RESUMEN
This study shows that prostaglandins in human FM55 melanoma cells and epidermal melanocytes are produced by COX-1. Prostaglandin production in FM55 melanoma cells was unrelated to that of melanin suggesting that the two processes can occur independently. Alpha-melanocyte-stimulating hormone, which had no effect on melanin production in FM55 cells, stimulated PGD(2) production in these cells without affecting PGE(2). While cAMP pathways may be involved in regulating PGD(2) production, our results suggest that alpha-MSH acts independently of cAMP, possibly by regulating the activity of lipocalin-type PGD synthase. This alpha-MSH-mediated effect may be associated with its role as an immune modulator.
Asunto(s)
Melaninas/metabolismo , Melanoma/metabolismo , Prostaglandinas D/metabolismo , Neoplasias Cutáneas/metabolismo , alfa-MSH/metabolismo , Línea Celular Tumoral , Células Cultivadas , AMP Cíclico/metabolismo , Ciclooxigenasa 1/metabolismo , Humanos , Oxidorreductasas Intramoleculares/metabolismo , Lipocalinas/metabolismo , Melanocitos/citología , Melanocitos/metabolismo , Melanoma/patología , Prostaglandina D2/metabolismo , Neoplasias Cutáneas/patologíaRESUMEN
Excessive ultraviolet radiation (UVR) exposure induces erythema, mediated in part by prostaglandin-E(2) (PGE(2)). While keratinocytes are a major PGE(2) source, epidermal melanocytes (EM) also express PGE(2)-production machinery. It is unclear whether EM-produced PGE(2) contributes to UVR-induced skin inflammation, and whether this is correlated with melanogenesis. Epidermal melanocytes were cultured from skin phototype-1 and -4 donors, followed by assessment of PGE(2) production and melanogenesis. Epidermal melanocytes expressed cytoplasmic phospholipase-A(2), cyclooxygenase-1, cytoplasmic prostaglandin-E synthase and microsomal prostaglandin-E synthase-1, -2. Epidermal melanocytes produced PGE(2) under basal conditions, which increased further after arachidonic acid stimulation. Epidermal melanocytes expressed cyclooxygenase-2 (COX-2) mRNA and a selective COX-2 inhibitor (NS-398) reduced PGE(2) production. Ultraviolet B-induced PGE(2) production was positively correlated with skin phototype-1, despite variability between individual EM donors. By contrast, there was no correlation between PGE(2) production by EM and their melanogenic status. Thus, EM may contribute to UVR-induced erythema, with role of donor skin phototype more important than their melanogenic status.
Asunto(s)
Dinoprostona/biosíntesis , Células Epidérmicas , Melaninas/biosíntesis , Melanocitos/metabolismo , Melanocitos/efectos de la radiación , Rayos Ultravioleta , Adulto , Ácido Araquidónico/farmacología , Femenino , Humanos , Peróxido de Hidrógeno/farmacología , Indometacina/farmacología , Oxidorreductasas Intramoleculares/metabolismo , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Queratinocitos/efectos de la radiación , Masculino , Melanocitos/efectos de los fármacos , Melanocitos/enzimología , Glicoproteínas de Membrana/metabolismo , Monofenol Monooxigenasa/metabolismo , Nitrobencenos , Oxidorreductasas/metabolismo , Sulfonamidas , Adulto JovenRESUMEN
Sunburn is a commonly occurring acute inflammatory process, with dermal vasodilatation and leukocyte infiltration as central features. Ultraviolet (UV) B-induced hydrolysis of membrane phospholipids releases polyunsaturated fatty acids, and their subsequent metabolism by cyclooxygenases (COXs) and lipoxygenases (LOXs) may produce potent eicosanoid mediators modulating different stages of the inflammation. Our objective was to identify candidate eicosanoids formed during the sunburn reaction in relation to its clinical and histological course. We exposed skin of healthy humans (n=32) to UVB and, for 72 h, examined expression of proinflammatory and anti-inflammatory eicosanoids using LC/ESI-MS/MS, and examined immunohistochemical expression of COX-2, 12-LOX, 15-LOX, and leukocyte markers, while quantifying clinical erythema. We show that vasodilatory prostaglandins (PGs) PGE(2), PGF(2alpha), and PGE(3) accompany the erythema in the first 24-48 h, associated with increased COX-2 expression at 24 h. Novel, potent leukocyte chemoattractants 11-, 12-, and 8-monohydroxy-eicosatetraenoic acid (HETE) are elevated from 4 to 72 h, in association with peak dermal neutrophil influx at 24 h, and increased dermal CD3(+) lymphocytes and 12- and 15-LOX expression from 24 to 72 h. Anti-inflammatory metabolite 15-HETE shows later expression, peaking at 72 h. Sunburn is characterized by overlapping sequential profiles of increases in COX products followed by LOX products that may regulate subsequent events and ultimately its resolution.
