RESUMEN
Epstein-Barr virus-induced gene 2 (EBI2, also known as GPR183) is a G-protein-coupled receptor that is required for humoral immune responses; polymorphisms in the receptor have been associated with inflammatory autoimmune diseases. The natural ligand for EBI2 has been unknown. Here we describe the identification of 7α,25-dihydroxycholesterol (also called 7α,25-OHC or 5-cholesten-3ß,7α,25-triol) as a potent and selective agonist of EBI2. Functional activation of human EBI2 by 7α,25-OHC and closely related oxysterols was verified by monitoring second messenger readouts and saturable, high-affinity radioligand binding. Furthermore, we find that 7α,25-OHC and closely related oxysterols act as chemoattractants for immune cells expressing EBI2 by directing cell migration in vitro and in vivo. A critical enzyme required for the generation of 7α,25-OHC is cholesterol 25-hydroxylase (CH25H). Similar to EBI2 receptor knockout mice, mice deficient in CH25H fail to position activated B cells within the spleen to the outer follicle and mount a reduced plasma cell response after an immune challenge. This demonstrates that CH25H generates EBI2 biological activity in vivo and indicates that the EBI2-oxysterol signalling pathway has an important role in the adaptive immune response.
Asunto(s)
Hidroxicolesteroles/farmacología , Receptores de Superficie Celular/inmunología , Animales , Formación de Anticuerpos/inmunología , Linfocitos B , Línea Celular , Movimiento Celular/efectos de los fármacos , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/inmunología , Humanos , Hidroxicolesteroles/química , Hígado/química , Ratones , Ratones Noqueados , Receptores Acoplados a Proteínas G , Ovinos , Linfocitos T/inmunologíaRESUMEN
TLQP-21, a peptide derived from VGF (non-acronymic) by proteolytic processing, has been shown to modulate energy metabolism, differentiation, and cellular response to stress. Although extensively investigated, the receptor for this endogenous peptide has not previously been described. This study describes the use of a series of studies that show G protein-coupled receptor-mediated biological activity of TLQP-21 signaling in CHO-K1 cells. Unbiased genome-wide sequencing of the transcriptome from responsive CHO-K1 cells identified a prioritized list of possible G protein-coupled receptors bringing about this activity. Further experiments using a series of defined receptor antagonists and siRNAs led to the identification of complement C3a receptor-1 (C3AR1) as a target for TLQP-21 in rodents. We have not been able to demonstrate so far that this finding is translatable to the human receptor. Our results are in line with a large number of physiological observations in rodent models of food intake and metabolic control, where TLQP-21 shows activity. In addition, the sensitivity of TLQP-21 signaling to pertussis toxin is consistent with the known signaling pathway of C3AR1. The binding of TLQP-21 to C3AR1 not only has effects on signaling but also modulates cellular functions, as TLQP-21 was shown to have a role in directing migration of mouse RAW264.7 cells.
Asunto(s)
Movimiento Celular/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Receptores de Complemento/metabolismo , Transducción de Señal/efectos de los fármacos , Animales , Secuencia de Bases , Células CHO , Cricetinae , Cricetulus , Estudio de Asociación del Genoma Completo , Células HEK293 , Humanos , Ratones , Datos de Secuencia Molecular , Factores de Crecimiento Nervioso/farmacología , Neuropéptidos/farmacología , Toxina del Pertussis/farmacología , Ratas , Receptores de Complemento/agonistas , Receptores de Complemento/genética , Especificidad de la Especie , Transcriptoma/efectos de los fármacosRESUMEN
BACKGROUND: The proton-activated G protein-coupled receptor GPR4 is expressed in many tissues including white adipose tissue. GPR4 is activated by extracellular protons in the physiological pH range (i.e. pH 7.7 - 6.8) and is coupled to the production of cAMP. METHODS: We examined mice lacking GPR4 and examined glucose tolerance and insulin sensitivity in young and aged mice as well as in mice fed with a high fat diet. Expression profiles of pro- and anti-inflammatory cytokines in white adipose tissue, liver and skeletal muscle was assessed. RESULTS: Here we show that mice lacking GPR4 have an improved intraperitoneal glucose tolerance test and increased insulin sensitivity. Insulin levels were comparable but leptin levels were increased in GPR4 KO mice. Gpr4-/- showed altered expression of PPARa, IL-6, IL-10, TNFa, and TGF-1b in skeletal muscle, white adipose tissue, and liver. High fat diet abolished the differences in glucose tolerance and insulin sensitivity between Gpr4+/+ and Gpr4-/- mice. In contrast, in aged mice (12 months old), the positive effect of GPR4 deficiency on glucose tolerance and insulin sensitivity was maintained. Liver and adipose tissue showed no major differences in the mRNA expression of pro- and anti-inflammatory factors between aged mice of both genotypes. CONCLUSION: Thus, GPR4 deficiency improves glucose tolerance and insulin sensitivity. The effect may involve an altered balance between pro- and anti-inflammatory factors in insulin target tissues.
