RESUMEN
Insulin-like peptide 5 (INSL5) is a member of the insulin-like family of peptides. It has been reported to be orexigenic in rodent models of obesity with impaired glucose metabolism. We attempted to confirm this property as a first step in establishing the ability of INSL5 to successfully integrate with other agents more proven in their ability to reverse obesity and improve metabolism. INSL5 was chemically synthesized by two alternative methods to a native form and one that was site-specifically conjugated to a 20â¯KDa polyethylene glycol (PEG) polymer. The pharmacology of each peptide was assessed by high-dose chronic administration in normal and obese mice. INSL5 failed to produce pharmacologically relevant effects on food intake, body weight or glucose control indicative of a negligible role of the peptide in the control of feeding and glucose metabolism.
Asunto(s)
Peso Corporal/efectos de los fármacos , Ingestión de Alimentos/efectos de los fármacos , Conducta Alimentaria/efectos de los fármacos , Glucosa/metabolismo , Obesidad/metabolismo , Hormonas Peptídicas/farmacología , Animales , Ratones , Ratones Obesos , Obesidad/tratamiento farmacológico , Obesidad/patología , Hormonas Peptídicas/síntesis química , Hormonas Peptídicas/químicaRESUMEN
Spexin (Spx), a novel neuropeptide, composed of 14 amino acid residues, is evolutionally conserved from fish to mammals. It has been suggested that Spx has pleiotropic functions in mammals. However, reports about Spx are very limited. To clarify the roles of Spx in the regulation of reproduction and food-intake in the spotted scat, the spx (ssspx) gene was cloned and analyzed. Analysis of the tissue distribution by RT-PCR showed that ssspx expression was widespread. During ovary development, expression of ssspx was found to be highest in phase II, moderate in phase III, and at its lowest level in phase IV. Ssspx expression was significantly down-regulated in the hypothalamus after treatment with E2 both in vitro and in vivo. A significant increase of ssspx was observed after 2 and 7â¯days of food deprivation. However, the ssspx transcript levels in the 7â¯day fasting group decreased significantly after refeeding 3â¯h after the scheduled feeding time. This suggests that ssSpx may be involved in the regulation of reproduction and food-intake in the spotted scat.
Asunto(s)
Perfilación de la Expresión Génica , Hormonas Peptídicas/genética , Perciformes/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , Estradiol/farmacología , Ayuno , Femenino , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Hipotálamo/efectos de los fármacos , Hipotálamo/metabolismo , Masculino , Ovario/efectos de los fármacos , Ovario/embriología , Ovario/metabolismo , Hormonas Peptídicas/química , Hormonas Peptídicas/metabolismo , Perciformes/metabolismo , Filogenia , Reproducción , Alineación de Secuencia , Distribución Tisular/efectos de los fármacosRESUMEN
Growth hormone (GH) release is a process that is well regulated by several factors, including GH secretagogues. GH can mediate the regulation of the fatty acid level and composition. The aim of this study was to determine the effect of a synthetic GH secretagogue peptide (A233) on the growth and fatty acid composition in tilapia (Oreochromis niloticus). To address this objective, we administrated a diet supplemented with A233 to juvenile tilapia for 60 days. The group fed with a diet supplemented with 600 µg of A233 per kg of feed increased in weight (4.81 ± 0.09 g) and specific growth rate (2.49 ± 0.03%/day) compared to the control diet group (3.63 ± 0.08 g, 2.07 ± 0.04%/day; respectively) (p < 0.001). In the muscle, the total lipids for the control diet group were higher than that in the group fed with 600 µg of A233 per kg feed; however, no differences were detected in the liver. In both tissues, the patterns of fatty acid composition and content were generally similar, with some exceptions. Tilapia fed with 600 µg of A233 per kg of feed showed, in liver and muscle, a significantly higher composition and content of n-3 polyunsaturated fatty acids (such as 20:5n-3, 22:5n-3, 22:6n-3) and n-3/n-6 PUFA than animals fed with the control diet. To our knowledge, this is the first report on the the effects of natural or synthetic GH secretagogues (GHS) on fatty acid composition, implying an increase in the nutritional quality of the tilapia.
