RESUMEN
In mammal's pineal glands, ATP interacts with the high-affinity P2Y1 and the low-affinity P2X7 receptors. ATP released from sympathetic nerve terminals potentiates noradrenaline-induced serotonin N-acetyltransferase (Snat) transcription, N-acetylserotonin (NAS), and melatonin (MLT) synthesis. Circulating melatonin impairs the expression of adhesion molecules in endothelial cells, blocking the migration of leukocytes. Acute defence response induced by pathogen- and danger/damage-associated molecular patterns (PAMPs and DAMPs) triggers the NF-κB pathway in pinealocytes and blocks the transcription of Snat. Therefore, the darkness hormone is not released, and neutrophils and monocytes migrate to the lesion sites. ATP released in high amounts from apoptotic and death cells was considered a DAMP, and the blockage of P2X7 receptors was tested as a new class of drugs for treating brain damage. However, this is not a simple equation. High ATP injected in a lateral ventricle blocked MLT, but not NAS, synthesis as it impairs the transcription of acetyl serotonin N-methyltransferase. NAS is released in the plasma and the cerebral spinal fluid. NAS also blocks the rolling and adhesion of leukocytes to endothelial cells. Otherwise, it is metabolised specifically in each brain area to provide the requested concentration of MLT as a neuroprotector. As observed in physiological conditions, high extracellular ATP, different from the other DAMPs, reports the environmental light/dark cycle rhythm because NAS substitutes MLT as the nocturnal chemical indicator, the darkness hormone. Thus, blocking the P2X7R should not be considered a universal therapy for improving acute strokes, as MLT and ATP are partners in health and disease.
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Melatonin is a highly conserved molecule found in prokaryotes and eukaryotes that acts as the darkness hormone, translating environmental lighting to the whole body, and as a moderator of innate and acquired defense, migration, and cell proliferation processes. This review evaluates the importance of pineal activity in monitoring PAMPs and DAMPs and in mounting an inflammatory response or innate immune response. Activation of the immune-pineal axis, which coordinates the pro-and anti-inflammatory phases of an innate immune response, is described. PAMPs and DAMPs promote the immediate suppression of melatonin production by the pineal gland, which allows leukocyte migration. Monocyte-derived macrophages, important phagocytes of microbes, and cellular debris produce melatonin locally and thereby initiate the anti-inflammatory phase of the acute inflammatory response. The role of locally produced melatonin in organs that directly contact the external environment, such as the skin and the gastrointestinal and respiratory tracts, is also discussed. In this context, as resident macrophages are self-renewing cells, we explore evidence indicating that, besides avoiding overreaction of the immune system, extra-pineal melatonin has a fundamental role in the homeostasis of organs and tissues.
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Inmunidad Innata , Macrófagos/inmunología , Melatonina/inmunología , Glándula Pineal/inmunología , Animales , Humanos , Inflamación/inmunologíaRESUMEN
Environmental pollution in the form of particulate matter <2.5 µm (PM2.5 ) is a major risk factor for diseases such as lung cancer, chronic respiratory infections, and major cardiovascular diseases. Our goal was to show that PM2.5 eliciting a proinflammatory response activates the immune-pineal axis, reducing the pineal synthesis and increasing the extrapineal synthesis of melatonin. Herein, we report that the exposure of rats to polluted air for 6 hours reduced nocturnal plasma melatonin levels and increased lung melatonin levels. Melatonin synthesis in the lung reduced lipid peroxidation and increased PM2.5 engulfment and cell viability by activating high-affinity melatonin receptors. Diesel exhaust particles (DEPs) promoted the synthesis of melatonin in a cultured cell line (RAW 264.7 cells) and rat alveolar macrophages via the expression of the gene encoding for AANAT through a mechanism dependent on activation of the NFκB pathway. Expression of the genes encoding AANAT, MT1, and MT2 was negatively correlated with cellular necroptosis, as disclosed by analysis of Gene Expression Omnibus (GEO) microarray data from the human alveolar macrophages of nonsmoking subjects. The enrichment score for antioxidant genes obtained from lung gene expression data (GTEx) was significantly correlated with the levels of AANAT and MT1 but not the MT2 melatonin receptor. Collectively, these data provide a systemic and mechanistic rationale for coordination of the pineal and extrapineal synthesis of melatonin by a standard damage-associated stimulus, which activates the immune-pineal axis and provides a new framework for understanding the effects of air pollution on lung diseases.