Asunto(s)
Resistencia a la Insulina/fisiología , Proteína Disulfuro Isomerasas/metabolismo , Tejido Adiposo Blanco/metabolismo , Envejecimiento , Animales , Dieta Alta en Grasa , Regulación de la Expresión Génica , Glucosa/metabolismo , Prueba de Tolerancia a la Glucosa , Homeostasis , Concentración de Iones de Hidrógeno , Interleucina-10/genética , Interleucina-6/genética , Hígado/metabolismo , Ratones , Ratones Noqueados , Músculo Esquelético/metabolismo , Obesidad/etiología , Obesidad/genética , PPAR alfa/genética , Proteína Disulfuro Isomerasas/genética , Protones , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
The G protein-coupled receptor GPR4 is activated by acidic pH and recent evidence indicates that it is expressed in endothelial cells. In agreement with these reports, we observe a high correlation of GPR4 mRNA expression with endothelial marker genes, and we confirm expression and acidic pH dependent function of GPR4 in primary human vascular endothelial cells. GPR4-deficient mice were generated; these are viable and fertile and show no gross abnormalities. However, these animals show a significantly reduced angiogenic response to VEGF (vascular endothelial growth factor), but not to bFGF (basic fibroblast growth factor), in a growth factor implant model. Accordingly, in two different orthotopic models, tumor growth is strongly reduced in mice lacking GPR4. Histological analysis of tumors indicates reduced tumor cell proliferation as well as altered vessel morphology, length and density. Moreover, GPR4 deficiency results in reduced VEGFR2 (VEGF Receptor 2) levels in endothelial cells, accounting, at least in part, for the observed phenotype. Our data suggest that endothelial cells sense local tissue acidosis via GPR4 and that this signal is required to generate a full angiogenic response to VEGF.
Asunto(s)
Células Endoteliales/metabolismo , Neoplasias/fisiopatología , Neovascularización Patológica/metabolismo , Receptores Acoplados a Proteínas G/deficiencia , Receptores Acoplados a Proteínas G/metabolismo , Animales , Western Blotting , Proliferación Celular , Biología Computacional , Cartilla de ADN/genética , Electroforesis en Gel de Poliacrilamida , Ensayo de Inmunoadsorción Enzimática , Femenino , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Células HeLa , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Neoplasias/metabolismo , ARN Interferente Pequeño/genética , Receptores Acoplados a Proteínas G/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factor A de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Bleomycin (BLM) induced lung injury is detectable in C57BL/6 mice using magnetic resonance imaging (MRI). We investigated the effects of the fibroblast activation protein (FAP) inhibitor, PT100, in this model. BLM (0.5mg/kg/day) was administered on days -7, -6, -5, -2, -1, 0 in the nostrils of male mice. PT100 (40µg/mouse) or vehicle (0.9%NaCl) was dosed per os twice daily from day 1-14. MRI was performed before BLM and at days 0, 7 and 14. After the last MRI acquisition, animals were euthanised and the lungs harvested for histological and quantitative real-time polymerase chain reaction (qRT-PCR) analyses. As evidenced longitudinally by MRI, the BLM-elicited lesions in the lungs of vehicle-treated mice progressed over time. In contrast, responses elicited by BLM did not progress in animals receiving PT100. Histology demonstrated significant less fibrosis in PT100- than in vehicle-treated, BLM-challenged mice. Significant correlation (R=0.91, P<0.001, N=24) was found between the volumes of BLM-induced lesions detected in vivo by MRI and the collagen content determined histologically (picrosirius staining). FAP was overexpressed in the lungs of BLM-challenged mice. Upon PT100 treatment, FAP expression was reduced. Significant differences in the MMP-12, MIP-1α, and MCP-3 mRNA expression levels in the lungs of PT100- compared to vehicle-treated mice were also revealed by qRT-PCR. The IBA-1 level determined histologically was higher in the lungs of PT100- compared to vehicle-treated mice. Taken together, these observations suggest that treatment with PT100 in this murine model of pulmonary fibrosis had an anti-fibro-proliferative effect and increased macrophage activation.
