RESUMO
Bovine milk is an essential supplement due to its rich energy- and nutrient-rich qualities. Caseins constitute the vast majority of the proteins in milk. Among these, ß-casein comprises around 37% of all caseins, and it is an important type of casein with several different variants. The A1 and A2 variants of ß-casein are the most researched genotypes due to the changes in their composition. It is accepted that the A2 variant is ancestral, while a point mutation in the 67th amino acid created the A1 variant. The digestion derived of both A1 and A2 milk is BCM-7. Digestion of A2 milk in the human intestine also forms BCM-9 peptide molecule. The opioid-like characteristics of BCM-7 are highlighted for their potential triggering effect on several diseases. Most research has been focused on gastrointestinal-related diseases; however other metabolic and nervous system-based diseases are also potentially triggered. By manipulating the mechanisms of these diseases, BCM-7 can induce certain situations, such as conformational changes, reduction in protein activity, and the creation of undesired activity in the biological system. Furthermore, the genotype of casein can also play a role in bone health, such as altering fracture rates, and calcium contents can change the characteristics of dietary products. The context between opioid molecules and BCM-7 points to a potential triggering mechanism for the central nervous system and other metabolic diseases discussed.
Assuntos
Caseínas , Endorfinas , Humanos , Animais , Caseínas/química , Caseínas/metabolismo , Caseínas/genética , Endorfinas/química , Endorfinas/metabolismo , Leite/química , Leite/metabolismo , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Fragmentos de Peptídeos/genética , Peptídeos Opioides/química , Peptídeos Opioides/metabolismo , BovinosRESUMO
Some new hemorphin-4 analogs with structures of Xxx-Pro-Trp-Thr-NH2 and Tyr-Yyy-Trp-Thr-NH2, where Xxx is 2-amino-3-(4-hydroxy-2,6-dimethylphenyl)propanoic acid or 2-amino-3-(4-dibenzylamino-2,6-dimethylphenyl)propanoic acid, and Yyy is (2S,4S)-4-amino-pyrrolidine-2-carboxylic acid, were synthesized and characterized by electrochemical and spectral analyses. In vivo anticonvulsant and antinociceptive activities of peptide derivatives were studied after intracerebroventricular injection in mice. The therapeutic effects of the modified peptides on seizures and pain in mice were evaluated to provide valuable insights into the potential applications of the novel compounds. Electrochemical characterization showed that the compounds behave as weak protolytes and that they are in a soluble, stable molecular form at physiological pH values. The antioxidant activity of the peptides was evaluated with voltammetric analyses, which were confirmed by applying the 2,2-Diphenyl-1-picrylhydrazyl method. The compounds showed satisfactory results regarding their structural stability, reaching the desired centers for the manifestation of biological activity without hydrolysis processes at 37°C and physiological pH. Dm-H4 and H4-P1 exhibited 100% and 83% potency to suppress the psychomotor seizures in the 6-Hz test compared to 67% activity of H4. Notably, only the H4-P1 had efficacy in blocking the tonic component in the maximal electroshock test with a potency comparable to H4. All investigated peptides containing unnatural conformationally restricted amino acids showed antinociceptive effects. The analogs Db-H4 and H4-P1 showed the most pronounced and long-lasting effect in both experimental models of pain induced by thermal and chemical stimuli. Dm-H4 produced a dose-dependent thermal antinociception and H4-P2 inhibited only formalin-induced pain behavior.