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Pulmón/metabolismo , Macrófagos Alveolares/metabolismo , Melatonina/metabolismo , Material Particulado/efectos adversos , Glándula Pineal/metabolismo , Receptores de Melatonina/metabolismo , Contaminación del Aire/efectos adversos , Animales , N-Acetiltransferasa de Arilalquilamina/metabolismo , Humanos , RatasRESUMEN
BACKGROUND/AIM: The nocturnal production of melatonin by the pineal gland is triggered by sympathetic activation of adrenoceptors and may be modulated by immunological signals. The effect of glucocorticoids on nocturnal melatonin synthesis is controversial; both stimulatory and inhibitory effects have been reported. During pathophysiological processes, an increased sympathetic tonus could result in different patterns of adrenoceptor activation in the pineal gland. Therefore, in this investigation, we evaluated whether the pattern of adrenergic stimulation of the pineal gland drives the direction of the glucocorticoid effect on melatonin production. METHODS: The corticosterone effect on the pineal hormonal production induced by ß-adrenoceptor or ß+α1-adrenoceptor activation was evaluated in cultured glands. We also investigated whether the in vivo lipopolysaccharide (LPS)-induced inhibition of melatonin is dependent on the interaction of glucocorticoids and the α1-adrenoceptor in adrenalectomized animals and on the in vivo blockade of glucocorticoid receptors (GRs) or the α1-adrenoceptor. RESULTS: Corticosterone potentiated ß-adrenoceptor-induced pineal melatonin synthesis, whilst corticosterone-dependent inhibition was observed when melatonin production was induced by ß+α1-adrenoceptors agonists. The inhibitory effect of corticosterone is mediated by GR, as it was abolished in the presence of a GR antagonist. Moreover, LPS-induced reduction in melatonin nocturnal plasma content was reversed by adrenalectomy and by antagonizing GR or α1-adrenoceptors. CONCLUSIONS: The dual effect of corticosterone on pineal melatonin synthesis is determined by the activation pattern of adrenoceptors (ß or ß+α1) in the gland during GR activation, suggesting that increased activation of the sympathetic system and the hypothalamic-pituitary-adrenal axis are necessary for the control of melatonin production during defense responses.
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Catecolaminas/metabolismo , Corticosterona/administración & dosificación , Melatonina/biosíntesis , Glándula Pineal/metabolismo , Receptores Adrenérgicos alfa 1/metabolismo , Receptores Adrenérgicos beta/metabolismo , Agonistas Adrenérgicos beta/administración & dosificación , Animales , Inflamación/metabolismo , Isoproterenol/administración & dosificación , Lipopolisacáridos , Masculino , Glándula Pineal/efectos de los fármacos , Ratas , Ratas WistarRESUMEN
Adenosine triphosphate (ATP) is released onto the pinealocyte, along with noradrenaline, from sympathetic neurons and triggers P2Y1 receptors that enhance ß-adrenergic-induced N-acetylserotonin (NAS) synthesis. Nevertheless, the biotransformation of NAS into melatonin, which occurs due to the subsequent methylation by acetylserotonin O-methyltransferase (ASMT; EC 2.1.1.4), has not yet been evaluated in the presence of purinergic stimulation. We therefore evaluated the effects of purinergic signaling on melatonin synthesis induced by ß-adrenergic stimulation. ATP increased NAS levels, but, surprisingly, inhibited melatonin synthesis in an inverse, concentration-dependent manner. Our results demonstrate that enhanced NAS levels, which depend on phospholipase C (PLC) activity (but not the induction of gene transcription), are a post-translational effect. By contrast, melatonin reduction is related to an ASMT inhibition of expression at both the gene transcription and protein levels. These results were independent of nuclear factor-kappa B (NF-kB) translocation. Neither the P2Y1 receptor activation nor the PLC-mediated pathway was involved in the decrease in melatonin, indicating that ATP regulates pineal metabolism through different mechanisms. Taken together, our data demonstrate that purinergic signaling differentially modulates NAS and melatonin synthesis and point to a regulatory role for ATP as a cotransmitter in the control of ASMT, the rate-limiting enzyme in melatonin synthesis. The endogenous production of melatonin regulates defense responses; therefore, understanding the mechanisms involving ASMT regulation might provide novel insights into the development and progression of neurological disorders since melatonin presents anti-inflammatory, neuroprotective, and neurogenic effects.