Asunto(s)
Ácidos Borónicos/farmacología , Dipéptidos/farmacología , Gelatinasas/antagonistas & inhibidores , Proteínas de la Membrana/antagonistas & inhibidores , Fibrosis Pulmonar/tratamiento farmacológico , Animales , Bleomicina/efectos adversos , Peso Corporal/efectos de los fármacos , Ácidos Borónicos/uso terapéutico , Dipéptidos/uso terapéutico , Modelos Animales de Enfermedad , Endopeptidasas , Regulación de la Expresión Génica/efectos de los fármacos , Imagen por Resonancia Magnética , Masculino , Ratones Endogámicos C57BL , Fibrosis Pulmonar/diagnóstico por imagen , Fibrosis Pulmonar/genética , Serina EndopeptidasasRESUMEN
GPR15 is an orphan G protein-coupled receptor (GPCR) that is found in lymphocytes. It functions as a co-receptor of simian immunodeficiency virus and HIV-2 and plays a role in the trafficking of T cells to the lamina propria in the colon and to the skin. We describe the purification from porcine colonic tissue extracts of an agonistic ligand for GPR15 and its functional characterization. In humans, this ligand, which we named GPR15L, is encoded by the gene C10ORF99 and has some features similar to the CC family of chemokines. GPR15L was found in some human and mouse epithelia exposed to the environment, such as the colon and skin. In humans, GPR15L was also abundant in the cervix. In skin, GPR15L was readily detected after immunologic challenge and in human disease, for example, in psoriatic lesions. Allotransplantation of skin from Gpr15l-deficient mice onto wild-type mice resulted in substantial graft protection, suggesting nonredundant roles for GPR15 and GPR15L in the generation of effector T cell responses. Together, these data identify a receptor-ligand pair that is required for immune homeostasis at epithelia and whose modulation may represent an alternative approach to treating conditions affecting the skin such as psoriasis.
Asunto(s)
Colon/inmunología , Mucosa Intestinal/inmunología , Receptores Acoplados a Proteínas G/inmunología , Piel/inmunología , Linfocitos T/inmunología , Aloinjertos , Animales , Colon/citología , Femenino , Humanos , Mucosa Intestinal/citología , Ratones , Receptores Acoplados a Proteínas G/genética , Piel/citología , Trasplante de Piel , Porcinos , Linfocitos T/citología , Inmunología del TrasplanteRESUMEN
BACKGROUND: A novel family of proton-sensing G protein-coupled receptors, including OGR1, GPR4, and TDAG8, was identified to be important for physiological pH homeostasis and inflammation. Thus, we determined the function of proton-sensing OGR1 in the intestinal mucosa. MTEHODS: OGR1 expression in colonic tissues was investigated in controls and patients with IBD. Expression of OGR1 upon cell activation was studied in the Mono Mac 6 (MM6) cell line and primary human and murine monocytes by real-time PCR. Ogr1 knockout mice were crossbred with Il-10 deficient mice and studied for more than 200 days. Microarray profiling was performed using Ogr1 and Ogr1 (WT) residential peritoneal macrophages. RESULTS: Patients with IBD expressed higher levels of OGR1 in the mucosa than non-IBD controls. Treatment of MM6 cells with TNF, led to significant upregulation of OGR1 expression, which could be reversed by the presence of NF-κB inhibitors. Kaplan-Meier survival analysis showed a significantly delayed onset and progression of rectal prolapse in female Ogr1/Il-10 mice. These mice displayed significantly less rectal prolapses. Upregulation of gene expression, mediated by OGR1, in response to extracellular acidification in mouse macrophages was enriched for inflammation and immune response, actin cytoskeleton, and cell-adhesion gene pathways. CONCLUSIONS: OGR1 expression is induced in cells of human macrophage lineage and primary human monocytes by TNF. NF-κB inhibition reverses the induction of OGR1 expression by TNF. OGR1 deficiency protects from spontaneous inflammation in the Il-10 knockout model. Our data indicate a pathophysiological role for pH-sensing receptor OGR1 during the pathogenesis of mucosal inflammation.