Assuntos
Convulsões , Animais , Camundongos , Masculino , Convulsões/tratamento farmacológico , Relação Estrutura-Atividade , Anticonvulsivantes/farmacologia , Anticonvulsivantes/síntese química , Anticonvulsivantes/química , Dor/tratamento farmacológico , Aminoácidos/química , Aminoácidos/farmacologia , Aminoácidos/síntese química , Antioxidantes/farmacologia , Antioxidantes/síntese química , Antioxidantes/química , Analgésicos/farmacologia , Analgésicos/síntese química , Analgésicos/química , Relação Dose-Resposta a Droga , Modelos Animais de Doenças , Peptídeos Opioides/farmacologia , Peptídeos Opioides/síntese química , Peptídeos Opioides/química , Analgésicos Opioides/farmacologia , Analgésicos Opioides/síntese química , Analgésicos Opioides/químicaRESUMO
Chronic wounds are a major healthcare challenge owing to their complex healing mechanism and number of impediments to the healing process, like infections, unregulated inflammation, impaired cellular functions, poor angiogenesis, and enhanced protease activity. Current topical care strategies, such as surgical debridement, absorption of exudates, drug-loaded hydrogels for infection and inflammation management, and exogenous supply of growth factors for angiogenesis and cell proliferation, slow the progression of wounds and reduce patient suffering but suffer from low overall cure rates. Therefore, we have developed a proteolytically stable, multifunctional nanoparticle loaded-peptide gel with inherent anti-inflammatory, antibacterial, and pro-angiogenic properties to provide a favorable wound healing milieu by restoring impaired cellular functions. We have fabricated a self-assembled, lauric acid-peptide conjugate gel, LA-LLys-DPhe-LLys-NH2, loaded with yttrium oxide (Y2O3) nanoparticles (NLG). Gel formed a nanofibrous structure, and nanoparticles were passively entrapped within the network. The surface morphology, stability, viscoelastic, and self-healing characteristics of gels were characterized. It showed a high stability against degradation by proteolytic enzymes and highly potent antibacterial activities against E. coli and S. aureus due to the presence of positively charged side chains of lysine in the peptide chain. It also exhibited an excellent antioxidant activity as well as ability to stimulate cell proliferation in murine fibroblast (L929) cells and human umbilical vein endothelial cells (HUVECs). The incorporation of nanoparticles promoted angiogenesis by upregulating pro-angiogenic genes, vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF2), and epidermal growth factor (EGFR), and the gel caused complete wound closure in cells. In summary, the Y2O3 nanoparticle-loaded lauric acid-peptide conjugate gel is able to elicit the desired tissue regeneration responses and, therefore, has a strong potential as a matrix for the treatment of chronic wounds.
Assuntos
Nanopartículas Metálicas , Humanos , Linhagem Celular , Animais , Nanopartículas Metálicas/química , Peptídeos Opioides/química , Di-Hidrotestosterona/química , Staphylococcus aureus/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Antibacterianos/química , Antibacterianos/farmacologia , Cicatrização , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Indutores da Angiogênese/química , Indutores da Angiogênese/farmacologia , Sobrevivência CelularRESUMO
Despite various advantages, opioid peptides have been limited in their therapeutic uses due to the main drawbacks in metabolic stability, blood-brain barrier permeability, and bioavailability. Therefore, extensive studies have focused on overcoming the problems and optimizing the therapeutic potential. Currently, numerous peptide-based drugs are being marketed thanks to new synthetic strategies for optimizing metabolism and alternative routes of administration. This tutorial review briefly introduces the history and role of natural opioid peptides and highlights the key findings on their structure-activity relationships for the opioid receptors. It discusses details on opioid peptidomimetics applied to develop therapeutic candidates for the treatment of pain from the pharmacological and structural points of view. The main focus is the current status of various mimetic tools and the successful applications summarized in tables and figures.
Assuntos
Peptídeos Opioides , Peptidomiméticos , Analgésicos Opioides/metabolismo , Analgésicos Opioides/farmacologia , Analgésicos Opioides/uso terapêutico , Descoberta de Drogas , Peptídeos Opioides/química , Peptídeos Opioides/farmacologia , Peptídeos Opioides/uso terapêutico , Peptidomiméticos/farmacologia , Peptidomiméticos/uso terapêutico , Receptores OpioidesRESUMO
The opioid system in the central nervous system regulates depressive-like behavior in animals. Opioid receptors and their endogenous ligands have been focused on as novel therapeutic targets for depression. We synthesized dermorphin (DRM)-dynorphin (DYN) analogs (DRM-DYN001-004) using the message-address concept concerning interactions with opioid receptors. It has previously been reported that DRM-DYN001, 003, and 004 have shown high affinities for µ- and κ-opioid receptors, whereas all analogs had a lower affinity for the δ-opioid receptor than for other receptors using a receptor binding assay. However, it remains unknown whether these analogs show antidepressant-like effects in mice. We examined the effects of DRM-DYN analogs on the duration of immobile behavior in a tail suspension test. Intracerebroventricular administration of DRM-DYN001 in mice shortened the duration of immobile behavior, but did not affect locomotion. The DRM-DYN001-induced antidepressant-like effect was inhibited by co-administration of naloxone (non-selective opioid receptor antagonist), naloxonazine (selective µ1-opioid receptor antagonist), or nor-BNI (κ-opioid receptor antagonist), but not naltrindole (δ-opioid receptor antagonist). These data suggest that DRM-DYN001 exerts an antidepressant-like effect via activation of the central µ1- and κ-opioid receptors in mice and may represent a new lead peptide for further investigation for the development of novel therapeutic approaches for depression.
