Melanocyte Migration Research Articles

Beta-catenin inhibits melanocyte migration but induces melanoma metastasis

The canonical Wnt signalling pathway induces the β-catenin/lymphoid enhancer factor transcription factors. It is activated in various cancers, most characteristically carcinomas, in which it promotes metastatic spread by increasing migration and/or invasion. The Wnt/β-catenin signalling pathway is frequently activated in melanoma, but the presence of β-catenin in the nucleus does not seem to be a sign of aggressiveness in these tumours. We found that, unlike its positive role in stimulating migration and invasion of carcinoma cells, β-catenin signalling decreased the migration of melanocytes and melanoma cell lines. In vivo, β-catenin signalling in melanoblasts reduced the migration of these cells, causing a white belly-spot phenotype. The inhibition by β-catenin of migration was dependent on MITF-M, a key transcription factor of the melanocyte lineage, and CSK, an Src-inhibitor. Despite reducing migration, β-catenin signalling promoted lung metastasis in the NRAS-driven melanoma murine model. Thus, β-catenin may have conflicting roles in the metastatic spread of melanoma, repressing migration while promoting metastasis. These results highlight that metastasis formation requires a series of successful cellular processes, any one of which may not be optimally efficient.

Atypical Lentiginous Nevus: A Clinical and Histopathologic Study of 14 Cases

BackgroundAtypical lentiginous nevus (of the elderly) is a peculiar form of dysplastic nevus. Clinically, this condition can resemble malignant melanoma and histologically, it has a lentiginous pattern with variable degrees of atypia and an absence of dermal nests. These features may lead to an erroneous diagnosis of lentigo maligna melanoma or lentiginous melanoma. Material and methodsWe reviewed 14 cases of atypical lentiginous nevus diagnosed at the dermatology department of Hospital General de Valencia in Valencia, Spain between December 2007 and March 2009. We studied the clinical and histopathologic features of the lesions after hematoxylin-eosin, Melan-A, and Ki-67 staining and compared our results to data reported in the literature. ResultsFour (28%) of the 14 patients (7 men, 7 women) were under 50 years of age. Clinically, most of the lesions (8/14) resembled atypical nevi and they were all located on the back. Histologically, they all had irregular lentiginous epidermal hyperplasia, with a proliferation of individual melanocytes only in the basal layer of the epidermis and an absence of dermal nests. Focal upward migration of melanocytes into the epidermis was present in just 4 cases. All the lesions had cellular atypia, which was moderate in 85% of cases. The Ki-67 proliferation index was low (<5%) in all the lesions analyzed. ConclusionsAtypical lentiginous nevi, which can be classified as atypical pigmented lesions with a lentiginous pattern, may clinically and histologically resemble melanoma. Our findings support earlier reports that both clinical and histologic findings may suggest a diagnosis of dysplastic nevus. All of the patients in our series are healthy and free of recurrence after 18 months or longer.

Membrane‐bound Kit ligand regulates melanocyte adhesion and survival, providing physical interaction with an intraepithelial niche

Morphogenesis, as illustrated by melanocyte migration and homing to the skin, requires cadherin adhesion, integrin-dependent migration and Kit-ligand growth factor signaling. However, it is not known how Kit ligand regulates integrin or cadherin-dependent intraepidermal melanocyte behavior. To answer this question, we developed specific 2-dimensional (2D) and 3D culture systems analyzing how soluble or immobilized Kit-ligand-regulated melanocyte migration on vitronectin and laminin, or within a monolayer of kidney epithelial cells. In a 2D system, soluble Kit ligand stimulated integrin-dependent melanoblast migration and chemotaxis and accelerated integrin turnover. In contrast, immobilized, but not soluble, Kit ligand, enhanced integrin-dependent melanocyte spreading on suboptimal laminin concentrations. In 3D, membrane-bound Kit ligand induced intraepithelial melanocyte proliferation, survival, and tight adhesion to epithelial cells, while cleavable Kit ligand was less effective. In contrast, melanocyte motility was independent of membrane-bound Kit ligand, but increased in the presence of the cleavable Kit-ligand isoform. Transmembrane-dimerization or basolateral-targeting mutants of Kit ligand altered intraepithelial melanocyte localization, survival, and adhesion to epithelial cells. These data and the identification of c-kit/Kit-ligand clusters at cell contacts suggest that membrane-bound Kit ligand captures cell surface-expressed c-kit, providing mechanical anchoring and survival signaling within intraepithelial niches, and thereby defining a new mechanism for melanocyte homeostasis and requirement for environmental niches.