Asunto(s)
Cíclidos/crecimiento & desarrollo , Cíclidos/metabolismo , Ácidos Grasos Omega-3/metabolismo , Hígado/efectos de los fármacos , Músculos/efectos de los fármacos , Hormonas Peptídicas/farmacología , Secretagogos/farmacología , Animales , Suplementos Dietéticos , Relación Dosis-Respuesta a Droga , Hígado/metabolismo , Músculos/metabolismo , Hormonas Peptídicas/administración & dosificación , Hormonas Peptídicas/química , Secretagogos/administración & dosificación , Secretagogos/químicaRESUMEN
In this brief review we summarize the current fndings relative to the discovery of a small peptide ligand, phoenixin (PNX). Using a bioinformatic approach, two novel peptides PNX-14 and PNX-20 containing 14 and 20 amino acids, respectively, were isolated from diverse tissues including the brain, heart, lung and stomach. Mass spectrometry analysis identified a major and minor peak corresponding to PNX-14 and PNX-20, in rat or mouse spinal cord extracts. With the use of a rabbit polyclonal antiserum, phoenixin immunoreactivity (irPNX) was detected in discrete areas of the rodent brain including several hypothalamic subnuclei and dorsal motor nucleus of the vagus. In addition, irPNX was detected in a population of sensory ganglion cells including dorsal root ganglion, nodose ganglion and trigeminal ganglion, and in cell processes densely distributed to the superficial layers of the dorsal horn, nucleus of the solitary tract and spinal trigeminal tract. irPNX cell processes were also detected in the skin and myenteric plexus, suggesting a brain-gut and/or brain-skin connection. Pharmacological studies show that PNX-14 injected subcutaneously to the nape of the neck of mice provoked dose-dependent repetitive scratching bouts directed to the back of the neck with the hindpaws. Our result suggests that the peptide PNX-14 and/or PNX-20, may serve as one of the endogenous signal molecules transducing itch sensation. Additionally, results from other laboratories show that exogenous PNX may affect a number of diverse behaviors such as memory formation, depression, reproduction, food-intake and anxiolytic-like behaviors.
Asunto(s)
Hormonas Hipotalámicas/fisiología , Hormonas Peptídicas/fisiología , Péptidos/fisiología , Secuencia de Aminoácidos , Animales , Humanos , Hormonas Hipotalámicas/administración & dosificación , Hormonas Hipotalámicas/química , Hipotálamo/metabolismo , Memoria/fisiología , Plexo Mientérico/metabolismo , Hormonas Peptídicas/administración & dosificación , Hormonas Peptídicas/química , Péptidos/administración & dosificación , Péptidos/química , Prurito/metabolismo , Médula Espinal/metabolismoRESUMEN
Phoenixin (PNX), a newly identified peptide cleaved from the C-terminus of protein C4orf52, mainly exists in two active isoforms, phoenixin-14 (PNX-14) amide and phoenixin-20 (PNX-20) amide that were first isolated from the rat hypothalamus and bovine heart, respectively. Initial studies demonstrated that PNX is a reproductive peptide, which affects the hypothalamus pituitary genital (HPG) axis through regulating the expression of kisspeptin, GnRH, GnRH receptor, LH and oestrus process. However, further studies indicated that PNX might play a wide range of roles in additional physiological process such as inhibiting visceral pain and eliciting pruritus, inducing anxiety, improving memory retention. Recently, Gpr173, also designated as SREB3, was identified as the cognate receptor of PNX. Whereas, the regulatory mechanism of PNX has not been fully clarified. This review aims to provide the current knowledge of PNX and propose some study directions for future research.