Asunto(s)
Eicosanoides/metabolismo , Quemadura Solar/fisiopatología , Adulto , Alprostadil/análogos & derivados , Alprostadil/biosíntesis , Complejo CD3/metabolismo , Ciclooxigenasa 2/biosíntesis , Dinoprost/biosíntesis , Dinoprostona/biosíntesis , Eritema/metabolismo , Femenino , Humanos , Ácidos Hidroxieicosatetraenoicos/metabolismo , Lipooxigenasa/metabolismo , Masculino , Persona de Mediana Edad , Infiltración Neutrófila , Piel/efectos de la radiación , Espectrometría de Masa por Ionización de Electrospray , Quemadura Solar/metabolismo , Espectrometría de Masas en Tándem , Rayos UltravioletaRESUMEN
Proopiomelanocortin (POMC) can be processed to ACTH and melanocortin peptides. However, processing is incomplete in some tissues, leading to POMC precursor release from cells. This study examined POMC processing in human skin and the effect of POMC on the melanocortin-1 receptor (MC-1R) and melanocyte regulation. POMC was secreted by both human epidermal keratinocytes (from 5 healthy donors) and matched epidermal melanocytes in culture. Much lower levels of alpha-MSH were secreted and only by the keratinocytes. Neither cell type released ACTH. Cell extracts contained significantly more ACTH than POMC, and alpha-MSH was detected only in keratinocytes. Nevertheless, the POMC processing components, prohormone convertases 1, 2 and regulatory protein 7B2, were detected in melanocytes and keratinocytes. In contrast, hair follicle melanocytes secreted both POMC and alpha-MSH, and this was enhanced in response to corticotrophin-releasing hormone (CRH) acting primarily through the CRH receptor 1. In cells stably transfected with the MC-1R, POMC stimulated cAMP, albeit with a lower potency than ACTH, alpha-MSH, and beta-MSH. POMC also increased melanogenesis and dendricity in human pigment cells. This release of POMC from skin cells and its functional activity at the MC-1R highlight the importance of POMC processing as a key regulatory event in the skin.
Asunto(s)
Queratinocitos/metabolismo , Melanocitos/citología , Melanocitos/metabolismo , Proopiomelanocortina/metabolismo , Piel/citología , Hormona Adrenocorticotrópica/análisis , Células Cultivadas , Humanos , Queratinocitos/química , Melanocortinas/análisis , Melanocitos/química , Proopiomelanocortina/fisiología , Receptor de Melanocortina Tipo 1 , alfa-MSH/análisisRESUMEN
The proopiomelanocortin (POMC)-derived peptides, ACTH and alpha-MSH, are the principal mediators of human skin pigmentation via their action at the melanocortin-1 receptor (MC-1R). Recent data have demonstrated the existence of a functionally active beta-endorphin/mu-opiate receptor system in both epidermal and hair follicle melanocytes, whereby beta-endorphin can regulate melanogenesis, dendricity, and proliferation in these cells. However, a role for ACTH and alpha-MSH in the regulation of the human follicular pigmentary unit has not been determined. This study was designed to examine the involvement of ACTH and the alpha-MSH/MC-1R system in human follicular melanocyte biology. To address this question we employed RT-PCR and immunohisto/cytochemistry, and a functional role for these POMC peptides was assessed in follicular melanocyte cultures. Human scalp hair follicle melanocytes synthesized and processed POMC. ACTH and alpha-MSH in association with their processing enzymes and MC-1R are expressed in human follicular melanocytes at the message level in vitro and at the protein level both in situ and in vitro. The expression of the POMC/MC-1R receptor system was confined only to subpopulations of poorly and moderately differentiated melanocytes. In addition, functional studies revealed that ACTH and alpha-MSH are able to promote follicular melanocyte differentiation by up-regulating melanogenesis, dendricity, and proliferation in less differentiated melanocyte subpopulations. Thus, these findings suggest a role for these POMC peptides in regulating human hair follicle melanocyte differentiation.
