Agouti protein, mahogunin, and attractin in pheomelanogenesis and melanoblast-like alteration of melanocytes: a cAMP-independent pathway

Melanocortin-1 receptor (MC1R) and its ligands, a-melanocyte stimulating hormone (aMSH) and agouti signaling protein (ASIP), regulate switching between eumelanin and pheomelanin synthesis in melanocytes. Here we investigated biological effects and signaling pathways of ASIP. Melan-a non agouti (a ⁄ a) mouse melanocytes produce mainly eumelanin, but ASIP combined with phenylthiourea and extra cysteine could induce over 200-fold increases in the pheomelanin to eumelanin ratio, and a tan-yellow color in pelletted cells.Moreover, ASIP-treated cells showed reduced proliferation and a melanoblast-like appearance, seen also in melanocyte lines from yellow (Ay ⁄ a and Mc1re ⁄ Mc1re) mice. However ASIP-YY, a C-terminal fragment of ASIP, induced neither biological nor pigmentary changes. As, like ASIP, ASIP-YY inhibited the cAMP rise induced by aMSH analog NDP-MSH, and reduced cAMP level without added MSH, the morphological changes and depigmentation seemed independent of cAMP signaling. Melanocytes genetically null for ASIP mediators attractin or mahogunin (Atrnmg-3J ⁄ mg-3J or Mgrn1md-nc ⁄ md-nc) also responded to both ASIP and ASIP-YY in cAMP level, while only ASIP altered their proliferation and (in part) shape. Thus, ASIP–MC1R signaling includes a cAMP-independent pathway through attractin and mahogunin, while the known cAMP-dependent component requires neither attractin nor mahogunin.Parts of the research were supported by Wellcome Trust program grants 064583 and 078327 to D.C.B. and E.V.S.; the Japan Society for the Promotion of Science KAKENHI (grants 20790808 to T.H. and 18591262 and 20591357 to K.W. and S.I.); a Grant-in-Aid from the Japanese Ministry of Health, Labour and Welfare (K.J.), the Spanish Ministry of Education and Science BFU2006-12185 (L.M.), the South West Academic Network (A.J.D. and E.V.S.), and NIH grant DK064265 (B.Y. and G.L.M.).Peer reviewe

Bird Integumentary Melanins: Biosynthesis, Forms, Function and Evolution

Melanins are the ubiquitous pigments distributed in nature. They are one of the main pigments responsible for colors in living cells. Birds are among the most diverse animals regarding melanin-based coloration, especially in the plumage, although they also pigment bare parts of the integument. This review is devoted to the main characteristics of bird melanins, including updated views of the formation and nature of melanin granules, whose interest has been raised in the last years for inferring the color of extinct birds and non-avian theropod dinosaurs using resistant fossil feathers. The molecular structure of the two main types of melanin, eumelanin and pheomelanin, and the environmental and genetic factors that regulate avian melanogenesis are also presented, establishing the main relationship between them. Finally, the special functions of melanin in bird feathers are also discussed, emphasizing the aspects more closely related to these animals, such as honest signaling, and the factors that may drive the evolution of pheomelanin and pheomelanin-based color traits, an issue for which birds have been pioneer study models.We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI).Ismael Galván was supported by a Ramón y Cajal Fellowship (ref. RYC-2012-10237) from the Spanish Ministry of Economy and Competitiveness. Francisco Solano belongs to the research group supported with the grant 19875/GERM/15 from Seneca Foundation, CARM, Spain.Peer reviewe

ヒト悪性黒色腫における細胞接着分子とインターロイキン2受容体の発現

This study investigated the expression of intercellular adhesion molecule-1 (ICAM-1), interleukin-2 receptor (IL-2R), integrins and E-cadherin (E-CD) in vivo and in vitro melanoma cells and their adhesion to extracellular matrix (ECM) as well as endothelial cells (HUVEC) in order to characterize the biological processes of melanoma invasion and metastasis. IL-2R was expressed in 21% of primary cutaneous melanoma, whereas 43% were positive in metastatic melanomas, suggesting that IL-2 may increase the growth of melanoma cells in certain subtypes. Integrin α2β1 was expressed highly in SK-MEL-118, and α2β1 was increased in a time- and dose-dependent manner by TPA treatment. Furthermore, the phorbol ester, TPA-induced α2β1 was mediated by calmodulin kinase, but not by protein kinase C (PKC). However, the adherence of SK-MEL-118 to HUVEC treated with TPA was decreased by 40%, suggesting that TPA affects a signaling mechanism. E-CD expression in primary melanoma did not show any correlation with the tumor invasion indexes. It was, however, indicated that primary melanoma cells adhere to each other through E-CD and that its expression correlates with invasion

正常ヒト表皮メラノサイト(NHEM)とヒト異色腫細胞株SK-MEL23の細胞増殖と分化に対する線維芽細胞培養液上清(FCM)の影響

Our goal is to study the role of dermal fibroblasts in the proliferation and differentiation of epidermal melanocytes using a reconstituted epidermal melanin unit (EMU). In order to achieve this goal, we have investigated the effect of fibroblast-conditioned medium (FCM) on the growth and melanogenesis of normal human epidermal melanocytes (NHEM) from neonatal foreskin using SK-MEL23 human melanoma cells as controls. NHEM and SK-MEL23 cells grown on type-I collagen gel alone revealed short dendrites. The culture on type-I collagen gel and fibroblasts resulted in prominent dendricities of NHEM. The FCM, which was likely to contain extracellular matrix (ECM) proteins and cytokines derived from fibroblasts, remarkably enhanced the dendricity of NHEM and their attachment to petri dishes/culture plates, but did so to a lesser degree with control SK-MEL23 cells. However, it did not affect the proliferation of both NHEM and SK-MEL23 cells. The FCM treatment decreased the tyrosinase activity of NHEM, while this decrease was not seen in melanoma cells. These studies suggest that cytokines and ECM proteins from dermal fibroblasts are important in regulating the functional and morphological differentiation of epidermal human melanocytes, and that this biological effect is much smaller in their neoplastic counter parts, melanoma cells