Asunto(s)
Hormonas Hipotalámicas/genética , Hormonas Peptídicas/genética , Péptidos/genética , Animales , Humanos , Hormonas Hipotalámicas/química , Hipotálamo/química , Hipotálamo/metabolismo , Hormonas Peptídicas/química , Péptidos/química , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Ratas , Reproducción/genéticaRESUMEN
The novel neuropeptide spexin (SPX) was discovered to activate galanin receptor 2 (GALR2) and 3 (GALR3) but not galanin receptor 1 (GALR1). Although GALR2 is known to display a function, particularly in anxiety, depression, and appetite regulation, the further determination of its function would benefit from a more stable and selective agonist that acts only at GALR2. In the present study, we developed a GALR2-specific agonist with increased stability in serum. As galanin (GAL) showed a low affinity to GALR3, the residues in SPX were replaced with those in GAL, revealing that particular mutations such as Gln5 â Asn, Met7 â Ala, Lys11 â Phe, and Ala13 â Pro significantly decreased potencies toward GALR3 but not toward GALR2. Quadruple (Qu) mutation of these residues still retained potency to GALR2 but totally abolished the potency to both GALR3 and GALR1. The first amino acid modifications or D-Asn1 substitution significantly increased the stability when they are incubated in 100% fetal bovine serum. Intracerebroventricular administration of the mutant peptide with D-Asn1 and quadruple substitution (dN1-Qu) exhibited an anxiolytic effect in mice. Taken together, the GALR2-specific agonist with increased stability can greatly help delineation of GALR2-mediated functions and be very useful for treatments of anxiety disorder.
Asunto(s)
Ansiolíticos/química , Receptor de Galanina Tipo 2/agonistas , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Animales , Ansiolíticos/farmacología , Evaluación Preclínica de Medicamentos , Estabilidad de Medicamentos , Células HEK293 , Humanos , Concentración 50 Inhibidora , Masculino , Ratones Endogámicos C57BL , Imitación Molecular , Hormonas Peptídicas/química , Estabilidad Proteica , Receptor de Galanina Tipo 2/metabolismo , Suero/químicaRESUMEN
A novel biomaterial formed by the immobilization of the Aminopeptidase from Aeromonas proteolytica (AAP) on synthetic Mg2+ and Al3+ ion-containing layered double hydroxide (LDH) particles was prepared. Immobilization of AAP on the LDH particles in a buffered, aqueous mixture is rapid such that the maximum loading capacity, 1×10(-9) moles of AAP/mg LDH, is achieved in a few minutes. X-ray powder diffraction of LDH samples before and after treatment with AAP indicates that the enzyme does not intercalate between the layers of LDH, but instead binds to the surface. Treatment of AAP/LDH with various amounts of salt in a buffered mixture demonstrates that between 15 and 20% of AAP can be removed from the LDH by washing the composite material in 0.2 M NaCl. However, the residual AAP remains bound to the LDH even at 1 M salt concentrations. A suspension of the AAP/LDH biomaterial in 10 mM Tricine buffered, aqueous solution (pH 8.0 and 25° C) catalyzes the hydrolysis of l-leucine-p-nitroanilide demonstrating that immobilized AAP remains available to substrate and retains its catalytic activity. Recycling experiments reveal that the AAP/LDH particles can be recovered and reused multiple times without appreciable loss of activity. This work provides the foundation for the development of materials that will function in the degradation or detection of peptide hormones or neurotoxins.