Asunto(s)
Enfermedades Inflamatorias del Intestino/metabolismo , Mucosa Intestinal/metabolismo , Intestinos/patología , Receptores Acoplados a Proteínas G/metabolismo , Equilibrio Ácido-Base/fisiología , Animales , Línea Celular , Femenino , Humanos , Enfermedades Inflamatorias del Intestino/patología , Interleucina-10 , Mucosa Intestinal/patología , Macrófagos Peritoneales/metabolismo , Masculino , Ratones , Ratones NoqueadosRESUMEN
Blood gas and tissue pH regulation depend on the ability of the brain to sense CO2 and/or H(+) and alter breathing appropriately, a homeostatic process called central respiratory chemosensitivity. We show that selective expression of the proton-activated receptor GPR4 in chemosensory neurons of the mouse retrotrapezoid nucleus (RTN) is required for CO2-stimulated breathing. Genetic deletion of GPR4 disrupted acidosis-dependent activation of RTN neurons, increased apnea frequency, and blunted ventilatory responses to CO2. Reintroduction of GPR4 into RTN neurons restored CO2-dependent RTN neuronal activation and rescued the ventilatory phenotype. Additional elimination of TASK-2 (K(2P)5), a pH-sensitive K(+) channel expressed in RTN neurons, essentially abolished the ventilatory response to CO2. The data identify GPR4 and TASK-2 as distinct, parallel, and essential central mediators of respiratory chemosensitivity.
Asunto(s)
Dióxido de Carbono/fisiología , Canales de Potasio de Dominio Poro en Tándem/fisiología , Receptores Acoplados a Proteínas G/fisiología , Respiración , Cuerpo Trapezoide/fisiología , Acidosis Respiratoria/genética , Acidosis Respiratoria/fisiopatología , Animales , Femenino , Eliminación de Gen , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Mutantes , Neuronas/metabolismo , Neuronas/fisiología , Canales de Potasio de Dominio Poro en Tándem/genética , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/genética , Cuerpo Trapezoide/citología , Cuerpo Trapezoide/metabolismoRESUMEN
Pulmonary fibrosis can be experimentally induced in small rodents by bleomycin. The antibiotic is usually administered via the intratracheal or intranasal routes. In the present study, we investigated the oropharyngeal aspiration of bleomycin as an alternative route for the induction of lung fibrosis in rats and mice. The development of lung injury was followed in vivo by ultrashort echo time magnetic resonance imaging (UTE-MRI) and by post-mortem analyses (histology of collagen, hydroxyproline determination, and qRT-PCR). In C57BL/6 mice, oropharyngeal aspiration of bleomycin led to more prominent lung fibrosis as compared to intranasal administration. Consequently, the oropharyngeal aspiration route allowed a dose reduction of bleomycin and, therewith, a model refinement. Moreover, the distribution of collagen after oropharyngeal aspiration of bleomycin was more homogenous than after intranasal administration: for the oropharyngeal aspiration route, fibrotic areas appeared all over the lung lobes, while for the intranasal route fibrotic lesions appeared mainly around the largest superior airways. Thus, oropharyngeal aspiration of bleomycin induced morphological changes that were more comparable to the human disease than the intranasal administration route did. Oropharyngeal aspiration of bleomycin led to a homogeneous fibrotic injury also in rat lungs. The present data suggest oropharyngeal aspiration of bleomycin as a less invasive means to induce homogeneous and sustained fibrosis in the lungs of mice and rats.