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Adenosina Trifosfato/farmacología , Melatonina/biosíntesis , Glándula Pineal/metabolismo , Acetilserotonina O-Metiltransferasa/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Femenino , Masculino , FN-kappa B/metabolismo , Ratas , Ratas Wistar , Receptores Purinérgicos P2Y1/metabolismo , Serotonina/análogos & derivados , Serotonina/metabolismo , Fosfolipasas de Tipo C/metabolismoRESUMEN
Increased plasma glucocorticoids (corticosterone - CORT, in amphibians) and melatonin (MEL) are associated with the daily activity phase and with environmental darkness, respectively. Besides, CORT and MEL also play pivotal immunomodulatory roles in several vertebrates. Herein we described the daily profile of plasma MEL and CORT for Rhinella icterica toads in captivity. Thereafter, we investigated the effects of lipopolysaccharide (LPS)-induced systemic inflammation on the production of CORT and MEL in the R. icterica. Captive toads showed CORT and MEL diurnal variation typical of nocturnal species, with increased values for CORT at ZT12 (18 h) and MEL peak at ZT18 (24 h). LPS-induced hormonal changes included increased plasma CORT and decreased ocular and plasma MEL when compared to those from toads treated with saline 2 h post-injection. Our results demonstrated the presence of a diurnal CORT and MEL variation in toads. We also showed the crosstalk between CORT and MEL during the toad's systemic inflammation in response to an immune challenge with LPS. Additionally, our results demonstrated that anuran eyes' MEL production might be regulated during the inflammatory processes.
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Lipopolisacáridos , Melatonina , Animales , Bufonidae , Ritmo Circadiano , Corticosterona , Lipopolisacáridos/toxicidad , Melatonina/farmacologíaRESUMEN
The pineal gland is a key player in surveillance and defense responses. In healthy conditions, nocturnal circulating melatonin (MEL) impairs the rolling and adhesion of leukocytes to the endothelial layer. Fungi, bacteria, and pro-inflammatory cytokines block nocturnal pineal MEL synthesis, facilitating leukocyte migration to injured areas. ATP is a cotransmitter of the noradrenergic signal and potentiates noradrenaline (NAd)-induced MEL synthesis via P2Y1 receptor (P2Y1R) activation. Otherwise, ATP low-affinity P2X7 receptor (P2X7R) activation impairs N-acetylserotonin (NAS) into MEL conversion in NAd incubated pineals. Here we mimicked a focal increase of ATP by injecting low (0.3 and 1.0 µg) and high (3.0 µg) ATP in the right lateral ventricle of adult rats. Nocturnal pineal activity mimicked the in culture data. Low ATP doses increased MEL output, while high ATP dose and the P2X7R agonist BzATP (15.0-50.0 ng) increased NAS pineal and blood content. In the brain, the response was structure-dependent. There was an increase in cortical and no change in cerebellar MEL. These effects were mediated by changes in the expression of coding genes to synthetic and metabolizing melatonergic enzymes. Thus, the pineal gland plays a role as a first-line structure to respond to the death of cells inside the brain by turning NAS into the darkness hormone.
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Melatonina , Glándula Pineal , Acetilserotonina O-Metiltransferasa/genética , Acetilserotonina O-Metiltransferasa/metabolismo , Adenosina Trifosfato/metabolismo , Adenosina Trifosfato/farmacología , Animales , Melatonina/farmacología , NAD/metabolismo , Norepinefrina/metabolismo , Norepinefrina/farmacología , Glándula Pineal/metabolismo , Ratas , Receptores Purinérgicos P2X7/metabolismo , Serotonina/análogos & derivadosRESUMEN
Hematopoietic stem and progenitor cells (HSPCs) tightly couple maintenance of the bone marrow (BM) reservoir, including undifferentiated long-term repopulating hematopoietic stem cells (LT-HSCs), with intensive daily production of mature leukocytes and blood replenishment. We found two daily peaks of BM HSPC activity that are initiated by onset of light and darkness providing this coupling. Both peaks follow transient elevation of BM norepinephrine and TNF secretion, which temporarily increase HSPC reactive oxygen species (ROS) levels. Light-induced norepinephrine and TNF secretion augments HSPC differentiation and increases vascular permeability to replenish the blood. In contrast, darkness-induced TNF increases melatonin secretion to drive renewal of HSPCs and LT-HSC potential through modulating surface CD150 and c-Kit expression, increasing COX-2/αSMA+ macrophages, diminishing vascular permeability, and reducing HSPC ROS levels. These findings reveal that light- and darkness-induced daily bursts of norepinephrine, TNF, and melatonin within the BM are essential for synchronized mature blood cell production and HSPC pool repopulation.