Asunto(s)
Aeromonas/enzimología , Óxido de Aluminio/química , Aminopeptidasas/química , Proteínas Bacterianas/química , Enzimas Inmovilizadas/química , Óxido de Magnesio/química , Neurotoxinas/química , Hormonas Peptídicas/químicaRESUMEN
pQDLDHVFLRFamide is a highly conserved crustacean neuropeptide with a structure that places it within the myosuppressin subfamily of the FMRFamide-like peptides. Despite its apparent ubiquitous conservation in decapod crustaceans, the paracrine and/or endocrine roles played by pQDLDHVFLRFamide remain largely unknown. We have examined the actions of this peptide on the cardiac neuromuscular system of the American lobster Homarus americanus using four preparations: the intact animal, the heart in vitro, the isolated cardiac ganglion (CG), and a stimulated heart muscle preparation. In the intact animal, injection of myosuppressin caused a decrease in heartbeat frequency. Perfusion of the in vitro heart with pQDLDHVFLRFamide elicited a decrease in the frequency and an increase in the amplitude of heart contractions. In the isolated CG, myosuppressin induced a hyperpolarization of the resting membrane potential of cardiac motor neurons and a decrease in the cycle frequency of their bursting. In the stimulated heart muscle preparation, pQDLDHVFLRFamide increased the amplitude of the induced contractions, suggesting that myosuppressin modulates not only the CG, but also peripheral sites. For at least the in vitro heart and the isolated CG, the effects of myosuppressin were dose-dependent (10(-9) to 10(-6) mol l(-1) tested), with threshold concentrations (10(-8)-10(-7) mol l(-1)) consistent with the peptide serving as a circulating hormone. Although cycle frequency, a parameter directly determined by the CG, consistently decreased when pQDLDHVFLRFamide was applied to all preparation types, the magnitudes of this decrease differed, suggesting the possibility that, because myosuppressin modulates the CG and the periphery, it also alters peripheral feedback to the CG.
Asunto(s)
Crustáceos/química , Corazón/efectos de los fármacos , Nephropidae/efectos de los fármacos , Nephropidae/fisiología , Sistema Nervioso/efectos de los fármacos , Neuropéptidos/farmacología , Hormonas Peptídicas/farmacología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , FMRFamida/farmacología , Ganglios de Invertebrados/efectos de los fármacos , Ganglios de Invertebrados/fisiología , Frecuencia Cardíaca/efectos de los fármacos , Datos de Secuencia Molecular , Contracción Miocárdica/efectos de los fármacos , Miocardio , Neuropéptidos/química , Neuropéptidos/genética , Hormonas Peptídicas/química , Perfusión , Reproducibilidad de los Resultados , Factores de TiempoRESUMEN
Transient increases in the cytoplasmic Ca(2+) concentration are key events that initiate many cellular signaling pathways in response to developmental and environmental cues in plants; however, only a few extracellular mediators regulating cytoplasmic Ca(2+) singling are known to date. To identify endogenous cell signaling peptides regulating cytoplasmic Ca(2+) signaling, Arabidopsis seedlings expressing aequorin were used for an in vivo luminescence assay for Ca(2+) changes. These seedlings were challenged with fractions derived from plant extracts. Multiple heat-stable, protease-sensitive peaks of calcium elevating activity were observed after fractionation of these extracts by high-performance liquid chromatography. Tandem mass spectrometry identified the predominant active molecule isolated by a series of such chromatographic separations as a 49-amino acid polypeptide, AtRALF1 (the rapid alkalinization factor protein family). Within 40 s of treatment with nanomolar concentrations of the natural or synthetic version of the peptides, the cytoplasmic Ca(2+) level increased and reached its maximum. Prior treatment with a Ca(2+) chelator or inhibitor of IP 3-dependent signaling partially suppressed the AtRALF1-induced Ca(2+) concentration increase, indicating the likely involvement of Ca(2+) influx across the plasma membrane as well as release of Ca(2+) from intracellular reserves. Ca(2+) imaging using seedlings expressing the FRET-based Ca(2+) sensor yellow cameleon (YC) 3.6 showed that AtRALF1 could induce an elevation in Ca(2+) concentration in the surface cells of the root consistent with the very rapid effects of addition of AtRALF1 on Ca(2+) levels as reported by aequorin. Our data support a model in which the RALF peptide mediates Ca(2+)-dependent signaling events through a cell surface receptor, where it may play a role in eliciting events linked to stress responses or the modulation of growth.