Oxidation Levels Differentially Impact Melanocytes: Low versus High Concentration of Hydrogen Peroxide Promotes Melanin Synthesis and Melanosome Transfer

Background: UVB light can generate potentially harmful hydrogen peroxide (H₂O₂) in vivo, but it can also promote the beneficial proliferation and migration of melanocytes. The successful use of UVB monotherapy for treatment of vitiligo suggests that H₂O₂ may have a biphasic effect on melanin synthesis and melanosome transfer. Objective: To study the beneficial role of H₂O₂ on melanogenesis and melanosome transport in living melanocytes and keratinocytes. Methods: A co-culture system model was constructed using the primary human melanocytes and keratinocytes. The MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay was used to determine cell proliferation, NaOH was used to determine the melanin content, and real-time PCR was used to determine tyrosinase expression. Western blot was used to determine Rab-27A and protease-activated receptor 2 (PAR-2) expression. Results: This study demonstrated that tyrosinase was activated by low concentrations of H₂O₂ (≤0.3 mM); however, this activity was downregulated by high concentrations of H₂O₂ (>0.3 mM). Activation of high levels of melanin synthesis was induced when cells were treated with low concentrations of H₂O₂ (0.3 mM). Further observation using an in vitro co-culture system of fluorescein (carboxyfluorescein diacetate succinimidyl ester, CFDA-SE)-labeled melanocytes and keratinocytes indicated that melanosome transfer occurred in normal human epidermal melanocytes. Fluorescence microscopy revealed increased melanosome transfer into keratinocytes treated with 0.3 mM H₂O₂ in the co-culture compared to the control. Examination of melanosomes in the keratinocytes by flow cytometry confirmed these results. Furthermore, treatment with H₂O₂ (0.3 mM) upregulated the expression of Rab-27A and PAR-2, significant proteins involved in melanosome transfer, according to Western blot. Conclusion: These results confirmed that low concentration levels of H₂O₂ play a major role in the regulation of human pigmentation by increasing melanin synthesis and melanosome transfer.

Phosphorylation of BRN2 Modulates Its Interaction with the Pax3 Promoter To Control Melanocyte Migration and Proliferation

MITF-M and PAX3 are proteins central to the establishment and transformation of the melanocyte lineage. They control various cellular mechanisms, including migration and proliferation. BRN2 is a POU domain transcription factor expressed in melanoma cell lines and is involved in proliferation and invasion, at least in part by regulating the expression of MITF-M and PAX3. The T361 and S362 residues of BRN2, both in the POU domain, are conserved throughout the POU protein family and are targets for phosphorylation, but their roles in vivo remain unknown. To examine the role of this phosphorylation, we generated mutant BRN2 in which these two residues were replaced with alanines (BRN2TS→BRN2AA). When expressed in melanocytes in vitro or in the melanocyte lineage in transgenic mice, BRN2TS induced proliferation and repressed migration, whereas BRN2AA repressed both proliferation and migration. BRN2TS and BRN2AA bound and repressed the MITF-M promoter, whereas PAX3 transcription was induced by BRN2TS but repressed by BRN2AA. Expression of the BRN2AA transgene in a Mitf heterozygous background and in a Pax3 mutant background enhanced the coat color phenotype. Our findings show that melanocyte migration and proliferation are controlled both through the regulation of PAX3 by nonphosphorylated BRN2 and through the regulation of MITF-M by the overall BRN2 level.