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Diferenciación Celular/efectos de la radiación , Oscuridad , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/efectos de la radiación , Luz , Animales , Células Cultivadas , Epigénesis Genética/genética , Células Madre Hematopoyéticas/metabolismo , Ratones , Ratones Endogámicos C57BL , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
The hippocampus is a brain region that is rich in nicotinic acetylcholine receptors (nAChRs), especially the α7 subtype. The hippocampus is severely affected in disorders that have a neuroinflammatory component, such as Alzheimer's disease, Parkinson's disease, and schizophrenia. Previous studies demonstrated both in vivo and in vitro that nicotine inhibits immunological responses, including those that are triggered by the inflammatory agent lipopolysaccharide (LPS), the endotoxin of Gram-negative bacteria. The present study investigated whether chronically administered nicotine interferes with the nuclear binding of nuclear factor-κB (NF-κB) and the expression of LPS-induced inflammatory response genes. The results indicated that chronic nicotine administration (0.1mg/kg, s.c., 14days) inhibited the LPS-induced nuclear binding of NF-κB and mRNA expression levels of Tnf, Il1b, Nos2, and Tlr4. The presence of both the selective α7 nAChR antagonist methyllycaconitine (MLA; 5.0mg/kg i.p., 14days) and the nonselective nAChR antagonist mecamylamine (Meca; 1.0mg/kg, s.c., 14days) reversed the inhibitory effects of nicotine. These results suggest that the chronic activation of α7- and αxßy-containing nAChRs reduces acute inflammatory responses in the brain.
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Hipocampo/efectos de los fármacos , Lipopolisacáridos/farmacología , FN-kappa B/metabolismo , Nicotina/farmacología , Agonistas Nicotínicos/farmacología , Transducción de Señal/efectos de los fármacos , Receptor Toll-Like 4/metabolismo , Aconitina/análogos & derivados , Aconitina/farmacología , Animales , Hipocampo/metabolismo , Inflamación/metabolismo , Masculino , Mecamilamina/farmacología , Ratas Wistar , Receptores Nicotínicos/metabolismoRESUMEN
Schistosomiasis is caused by an intravascular parasite and linked to phenotypic changes in endothelial cells that favor inflammation. Endothelial cells express P2Y1 receptors (P2Y1R), and their activation by ADP favors leukocyte adhesion to the endothelial monolayer. We aimed to evaluate the influence of schistosomiasis upon endothelial purinergic signaling-mediated leukocyte adhesion. Mesenteric endothelial cells and mononuclear cells from control and Schistosoma mansoni-infected mice were used in co-culture. P2Y1R levels were similar in both groups. Basal leukocyte adhesion was higher in the infected than in the control group; leukocyte adhesion increased after treatment with the P2Y1R agonist 2-MeSATP in both groups, though it only marginally increased in the infected group. Pre-incubation with the selective P2Y1R antagonist MRS2179 (0.3µM) prevented the agonist effect. However, in the infected group it also reduced the basal leukocyte adhesion, suggesting endothelial cell pre-activation. The endothelial expressions of NTPDases 2 and 3 were significantly increased in the infected group, increasing extracellular ATP hydrolysis and ADP formation by endothelial cells. Therefore, mesenteric endothelial cells are primed by schistosomiasis to a pro-inflammatory phenotype characterized by an increased expression of NTPDases 2 and 3, favoring ADP accumulation and mononuclear cell adhesion, possibly contributing to mesenteric inflammation and schistosomiasis morbidity.
