RESUMEN
A relatively new pharmacological target in obesity treatment has been the preproglucagon (PPG) signalling, predominantly with glucagon-like peptide (GLP) 1 receptor agonists. As far as the PPG role within the digestive system is well recognised, its actions in the brain remain understudied. Here, we investigated PPG signalling in the Dorsomedial Hypothalamus (DMH), a structure involved in feeding regulation and metabolism, using in situ hybridisation, electrophysiology, and immunohistochemistry. Our experiments were performed on animals fed both control, and high-fat diet (HFD), uncovering HFD-mediated alterations. First, sensitivity to exendin-4 (Exn4, a GLP1R agonist) was shown to increase under HFD, with a higher number of responsive neurons. The amplitude of the response to both Exn4 and oxyntomodulin (Oxm) was also altered, diminishing its relationship with the cells' spontaneous firing rate. Not only neuronal sensitivity, but also GLP1 presence, and therefore possibly release, was influenced by HFD. Immunofluorescent labelling of the GLP1 showed changes in its density depending on the metabolic state (fasted/fed), but this effect was eliminated by HFD feeding. Interestingly, these dietary differences were absent after a period of restricted feeding, allowing for an anticipation of the alternating metabolic states, which suggests possible prevention of such outcome.
Asunto(s)
Dieta Alta en Grasa , Hipotálamo , Proglucagón , Transducción de Señal , Animales , Ratas , Hipotálamo/fisiología , Proglucagón/metabolismo , Ratas Sprague-Dawley , Masculino , Receptor del Péptido 1 Similar al Glucagón/genética , Receptor del Péptido 1 Similar al Glucagón/metabolismo , Receptor del Péptido 2 Similar al Glucagón/genética , Receptor del Péptido 2 Similar al Glucagón/metabolismo , ARN Mensajero/metabolismo , Neuronas/metabolismo , Sinapsis , Fibras Nerviosas/metabolismo , Electrofisiología , Proteínas Proto-Oncogénicas c-fos/metabolismo , Respuesta de Saciedad , Conducta AlimentariaRESUMEN
BACKGROUND: The measurement of proglucagon-derived peptides (PGDPs) is a challenging task mainly due to major overlaps in their molecular sequence in addition to their low circulating levels. Here, we present the technical characteristics of novel ELISA assays measuring C-peptide and all six PGDPs including, for the first time, major proglucagon fragment (MPGF), and we validate them by performing a pilot in vivo cross-over randomized clinical trial on whether coffee consumption may affect levels of circulating PGDPs. METHODS: The performance and technical characteristics of novel ELISA assays from Ansh measuring GLP-1, GLP-2, oxyntomodulin, glicentin, glucagon, MPGF and C-peptide were first evaluated in vitro in procured samples from a commercial vendor as well as in deidentified human samples from three previously performed clinical studies. Their performance was further evaluated in vivo in the context of a cross-over randomized controlled trial, in which 33 subjects consumed in random order and together with a standardized meal, 200 ml of either (a) instant coffee with 3 mg/kg caffeine, or (b) instant coffee with 6 mg/kg caffeine, (c) or water. RESULTS: All assays demonstrated high accuracy (spike and recovery and average linearity recovery ±15%), precision (inter-assay CV ≤ 6.4%), specificity (no significant cross-reactivities) and they were sensitive in low concentrations. Measurements of glicentin in archived random human samples using the Ansh assay correlated strongly with the glicentin measurements of Mercodia assay (r = 0.968) and of GLP-1 modestly with Millipore GLP-1 assay (r = 0.440). Oxyntomodulin, glicentin and glucagon concentrations were 2-5 fold higher in plasma compared to serum and serum concentrations correlated modestly (for oxyntomodulin and glicentin) or poorly (for glucagon) with the plasma concentrations. The evaluated assays detected a postprandial increase of gut-secreted PGDPs (GLP-1, GLP-2, oxyntomodulin and glicentin) and a postprandial decrease of pancreas-secreted PGDPs (glucagon, MPGF) in response to consuming coffee in comparison to consuming water with breakfast (enter here composition of breakfast). Only coffee consumption at the high dose alter levels of gut-secreted PGDPs and both at low and high dose to lower levels of pancreas-secreted PGDPs compared to water consumption during breakfast. CONCLUSION: Accurate, precise and specific measurement of six PGDPs is possible with novel assays. A randomized controlled trial demonstrated in vivo utility of those assays and supports the notion that coffee may exert part of its beneficial effects on glucose homeostasis in the short term through the regulation of PGDPs.