Asunto(s)
Proteínas de Arabidopsis/química , Arabidopsis/química , Señalización del Calcio/fisiología , Calcio/química , Hormonas Peptídicas/química , Péptidos/química , Plantones/química , Transducción de Señal/fisiología , Aequorina , Secuencia de Aminoácidos , Animales , Arabidopsis/citología , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/aislamiento & purificación , Proteínas de Arabidopsis/fisiología , Calcio/fisiología , Citoplasma/química , Citoplasma/fisiología , Ligandos , Proteínas Luminiscentes/metabolismo , Proteínas Luminiscentes/fisiología , Datos de Secuencia Molecular , Hormonas Peptídicas/aislamiento & purificación , Hormonas Peptídicas/fisiología , Péptidos/aislamiento & purificación , Péptidos/fisiología , Escifozoos , Plantones/citología , Plantones/crecimiento & desarrolloRESUMEN
Stanniocalcin is a glycoprotein hormone important in the maintenance of calcium and phosphate homeostasis in fish. Two related mammalian stanniocalcin genes, STC1 and STC2, were found to be expressed in various tissues as paracrine regulators. We have demonstrated the existence of a second stanniocalcin gene in fish, designated fish STC2, with only 30% identity to fish STC1. However, phylogenetic analysis and comparison of the genomic structure of STC genes in vertebrates indicated that STC1 and STC2 genes were probably derived from a common ancestor gene. Based on the prominent expression of mammalian STC1 in the ovary, we tested STC2 expression in rat ovary and the regulation of STC2 expression by gonadotropins. Treatment of immature rats with pregnant mare serum gonadotropin increased STC2 transcripts, whereas subsequent treatment with human chorionic gonadotropin suppressed STC2 expression. Real-time PCR analyses also demonstrated that STC2 is expressed mainly in thecal layers. In situ hybridization studies also revealed that STC2 is expressed in thecal cell layers of antral and preovulatory follicles after gonadotropin stimulation. To elucidate the physiological functions of STC2, recombinant human and fish STC2 proteins were generated and found to be N-glycosylated homodimers. In cultured granulosa cells, treatment with human or fish STC2 suppressed FSH-induced progesterone, but not estradiol or cAMP, production. The STC2 suppression of progesterone production was associated with the inhibition of FSH-induced CYP11A and 3beta-hydroxysteroid dehydrogenase expression. Thus, STC2 is a functional homodimeric glycoprotein, and thecal cell-derived STC2 could play a paracrine role during follicular development.
Asunto(s)
Glicoproteínas/genética , Glicoproteínas/metabolismo , Ovario/metabolismo , Comunicación Paracrina , Hormonas Peptídicas/genética , Ratas/metabolismo , Pez Cebra/genética , Animales , Células Cultivadas , Gonadotropina Coriónica/farmacología , Clonación Molecular , AMP Cíclico/biosíntesis , ADN Complementario , Estradiol/biosíntesis , Femenino , Hormona Folículo Estimulante/farmacología , Glicoproteínas/química , Glicoproteínas/farmacología , Gonadotropinas Equinas/farmacología , Células de la Granulosa/efectos de los fármacos , Células de la Granulosa/metabolismo , Péptidos y Proteínas de Señalización Intercelular , Datos de Secuencia Molecular , Hormonas Peptídicas/química , Hormonas Peptídicas/metabolismo , Progesterona/biosíntesis , ARN Mensajero/metabolismo , Ratas Sprague-Dawley , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacología , Células Tecales/efectos de los fármacos , Células Tecales/metabolismo , Pez Cebra/metabolismoRESUMEN
Recent identification of novel appetite-regulating hormones has revealed the complex interactions of these humoral factors in the regulation of feeding behavior in mammals. One of these hormones, ghrelin, a natural ligand of the orphan receptor GHS-R, purified from stomach, is able to stimulate growth hormone release from pituitary cells. Ghrelin is a 28 amino acid peptide containing an n-octanoylated serine 3 residue that is essential for its activity. Ghrelin stimulates appetite by acting on the hypothalamic arcuate nucleus, the region known to control food intake. As an orexigenic peptide, ghrelin is therefore an endogenous regulator of feeding behavior from the peripheral tissues to the central nervous system.