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Adenosina Trifosfatasas/metabolismo , Adhesión Celular , Células Endoteliales/enzimología , Leucocitos/metabolismo , Mesenterio/irrigación sanguínea , Receptores Purinérgicos P2Y1/metabolismo , Schistosoma mansoni/patogenicidad , Esquistosomiasis/enzimología , Adenosina Difosfato/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Células Cultivadas , Técnicas de Cocultivo , Modelos Animales de Enfermedad , Células Endoteliales/efectos de los fármacos , Células Endoteliales/parasitología , Interacciones Huésped-Patógeno , Humanos , Hidrólisis , Leucocitos/efectos de los fármacos , Leucocitos/parasitología , Masculino , Ratones , Fenotipo , Agonistas del Receptor Purinérgico P2Y/farmacología , Receptores Purinérgicos P2Y1/efectos de los fármacos , Esquistosomiasis/parasitología , Transducción de Señal , Regulación hacia ArribaRESUMEN
Drug dependence is a major health problem in adults and has been recognized as a significant problem in adolescents. We previously demonstrated that repeated treatment with a behaviorally sensitizing dose of ethanol in adult mice induced tolerance or no sensitization in adolescents and that repeated ethanol-treated adolescents expressed lower Fos and Egr-1 expression than adult mice in the prefrontal cortex (PFC). In the present work, we investigated the effects of acute and repeated ethanol administration on cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB) DNA-binding activity using the electrophoretic mobility shift assay (EMSA) and the phosphorylated CREB (pCREB)/CREB ratio using immunoblotting in both the PFC and hippocampus in adolescent and adult mice. Adult mice exhibited typical locomotor sensitization after 15 days of daily treatment with 2.0 g/kg ethanol, whereas adolescent mice did not exhibit sensitization. Overall, adolescent mice displayed lower CREB binding activity in the PFC compared with adult mice, whereas opposite effects were observed in the hippocampus. The present results indicate that ethanol exposure induces significant and differential neuroadaptive changes in CREB DNA-binding activity in the PFC and hippocampus in adolescent mice compared with adult mice. These differential molecular changes may contribute to the blunted ethanol-induced behavioral sensitization observed in adolescent mice.
RESUMEN
Melatonin, the darkness hormone, synchronizes several physiological functions to light/dark cycle. Besides the awake/sleep cycle that is intuitively linked to day/night, daily variations in memory acquisition and innate or acquired immune responses are some of the major activities linked to melatonin rhythm. The daily variation of these complex processes is due to changes in specific mechanisms. In the last years we focused on the influence of melatonin on the expression and function of nicotinic acetylcholine receptors (nAChRs). Melatonin, either "in vivo" or "in vitro", increases, in a selective manner, the efficiency of alpha-bungarotoxin (alpha-BTX)-sensitive nAChRs. Melatonin's effect on receptors located in rat sympathetic nerve terminals, cerebellum, skeletal muscle and chick retina, was tested. We observed that melatonin is essential for the development of alpha-BTX-sensitive nAChRs, and important for receptor maintenance in aging models. Taking into account that both melatonin and alpha-7 nAChRs (one of the subtypes sensitive to alpha-BTX) are involved in the development of Alzheimer's disease, here we discuss the possibility of a therapeutic strategy focused on both melatonin replacement and its potential association with cholinergic drugs.
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Colinérgicos/administración & dosificación , Melatonina/administración & dosificación , Melatonina/fisiología , Receptores Nicotínicos/fisiología , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/metabolismo , Animales , Quimioterapia Combinada , Humanos , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiologíaRESUMEN
In mammals, the production of melatonin by the pineal gland is mainly controlled by the suprachiasmatic nuclei (SCN), the master clock of the circadian system. We have previously shown that agents involved in inflammatory responses, such as cytokines and corticosterone, modulate pineal melatonin synthesis. The nuclear transcription factor NFKB, detected by our group in the rat pineal gland, modulates this effect. Here, we evaluated a putative constitutive role for the pineal gland NFKB pathway. Male rats were kept under 12 h:12 h light-dark (LD) cycle or under constant darkness (DD) condition. Nuclear NFKB was quantified by electrophoretic mobility shift assay on pineal glands obtained from animals killed throughout the day at different times. Nuclear content of NFKB presented a daily rhythm only in LD-entrained animals. During the light phase, the amount of NFKB increased continuously, and a sharp drop occurred when lights were turned off. Animals maintained in a constant light environment until ZT 18 showed diurnal levels of nuclear NFKB at ZT15 and ZT18. Propranolol (20 mg/kg, i.p., ZT 11) treatment, which inhibits nocturnal sympathetic input, impaired nocturnal decrease of NFKB only at ZT18. A similar effect was observed in free-running animals, which secreted less nocturnal melatonin. Because melatonin reduces constitutive NFKB activation in cultured pineal glands, we propose that this indolamine regulates this transcription factor pathway in the rat pineal gland, but not at the LD transition. The controversial results regarding the inhibition of pineal function by constant light or blocking sympathetic neurotransmission are discussed according to the hypothesis that the prompt effect of lights-off is not mediated by noradrenaline, which otherwise contributes to maintaining low levels of nuclear NFKB at night. In summary, we report here a novel transcription factor in the pineal gland, which exhibits a constitutive rhythm dependent on environmental photic information.