Asunto(s)
Glucagón , Oxintomodulina , Péptido C , Cafeína , Café , Glicentina , Péptido 1 Similar al Glucagón , Humanos , Péptidos , Proglucagón , AguaRESUMEN
Stress responses are coordinated by widespread neural circuits. Homeostatic and psychogenic stressors activate preproglucagon (PPG) neurons in the caudal nucleus of the solitary tract (cNTS) that produce glucagon-like peptide-1; published work in rodents indicates that these neurons play a crucial role in stress responses. While the axonal targets of PPG neurons are well established, their afferent inputs are unknown. Here we use retrograde tracing with cholera toxin subunit b to show that the cNTS in male and female mice receives axonal inputs similar to those reported in rats. Monosynaptic and polysynaptic inputs specific to cNTS PPG neurons were revealed using Cre-conditional pseudorabies and rabies viruses. The most prominent sources of PPG monosynaptic input include the lateral (LH) and paraventricular (PVN) nuclei of the hypothalamus, parasubthalamic nucleus, lateral division of the central amygdala, and Barrington's nucleus (Bar). Additionally, PPG neurons receive monosynaptic vagal sensory input from the nodose ganglia and spinal sensory input from the dorsal horn. Sources of polysynaptic input to cNTS PPG neurons include the hippocampal formation, paraventricular thalamus, and prefrontal cortex. Finally, cNTS-projecting neurons within PVN, LH, and Bar express the activation marker cFOS in mice after restraint stress, identifying them as potential sources of neurogenic stress-induced recruitment of PPG neurons. In summary, cNTS PPG neurons in mice receive widespread monosynaptic and polysynaptic input from brain regions implicated in coordinating behavioral and physiological stress responses, as well as from vagal and spinal sensory neurons. Thus, PPG neurons are optimally positioned to integrate signals of homeostatic and psychogenic stress.SIGNIFICANCE STATEMENT Recent research has indicated a crucial role for glucagon-like peptide-1-producing preproglucagon (PPG) neurons in regulating both appetite and behavioral and autonomic responses to acute stress. Intriguingly, the central glucagon-like peptide-1 system defined in rodents is conserved in humans, highlighting the translational importance of understanding its anatomical organization. Findings reported here indicate that PPG neurons receive significant monosynaptic and polysynaptic input from brain regions implicated in autonomic and behavioral responses to stress, as well as direct input from vagal and spinal sensory neurons. Improved understanding of the neural pathways underlying the recruitment of PPG neurons may facilitate the development of novel therapies for the treatment of stress-related disorders.
Asunto(s)
Neuronas/fisiología , Proglucagón/fisiología , Sinapsis/fisiología , Nervio Vago/fisiología , Animales , Axones/fisiología , Femenino , Hipotálamo/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Vías Nerviosas/fisiología , Neuronas Aferentes/fisiología , Células del Asta Posterior/fisiología , Reflejo Monosináptico/fisiología , Restricción Física , Núcleo Solitario/citología , Núcleo Solitario/fisiología , Estrés Psicológico/fisiopatología , Tálamo/fisiologíaRESUMEN
Obesity is one of the major public health issues, and its prevalence is steadily increasing all the world over. The endocannabinoid system (ECS) has been shown to be involved in the intake of palatable food via activation of cannabinoid 1 receptor (CB1R). However, the involvement of lingual CB1R in the orosensory perception of dietary fatty acids has never been investigated. In the present study, behavioral tests on CB1R-/- and wild type (WT) mice showed that the invalidation of Cb1r gene was associated with low preference for solutions containing rapeseed oil or a long-chain fatty acid (LCFA), such as linoleic acid (LA). Administration of rimonabant, a CB1R inverse agonist, in mice also brought about a low preference for dietary fat. No difference in CD36 and GPR120 protein expressions were observed in taste bud cells (TBC) from WT and CB1R-/- mice. However, LCFA induced a higher increase in [Ca2+]i in TBC from WT mice than that in TBC from CB1R-/- mice. TBC from CB1R-/- mice also exhibited decreased Proglucagon and Glp-1r mRNA and a low GLP-1 basal level. We report that CB1R is involved in fat taste perception via calcium signaling and GLP-1 secretion.