Sema4D, the Ligand for Plexin B1, Suppresses c-Met Activation and Migration and Promotes Melanocyte Survival and Growth

Semaphorins are secreted and membrane bound proteins involved in neural pathfinding, organogenesis, and tumor progression, through Plexin and neuropilins receptors. We recently reported that Plexin B1, the Semaphorin 4D receptor, is a tumor suppressor protein for melanoma, in part, through inhibition of the oncogenic c-Met tyrosine kinase receptor. In this report we show that Sema4D is a protective paracrine factor for normal human melanocyte survival in response to ultraviolet irradiation, that it stimulates proliferation, and regulates the activity of the c-Met receptor. c-Met receptor signaling stimulates melanocyte migration, in part through down-regulation of the cell adhesion molecule E-cadherin. Sema4D suppressed activation of c-Met in response to its ligand hepatocyte growth factor (HGF), and partially blocked the suppressive effects of HGF on E-cadherin expression in melanocytes and HGF-dependent migration. These data demonstrate a role for Plexin B1 in maintenance of melanocyte survival and proliferation in the skin, and suggest that Semaphorin 4D and Plexin B1 act cooperatively with HGF and c-Met to regulate c-Met dependent effects in human melanocytes. Because our data show that Plexin B1 is profoundly down-regulated by UVB in melanocytes, loss of Plexin B1 may accentuate HGF dependent effects on melanocytes, including melanocyte migration.

Extended long-segment Hirschsprungs’ disease in the Waardenburg–Shah syndrome

Hypopigmentation, either a white forelock or changes in the eyebrows and/or isochromia irides, associated with signs of bowel obstruction should alert the paediatrician and paediatric surgeons to the possibility of aganglionosis. We report a case of Waardenburg–Shah syndrome, which is a very rare congenital disorder with variable clinical expression, characterized by Hirschsprungs’ disease and abnormal melanocyte migration, resulting in pigmentary abnormalities and sensorineural deafness. Our patient had an exceptional association of extended long-segment aganglionosis and underwent a proximal ileostomy but did not survive until a definitive procedure was performed. Long-term prognosis of such children is associated with high morbidity and mortality. Keywords : extended long-segment Hirschsprungs’ disease, Waardenburg–Shah Syndrome, white forelock

Hair repigmentation associated with the use of lenalidomide: Graying may not be an irreversible process!

We report the first case of progressive hair repigmentation associated with the use of lenalidomide in an elderly patient with multiple myeloma. The influence of lenalidomide on follicular melanogenesis may involve removing the inhibitory influences of some cytokines such as IL-1, IL-6 and TNF-α. In addition, certain endocrine effects of lenalidomide on the hypophyseal-adrenal axis could explain its action on hair pigmentation. We further hypothesize that lenalidomide may be capable of stimulating migration and/or differentiation of melanocytes to promote repigmentation of gray hair follicles. Pending the clarification of how hair repigmentation occurs with lenalidomide, our observation materializes the concept that hair graying may not be an irreversible process, which opens avenues for targeted therapeutics in the fields of cosmetics and anti-aging medicine.

Nevus lentiginoso atípico. Estudio clínico-patológico de 14 casos

BackgroundAtypical lentiginous nevus (of the elderly) is a peculiar form of dysplastic nevus. Clinically, this condition can resemble malignant melanoma and histologically, it has a lentiginous pattern with variable degrees of atypia and an absence of dermal nests. These features may lead to an erroneous diagnosis of lentigo maligna melanoma or lentiginous melanoma. Material and methodsWe reviewed 14 cases of atypical lentiginous nevus diagnosed at the dermatology department of Hospital General de Valencia in Valencia, Spain between December 2007 and March 2009. We studied the clinical and histopathologic features of the lesions after hematoxylin-eosin, Melan-A, and Ki-67 staining and compared our results to data reported in the literature. ResultsFour (28%) of the 14 patients (7 men, 7 women) were under 50 years of age. Clinically, most of the lesions (8/14) resembled atypical nevi and they were all located on the back. Histologically, they all had irregular lentiginous epidermal hyperplasia, with a proliferation of individual melanocytes only in the basal layer of the epidermis and an absence of dermal nests. Focal upward migration of melanocytes into the epidermis was present in just 4 cases. All the lesions had cellular atypia, which was moderate in 85% of cases. The Ki-67 proliferation index was low (<5%) in all the lesions analyzed. ConclusionsAtypical lentiginous nevi, which can be classified as atypical pigmented lesions with a lentiginous pattern, may clinically and histologically resemble melanoma. Our findings support earlier reports that both clinical and histologic findings may suggest a diagnosis of dysplastic nevus. All of the patients in our series are healthy and free of recurrence after 18 months or longer.