Asunto(s)
Bleomicina/administración & dosificación , Pulmón/patología , Imagen por Resonancia Magnética , Boca/patología , Faringe/patología , Fibrosis Pulmonar/inducido químicamente , Administración Intranasal , Administración Oral , Animales , Cromatografía Líquida de Alta Presión , Humanos , Procesamiento de Imagen Asistido por Computador , Espectrometría de Masas , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Fibrosis Pulmonar/patología , Ratas , Ratas Sprague-Dawley , Succión , Factores de TiempoRESUMEN
Generation of gain-of-function transgenic mice by targeting the Rosa26 locus has been established as an alternative to classical transgenic mice produced by pronuclear microinjection. However, targeting transgenes to the endogenous Rosa26 promoter results in moderate ubiquitous expression and is not suitable for high expression levels. Therefore, we now generated a modified Rosa26 (modRosa26) locus that combines efficient targeted transgenesis using recombinase-mediated cassette exchange (RMCE) by Flipase (Flp-RMCE) or Cre recombinase (Cre-RMCE) with transgene expression from exogenous promoters. We silenced the endogenous Rosa26 promoter and characterized several ubiquitous (pCAG, EF1α and CMV) and tissue-specific (VeCad, αSMA) promoters in the modRosa26 locus in vivo. We demonstrate that the ubiquitous pCAG promoter in the modRosa26 locus now offers high transgene expression. While tissue-specific promoters were all active in their cognate tissues they additionally led to rare ectopic expression. To achieve high expression levels in a tissue-specific manner, we therefore combined Flp-RMCE for rapid ES cell targeting, the pCAG promoter for high transgene levels and Cre/LoxP conditional transgene activation using well-characterized Cre lines. Using this approach we generated a Cre/LoxP-inducible reporter mouse line with high EGFP expression levels that enables cell tracing in live cells. A second reporter line expressing luciferase permits efficient monitoring of Cre activity in live animals. Thus, targeting the modRosa26 locus by RMCE minimizes the effort required to target ES cells and generates a tool for the use exogenous promoters in combination with single-copy transgenes for predictable expression in mice.
Asunto(s)
Sitios Genéticos/genética , Integrasas/metabolismo , Mutagénesis Insercional/métodos , Regiones Promotoras Genéticas/genética , Proteínas/genética , Transgenes/genética , Animales , Sitios de Ligazón Microbiológica/genética , Células Madre Embrionarias/metabolismo , Genes Reporteros/genética , Proteínas Fluorescentes Verdes/metabolismo , Luciferasas/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Transgénicos , Especificidad de Órganos/genética , ARN no TraducidoRESUMEN
The present study investigated the phenotype of heterozygous and homozygous neuropeptide S receptor (Npsr) deficient C57BL/6 mice in NPS- and cocaine induced hyperactivity, spontaneous and reactive locomotor activity, elevated plus maze, conditioned fear, and prepulse inhibition of the acoustic startle response. In Npsr-deficient mice, a strong reduction of spontaneous locomotor activity and of the startle magnitude was observed; heterozygous mice had an intermediate phenotype. In the other experiments, Npsr deficiency leads to no or only a very modest phenotype. These results support an important role of neuropeptide S in regulating locomotor activity.
Asunto(s)
Actividad Motora/genética , Receptores Acoplados a Proteínas G/fisiología , Reflejo de Sobresalto/genética , Estimulación Acústica , Animales , Cocaína/administración & dosificación , Cocaína/farmacología , Condicionamiento Clásico/efectos de los fármacos , Condicionamiento Clásico/fisiología , Pérdida de Tono Postural/efectos de los fármacos , Pérdida de Tono Postural/fisiología , Infusiones Intraventriculares , Inhibición Psicológica , Aprendizaje por Laberinto/efectos de los fármacos , Aprendizaje por Laberinto/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Actividad Motora/efectos de los fármacos , Actividad Motora/fisiología , Neuropéptidos/administración & dosificación , Neuropéptidos/farmacología , Receptores Acoplados a Proteínas G/efectos de los fármacos , Receptores Acoplados a Proteínas G/genética , Reflejo de Sobresalto/efectos de los fármacos , Reflejo de Sobresalto/fisiologíaRESUMEN
Melanin-concentrating hormone (MCH) is a cyclic neuropeptide of nineteen amino acids in mammals. Its involvement in the feeding behaviour has been well established during the last few years. A first receptor subtype, now termed MCHIR, was discovered in 1999, following the desorphanisation of the SLCI orphan receptor, using either reverse pharmacology or systematic screening of agonist candidates. A second MCH receptor, MCH2R, has been discovered recently, by several groups working on data mining of genomic banks. The molecular pharmacology of these two receptors is only described on the basis of the action of peptides derived from MCH. The present review tentatively summarizes the knowledge on these two receptors and presents the first attempts to discover new classes of antagonists that might have major roles in the control of obesity and feeding behaviour.