Asunto(s)
Ácidos Grasos , Preferencias Alimentarias , Obesidad/genética , Receptor Cannabinoide CB1/genética , Papilas Gustativas/metabolismo , Percepción del Gusto/genética , Gusto/genética , Animales , Antígenos CD36/genética , Antígenos CD36/metabolismo , Señalización del Calcio/genética , Antagonistas de Receptores de Cannabinoides/farmacología , Grasas de la Dieta , Péptido 1 Similar al Glucagón/metabolismo , Receptor del Péptido 1 Similar al Glucagón/genética , Receptor del Péptido 1 Similar al Glucagón/metabolismo , Ácido Linoleico , Masculino , Ratones Noqueados , Obesidad/etiología , Proglucagón/genética , Proglucagón/metabolismo , ARN Mensajero/metabolismo , Aceite de Brassica napus , Receptor Cannabinoide CB1/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Rimonabant/farmacologíaRESUMEN
OBJECTIVE: To investigate the beneficial role of prebiotics on endothelial dysfunction, an early key marker of cardiovascular diseases, in an original mouse model linking steatosis and endothelial dysfunction. DESIGN: We examined the contribution of the gut microbiota to vascular dysfunction observed in apolipoprotein E knockout (Apoe-/-) mice fed an n-3 polyunsaturated fatty acid (PUFA)-depleted diet for 12â weeks with or without inulin-type fructans (ITFs) supplementation for the last 15â days. Mesenteric and carotid arteries were isolated to evaluate endothelium-dependent relaxation ex vivo. Caecal microbiota composition (Illumina Sequencing of the 16S rRNA gene) and key pathways/mediators involved in the control of vascular function, including bile acid (BA) profiling, gut and liver key gene expression, nitric oxide and gut hormones production were also assessed. RESULTS: ITF supplementation totally reverses endothelial dysfunction in mesenteric and carotid arteries of n-3 PUFA-depleted Apoe-/- mice via activation of the nitric oxide (NO) synthase/NO pathway. Gut microbiota changes induced by prebiotic treatment consist in increased NO-producing bacteria, replenishment of abundance in Akkermansia and decreased abundance in bacterial taxa involved in secondary BA synthesis. Changes in gut and liver gene expression also occur upon ITFs suggesting increased glucagon-like peptide 1 production and BA turnover as drivers of endothelium function preservation. CONCLUSIONS: We demonstrate for the first time that ITF improve endothelial dysfunction, implicating a short-term adaptation of both gut microbiota and key gut peptides. If confirmed in humans, prebiotics could be proposed as a novel approach in the prevention of metabolic disorders-related cardiovascular diseases.
