). However, Western blot analysis using anti-tyrosinase antibody suggested that there was no change of tyrosinase protein expression between SCF-treated and non-treated cells. We therefore suggest that protein modulation of tyrosinase (e.g. phosphorylation) plays an important role in c-Kit-induced melanogenesis.
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PMID:Coordinated mRNA expression of c-Kit with tyrosinase and TRP-1 in melanin pigmentation of normal and malignant human melanocytes and transient activation of tyrosinase by Kit/SCF-R. 854 20

During development, the interaction of stem cell factor (SCF) with its receptor, KIT, is critical for the survival of melanocytes. Limited in vivo human studies have suggested a possible activating role of SCF on adult human melanocytes. In order to study the impact of this pathway on normal melanocyte homeostasis, human skin xenografts were treated with serial injections of recombinant human SCF or a KIT-inhibitory antibody (K44.2). On histologic evaluation, SCF injection increased, whereas KIT inhibition decreased the number, size, and dendricity of melanocytes. Immunohistochemical expression of melanocyte differentiation antigens, including tyrosinase-related-protein-1 and gp100/pmel17, was markedly increased by treatment with SCF, and decreased by K44.2 treatment. The number of Ki67-positive melanocytes was increased in the SCF-treated tissue, suggesting a direct proliferative effect of SCF; conversely, treatment with K44.2 resulted in melanocyte loss, which did not appear reversible with prolonged treatment. These findings demonstrate that the SCF/KIT pathway remains critical in adult human skin, and that pharmacologic modulation of this single pathway can control cutaneous melanocyte homeostasis.
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PMID:The SCF/KIT pathway plays a critical role in the control of normal human melanocyte homeostasis. 1041 34

Dendritic cells (DCs) are professional antigen-presenting cells (APCs) that can be used for vaccination purposes, to induce a specific T-cell response in vivo against melanoma-associated antigens. We have shown that the sequential use of early-acting hematopoietic growth factors, stem cell factor, IL-3 and IL-6, followed by differentiation with IL-4 and granulocyte-macrophage colony-stimulating factor allows the in vitro generation of large numbers of immature DCs from CD34(+) peripheral blood progenitor cells. Maturation to interdigitating DCs could specifically be induced within 24 hr by addition of TNF-alpha. Here, we report on a phase I clinical vaccination trial in melanoma patients using peptide-pulsed DCs. Fourteen HLA-A1(+) or HLA-A2(+) patients received at least 4 i.v. infusions of 5 x 10(6) to 5 x 10(7) DCs pulsed with a pool of peptides including either MAGE-1, MAGE-3 (HLA-A1) or Melan-A, gp100, tyrosinase (HLA-A2), depending on the HLA haplotype. A total of 83 vaccinations were performed. Clinical side effects were mild and consisted of low-grade fever (WHO grade I-II). Clinical and immunological responses consisted of anti-tumor responses in 2 patients, increased melanoma peptide-specific delayed-type hypersensitivity reactions in 4 patients, significant expansion of Melan-A- and gp100-specific cytotoxic T lymphocytes in the peripheral blood lymphocytes of 1 patient after vaccination and development of vitiligo in another HLA-A2(+) patient. Our data indicate that the vaccination of peptide-pulsed DCs is capable of inducing clinical and systemic tumor-specific immune responses without provoking major side effects.
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PMID:Phase I study in melanoma patients of a vaccine with peptide-pulsed dendritic cells generated in vitro from CD34(+) hematopoietic progenitor cells. 1076 Aug 27

Stem cell factor (SCF) and endothelin-3 (ET3) are both necessary for melanocyte development. In order to obtain immortal cell populations of melanoblasts that can survive without feeder cells, we first obtained an immortal cell population of neural crest cells (NCCs) from Sl/+ and +/+ mice of strain WB by incubating with a culture medium supplemented with SCF and ET3, and then we designated them as NCC-SE3 cells. NCC-SE3 cells were bipolar, polygonal, or round in shape and possessed melanosomes of stages I-III (mainly stage I). They were positive to dihydroxyphenylalanine (DOPA) reaction and expressed KIT (a receptor tyrosine kinase), tyrosinase, tyrosinase-related protein-1 (TRP1), tyrosinase-related protein-2 (TRP2), and endothelin-B receptor (ETRB) as determined by immunostaining. We next cultured NCC-SE3 cells by changing culture medium from the one supplemented with SCF + ET3 to the one supplemented with SCF or ET3. NCC-SE3 cells cultured with ET3 alone, designated as NCC-E3 cells, were bipolar in shape and had mainly stage II melanosomes and expressed the same proteins as did NCC-SE3 cells. However, NCC-SE3 cells cultured with SCF alone, designated as NCC-S4.1 cells, were polygonal in shape and had mainly stage I melanosomes. They are thought to be more immature because they were positive to KIT, TRP1, and TRP2, but not to ETR(B), tyrosinase, and DOPA reaction. When 12-O-tetradecanoylphorbol 13-acetate and cholera toxin were added to the culture medium, NCC-S4.1 cells changed shape from polygonal to bipolar and became DOPA-positive. This suggests that NCC-S4.1 cells are melanoblasts that have the potential to differentiate into melanocytes. These cell populations will be extremely useful to study factors that affect melanocyte development and melanogenesis.
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PMID:Stem cell factor and/or endothelin-3 dependent immortal melanoblast and melanocyte populations derived from mouse neural crest cells. 1104 61