Asunto(s)
Apetito/fisiología , Mucosa Gástrica/metabolismo , Hormonas Gastrointestinales/fisiología , Hormona de Crecimiento Humana/metabolismo , Hormonas Peptídicas/fisiología , Secuencia de Aminoácidos , Animales , Anorexia/fisiopatología , Ghrelina , Humanos , Hipotálamo/fisiología , Datos de Secuencia Molecular , Obesidad/fisiopatología , Hormonas Peptídicas/química , Hormonas Peptídicas/genéticaRESUMEN
2-(2-Amino-2-methyl-propionylamino)-5-phenyl-pentanoic acid [1-[1-(4-methoxy-phenyl)-1-methyl-2-oxo-2-pyrrolidin-1-yl-ethyl]-1H-imidazol-4-yl]-amide (LY444711, 6) is an orally active ghrelin agonist that binds with high affinity to and is a potent activator of the growth hormone secretagogue receptor 1a (GHS-R1a) receptor. In rat models of feeding behavior and pharmacology, 6 creates a positive energy balance and induces adiposity by stimulating food consumption and sparing fat utilization. As an orally active ghrelin agonist, 6 represents a new pharmacological tool to investigate the orexigenic role of ghrelin in regulating energy homeostasis.
Asunto(s)
Tejido Adiposo/efectos de los fármacos , Ingestión de Alimentos/efectos de los fármacos , Receptores Acoplados a Proteínas G/agonistas , Tejido Adiposo/fisiología , Administración Oral , Animales , Evaluación Preclínica de Medicamentos , Ingestión de Alimentos/fisiología , Ghrelina , Masculino , Hormonas Peptídicas/química , Hormonas Peptídicas/farmacología , Ratas , Ratas Long-Evans , Receptores Acoplados a Proteínas G/fisiología , Receptores de GhrelinaRESUMEN
Calcitonin, calcitonin gene-related peptide (CGRP), adrenomedullin (ADM), and amylin belong to a unique group of peptide hormones important for homeostasis in diverse tissues. Calcitonin is essential for calcium balance, whereas CGRP and ADM are important for neurotransmission and cardiovascular and respiratory regulation. Based on phylogenetic analysis, we identified intermedin as a novel member of the calcitonin/CGRP peptide family. Analysis of intermedin expression indicated that intermedin is expressed primarily in the pituitary and gastrointestinal tract. Intermedin increased cAMP production in SK-N-MC and L6 cells expressing endogenous CGRP receptors and competed with labeled CGRP for binding to its receptors in these cells. In addition, treatment of 293T cells expressing recombinant calcitonin receptor-like receptor (CRLR) and one of the three receptor activity-modifying proteins (RAMPs) showed that a CRLR/RAMP receptor complex is required for intermedin signaling. In contrast to CGRP and ADM, which exhibited a preferential stimulation of CRLR when co-expressed with RAMP1 and RAMP2 or RAMP3, respectively, intermedin represents a nonselective agonist for the RAMP coreceptors. In vivo studies demonstrated that intermedin treatment led to blood pressure reduction in both normal and spontaneously hypertensive rats via interactions with the CRLR/RAMP receptor complexes. Furthermore, in vivo treatment in mice with intermedin led to suppression of gastric emptying activity and food intake. Thus, identification of intermedin as a novel member of the calcitonin/CGRP peptide family capable of signaling through CRLR/RAMP receptor complexes provides an additional player in the regulation of peripheral tissues by CRLR and will allow development of new therapeutic agents for pathologies associated with diverse vascular and gastrointestinal disorders.