Endothelin‐1 enhances the proliferation of normal human melanocytes in a paradoxical manner from the TNF‐α‐inhibited condition, but tacrolimus promotes exclusively the cellular migration without proliferation: a proposed action mechanism for combination therapy of phototherapy and topical tacrolimus in vitiligo treatment

Vitiligo is an acquired pigmentary disorder caused by the destruction of melanocytes. Two of the major theories regarding the pathogenesis of vitiligo are the autoimmune theory and autocytotoxicity theory, but, the precise pathogenetic mechanism is still not clarified. We investigated the effects of ET-1, tacrolimus and tumour necrosis factor-α (TNF-α) on proliferation and migration of cultured normal human melanocytes (NHMs). We also sought to clarify the theoretical rationale underlying the topical tacrolimus monotherapy or tacrolimus-UV combination therapy as tools for vitiligo treatment. The effects of ET-1, tacrolimus and TNF-α on proliferation/migration of cultured NHMs were investigated by MTT assay/Boyden chamber transwell migration assay. We also examined roles of CXC-chemokine receptor II (CXCR II) and matrix metalloproteinases (MMPs) in such conditions. ET-1 exerted a stimulatory effect on melanocyte proliferation and migration, but, tacrolimus exerted a stimulatory effect only on melanocyte migration higher than ET-1. TNF-α inhibited melanocyte proliferation in a dose-dependent manner. Paradoxically, TNF-α-pretreated NHMs exhibited an enhanced proliferative efficiency after being switched to ET-1. We found CXCRII was highly expressed in TNF-α-incubated melanocytes than the agents-free control, and ET-1 treatment after TNF-α preincubation showed the higher levels of CXCRII expression than the condition incubated with TNF-α alone. Moreover, the greater activities of MMP-2 and MMP-9 induced by tacrolimus than ET-1, reflected tacrolimus would enhance migration stimulatory effect in cultured NHMs. Topical tacrolimus can be used an effective agent for vitiligo treatment as monotherapy, maybe due to its migration stimulatory action or TNF-α inhibitory property, and also as a component in combination therapy with UV treatment, considering the more upregulated MMPs activities are induced and the more effective migrations are feasible by itself than ET-1.

The effects of NB-UVB on the hair follicle-derived neural crest stem cells differentiating into melanocyte lineage in vitro

Narrow-band UVB (NB-UVB) is an effective therapeutic option in the treatment of vitiligo. Despite the apparent clinical efficacy, the underlying mechanism of how topical NB-UVB induces repigmentation in vitiligo has not been clearly elucidated. To investigate the effects of NB-UVB on the maturation of melanocyte lineage differentiated from hair follicle-derived neural crest stem cells (HF-NCSCs) in vitro. HF-NCSCs were isolated from mouse whisker follicles. The isolated cells were multipotent and expressed embryonic NCSC biomarkers. The effects of NB-UVB on development and differentiation of HF-NCSCs were evaluated. We assessed cell viability, melanogenesis and migration of melanocytes derived from HF-NCSCs after NB-UVB radiation. Tyrosinase, Tyrp1, Dct, Kit, Mc1R, Fzd4, NT3R, Ednra, EP1, TGFβR, Sox10, Mitf, Lef1 and Pax3 gene expression was measured by quantitative RT-PCR, while Tyrosinase, Sox10 and Mitf protein expression were measured by Western blot analysis. Cell migration was measured by Boyden chamber transwell assay. NB-UVB increased the expression of tyrosinase during melanocytic differentiation from mouse HF-NCSCs, however, NB-UVB inhibited proliferation of melanocytes derived from HF-NCSCs. Mechanistically, increased melanocyte maturation after NB-UVB treatment was resulted from increased expression of several key melanogenic factors, including Sox10, Kit and Mc1R, which play a critical role to promote tyrosinase expression. Furthermore, the migration of the HF-NCSCs-derived melanocytes was downregulated as NB-UVB doses increased. However, the migration of HF-NCSCs was upregulated under 0.4J NB-UVB radiation. Those data provide in vitro evidence demonstrating some direct effects of NB-UVB on pigmentation of melanocyte lineage differentiated from HF-NCSCs, and may provide a possible mechanism for the effect of NB-UVB in vitiligo.