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Ritmo Circadiano/fisiología , FN-kappa B/metabolismo , Glándula Pineal/metabolismo , Animales , Secuencia de Bases , Núcleo Celular/metabolismo , Ritmo Circadiano/genética , Masculino , Melatonina/biosíntesis , Melatonina/sangre , FN-kappa B/química , FN-kappa B/genética , Subunidad p50 de NF-kappa B/metabolismo , Sondas de Oligonucleótidos/genética , Fotoperiodo , Subunidades de Proteína , Ratas , Ratas Wistar , Factor de Transcripción ReIA/metabolismoRESUMEN
The venom gland of viperid snakes has a central lumen where the venom produced by secretory cells is stored. When the venom is lost from the gland, the secretory cells are activated and new venom is produced. The production of new venom is triggered by the action of noradrenaline on both alpha(1)- and beta-adrenoceptors in the venom gland. In this study, we show that venom removal leads to the activation of transcription factors NFkappaB and AP-1 in the venom gland. In dispersed secretory cells, noradrenaline activated both NFkappaB and AP-1. Activation of NFkappaB and AP-1 depended on phospholipase C and protein kinase A. Activation of NFkappaB also depended on protein kinase C. Isoprenaline activated both NFkappaB and AP-1, and phenylephrine activated NFkappaB and later AP-1. We also show that the protein composition of the venom gland changes during the venom production cycle. Striking changes occurred 4 and 7 days after venom removal in female and male snakes, respectively. Reserpine blocks this change, and the administration of alpha(1)- and beta-adrenoceptor agonists to reserpine-treated snakes largely restores the protein composition of the venom gland. However, the protein composition of the venom from reserpinized snakes treated with alpha(1)- or beta-adrenoceptor agonists appears normal, judging from SDS-PAGE electrophoresis. A sexual dimorphism in activating transcription factors and activating venom gland was observed. Our data suggest that the release of noradrenaline after biting is necessary to activate the venom gland by regulating the activation of transcription factors and consequently regulating the synthesis of proteins in the venom gland for venom production.
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Bothrops/fisiología , Venenos de Crotálidos/metabolismo , Regulación de la Expresión Génica/fisiología , Sistema Nervioso Simpático/fisiología , Factores de Transcripción/metabolismo , Animales , Venenos de Crotálidos/química , Femenino , Masculino , FN-kappa B/genética , FN-kappa B/metabolismo , Proteínas/genética , Proteínas/metabolismo , Receptores Adrenérgicos/genética , Receptores Adrenérgicos/metabolismo , Reserpina , Factor de Transcripción AP-1/genética , Factor de Transcripción AP-1/metabolismoRESUMEN
Protein degradation by the ubiquitin proteasome system releases large amounts of oligopeptides within cells. To investigate possible functions for these intracellularly generated oligopeptides, we fused them to a cationic transactivator peptide sequence using reversible disulfide bonds, introduced them into cells, and analyzed their effect on G protein-coupled receptor (GPCR) signal transduction. A mixture containing four of these peptides (20-80 microm) significantly inhibited the increase in the extracellular acidification response triggered by angiotensin II (ang II) in CHO-S cells transfected with the ang II type 1 receptor (AT1R-CHO-S). Subsequently, either alone or in a mixture, these peptides increased luciferase gene transcription in AT1R CHO-S cells stimulated with ang II and in HEK293 cells treated with isoproterenol. These peptides without transactivator failed to affect GPCR cellular responses. All four functional peptides were shown in vitro to competitively inhibit the degradation of a synthetic substrate by thimet oligopeptidase. Overexpression of thimet oligopeptidase in both CHO-S and HEK293 cells was sufficient to reduce luciferase activation triggered by a specific GPCR agonist. Moreover, using individual peptides as baits in affinity columns, several proteins involved in GPCR signaling were identified, including alpha-adaptin A and dynamin 1. These results suggest that before their complete degradation, intracellular peptides similar to those generated by proteasomes can actively affect cell signaling, probably representing additional bioactive molecules within cells.