Asunto(s)
Péptido Relacionado con Gen de Calcitonina/química , Calcitonina/química , Hormonas Peptídicas/química , Hormonas Peptídicas/fisiología , Receptores de Péptido Relacionado con el Gen de Calcitonina/química , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Presión Sanguínea , Northern Blotting , Línea Celular , Línea Celular Tumoral , Clonación Molecular , AMP Cíclico/metabolismo , ADN Complementario/metabolismo , Relación Dosis-Respuesta a Droga , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Datos de Secuencia Molecular , Péptidos/química , Filogenia , Unión Proteica , Ratas , Ratas Endogámicas SHR , Ratas Sprague-Dawley , Homología de Secuencia de Aminoácido , Transducción de Señal , Factores de Tiempo , Distribución Tisular , TransfecciónRESUMEN
The purpose of this research was to assess the acylation reactions of peptides, salmon calcitonin (sCT), human parathyroid hormone 1-34 (hPTH1-34) and leuprolide, in poly(lactic-co-glycolic acid) (PLGA) microspheres. Capillary electrophoresis (CE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) were used for determining and monitoring peptide acylation and quantitating acylation products in the degrading PLGA microspheres. In the degrading PLGA microspheres of sCT and hPTH1-34, the acylation products were observed and determined to be adducts with glycolic acid units from degradable PLGA polymer by MALDI-TOF MS. In the microsphere of leuprolide, however, the acylation product was not observed even after 28 days of incubation at the release medium, which represents the different stabilities among peptides according to the primary structure. As the leuprolide contains tyrosine and serine having hydroxyl group of nucleophilic amino acids, the acylation reaction of peptide is shown to be mainly due to the primary amino groups of N-terminus or lysine residue. The complementary use of CE and MALDI-TOF MS will be useful for searching the counter measures as well as determining the peptide acylation in the manufactured formulations on the market.
Asunto(s)
Ácido Láctico/química , Hormonas Peptídicas/química , Ácido Poliglicólico/química , Polímeros/química , Acilación , Secuencia de Aminoácidos , Animales , Calcitonina/química , Portadores de Fármacos/química , Electroforesis Capilar , Glicolatos/química , Humanos , Leuprolida/química , Lisina/química , Microesferas , Datos de Secuencia Molecular , Hormona Paratiroidea/química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Salmón , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
Urotensin II (UII) has been reported as the most potent known vasoconstrictor. While rat and mouse orthologs of UII precursor protein have been reported, only the tentative structures of UII peptides of these animals have been demonstrated, since prepro-UII proteins lack typical processing sites for their mature peptides. In the present study, we isolated a novel peptide, UII-related peptide (URP), from the extract of the rat brain as the sole immunoreactive substance to anti-UII antibody; the amino acid sequence of the peptide was determined as ACFWKYCV. cDNAs encoding rat, mouse, and human precursor proteins for URP were cloned and revealed that the sequences of mouse and human URP peptides are the same as that for rat URP. Prepro-URP gene is expressed in several rat tissues such as those of the thymus, spleen, testis, and spinal cord, although with lower levels than the prepro-UII gene. In the human, the prepro-URP gene is expressed comparably to prepro-UII in several tissues except the spinal cord. URP was found to bind and activate the human or rat UII receptors (GPR14) and showed a hypotensive effect when administered to anesthetized rats. These results suggest that URP is the endogenous and functional ligand for UII receptor in the rat and mouse, and possibly in the human. We also describe the preparation of specific monoclonal antibodies raised against UII peptide and the establishment of a highly sensitive enzyme immunoassay system for UII peptides.