Asunto(s)
Hormonas Hipotalámicas/metabolismo , Melaninas/metabolismo , Hormonas Hipofisarias/metabolismo , Receptores de la Hormona Hipofisaria/metabolismo , Secuencia de Aminoácidos/fisiología , Animales , Conducta Alimentaria/fisiología , Humanos , Hormonas Hipotalámicas/química , Hormonas Hipotalámicas/genética , Melaninas/química , Melaninas/genética , Datos de Secuencia Molecular , Hormonas Hipofisarias/química , Hormonas Hipofisarias/genética , Receptores de la Hormona Hipofisaria/química , Receptores de la Hormona Hipofisaria/genética , Homología de Secuencia de AminoácidoRESUMEN
Melanin-concentrating hormone (MCH) is a cyclic neuropeptide, with a major role in stimulation of feeding behavior in mammals. MCH signals in the brain occur via two seven-transmembrane G protein-coupled receptors, namely MCH1 (SLC-1, MCH(1), MCH-R1, or MCH-1R) and MCH2 (SLT, MCH(2), MCH-R2, or MCH-2R). In this study, we demonstrate that the pro-MCH(131-165) peptide neuropeptide-glutamic acid-isoleucine (NEI)-MCH is more potent than MCH in stimulating feeding in the rat. Using rat MCH1-expressed human embryonic kidney 293 cells, we show that NEI-MCH exhibits 5-fold less affinity in a binding assay and 2-fold less potency in a cAMP assay than MCH. A similar 7- to 8-fold shift in potency was observed in a Ca(2+)(i) assay using rat MCH1 or human MCH2-transfected Chinese hamster ovary cell models. This demonstrates that NEI-MCH is not a better agonist than MCH at either of the MCH receptors. Then, we compared the proteolysis resistance of MCH and NEI-MCH to rat brain membrane homogenates and purified proteases. Kinetics of peptide degradation using brain extracts indicated a t(1/2) of 34.8 min for MCH and 78.5 min for NEI-MCH with a specific pattern of cleavage of MCH but not NEI-MCH by exo- and endo-proteases. Furthermore, MCH was found highly susceptible to degradation by aminopeptidase M and endopeptidase 24.11, whereas NEI-MCH was fully resistant to proteolysis by these enzymes. Therefore, our results strongly suggest that reduced susceptibility to proteases of NEI-MCH compared with MCH account for its enhanced activity in feeding behavior. NEI-MCH represents therefore the first MCH natural functional "superagonist" so far described.
Asunto(s)
Estimulantes del Apetito/farmacología , Ventrículos Cerebrales/fisiología , Conducta Alimentaria/efectos de los fármacos , Hormonas Hipotalámicas/farmacología , Fragmentos de Péptidos/farmacología , Precursores de Proteínas/farmacología , Secuencia de Aminoácidos , Animales , Estimulantes del Apetito/administración & dosificación , Células CHO , Señalización del Calcio/efectos de los fármacos , Señalización del Calcio/fisiología , Línea Celular , Ventrículos Cerebrales/efectos de los fármacos , Cricetinae , AMP Cíclico/farmacología , Humanos , Hormonas Hipotalámicas/administración & dosificación , Hormonas Hipotalámicas/química , Inyecciones Intraventriculares , Riñón , Cinética , Masculino , Datos de Secuencia Molecular , Fragmentos de Péptidos/administración & dosificación , Precursores de Proteínas/administración & dosificación , Precursores de Proteínas/química , Ratas , Ratas Wistar , TransfecciónRESUMEN
Melanin-concentrating hormone (MCH) is a cyclic peptide, mainly involved in the regulation of skin pigmentation in teleosts and feeding behavior in mammals. The human keratinocyte SVK14 cell line has been previously shown to express binding sites for the MCH analog [125I]-[Phe13,3-iodo-Tyr19]MCH. We report here that: (1) this binding site similarly recognized [125I]-[3-iodo-Tyr13]MCH; (2) its pharmacological profile clearly differed from those observed at the two human MCH receptor subtypes, MCH1-R and MCH2-R; (3) MCH did not induce any effect on second messenger systems (including cAMP, calcium, and MAP kinase signaling pathways), and (4) no mRNAs corresponding to the MCH receptors were found. In conclusion, the binding site characterized in the SVK14 cell line is distinct from the MCH1 and MCH2 receptors and deserves therefore further investigation.