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Oligopéptidos/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Receptor de Angiotensina Tipo 1/metabolismo , Transducción de Señal/fisiología , Subunidades alfa de Complejo de Proteína Adaptadora/genética , Subunidades alfa de Complejo de Proteína Adaptadora/metabolismo , Angiotensina II/farmacología , Animales , Células CHO , Cricetinae , Cricetulus , Dinamina I/genética , Dinamina I/metabolismo , Expresión Génica , Humanos , Metaloendopeptidasas/biosíntesis , Metaloendopeptidasas/genética , Oligopéptidos/genética , Complejo de la Endopetidasa Proteasomal/genética , Ratas , Receptor de Angiotensina Tipo 1/genética , Transducción de Señal/efectos de los fármacos , Transfección , Ubiquitina/genética , Ubiquitina/metabolismo , Vasoconstrictores/farmacologíaRESUMEN
The time course of the innate immunological response involves a pro-inflammatory phase followed by an anti-inflammatory phase. Pro-inflammatory responses serve as a defense against several stressor conditions, and sequential processes that shut down these responses are necessary to avoid exacerbation or the development of chronic diseases. In the present review, we put together recent data that show that the pineal gland is a player in bidirectional control of the inflammatory response. Healthy organisms stay in standby mode, ready to react. The nocturnal melatonin surge impairs the rolling and adherence of leukocytes to endothelial layers, limiting cell migration, and stimulates nocturnal production of IL-2 by T helper lymphocytes, exerting an immunostimulatory effect. Otherwise, the release of TNF-alpha from activated macrophages suppresses the nocturnal melatonin surge, allowing a full cell migration and inhibiting IL-2 production. In sequence, activated mononuclear and polymorphonuclear cells produce melatonin in a paracrine manner at the site of injury, which scavenges free radicals and collaborates to resolve the inflammatory response. The sequential diminution of TNF-alpha production is followed by the recovery of the nocturnal melatonin surge and IL-2 production. In summary, the immune-pineal axis, implicated in the sequential involvement of the melatonin produced by the pineal gland and immune-competent cells, is an integral participant of the innate immune response.
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Inmunidad Innata/inmunología , Neuroinmunomodulación/inmunología , Sistemas Neurosecretores/inmunología , Glándula Pineal/metabolismo , Animales , Humanos , Interleucina-2/inmunología , Interleucina-2/metabolismo , Melatonina/inmunología , Melatonina/metabolismo , Comunicación Paracrina/inmunología , Glándula Pineal/inmunología , Linfocitos T Colaboradores-Inductores/inmunología , Linfocitos T Colaboradores-Inductores/metabolismo , Factor de Necrosis Tumoral alfa/inmunología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
A retino-hypothalamic-sympathetic pathway drives the nocturnal surge of pineal melatonin production that determines the synchronization of pineal function with the environmental light/dark cycle. In many studies, melatonin has been implicated in the modulation of the inflammatory response. However, scant information on the feedback action of molecules present in the blood on the pineal gland during the time course of an inflammatory response is available. Here we analyzed the effect of tumor necrosis factor-alpha (TNF-alpha) and corticosterone on the transcription of the Aa-nat, hiomt and 14-3-3 protein genes in denervated pineal glands of rats stimulated for 5 hr with norepinephrine, using real-time reverse transcription-polymerase chain reaction. The transcription of Aa-nat, a gene encoding the key enzyme in melatonin biosynthesis, together with the synthesis of the melatonin precursor N-acetylserotonin, was inhibited by TNF-alpha. This inhibition was transient, and a preincubation of TNF-alpha for more than 24 hr had no detectable effect. In fact, a protein(s) transcribed, later on, as shown by cycloheximide, was responsible for the reversal of the inhibition of Aa-nat transcription. In addition, corticosterone induced a potentiation of norepinephrine-induced Aa-nat transcription even after 48 hr of incubation. These data support the hypothesis that the nocturnal surge in melatonin is impaired at the beginning of an inflammatory response and restored either during the shutdown of an acute response or in a chronic inflammatory pathology. Here, we introduce a new molecular pathway involved in the feedback of an inflammatory response on pineal activity, and provide a molecular basis for understanding the expression of circadian timing in injured organisms.