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Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/fisiopatología , Fructanos/farmacología , Microbioma Gastrointestinal/efectos de los fármacos , Prebióticos , Aminopeptidasas/genética , Animales , Péptidos Catiónicos Antimicrobianos/genética , Bacterias/efectos de los fármacos , Ácidos y Sales Biliares/biosíntesis , Ácidos y Sales Biliares/sangre , Arterias Carótidas/fisiología , Ciego/microbiología , Suplementos Dietéticos , Modelos Animales de Enfermedad , Ácidos Grasos Omega-3/administración & dosificación , Ácidos Grasos Omega-3/deficiencia , Expresión Génica/efectos de los fármacos , Péptido 1 Similar al Glucagón/biosíntesis , Masculino , Arterias Mesentéricas/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados para ApoE , Neurotensina/genética , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa/metabolismo , Transportadores de Anión Orgánico Sodio-Dependiente/genética , Proglucagón/genética , Simportadores/genética , VasodilataciónRESUMEN
Short bowel syndrome (SBS) patients developing hyperphagia have a better outcome. Gastrointestinal endocrine adaptations help to improve intestinal functions and food behaviour. We investigated neuroendocrine adaptations in SBS patients and rat models with jejuno-ileal (IR-JI) or jejuno-colonic (IR-JC) anastomosis with and without parenteral nutrition. Circulating levels of ghrelin, PYY, GLP-1, and GLP-2 were determined in SBS rat models and patients. Levels of mRNA for proglucagon, PYY and for hypothalamic neuropeptides were quantified by qRT-PCR in SBS rat models. Histology and immunostaining for Ki67, GLP-1 and PYY were performed in SBS rats. IR-JC rats, but not IR-JI, exhibited significantly higher crypt depths and number of Ki67-positive cells than sham. Fasting and/or postprandial plasma ghrelin and PYY concentrations were higher, or tend to be higher, in IR-JC rats and SBS-JC patients than in controls. Proglucagon and Pyy mRNA levels were significantly enhanced in IR-JC rats. Levels of mRNA coding hypothalamic orexigenic NPY and AgRP peptides were significantly higher in IR-JC than in sham rats. We demonstrate an increase of plasma ghrelin concentrations, major changes in hypothalamic neuropeptides levels and greater induction of PYY in SBS-JC rats and patients suggesting that jejuno-colonic continuity creates a peculiar environment promoting further gut-brain adaptations.
Asunto(s)
Proteína Relacionada con Agouti/metabolismo , Colon/patología , Ghrelina/sangre , Hipotálamo/metabolismo , Yeyuno/patología , Neuropéptido Y/metabolismo , Síndrome del Intestino Corto/metabolismo , Adulto , Anciano , Anastomosis Quirúrgica , Animales , Modelos Animales de Enfermedad , Conducta Alimentaria , Femenino , Péptido 1 Similar al Glucagón/sangre , Péptido 2 Similar al Glucagón/sangre , Humanos , Hiperfagia/metabolismo , Mucosa Intestinal/metabolismo , Antígeno Ki-67/metabolismo , Masculino , Persona de Mediana Edad , Péptido YY/sangre , Proglucagón/metabolismo , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
Selective GLP-1 secretagogues represent a novel potential therapy for type 2 diabetes mellitus. This study examined the GLP-1 secretory activity of the ethnomedicinal plant, Fagonia cretica, which is postulated to possess anti-diabetic activity. After extraction and fractionation extracts and purified compounds were tested for GLP-1 and GIP secretory activity in pGIP/neo STC-1 cells. Intracellular levels of incretin hormones and their gene expression were also determined. Crude F. cretica extracts stimulated both GLP-1 and GIP secretion, increased cellular hormone content, and upregulated gene expression of proglucagon, GIP and prohormone convertase. However, ethyl acetate partitioning significantly enriched GLP-1 secretory activity and this fraction underwent bioactivity-guided fractionation. Three isolated compounds were potent and selective GLP-1 secretagogues: quinovic acid (QA) and two QA derivatives, QA-3ß-O-ß-D-glycopyranoside and QA-3ß-O-ß-D-glucopyranosyl-(28â1)-ß-D-glucopyranosyl ester. All QA compounds activated the TGR5 receptor and increased intracellular incretin levels and gene expression. QA derivatives were more potent GLP-1 secretagogues than QA. This is the first time that QA and its naturally-occurring derivatives have been shown to activate TGR5 and stimulate GLP-1 secretion. These data provide a plausible mechanism for the ethnomedicinal use of F. cretica and may assist in the ongoing development of selective GLP-1 agonists.