Stem cell factor (SCF) and endothelin 3 (EDN3) are both necessary for melanocyte development. We have established an immortal cell population of neural crest cells from C57BL/6 mice, cultivating them with SCF, EDN3 and 15% fetal calf serum without feeder cells, and have designated that line as C57NCC SE. C57NCC SE consists of a population of melanocytes in various stages of differentiation. We used a single-cell cloning method, in which only one cell is transferred to each new culture plate, and succeeded in establishing an immortal cell line named NCCmelan5. All NCCmelan5 cells were positive for KIT (SCF receptor), HMB45 (human melanosomal antigen), tyrosinase-related protein-1 (TYRP1), tyrosinase-related protein-2 (TYRP2), tyrosinase and endothelin receptor B (EDNRB) and all could oxidize 3,4-dihydroxyphenylalanine (DOPA) to form melanin. Measurement of their DNA content revealed that 88.6% of the cells were in the G0-G1 phase, suggesting that they retained normal DNA ploidy. Thus, NCCmelan5 cells have the characteristics of mature melanocytes except that they are immortal; these cells may prove useful to study factors that directly affect melanogenesis and melanocyte development without the influence of feeder cells. It is clear that our attempt to establish immortal cell lines from murine neural crest cells would have never been successful without the addition of SCF and EDN3, since C57NCC SE and NCCmelan5 cells require those factors to proliferate.
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PMID:Establishment and characterization of a mouse neural crest derived cell line (NCCmelan5). 1154 10

Dermatofibromas have an increased brownish color due to hyperpigmentation of the overlying skin. To determine paracrine factors involved in the epidermal hyperpigmentation, we have studied the expression of cytokines in lesional and nonlesional dermatofibroma skin at the transcriptional and protein levels using reverse transcription polymerase chain reaction and immunohistochemistry, respectively. The number of tyrosinase immuno-positive melanocytes in the pigmented dermatofibroma epidermis is significantly increased (2-fold) compared with nonlesional normal epidermis. Reverse transcription polymerase chain reaction analysis of mRNAs encoding stem cell factor and hepatocyte growth factor demonstrated that there is an accentuated expression of stem cell factor and hepatocyte growth factor transcripts in the lesional dermatofibroma dermis compared with the nonlesional dermis, although there is no difference in their expression between the lesional and nonlesional epidermis. In contrast, mRNA transcripts encoding endothelin-1, growth-related oncogene alpha, and basic fibroblast growth factor are not increased in lesional epidermis or in dermis relative to nonlesional skin. In parallel, immunohistochemical analysis using antibodies to stem cell factor and hepatocyte growth factor reveal a marked immunostaining in growing fibroblastic tumor cells in the dermatofibroma lesions with no detectable staining in the nonlesional dermis, but there is no difference in their immunostaining between the lesional and nonlesional epidermis. Interestingly, and consistent with the increased expression of stem cell factor in lesional dermatofibroma dermis, toluidine blue staining in the dermis revealed a 5-fold increase in the number of mast cells, an indication of their longevity or accumulation induced by stem cell factor. These findings suggest an important role of fibroblastic tumor cell-derived stem cell factor in the mechanism involved in the hyperpigmentation of the dermatofibroma epidermis.
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PMID:The mechanism of epidermal hyperpigmentation in dermatofibroma is associated with stem cell factor and hepatocyte growth factor expression. 1156 69