Asunto(s)
Folículo Piloso/citología , Melanocitos/citología , Melanocitos/fisiología , Proopiomelanocortina/genética , alfa-MSH/genética , Hormona Adrenocorticotrópica/metabolismo , Adulto , Diferenciación Celular/fisiología , Células Cultivadas , Femenino , Humanos , Persona de Mediana Edad , Proteínas del Tejido Nervioso/genética , Proteína 7B2 Secretora Neuroendocrina , Hormonas Hipofisarias/genética , Proopiomelanocortina/metabolismo , Proproteína Convertasa 1/genética , Proproteína Convertasa 2/genética , ARN Mensajero/análisis , Receptor de Melanocortina Tipo 1/genética , Cuero Cabelludo/citología , alfa-MSH/metabolismoRESUMEN
Prostaglandins are potent mediators of the inflammatory response and are also involved in cancer development. In this study, we show that human melanocytes and FM55 melanoma cells express cyclooxygenase-1 and -2 (COX-1 and -2) and thus have the capability to produce prostaglandins. The FM55 cells produced predominantly PGE2 and PGF2alpha, whereas the HaCaT keratinocyte cell line produced mainly PGE2. The anti-inflammatory peptide, alpha-melanocyte stimulating hormone (alpha-MSH), reduced prostaglandin production in FM55 and HaCaT cells and reversed the effect of the pro-inflammatory cytokine TNF-alpha in the former. These results indicate that melanocytes produce prostaglandins and that alpha-MSH, by inhibiting this response, may play an important role in regulating inflammatory responses in the skin.
Asunto(s)
Melanocitos/metabolismo , Prostaglandinas/biosíntesis , alfa-MSH/metabolismo , Línea Celular , Supervivencia Celular , Células Cultivadas , Cromatografía Líquida de Alta Presión , Ciclooxigenasa 1 , Ciclooxigenasa 2 , Humanos , Inflamación , Isoenzimas/metabolismo , Queratinocitos/metabolismo , Proteínas de la Membrana , Fenotipo , Prostaglandina-Endoperóxido Sintasas/metabolismo , Prostaglandinas/metabolismo , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
The pro-opiomelanocortin (POMC)-derived peptides, alpha-melanocyte-stimulating hormone, and adrenocorticotropic hormone, are important mediators of human skin pigmentation via action at the melanocortin-1 receptor. Recent data suggests that such a regulatory role also exists for the endogenous opiate, beta-endorphin (beta-END). A role for this beta-END in the regulation of follicular pigmentation, however, has not been determined. This study was designed to examine the involvement of the beta-END/mu-opiate receptor system in human follicular melanocyte biology. We employed RT-PCR, and immunohisto/cytochemistry and immunoelectron microscopy using beta-END and mu-opiate receptor specific antibodies and a functional role for beta-END was assessed by direct stimulation with the peptide. This study has demonstrated that human hair follicle melanocytes (HFM) express mRNA for the mu-opiate receptor and POMC. Furthermore, beta-END and its high affinity mu-opiate receptor are expressed at the protein level in glycoprotein100-positive follicular melanocytes and as a function of their anatomic location and differentiation status during the hair growth cycle. Functional studies revealed that beta-END is a modifier of HFM phenotype via its ability to upregulate melanogenesis, dendricity, and proliferation. These findings suggest a new regulatory role for beta-END in human HFM biology, providing a new research direction into the fundamental regulation of human hair pigmentation.