Asunto(s)
Células Enteroendocrinas/efectos de los fármacos , Polipéptido Inhibidor Gástrico/agonistas , Péptido 1 Similar al Glucagón/agonistas , Hipoglucemiantes/farmacología , Proglucagón/agonistas , Zygophyllaceae/química , Línea Celular , Células Enteroendocrinas/citología , Células Enteroendocrinas/metabolismo , Polipéptido Inhibidor Gástrico/biosíntesis , Polipéptido Inhibidor Gástrico/genética , Polipéptido Inhibidor Gástrico/metabolismo , Regulación de la Expresión Génica , Péptido 1 Similar al Glucagón/biosíntesis , Péptido 1 Similar al Glucagón/genética , Péptido 1 Similar al Glucagón/metabolismo , Glicósidos/aislamiento & purificación , Glicósidos/farmacología , Humanos , Hipoglucemiantes/aislamiento & purificación , Incretinas/agonistas , Incretinas/genética , Incretinas/metabolismo , Componentes Aéreos de las Plantas/química , Extractos Vegetales/química , Extractos Vegetales/farmacología , Proglucagón/biosíntesis , Proglucagón/genética , Proproteína Convertasas/genética , Proproteína Convertasas/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Transducción de Señal , Triterpenos/aislamiento & purificación , Triterpenos/farmacologíaRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: The radix of Acorus calamus L. (AC) is widely used in diabetes therapies in traditional folk medicine from America and Indonesia, and we have previously reported that the ethyl acetate fraction of AC (ACE) acts as an antidiabetic through insulin sensitizing, insulin releasing and alpha-glucosidase inhibitory activities. The present study is designed to investigate the effects and molecular mechanisms of ACE on glucagon-like peptide-1 (GLP-1) expression and secretion related to its hypoglycemic effects. MATERIALS AND METHODS: The hypoglycemic effect of ACE (100mg/kg, i.g.) was confirmed by testing blood glucose levels or via oral glucose tolerance test (OGTT) in streptozotocin (STZ) induced hyperglycemic mice, db/db diabetic mice and diet-induced obese (DIO) mice. Plasma insulin, GLP-1 levels and intestinal GLP-1 related gene expression were determined in STZ-induced and db/db diabetic mice. The in vitro effects of ACE (12.5µg/ml) on the expression and secretion of GLP-1 were detected in NCI-H716 intestinal L-cells, and the correlation between ACE and molecules in the Wnt signaling pathway was further explored. RESULTS: ACE (100mg/kg) significantly lowered fasting blood glucose in STZ-induced and db/db diabetic mice and improved the OGTT in DIO mice. Insulin releasing and islet protective effects, along with the increased secretion of GLP-1, were observed. The expression of proglucagon gene (gcg) and post-translational processing gene prohormone convertase 3 (pc3) and the GLP-1 content in the culture medium of L-cells notably increased after the ACE treatment (12.5µg/ml). At the same time, ß-catenin nuclear translocation occurred, and its downstream protein cyclin D1 was activated, showing the involvement of Wnt signaling. CONCLUSIONS: ACE might activate Wnt signaling to increase the gene expression of gcg and pc3 and exert incretin effects, including insulinotropic and islet protection, to lower blood glucose levels via elevated GLP-1 secretion either directly or indirectly.
Asunto(s)
Acorus/química , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/metabolismo , Péptido 1 Similar al Glucagón/metabolismo , Hipoglucemiantes/farmacología , Animales , Glucemia/efectos de los fármacos , Prueba de Tolerancia a la Glucosa/métodos , Insulina/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Obesos , Proglucagón/metabolismo , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Vía de Señalización Wnt/efectos de los fármacos , beta Catenina/metabolismoRESUMEN
The gut microbiota contributes to fat mass and the susceptibility to obesity. However, the underlying mechanisms are not completely understood. To investigate whether the gut microbiota affects hypothalamic and brainstem body fat-regulating circuits, we compared gene expression of food intake-regulating neuropeptides between germ-free and conventionally raised (CONV-R) mice. We found that CONV-R mice had decreased expression of the antiobesity neuropeptide glucagon-like peptide-1 (GLP-1) precursor proglucagon (Gcg) in the brainstem. Moreover, in both the hypothalamus and the brainstem, CONV-R mice had decreased expression of the antiobesity neuropeptide brain-derived neurotrophic factor (Bdnf). CONV-R mice had reduced expression of the pro-obesity peptides neuropeptide-Y (Npy) and agouti-related protein (Agrp), and increased expression of the antiobesity peptides proopiomelanocortin (Pomc) and cocaine- and amphetamine-regulated transcript (Cart) in the hypothalamus. The latter changes in neuropeptide expression could be secondary to elevated fat mass in CONV-R mice. Leptin treatment caused less weight reduction and less suppression of orexigenic Npy and Agrp expression in CONV-R mice compared with germ-free mice. The hypothalamic expression of leptin resistance-associated suppressor of cytokine signaling 3 (Socs-3) was increased in CONV-R mice. In conclusion, the gut microbiota reduces the expression of 2 genes coding for body fat-suppressing neuropeptides, Gcg and Bdnf, an alteration that may contribute to fat mass induction by the gut microbiota. Moreover, the presence of body fat-inducing gut microbiota is associated with hypothalamic signs of Socs-3-mediated leptin resistance, which may be linked to failed compensatory body fat reduction.