Assuntos
Antidepressivos , Dinorfinas , Peptídeos Opioides , Receptores Opioides kappa , Animais , Antidepressivos/administração & dosagem , Antidepressivos/farmacologia , Dinorfinas/administração & dosagem , Dinorfinas/química , Camundongos , Antagonistas de Entorpecentes/farmacologia , Peptídeos Opioides/administração & dosagem , Peptídeos Opioides/química , Receptores Opioides , Receptores Opioides kappa/metabolismoRESUMO
Activated opioid receptors transmit internal signals through two major pathways: the G-protein-mediated pathway, which exerts analgesia, and the ß-arrestin-mediated pathway, which leads to unfavorable side effects. Hence, G-protein-biased opioid agonists are preferable as opioid analgesics. Rubiscolins, the spinach-derived naturally occurring opioid peptides, are selective δ opioid receptor agonists, and their p.o. administration exhibits antinociceptive effects. Although the potency and effect of rubiscolins as G-protein-biased molecules are partially confirmed, their in vitro profiles remain unclear. We, therefore, evaluated the properties of rubiscolins, in detail, through several analyses, including the CellKeyTM assay, cADDis® cAMP assay, and PathHunter® ß-arrestin recruitment assay, using cells stably expressing µ, δ, κ, or µ/δ heteromer opioid receptors. In the CellKeyTM assay, rubiscolins showed selective agonistic effects for δ opioid receptor and little agonistic or antagonistic effects for µ and κ opioid receptors. Furthermore, rubiscolins were found to be G-protein-biased δ opioid receptor agonists based on the results obtained in cADDis® cAMP and PathHunter® ß-arrestin recruitment assays. Finally, we found, for the first time, that they are also partially agonistic for the µ/δ dimers. In conclusion, rubiscolins could serve as attractive seeds, as δ opioid receptor-specific agonists, for the development of novel opioid analgesics with reduced side effects.
Assuntos
Peptídeos Opioides/farmacologia , Receptores Opioides delta/agonistas , Transdução de Sinais/efeitos dos fármacos , Spinacia oleracea/química , Proteínas de Ligação ao GTP/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Estrutura Molecular , Peptídeos Opioides/química , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/farmacologia , Receptores Opioides mu/metabolismo , Ribulose-Bifosfato Carboxilase/química , Ribulose-Bifosfato Carboxilase/farmacologia , beta-Arrestinas/metabolismoRESUMO
Opioids are the most effective analgesics, with most clinically available opioids being agonists to the µ-opioid receptor (MOR). The MOR is also responsible for their unwanted effects, including reward and opioid misuse leading to the current public health crisis. The imperative need for safer, non-addictive pain therapies drives the search for novel leads and new treatment strategies. In this study, the recently discovered MOR/nociceptin (NOP) receptor peptide hybrid KGNOP1 (H-Dmt-D-Arg-Aba-ß-Ala-Arg-Tyr-Tyr-Arg-Ile-Lys-NH2) was evaluated following subcutaneous administration in mouse models of acute (formalin test) and chronic inflammatory pain (Complete Freund's adjuvant-induced paw hyperalgesia), liabilities of spontaneous locomotion, conditioned place preference, and the withdrawal syndrome. KGNOP1 demonstrated dose-dependent antinociceptive effects in the formalin test, and efficacy in attenuating thermal hyperalgesia with prolonged duration of action. Antinociceptive effects of KGNOP1 were reversed by naltrexone and SB-612111, indicating the involvement of both MOR and NOP receptor agonism. In comparison with morphine, KGNOP1 was more potent and effective in mouse models of inflammatory pain. Unlike morphine, KGNOP1 displayed reduced detrimental liabilities, as no locomotor impairment nor rewarding and withdrawal effects were observed. Docking of KGNOP1 to the MOR and NOP receptors and subsequent 3D interaction pattern analyses provided valuable insights into its binding mode. The mixed MOR/NOP receptor peptide KGNOP1 holds promise in the effort to develop new analgesics for the treatment of various pain states with fewer MOR-mediated side effects, particularly abuse and dependence liabilities.