As lysosphingolipids have multiple bio-modulator functions in various types of cells, we measured the biological effects of sphingosylphosphorylcholine (SPC) on cultured human melanocytes to determine whether these lysosphingolipids have the potential to activate these cells. The addition of SPC to cultured human melanocytes significantly stimulated DNA synthesis assessed by [3H]thymidine and melanogenesis assessed by the release of [3H]H2O (tyrosinase activity), the incorporation of [14C]thiouracil (melanin synthesis) and dopa-oxidase activity. Reverse transcription-polymerase chain reaction of RNA isolated from human melanocytes exposed to SPC revealed an upregulation of mRNA transcripts for tyrosinase, microphthalmia-associated transcription factor-M, endothelin B receptor and the stem cell factor receptor, c-kit. An increase in expression of tyrosinase and c-kit proteins was also demonstrated by Western blot analysis. This stimulation of melanogenesis by SPC was associated with a marked increase in the phosphorylation of extracellular signal-regulated kinase 1/2. These results suggest that SPC may be a melanogenic stimulator of human melanocytes inducing the coordinated upregulated expression of various melanogenic molecules, including c-kit.
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PMID:Sphingosylphosphorylcholine is a Melanogenic Stimulator for Human Melanocytes. 1462 25

To evaluate the etiologic role of ultraviolet (UV) radiation in acquired dermal melanocytosis (ADM), we investigated the effects of UVA and UVB irradiation on the development and differentiation of melanocytes in primary cultures of mouse neural crest cells (NCC) by counting the numbers of cells positive for KIT (the receptor for stem cell factor) and for the L-3,4-dihydroxyphenylalanine (DOPA) oxidase reaction. No significant differences were found in the number of KIT- or DOPA-positive cells between the UV-irradiated cultures and the non-irradiated cultures. We then examined the effects of UV light on KIT-positive cell lines derived from mouse NCC cultures. Irradiation with UVA but not with UVB inhibited the tyrosinase activity in a tyrosinase-positive cell line (NCCmelan5). Tyrosinase activity in the cells was markedly enhanced by treatment with alpha-melanocyte-stimulating hormone (alpha-MSH), but that stimulation was inhibited by UVA or by UVB irradiation. Irradiation with UVA or UVB did not induce tyrosinase activity in a tyrosinase-negative cell line (NCCmelb4). Levels of KIT expression in NCCmelan5 cells and in NCCmelb4 cells were significantly decreased after UV irradiation. Phosphorylation levels of extracellular signal-regulated kinase 1/2 in cells stimulated with stem cell factor were also diminished after UV irradiation. These results suggest that UV irradiation does not stimulate but rather suppresses mouse NCC. Thus if UV irradiation is a causative factor for ADM lesions, it would not act directly on dermal melanocytes but may act in indirect manners, for instance, via the overproduction of melanogenic cytokines such as alpha-MSH and/or endothelin-1.
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PMID:Effects of ultraviolet light on melanocyte differentiation: studies with mouse neural crest cells and neural crest-derived cell lines. 1501 4

Little is known about the mechanisms involved in the dysfunction of melanocytes in vitiligo epidermis. It is hypothesized that some cytokine/receptor interactions may be affected, resulting in dysfunction and/or loss of melanocytes. This study has compared the expression of endothelin (ET)-1, the ET-1 receptor (ET(B)R), granulocyte macrophage colony stimulating factor (GM-CSF), stem cell factor (SCF), the SCF receptor (KIT protein), tyrosinase, and S100 alpha between lesional and non-lesional vitiligo epidermis. Analysis by reverse transcription-polymerase chain reaction (RT-PCR) and by western blotting for ET-1 and SCF unexpectedly demonstrated up-regulated expression of these cytokines in lesional vitiligo epidermis. Immunohistochemistry with antibodies to melanocyte markers revealed that at the edge of the lesional epidermis, melanocytes remain and express tyrosinase, S100 alpha and ET(B)R, but not KIT protein or melanocyte-specific microphthalmia-associated transcription factor (MITF-M). Quantitation of the staining revealed a slight or moderate decrease in the number of S100 alpha, tyrosinase, and ET(B)R-positive cells at the edge of the lesional epidermis. In contrast, the number of cells expressing KIT protein was markedly decreased at the edge of the lesional epidermis compared with the non-lesional epidermis. At the centre of the lesional epidermis, there was complete loss of melanocytes expressing KIT protein, S100 alpha, ET(B)R, and/or tyrosinase. Western blotting revealed down-regulated expression of c-kit and MITF-M proteins at the edge of the lesional epidermis in vitiligo. These findings suggest that reduction in the expression of KIT protein by melanocytes and its downstream effectors, including MITF-M, may be associated with the dysfunction and/or loss of melanocytes in vitiligo epidermis.
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PMID:Mechanisms underlying the dysfunction of melanocytes in vitiligo epidermis: role of SCF/KIT protein interactions and the downstream effector, MITF-M. 1509 74


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