Asunto(s)
Factor Neurotrófico Derivado del Encéfalo/metabolismo , Sistema Nervioso Central/metabolismo , Regulación hacia Abajo , Intestinos/microbiología , Leptina/metabolismo , Neuronas/metabolismo , Proglucagón/metabolismo , Adiposidad , Proteína Relacionada con Agouti/genética , Proteína Relacionada con Agouti/metabolismo , Animales , Tronco Encefálico/metabolismo , Factor Neurotrófico Derivado del Encéfalo/genética , Vida Libre de Gérmenes , Hipotálamo/metabolismo , Inyecciones Intraperitoneales , Leptina/administración & dosificación , Leptina/sangre , Leptina/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Neuropéptido Y/genética , Neuropéptido Y/metabolismo , Proopiomelanocortina/genética , Proopiomelanocortina/metabolismo , Proglucagón/genética , Proteínas Recombinantes/administración & dosificación , Proteínas Recombinantes/metabolismo , Proteína 3 Supresora de la Señalización de Citocinas , Proteínas Supresoras de la Señalización de Citocinas/genética , Proteínas Supresoras de la Señalización de Citocinas/metabolismoRESUMEN
The type 2 diabetes risk gene TCF7L2 is the effector of the Wnt signaling pathway. We found previously that in gut endocrine L-cell lines, TCF7L2 controls transcription of the proglucagon gene (gcg), which encodes the incretin hormone glucagon-like peptide-1 (GLP-1). Whereas peripheral GLP-1 stimulates insulin secretion, brain GLP-1 controls energy homeostasis through yet-to-be defined mechanisms. We aim to determine the metabolic effect of a functional knockdown of TCF7L2 by generating transgenic mice that express dominant-negative TCF7L2 (TCF7L2DN) specifically in gcg-expressing cells. The gcg-TCF7L2DN transgenic mice showed reduced gcg expression in their gut and brain, but not in pancreas. Defects in glucose homeostasis were observed in these mice, associated with attenuated plasma insulin levels in response to glucose challenge. The defect in glucose disposal was exacerbated with high-fat diet. Brain Wnt activity and feeding-mediated hypothalamic AMP-activated protein kinase (AMPK) repression in these mice were impaired. Peripheral injection of the cAMP-promoting agent forskolin increased brain ß-cat Ser675 phosphorylation and brain gcg expression and restored feeding-mediated hypothalamic AMPK repression. We conclude that TCF7L2 and Wnt signaling control gut and brain gcg expression and glucose homeostasis and speculate that positive cross-talk between Wnt and GLP-1/cAMP signaling is an underlying mechanism for brain GLP-1 in exerting its metabolic functions.