Assuntos
Oligopeptídeos/genética , Peptídeos Opioides/química , Receptores Opioides mu/metabolismo , Dor Aguda/tratamento farmacológico , Analgésicos , Animais , Comportamento Animal , Células CHO , Cricetinae , Cricetulus , Cicloeptanos/farmacologia , Humanos , Hiperalgesia/tratamento farmacológico , Técnicas In Vitro , Inflamação/tratamento farmacológico , Masculino , Camundongos , Modelos Moleculares , Simulação de Acoplamento Molecular , Morfina/química , Morfina/farmacologia , Movimento/efeitos dos fármacos , Naloxona/farmacologia , Naltrexona/farmacologia , Manejo da Dor , Piperidinas/farmacologia , NociceptinaRESUMO
BACKGROUND: Dermorphin is a heptapeptide with an analgesic potential higher than morphine that does not present the same risk for the development of tolerance. These pharmacological features make dermorphin a potential doping agent in competitive sports and it is already prohibited for racehorses. For athletes, the development of an efficient strategy to monitor for its abuse necessitates an investigation of the metabolism of dermorphin in humans. METHODS: Here, human liver microsomes and zebrafish were utilized as model systems of human metabolism to evaluate the presence and kinetics of metabolites derived from dermorphin. Five hours after its administration, the presence of dermorphin metabolites could be detected in both models by liquid chromatography coupled to highresolution mass spectrometry. RESULTS: Although the two models showed common results, marked differences were also observed in relation to the formed metabolites. Six putative metabolites, based on their exact masses of m/z 479.1915, m/z 501.1733, m/z 495.1657, m/z 223.1073, m/z 180.1017 and m/z 457.2085, are proposed to represent the metabolic pattern of dermorphin. The major metabolite generated from the administration of dermorphin in both models was YAFG-OH (m/z 457.2085), which is the N-terminal tetrapeptide previously identified from studies on rats. CONCLUSION: Its extensive characterization and commercial availability suggest that it could serve as a primary target analyte for the detection of dermorphin misuse. The metabolomics approach also allowed the assignment of other confirmatory metabolites.
Assuntos
Microssomos Hepáticos/metabolismo , Peptídeos Opioides/metabolismo , Animais , Humanos , Cinética , Peptídeos Opioides/química , Peixe-ZebraRESUMO
The discovery of enzyme-derived d-amino acid-containing peptides (DAACPs) that have physiological importance in the metazoan challenges previous assumptions about the homochirality of animal proteins while simultaneously revealing new analytical challenges in the structural and functional characterization of peptides. Most known DAACPs have been identified though laborious activity-guided purification studies or by homology to previously identified DAACPs. Peptide characterization experiments are increasingly dominated by high throughput mass spectrometry-based peptidomics, with stereochemistry rarely considered due to the technical challenges of identifying l/d isomerization. This review discusses the prevalence of enzyme-derived DAACPs among animals and the physiological consequences of peptide isomerization. Also highlighted are the analytical methods that have been applied for structural characterization/discovery of DAACPs, including results of several recent studies using non-targeted discovery methods for revealing novel DAACPs, strongly suggesting that more DAACPs remain to be uncovered.