Constitutional Intraepidermal Ascent of Melanocytes

Transepidermal melanocytic migration (TEM) is an important diagnostic criterion for malignancy, especially in association with cytologic atypia. However, TEM may also be observed in benign melanocytic tumors, such as Spitz nevus, acral nevi, or nevi in infancy. We discuss the value of TEM for the diagnosis of melanocytic tumors in a young patient previously diagnosed as having 11 cutaneous melanomas. A 17-year-old patient with a history of 11 cutaneous melanomas diagnosed in the past 3 years by different expert dermatopathologists presented in our department. The previous histological diagnoses of melanoma were mainly based on the presence of important TEM. A reevaluation of all histological specimens in light of the clinical context and the lack of genomic aberrations as detected by array-comparative genomic hybridization led to a revision of the previous diagnoses. The striking TEM observed represents, in our opinion, a constitutional element of the melanocytic nevi in this patient and not a marker of malignancy. Awareness of this finding is important to avoid overdiagnosis of melanoma in cases of melanocytic nevi.

The mRNA expression profile of cytokines connected to the regulation of melanocyte functioning in vitiligo skin biopsy samples and peripheral blood mononuclear cells

The expression pattern of several genes associated with different processes in melanocytes, including melanogenesis, is changed in vitiligo patients. We evaluated possible changes in the expression of interleukin (IL)-10 family cytokines (IL26, IL-28A, IL28B, IL29), their receptor subunits (IL20RB, IL22RA2, IL28RA), and genes potentially related to functioning of melanocytes (MDM1, IFNA1, IFNB1, IFNG, and ICAM1) in the case of vitiligo. We observed mRNA expression in vitiligo patients' and controls' skin and peripheral blood mononuclear cells using quantitative real-time polymerase chain reaction. The mRNA expression pattern of IL20RB, IL22RA2, IL-28A, IL28B, IL28RA, MDM1, IFNA1, IFNB1, IFNG, and ICAM1 changed in vitiligo skin and/or peripheral blood mononuclear cells (PBMC) compared with controls. All of these genes may potentially be involved in vitiligo pathogenesis through controlling or participating in different pathways that regulate survival/apoptosis, development and migration of melanocytes, and melanogenesis. This study presents additional support for our previous findings about the importance of IL-10 family cytokines in vitiligo, in particular the possible involvement of IL-22. Further studies should be considered.

Migration of human melanocytes into keratinocyte monolayers in vitro

Migration of human melanocytes into keratinocyte monolayers in vitro

Co-culture of Melanocytes with Adipose-derived Stem Cells as a Potential Substitute for Co-culture with Keratinocytes

Cell-to-cell interactions between melanocytes and keratinocytes increase the proliferation and migration of melanocytes. In fact, mixed keratinocyte and melanocyte cultures have been used for autologous cell transplantation for treatment of vitiligo. However, this may require taking an amount of skin tissue large enough to leave scars. In this study, the in vitro effect of adipose-derived stem cells (ADSCs) on proliferation, differentiation and migration of melanocytes was compared with that of keratinocytes using immunohistochemistry and a Boyden chamber migration assay. The proliferation and migration of melanocytes was significantly stimulated by co-culture with ADSCs compared with melanocyte monocultures, al-though the effect of ADSCs was less powerful than that of keratinocytes. This may be related to increases in stem cell factor and basic fibroblast growth factor, growth factors for melanocytes, produced by the ADSCs. The ratios of melanocytes stained with antibodies against Trp-2, E-cadherin and N-cadherin were significantly increased by co-culturing with ADSCs compared with co-culturing with keratinocytes as well as melanocyte monocultures. The proportion of less-pigmented melanocytes was also increased and sustained for a longer duration in the presence of ADSCs. Our data show that co-culturing with ADSCs results in increased melanocyte proliferation and migration while reducing differentiation, and could provide a means to treat disorders such as vitiligo.