Asunto(s)
Ritmo Circadiano/efectos de los fármacos , Ritmo Circadiano/fisiología , Inmunidad/fisiología , Melatonina/biosíntesis , Glándula Pineal/efectos de los fármacos , Glándula Pineal/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Animales , Retroalimentación Fisiológica/efectos de los fármacos , Femenino , Masculino , Norepinefrina/farmacología , Glándula Pineal/inmunología , Ratas , Ratas Wistar , Técnicas de Cultivo de TejidosRESUMEN
In chronically inflamed animals, adrenal hormones exert a positive control on the secretion of melatonin by the pineal gland. In this paper, the mechanism of corticosterone as a modulator of melatonin and N-acetylserotonin (NAS) was determined. Rat pineal glands in culture, stimulated for 5 hr with noradrenaline (10 nm), were previously incubated with corticosterone (1.0 nm-1.0 microm) for 48 hr in the presence or absence of the glucocorticoid receptor (GR) antagonist, mifepristone (1.0 microm), the proteasome inhibitor, N-acetyl-leucinyl-leucinyl-norleucinal-H (ALLN, 12.5 microm) or the antagonist of the nuclear factor kappa B (NFkappaB), pyrrolidinedithiocarbamate (PDTC, 12.5 microm). Corticosterone potentiated noradrenaline-induced melatonin and NAS production in a bell-shaped manner. The increase in NAS (12.9 +/- 2.7, n=6 versus 34.3 +/- 8.3 ng per pineal) and melatonin (16.3 +/- 2.0, n=6 versus 44.3 +/- 12.9 ng per pineal) content induced by 1 microm corticosterone was blocked by mifepristone, and mimicked by ALLN and PDTC. The presence of GRs was shown by [3H]-dexamethasone binding (0.30 +/- 0.09 pmol/mg protein) and corticosterone inhibition of NFkappaB nuclear translocation was demonstrated by electromobility shift assay. Therefore, corticosterone potentiates noradrenaline-induced melatonin and NAS production through GR inhibition of NFkappaB nuclear translocation. To the best of our knowledge, this is the first time that this relevant pathway for passive and acquired immune response is shown to modulate melatonin production in pineal gland.
Asunto(s)
Corticosterona/farmacología , Melatonina/biosíntesis , FN-kappa B/antagonistas & inhibidores , Norepinefrina/farmacología , Glándula Pineal/efectos de los fármacos , Glándula Pineal/metabolismo , Serotonina/análogos & derivados , Animales , Secuencia de Bases , ADN/genética , ADN/metabolismo , Dexametasona/metabolismo , Femenino , Antagonistas de Hormonas/farmacología , Leupeptinas/farmacología , Masculino , Mifepristona/farmacología , FN-kappa B/genética , Técnicas de Cultivo de Órganos , Pirrolidinas/farmacología , Ratas , Ratas Wistar , Receptores de Glucocorticoides/antagonistas & inhibidores , Serotonina/biosíntesis , Tiocarbamatos/farmacologíaRESUMEN
Funçäo cognitiva de pacientes em monoterapia específica para sídrome epiléptica tem sido pouco avaliada. Estudamos: 7 pacientes com epilepsias localizadas sintomáticas (SEL) utilizando fenitoína, com 30 ñ 12 (média ñ desvio padräo) anos de idade; 8 com epilepsias generalizadas idiopáticas utilizando valproato de sódio, com 18 ñ 4 anos; 16 com SEL utilizando carbamazepina, com 28 ñ 11 anos; 35 controles sadios, com 27 ñ 11 anos. Todos tinham inteligência normal, educaçäo apropriada para a idade e vidas produtivas na sociedade. Dois dos pacientes utilizando carbamazepina e um valproato de sódio tinham menos de 5 crises parciais, de ausência ou mioclônicas ao mês. Os outros pacientes tinham crises controladas há mais de 6 meses. As concentraçöes séricas de carbamazepina, fenitoína e valproato tiveram performanece pior que controles em memória imediata; o grupo de carbamazepina foi pior que controles no teste de Stroop (p < 0.01) Os resultados indicam efeitos discretos das síndromes epilépticas, de fenitoína, carbamazepina e valproato de sódio na cogniçäo de pacientes com epilepsia eminentemente controlada e vida produtiva na comunidade. O déficit de pacientes com epilepsia crônica sob regimes politerapêuticos deve ser multifatorial. Estudos futuros devem controlar, em seu planejamento, fatores causais possíveis para que seus resultados sejam relevantes