Assuntos
Peptídeos Catiônicos Antimicrobianos/fisiologia , Conotoxinas/química , Oligopeptídeos/fisiologia , Peptídeos Opioides/fisiologia , Peptídeos Cíclicos/fisiologia , Aminoácidos/química , Aminoácidos/metabolismo , Anfíbios/classificação , Anfíbios/fisiologia , Animais , Peptídeos Catiônicos Antimicrobianos/química , Artrópodes/classificação , Artrópodes/fisiologia , Mamíferos/classificação , Mamíferos/fisiologia , Moluscos/classificação , Moluscos/fisiologia , Oligopeptídeos/química , Peptídeos Opioides/química , Peptídeos Cíclicos/química , Filogenia , EstereoisomerismoRESUMO
Nociceptin receptor (NOP) belongs to the family of opioid receptors but was discovered and characterized much later than the so called classical opioid receptors, µ, δ and κ (or MOP, DOP and KOP, resp.). Nociceptin/orphanin FQ (N/OFQ) is the endogenous ligand of this receptor and it controls numerous important functions in the central nervous system and in the periphery, so its analogs may be developed as innovative drugs for the treatment of a variety of conditions and pathological states. Availability of potent and selective ligands with high affinity to NOP receptor is essential to fully understand the role of NOP-N/OFQ system in the body, which in turn may lead to designing novel therapeutics. Here, we have focused on reviewing the structure of potent peptide-based agonists, antagonists, biased analogs and bivalent ligands that target NOP receptor.
Assuntos
Descoberta de Drogas , Peptídeos Opioides/química , Receptores Opioides/metabolismo , Sequência de Aminoácidos , Humanos , Ligantes , Antagonistas de Entorpecentes/química , Antagonistas de Entorpecentes/metabolismo , Peptídeos Opioides/metabolismo , Receptores Opioides/agonistas , Receptores Opioides/química , Relação Estrutura-Atividade , Receptor de Nociceptina , NociceptinaRESUMO
In the present contribution, we analyze the influence that C-terminal extension of short opioid peptide sequences by organic fragments has on receptor affinity, in vivo analgesic activity, and antimelanoma properties. The considered fragments were based on either N-acylhydrazone (NAH) or N'-acylhydrazide motifs combined with the 3,5-bis(trifluoromethyl)phenyl moiety. Eleven novel compounds were synthesized and subject to biological evaluation. The analyzed compounds exhibit a diversified range of affinities for the µ opioid receptor (MOR), rather low δ opioid receptor (DOR) affinities, and no appreciable neurokinin-1 receptor binding. In three out of four pairs, N-acylhydrazone-based derivatives bind MOR better than their N'-acylhydrazide counterparts. The best of the novel derivatives have similar low nanomolar MOR binding affinity as the reference opioids, such as morphine and biphalin. The obtained order of MOR affinities was compared to the results of molecular docking. In vivo, four tested compounds turned out to be relatively strong analgesics. Finally, the NAH-based analogues reduce the number of melanoma cells in cell culture, while their N'-acylhydrazide counterparts do not. The antimelanoma properties are roughly correlated to the lipophilicity of the compounds.
Assuntos
Analgésicos , Citotoxinas , Hidrazonas/química , Melanoma/tratamento farmacológico , Simulação de Acoplamento Molecular , Peptídeos Opioides , Analgésicos/síntese química , Analgésicos/química , Analgésicos/farmacologia , Animais , Linhagem Celular Tumoral , Citotoxinas/síntese química , Citotoxinas/química , Citotoxinas/farmacologia , Humanos , Masculino , Melanoma/metabolismo , Melanoma/patologia , Peptídeos Opioides/síntese química , Peptídeos Opioides/química , Peptídeos Opioides/farmacologia , Ratos , Ratos Wistar , Receptores Opioides mu/agonistas , Receptores Opioides mu/metabolismoRESUMO
Endogenous opioid peptides and prescription opioid drugs modulate pain, anxiety and stress by activating opioid receptors, currently classified into four subtypes. Here we demonstrate that ACKR3/CXCR7, hitherto known as an atypical scavenger receptor for chemokines, is a broad-spectrum scavenger of opioid peptides. Phylogenetically, ACKR3 is intermediate between chemokine and opioid receptors and is present in various brain regions together with classical opioid receptors. Functionally, ACKR3 is a scavenger receptor for a wide variety of opioid peptides, especially enkephalins and dynorphins, reducing their availability for the classical opioid receptors. ACKR3 is not modulated by prescription opioids, but we show that an ACKR3-selective subnanomolar competitor peptide, LIH383, can restrain ACKR3's negative regulatory function on opioid peptides in rat brain and potentiate their activity towards classical receptors, which may open alternative therapeutic avenues for opioid-related disorders. Altogether, our results reveal that ACKR3 is an atypical opioid receptor with cross-family ligand selectivity.