Asunto(s)
Folículo Piloso/fisiología , Melanocitos/fisiología , Pigmentación de la Piel/fisiología , betaendorfina/fisiología , Adulto , División Celular/fisiología , Tamaño de la Célula/fisiología , Células Cultivadas , Femenino , Expresión Génica/fisiología , Folículo Piloso/citología , Folículo Piloso/crecimiento & desarrollo , Humanos , Melanocitos/citología , Melanosomas/fisiología , Persona de Mediana Edad , Proopiomelanocortina/genética , Proopiomelanocortina/metabolismo , Receptores Opioides/genética , Receptores Opioides/metabolismo , Receptores Opioides mu/genéticaRESUMEN
Melanocortin receptor type 1 (MC-1R) is an important control point for ultraviolet ray (UVR)-induced tanning response in the skin. In this study, we show that p-locus is a downstream target for MC-1R signaling. The expression of p-locus was up-regulated by alpha-MSH as well as db-cAMP, a synthetic analogue of cAMP that mimics activation of MC-1R. Furthermore, p-locus transcript abundance was significantly increased in epidermal melanocytes of white skin with facultative (UVR-induced) pigmentation. Because p-locus product is essential for pigmentation and also has been shown to be highly polymorphic in human population, we propose that the pigmentary response to the melanocortin peptides/UVR would be affected not only by MC-1R mutations but also by the functionality of p-locus product. These factors together could account for the many different levels of tanning ability seen in the white population.
Asunto(s)
Regulación de la Expresión Génica , Receptores de Corticotropina/metabolismo , Transducción de Señal/fisiología , Pigmentación de la Piel/fisiología , alfa-MSH/metabolismo , Bucladesina/metabolismo , Humanos , Receptores de Corticotropina/genética , Receptores de Melanocortina , Piel/citología , Piel/metabolismo , Piel/efectos de la radiación , Rayos UltravioletaRESUMEN
beta-Endorphin is an opioid peptide cleaved from the precursor pro-hormone pro-opiomelanocortin, from which other peptides such as adrenocorticotropic hormone, beta-lipotropic hormone, and alpha-melanocyte-stimulating hormone are also derived. alpha-Melanocyte-stimulating hormone and adrenocorticotropic hormone are well documented to regulate human skin pigmentation via action at the melanocortin-1 receptor. Whereas plasma beta-endorphin is reported to increase after exposure to ultraviolet radiation, to date a functional role for beta-endorphin in the regulation of human epidermal melanocyte biology has not been demonstrated. This study was designed to examine the involvement of the beta-endorphin/mu-opiate receptor system in human epidermal melanocytes. To address this question we employed reverse transcription-polymerase chain reaction, and immunohistochemistry/cytochemistry and immunoelectron microscopy using beta-endorphin and mu-opiate receptor specific antibodies. A functional role for beta-endorphin was assessed in epidermal melanocyte cultures by direct stimulation with the peptide. This study demonstrated the expression of mu-opiate receptor mRNA in cultured epidermal melanocytes, as well as mRNA for pro-opiomelanocortin. In addition, we have shown that beta-endorphin and mu-opiate receptor are expressed at the protein level in situ in glycoprotein100-positive melanocytes. The expression of both beta-endorphin and mu-opiate receptor correlated positively with their differentiation status in vitro. Furthermore, immunoelectron microscopy studies revealed an association of beta-endorphin with melanosomes. Functional studies showed that beta-endorphin has potent melanogenic, mitogenic, and dendritogenic effects in cultured epidermal melanocytes deprived of any exogenous supply of pro-opiomelanocortin peptides. Thus, we report that human epidermal melanocytes express a fully functioning beta-endorphin/mu-opiate receptor system. In the absence of any data showing cross-talk between the mu-opiate receptor and the melanocortin-1 receptor, we conclude that the beta-endorphin/mu-opiate receptor system participates in the regulation of skin pigmentation.