Asunto(s)
Encéfalo/metabolismo , Hormonas Peptídicas/química , Hormonas Peptídicas/fisiología , Urotensinas/metabolismo , Secuencia de Aminoácidos , Animales , Presión Sanguínea , Células CHO , Calcio/metabolismo , Clonación Molecular , Cricetinae , ADN Complementario/metabolismo , Relación Dosis-Respuesta a Droga , Humanos , Técnicas para Inmunoenzimas , Péptidos y Proteínas de Señalización Intracelular , Ligandos , Masculino , Ratones , Datos de Secuencia Molecular , Péptidos/química , Reacción en Cadena de la Polimerasa , Unión Proteica , Ratas , Ratas Wistar , Homología de Secuencia de Aminoácido , Factores de Tiempo , Distribución TisularRESUMEN
In order to develop an effective pharmacological treatment for obesity, an endogenous factor that promotes a positive energy balance by increasing appetite and decreasing fat oxidation could represent the drug target scientists have been looking for. The recently discovered gastric endocrine agent ghrelin, which appears to be the only potent hunger-inducing factor to naturally circulate in our blood stream, was discovered in 1999. Since then the acylated peptide hormone ghrelin has evolved from an endogenous growth hormone secretagogue to a regulator of energy balance to a pleiotropic hormone with multiple sources, numerous target tissues and most likely several physiological functions. Although neither the exact mechanism of action by which ghrelin increases food intake and adiposity is known, nor the putatively differential effects of brain-derived and stomach-derived ghrelin on energy homeostasis have been determined, blocking or neutralizing ghrelin action still seems one of the more reasonable pharmacological approaches to reverse a chronically positive energy balance. However, based on growing experience with compounds targeting the neuroendocrine regulation of energy balance, it is quite possible that a ghrelin antagonist will either fail to cure obesity due to the existence of compensatory mechanisms or undesired effects might reveal the true biological function of ghrelin (e.g. cardiovascular mechanisms, anti-proliferative effects, reproduction).
Asunto(s)
Obesidad/metabolismo , Hormonas Peptídicas/metabolismo , Receptores Acoplados a Proteínas G , Tejido Adiposo/patología , Secuencia de Aminoácidos , Animales , Fármacos Antiobesidad/química , Fármacos Antiobesidad/farmacología , Mucosa Gástrica/metabolismo , Ghrelina , Humanos , Hipotálamo/metabolismo , Datos de Secuencia Molecular , Obesidad/tratamiento farmacológico , Hormonas Peptídicas/sangre , Hormonas Peptídicas/química , Receptores de Superficie Celular/agonistas , Receptores de Superficie Celular/antagonistas & inhibidores , Receptores de Superficie Celular/fisiología , Receptores de GhrelinaRESUMEN
The genes for parathyroid hormone-related protein (PTHrP) have been cloned in two teleost fishes, cDNA of sea bream (Sparus aurata) and genomic DNA of puffer fish (Fugu rubripes). The gene sequences show that there is significant conservation of amino acid identity, with specific domains most highly conserved. The N-terminus, responsible for bone matrix lysis in mammals and chickens, is present in the fish genes with 52% sequence identity to higher vertebrate PTHrP peptides; the nuclear transporter region shares 73% identity, and the RNA-binding sequence is 65% identical. However, the peptides are shorter then mammalian PTHrP, lacking the C-terminus responsible for inhibition of osteoclast lytic activity, but they have an additional inserted sequence between amino acids 38 and 54 that is not present in higher vertebrate PTHrPs. The N-terminus 1-38 Fugu PTHrP proved to be hypercalcaemic in larval Sparus, suggesting that it may be a physiological regulator of calcium homeostasis in fish. Using homologous nucleotide probes for in situ hybridisation and reverse-transcription polymerase chain reaction (RT-PCR) of extracted RNA, PTHrP gene expression has been widely found in both developing and adult fish. Antiserum to the fish insert sequence demonstrated transcription of PTHrP in all stages of Sparus development, and also detected the same epitope in tissues of developing frog (Rana temporaria), indicating that this has been retained during evolution of the amphibia.