Assuntos
Peptídeos Opioides/química , Receptores CXCR/metabolismo , Analgésicos Opioides/química , Animais , Encéfalo/metabolismo , Linhagem Celular Tumoral , Sistema Nervoso Central/efeitos dos fármacos , Quimiocinas/metabolismo , Humanos , Ligantes , Sistema de Sinalização das MAP Quinases , Masculino , Fosforilação , Ratos , Ratos Wistar , Transdução de Sinais , Relação Estrutura-Atividade , beta-Arrestina 1/metabolismoRESUMO
Hemorphins are endogenous peptides, 4-10 amino acids long, belonging to the family of atypical opioid peptides released during the sequential cleavage of hemoglobin protein. Hemorphins have been shown to exhibit diverse therapeutic effects in both human and animal models. However, the precise cellular and molecular mechanisms involved in such effects remain elusive. In this review, we summarize and propose potential mechanisms based on studies that investigated the biological activity of hemorphins of different lengths on multiple therapeutic targets. Special emphasis is given to molecular events related to renin-angiotensin system (RAS), opioid receptors and insulin-regulated aminopeptidase receptor (IRAP). This review provides a comprehensive coverage of the molecular mechanisms that underpin the therapeutic potential of hemorphins. Furthermore, it highlights the role of various hemorphin residues in pathological conditions, which could be explored further for therapeutic purposes.
Assuntos
Peptídeos Opioides/fisiologia , Peptídeos Opioides/uso terapêutico , Animais , Humanos , Proteína Antagonista do Receptor de Interleucina 1/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Peptídeos Opioides/química , Receptores Opioides/efeitos dos fármacos , Sistema Renina-Angiotensina/efeitos dos fármacosRESUMO
All life forms typically possess homochirality, with rare exceptions. In the case of peptides and proteins, only L-amino acids are known to be encoded by genes. Nevertheless, D-amino acids have been identified in a variety of peptides, synthesized by animal cells. They include neuroexcitatory and neuroprotective peptides, cardioexcitatory peptides, hyperglycemic hormones, opioid peptides, antimicrobial peptides, natriuretic and defensin-like peptides, and fibrinopeptides. This article is a review of their occurrence, structure and bioactivity. It further explores the pharmacology and potential medical applications of some of the peptides.
Assuntos
Aminoácidos/química , Conotoxinas/química , Hormônios de Invertebrado/síntese química , Proteínas do Tecido Nervoso/química , Peptídeos Opioides/química , Proteínas Citotóxicas Formadoras de Poros/química , Sequência de Aminoácidos , Aminoácidos/metabolismo , Animais , Fármacos Cardiovasculares/química , Fármacos Cardiovasculares/metabolismo , Fármacos Cardiovasculares/farmacologia , Conotoxinas/biossíntese , Conotoxinas/farmacologia , Crustáceos/química , Crustáceos/metabolismo , Fibrinopeptídeo A/biossíntese , Fibrinopeptídeo A/química , Fibrinopeptídeo A/farmacologia , Humanos , Hormônios de Invertebrado/biossíntese , Hormônios de Invertebrado/química , Hormônios de Invertebrado/farmacologia , Moluscos/química , Moluscos/metabolismo , Peptídeos Natriuréticos/biossíntese , Peptídeos Natriuréticos/química , Peptídeos Natriuréticos/farmacologia , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/farmacologia , Peptídeos Opioides/biossíntese , Peptídeos Opioides/farmacologia , Proteínas Citotóxicas Formadoras de Poros/biossíntese , Proteínas Citotóxicas Formadoras de Poros/farmacologia , Especificidade da Espécie , Aranhas/química , Aranhas/metabolismo , EstereoisomerismoRESUMO
Opioid peptides, derived from PENK, POMC, PDYN and PNOC precursors, together with their receptors (DOR, MOR, KOR and ORL1), constitute the opioid system and are suggested to participate in multiple physiological/pathological processes in vertebrates. However, the question whether an opioid system exists and functions in non-mammalian vertebrates including birds remains largely unknown. Here, we cloned genes encoding opioid system from the chicken brain and examined their functionality and tissue expression. As in mammals, 6 opioid peptides encoded by PENK (Met-enkephalin and Leu-enkephalin), POMC (ß-endorphin), PDYN (dynorphin-A and dynorphin-B) and PNOC (nociceptin) precursors and four opioid receptors were found to be highly conserved in chickens. Using pGL3-CRE-luciferase