Plexin B1 inhibits MET through direct association and regulates Shp2 expression in melanocytes

Plexin B1, the receptor for Semaphorin 4D (Sema4D), is expressed by melanocytes in the skin. We recently showed that Sema4D suppresses activation of the hepatocyte growth factor receptor, MET, in melanocytes, and that knockdown of Plexin B1 results in activation of MET. MET signaling mediates proliferation, survival and migration in melanocytes, and its activation is associated with transformation of melanocytes to melanoma. In this report we investigated the mechanism by which Plexin B1 inhibits MET activation. Our results show that Plexin B1 and MET exist as an oligomeric receptor-receptor complex in melanocytes, and that receptor association is increased by Sema4D. MET and Plexin B1 receptor complexes were identified along the cell body of melanocytes, and Sema4D increased receptor association on dendrites, suggesting that Sema4D regulates MET-dependent processes at precise locations on the melanocyte. Despite activation of MET, Plexin B1 knockdowns proliferated slowly and showed increased apoptosis compared with controls. Shp2, a non-receptor protein tyrosine phosphatase, translates growth and survival signals from MET and other receptor tyrosine kinases. Plexin B1 knockdowns had markedly lower levels of Shp2 compared with controls, and Sema4D upregulated Shp2 expression at the protein and message level in normal melanocytes. Functional studies showed that blockade of Shp2 activity abrogated MET-dependent activation of Erk1/Erk2 and Akt in melanocytes. These results suggest a complex role for Sema4D and Plexin B1 in orchestrating signaling from the MET receptor in melanocytes. Because Shp2 is a downstream adaptor protein for multiple receptors, Sema4D may control the effects of several growth factors on melanocytes through regulation of Shp2.

Melanocyte Colonization and Pigmentation of Breast Carcinoma: Pathological and Clinical Aspects

Introduction. Melanocyte colonization of breast carcinoma by nonneoplastic melanocytes of epidermal origin is a rare and serious condition first described in 1977. We report on the exceptional clinical and pathological features of this migration phenomenon in a 74-year-old patient. Discussion. The pathogenesis by which melanocyte migration takes place is not known, but a breached basement membrane is considered essential. Conclusion. Histological examination and additional staining of skin are essential to differentiate breast cancer melanosis from malignant melanoma.

Ultraviolet Radiation and the Slug Transcription Factor Induce Proinflammatory and Immunomodulatory Mediator Expression in Melanocytes

Despite extensive investigation, the precise contribution of the ultraviolet radiation (UVR) component of sunlight to melanoma etiology remains unclear. UVR induces keratinocytes to secrete proinflammatory and immunomodulatory mediators that promote inflammation and skin tumor development; expression of the slug transcription factor in keratinocytes is required for maximal production of these mediators. In the present studies we examined the possibility that UVR-exposed melanocytes also produce proinflammatory mediators and that Slug is important in this process. Microarray studies revealed that both UVR exposure and Slug overexpression altered transcription of a variety of proinflammatory mediators by normal human melanocytes; some of these mediators are also known to stimulate melanocyte growth and migration. There was little overlap in the spectra of cytokines produced by the two stimuli. However IL-20 was similarly induced by both stimuli and the NFκB pathway appeared to be important in both circumstances. Further exploration of UVR-induced and Slug-dependent pathways of cytokine induction in melanocytes may reveal novel targets for melanoma therapy.

Coexisting Malignant Melanoma and Blue Nevus of the Uterine Cervix: An Unusual Combination

Malignant melanoma (MM) and blue nevi of the uterine cervix are an extremely rare neoplasm, probably derived from embryologic migration of melanocytes from the neural crest. MM displays aggressive behavior with a poor prognosis. We report the case of a 76-year-old postmenopausal woman abnormal vaginal bleeding. She underwent a hysterectomy and bilateral salpingo-oophorectomy with paraaortic-iliac lymphadenectomy. Histopathological and immunohistochemical studies were consistent with the diagnosis of MM and blue nevi in the uterine cervix. Although it is extremely rare, this case suggests that MM of the uterine cervix should be considered in the differential diagnosis of undifferentiated neoplasm. Early diagnosis is essential in order to warrant a better prognosis, although there are no cases of cure described.

P-REX1, a Rac guanine exchange factor, links melanocyte development and melanoma progression

P-REX1, a Rac guanine exchange factor, links melanocyte development and melanoma progression