Asunto(s)
Células Epidérmicas , Melanocitos/fisiología , betaendorfina/fisiología , División Celular/efectos de los fármacos , Células Cultivadas , Dendritas/ultraestructura , Humanos , Queratinocitos/metabolismo , Melaninas/biosíntesis , Melanocitos/efectos de los fármacos , Melanocitos/ultraestructura , Melanosomas/metabolismo , Proopiomelanocortina/genética , ARN Mensajero/metabolismo , Receptores Opioides mu/genética , Distribución Tisular , betaendorfina/metabolismo , betaendorfina/farmacologíaRESUMEN
In this study, we have demonstrated the presence of melanin-concentrating hormone (MCH) and melanin-concentrating hormone receptor (MCHR1) transcripts in human skin. Sequence analysis confirmed that the transcripts of both genes were identical to those previously found in human brain. In culture, endothelial cells showed pro-MCH expression whereas no signal was found in keratinocytes, melanocytes, and fibroblasts. MCHR1 expression was restricted to melanocytes and melanoma cells. Stimulation of cultured human melanocytes with MCH reduced the alpha-MSH-induced increase in cAMP production. Furthermore, the melanogenic actions of alpha-MSH were inhibited by MCH. We propose that the MCH/MCHR1 signalling system is present in human skin and may have a role with the melanocortins in regulating the melanocyte.
Asunto(s)
Hormonas Hipotalámicas/biosíntesis , Hormonas Hipotalámicas/fisiología , Melaninas/biosíntesis , Melaninas/fisiología , Hormonas Hipofisarias/biosíntesis , Hormonas Hipofisarias/fisiología , Receptores de la Hormona Hipofisaria/biosíntesis , Receptores de la Hormona Hipofisaria/fisiología , Piel/metabolismo , Células Cultivadas , AMP Cíclico/biosíntesis , Relación Dosis-Respuesta a Droga , Humanos , Hormonas Hipotalámicas/genética , Hormonas Hipotalámicas/farmacología , Melaninas/genética , Melaninas/farmacología , Melanocitos/efectos de los fármacos , Melanocitos/metabolismo , Hormonas Hipofisarias/genética , Hormonas Hipofisarias/farmacología , ARN Mensajero/biosíntesis , Receptores de la Hormona Hipofisaria/genética , Piel/citología , Transcripción Genética , Células Tumorales Cultivadas , alfa-MSH/antagonistas & inhibidoresRESUMEN
Melanocytes are cells of neural crest origin. In the human epidermis, they form a close association with keratinocytes via their dendrites. Melanocytes are well known for their role in skin pigmentation, and their ability to produce and distribute melanin has been studied extensively. One of the factors that regulates melanocytes and skin pigmentation is the locally produced melanocortin peptide alpha-MSH. The effects of alpha-MSH on melanogenesis are mediated via the MC-1R and tyrosinase, the rate-limiting enzyme in the melanogenesis pathway. Binding of alpha-MSH to its receptor increases tyrosinase activity and eumelanin production, which accounts for the skin-darkening effect of alpha-MSH. Other alpha-MSH-related melanocortin peptides, such as ACTH1-17 and desacetylated alpha-MSH, are also agonists at the MC-1R and could regulate melanocyte function. Recent evidence shows that melanocytes have other functions in the skin in addition to their ability to produce melanin. They are able to secrete a wide range of signal molecules, including cytokines, POMC peptides, catecholamines, and NO in response to UV irradiation and other stimuli. Potential targets of these secretory products are keratinocytes, lymphocytes, fibroblasts, mast cells, and endothelial cells, all of which express receptors for these signal molecules. Melanocytes may therefore act as important local regulators of a range of skin cells. It has been shown that alpha-MSH regulates NO production from melanocytes, and it is possible that the melanocortins regulate the release of other signalling molecules from melanocytes. Therefore, the melanocortin signaling system is one of the important regulators of skin homeostasis.