and pGL4-SRE-luciferase reporter systems, we demonstrated that chicken opioid receptors (cDOR, cMOR, cKOR and cORL1) expressed in CHO cells, could be differentially activated by chicken opioid peptides, and resulted in the inhibition of cAMP/PKA and activation of MAPK/ERK signaling pathways. cDOR is potently activated by Met-enkephalin and Leu-enkephalin, and cKOR is potently activated by dynorphin-A, dynorphin-B and nociceptin, whereas cORL1 is specifically activated by nociceptin. Unlike cDOR, cKOR and cORL1, cMOR is moderately/weakly activated by enkephalins and other opioid peptides. These findings suggest the ligand-receptor pair in chicken opioid system is similar, but not identical to, that in mammals. Quantitative real-time PCR revealed that the opioid system is mainly expressed in chicken central nervous system including the hypothalamus. Collectively, our data will help to facilitate the better understanding of the conserved roles of opioid system across vertebrates.
Assuntos
Encéfalo/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Sistema de Sinalização das MAP Quinases , Peptídeos Opioides/metabolismo , Receptores Opioides/metabolismo , Sequência de Aminoácidos , Animais , Células Cultivadas , Galinhas , Clonagem Molecular/métodos , Cricetinae , Proteínas Quinases Dependentes de AMP Cíclico/genética , DNA Complementar/genética , Peptídeos Opioides/química , Peptídeos Opioides/genética , Receptores Opioides/química , Receptores Opioides/genética , Homologia de Sequência , Distribuição TecidualRESUMO
In this work the conformation of dermorphin, Tyr-D-Ala-Phe-Gly-Tyr-Pro-Ser-NH2, an opioid peptide and its analogues with different stereochemistry of alanine and different C-terminus is studied in aqueous and membrane environments. Using two-dimensional NMR techniques we demonstrate that in D2O/H2O peptides with D-alanine have extended conformation, while for the L-isomers more compact conformation is preferred. The analysis of ROESY HR MAS spectra of the peptides interacting with the DMPC bilayer indicates that both stereoisomers have still more extended conformation compared to aqueous phase, as shown by much weaker intermolecular interactions. The influence of Ala residue stereochemistry is also reflected in the interactions of the studied peptides with model membranes, as shown by the 31P NMR static spectra, in which the shapes of the phosphorus NMR signals originating from D-isomers correspond to spherically shaped vesicles in the presence of external magnetic field, in comparison to a more elongated ones observed for L-isomers, while TEM photographs shows that upon addition of D-isomers larger lipid vesicles are formed, in contrast to smaller ones for L-isomers. The location of aromatic fragments of dermorphins in the membrane is determined based on static 2H NMR and 1H1H RFDR MAS experiments. All aromatic rings were found to be inserted in the hydrophobic part of the bilayer, with the exception of the Tyr5 rings of D-Ala dermorphins. The influence of the C-terminal modification was found to be almost imperceptible.
Assuntos
Bicamadas Lipídicas/química , Peptídeos Opioides/química , Fosfolipídeos/metabolismo , Aminoácidos Aromáticos , Dimiristoilfosfatidilcolina/metabolismo , Interações Hidrofóbicas e Hidrofílicas , Conformação Molecular , Peptídeos Opioides/metabolismo , EstereoisomerismoRESUMO
Beyond being a source of key nutrients, bovine milk influences physiological functions by synthesising bioactive peptides during the process of digestion. Some of the claimed negative health outcomes associated with milk consumption, such as cardiovascular diseases and type 1 diabetes may be attributed to an opioid peptide, beta-casomorphin-7 (BCM-7), derived from A1 beta-casein. BCM-7 exerts its function by binding to the µ-opioid receptors in the body. It is hypothesised that activation of the µ-opioid receptors in the gut can alter gut microbial composition, impair gut barrier integrity and bile acid metabolism, in addition to increasing gastrointestinal transit time and gut inflammation. Further, it is hypothesised that BCM-7 may influence fractures and obesity via µ-opioid receptor pathways. In conclusion, it appears that BCM-7 might have multiple functions pertinent to human health; however, the evidence is limited and warrants further pre-clinical and clinical studies for hypothesis confirmation.