N-propionyl-4-S-Cysteaminylphenolのアポトーシスを介した色素細胞障害効果

Phenolic thioether amines, 4-S-cysteaminylphenol (4-S-CAP) and its N-acetyl derivative, N-acetyl-4-S-CAP, are tyrosinase substrates and potent depigmenting agents for skin and hair follicles. They also possess an antitumor effect on melanoma. However, it is not clear if their biological activity derives from a cytostatic or a cytocidal effect on melanocytes. We have recently synthesized a new derivative of 4-S-CAP, N-propionyl-4-S-CAP (NPr-CAP), in order to develop a better targeted chemotherapeutic or depigmenting agent. Structurally, NPr-CAP is more lipophilic than N-acetyl-4-S-CAP, hence it is expected to have better penetration into the cells and a more potent pharmacological effect. This study is aimed to identify the selectivity and potency of the biological effect of NPr-CAP on melanocytotoxicity. NPr-CAP was found to be a good tyrosinase substrate. The i. p. administration of NPr-CAP caused dose-dependent depigmentation of black hair follicles in C57 mice. However, the black hair follicles replaced the depigmented ones in the next hair cycle, suggesting the existence of dormant or drug-resistant melanocytes. Follicular melanocytes stained positively with fluorescein isothiocyanate by TUNEL (TdT-mediated dUTP-biotin nick end labeling) method, and by using the electron microscope, showed apoptotic changes as early as 12 h post-administration of NPr-CAP. However, surrounding keratinocytes and fibroblasts revealed no changes. NPr-CAP treatment induced irreversible inhibition of cell proliferation of melan-a2, an immortal melanocyte line of C57 black mice, but this did not occur in the control albino melanocyte line, melan-c. The agarose gel electrophoresis of DNA from drug-treated melan-a2 cells showed the nucleosomal DNA ladder pattern. The MTT assay and trypan blue test indicated that NPr-CAP can cause cell death of tyrosinase-positive melan-a2 cells, whereas, it transiently inhibits the proliferation of tyrosinase-negative melan-c cells. Thus, we propose that the melanocytotoxicity of NPr-CAP is selective and causes apoptosis in melanocytes through the interaction with active tyrosinase. (Received February 27, 1998 and accepted March 10, 1998

Growth Inhibition of Re-Challenge B16 Melanoma Transplant by Conjugates of Melanogenesis Substrate and Magnetite Nanoparticles as the Basis for Developing Melanoma-Targeted Chemo-Thermo-Immunotherapy

Melanogenesis substrate, N-propionyl-cysteaminylphenol (NPrCAP), is selectively incorporated into melanoma cells and inhibits their growth by producing cytotoxic free radicals. Magnetite nanoparticles also disintegrate cancer cells and generate heat shock protein (HSP) upon exposure to an alternating magnetic field (AMF). This study tested if a chemo-thermo-immunotherapy (CTI therapy) strategy can be developed for better management of melanoma by conjugating NPrCAP on the surface of magnetite nanoparticles (NPrCAP/M). We examined the feasibility of this approach in B16 mouse melanoma and evaluated the impact of exposure temperature, frequency, and interval on the inhibition of re-challenged melanoma growth. The therapeutic protocol against the primary transplanted tumor with or without AMF exposure once a day every other day for a total of three treatments not only inhibited the growth of the primary transplant but also prevented the growth of the secondary, re-challenge transplant. The heat-generated therapeutic effect was more significant at a temperature of 43°C than either 41°C or 46°C. NPrCAP/M with AMF exposure, instead of control magnetite alone or without AMF exposure, resulted in the most significant growth inhibition of the re-challenge tumor and increased the life span of the mice. HSP70 production was greatest at 43°C compared to that with 41°C or 46°C. CD8+T cells were infiltrated at the site of the re-challenge melanoma transplant

Rab27a: A Key to Melanosome Transport in Human Melanocytes

Normal pigmentation depends on the uniform distribution of melanin-containing vesicles, the melanosomes, in the epidermis. Griscelli syndrome (GS) is a rare autosomal recessive disease, characterized by an immune deficiency and a partial albinism that has been ascribed to an abnormal melanosome distribution. GS maps to 15q21 and was first associated with mutations in the myosin-V gene. However, it was demonstrated recently that GS can also be caused by a mutation in the Rab27a gene. These observations prompted us to investigate the role of Rab27a in melanosome transport. Using immunofluorescence and immunoelectron microscopy studies, we show that in normal melanocytes Rab27a colocalizes with melanosomes. In melanocytes isolated from a patient with GS, we show an abnormal melanosome distribution and a lack of Rab27a expression. Finally, reexpression of Rab27a in GS melanocytes restored melanosome transport to dendrite tips, leading to a phenotypic reversion of the diseased cells. These results identify Rab27a as a key component of vesicle transport machinery in melanocytes