Assuntos
Trato Gastrointestinal/fisiologia , Leite/química , Obesidade/metabolismo , Peptídeos Opioides/química , Analgésicos Opioides , Animais , Ácidos e Sais Biliares , Osso e Ossos/metabolismo , Caseínas/química , Bovinos , Bases de Dados Factuais , Endorfinas/química , Microbioma Gastrointestinal/fisiologia , Trato Gastrointestinal/imunologia , Trato Gastrointestinal/microbiologia , Homeostase , Humanos , Inflamação , Fragmentos de Peptídeos/química , Receptores Opioides muRESUMO
Insulin-degrading enzyme (IDE) was applied to catalyze hydrolysis of Nociceptin/Orphanin 1-16 (OFQ/N) to show the involvement of the enzyme in degradation of neuropeptides engaged in pain transmission. Moreover, IDE degradative action towards insulin (Ins) was inhibited by the OFQ/N fragments, suggesting a possible regulatory mechanism in the central nervous system. It has been found that OFQ/N and Ins affect each other degradation by IDE, although in a different manner. Indeed, while the digestion of OFQ/N is significantly affected by the presence of Ins, the kinetic profile of the Ins hydrolysis is not affected by the presence of OFQ/N. However, the main hydrolytic fragments of OFQ/N produced by IDE exert inhibitory activity towards the IDE-mediated Ins degradation. Here, we present the results indicating that, besides Ins, IDE cleaves neuropeptides and their released fragments act as inhibitors of IDE activity toward Ins. Having in mind that IDE is present in the brain, which also contains Ins receptors, it cannot be excluded that this enzyme indirectly participates in neural communication of pain signals and that neuropeptides involved in pain transmission may contribute to the regulation of IDE activity. Finally, preliminary results on the metabolism of OFQ/N, carried out in the rat spinal cord homogenate in the presence of various inhibitors specific for different classes of proteases, show that OFQ/N proteolysis in rat spinal cord could be due, besides IDE, also to a cysteine protease not yet identified.
Assuntos
Insulina/metabolismo , Insulisina/metabolismo , Peptídeos Opioides/metabolismo , Medula Espinal/metabolismo , Sequência de Aminoácidos , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Cromatografia Líquida/métodos , Insulina/química , Insulisina/antagonistas & inibidores , Espectrometria de Massas/métodos , Neuropeptídeos/química , Neuropeptídeos/metabolismo , Neuropeptídeos/farmacologia , Peptídeos Opioides/química , Dor/prevenção & controle , Medição da Dor/métodos , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/farmacologia , Ratos , Receptor de Insulina/metabolismo , Medula Espinal/efeitos dos fármacos , NociceptinaRESUMO
Acid-sensing ion channel (ASIC) channels belong to the family of ligand-gated ion channels known as acid-sensing (proton-gated) ion channels. Only a few activators of ASICs are known. These are exogenous and endogenous molecules that cause a persistent, slowly desensitized current, different from an acid-induced current. Here we describe a novel endogenous agonist of ASICs-peptide nocistatin produced by neuronal cells and neutrophils as a part of prepronociceptin precursor protein. The rat nocistatin evoked currents in X. laevis oocytes expressing rat ASIC1a, ASIC1b, ASIC2a, and ASIC3 that were very similar in kinetic parameters to the proton-gated response. Detailed characterization of nocistatin action on rASIC1a revealed a proton-like dose-dependence of activation, which was accompanied by a dose-dependent decrease in the sensitivity of the channel to the protons. The toxin mambalgin-2, antagonist of ASIC1a, inhibited nocistatin-induced current, therefore the close similarity of mechanisms for ASIC1a activation by peptide and protons could be suggested. Thus, nocistatin is the first endogenous direct agonist of ASICs. This data could give a key to understanding ASICs activation regulation in the nervous system and also could be used to develop new drugs to treat pathological processes associated with ASICs activation, such as neurodegeneration, inflammation, and pain.