Asunto(s)
Encéfalo/metabolismo , Tracto Gastrointestinal/metabolismo , Regulación de la Expresión Génica , Glucosa/metabolismo , Proglucagón/metabolismo , Proteína 2 Similar al Factor de Transcripción 7/metabolismo , Vía de Señalización Wnt , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Encéfalo/citología , Encéfalo/efectos de los fármacos , Línea Celular , Colforsina/farmacología , AMP Cíclico/agonistas , AMP Cíclico/metabolismo , Tracto Gastrointestinal/citología , Regulación de la Expresión Génica/efectos de los fármacos , Péptido 1 Similar al Glucagón/metabolismo , Homeostasis/efectos de los fármacos , Hipotálamo/citología , Hipotálamo/efectos de los fármacos , Hipotálamo/metabolismo , Masculino , Ratones , Ratones Transgénicos , Especificidad de Órganos , Fosforilación/efectos de los fármacos , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Proteínas Recombinantes de Fusión/metabolismo , Proteína 2 Similar al Factor de Transcripción 7/genética , Vía de Señalización Wnt/efectos de los fármacosRESUMEN
The proglucagon gene is expressed not only in the pancreas and intestine but also in the hypothalamus. Proglucagon-derived peptides have emerged as potential regulators of energy homeostasis. Whether leptin, insulin, or cAMP activation controls proglucagon gene expression in the hypothalamus is not known. A key reason for this has been the inaccessibility of hypothalamic proglucagon-expressing neurons and the lack of suitable neuronal cell lines. Herein we describe the mechanisms involved in the direct regulation of the proglucagon gene by insulin, leptin, and cAMP in hypothalamic cell models. Insulin, through an Akt-dependent manner, significantly induced proglucagon mRNA expression by 70% in adult-derived mHypoA-2/10 neurons and significantly suppressed it by 45% in embryonic-derived mHypoE-39 neurons. Leptin, via the Janus kinase-2/ signal transducer and activator of transcription-3 pathway, caused an initial increase by 66 and 43% at 1 h followed by a decrease by 45 and 34% at 12 h in mHypoA-2/10 and mHypoE-39 cells, respectively. Furthermore, cAMP activation by forskolin up-regulated proglucagon expression by 87% in mHypoE-39 neurons and increased proglucagon mRNA, through Epac activation, in the mHypoE-20/2 neurons. Specific regions of the proglucagon promoter were regulated by cAMP signaling, as determined by transient transfections, whereas mRNA stability assays demonstrate that insulin and leptin increase proglucagon mRNA stability in the adult cells. These findings suggest that insulin, leptin, and cAMP act directly, but differentially, on specific hypothalamic neurons to regulate proglucagon gene expression. Because proglucagon-derived peptides are potential regulators of energy homeostasis, an understanding of hypothalamic proglucagon neurons is important to further expand our knowledge of alternative feeding circuits.
Asunto(s)
AMP Cíclico/fisiología , Hipotálamo/citología , Insulina/fisiología , Leptina/fisiología , Neuronas/metabolismo , Proglucagón/genética , Androstadienos/farmacología , Animales , Células Cultivadas , Cromonas/farmacología , AMP Cíclico/farmacología , Regulación de la Expresión Génica , Humanos , Insulina/farmacología , Janus Quinasa 2/antagonistas & inhibidores , Janus Quinasa 2/metabolismo , Leptina/farmacología , Ratones , Morfolinas/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3 , Proglucagón/metabolismo , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas c-akt/metabolismo , Estabilidad del ARN , Ratas , Reacción en Cadena en Tiempo Real de la Polimerasa , Factor de Transcripción STAT3/metabolismo , Transducción de Señal , Transcripción Genética , Transcriptoma , Triterpenos/farmacología , WortmaninaRESUMEN
Recent studies have provided new evidence that alterations in the composition of the gut microbiota--known as dysbiosis--participate in the development of obesity. The aim of the present study was to investigate the ability of chitin-glucan (CG) from a fungal source to modulate both the gut microbiota and glucose and lipid metabolism in high-fat (HF) diet-induced obese mice. Supplementation of the HF diet with fungal CG (10% w/w) induced caecal enlargement with prominent changes in gut microbiota: it restored the number of bacteria from clostridial cluster XIVa including Roseburia spp., which were decreased due to HF feeding. Furthermore, CG treatment significantly decreased HF-induced body weight gain, fat mass development, fasting hyperglycemia, glucose intolerance, hepatic triglyceride accumulation and hypercholesterolemia, independently of the caloric intake. All those parameters were negatively correlated with specific bacteria of clostridial cluster XIVa, i.e., Roseburia spp. (Pearson's correlations analysis). In contrast to prebiotics that more specifically target the bifidobacteria species, CG effects on obesity appear to be independent of the incretin glucagon-like peptide 1 (GLP-1) production, since portal GLP-1 and proglucagon (its precursor) expression were not modified by the dietary intervention. In conclusion, our findings support the view that chronic consumption of CG has potential beneficial effects with respect to the development of obesity and associated metabolic diabetes and hepatic steatosis, through a mechanism related to the restoration of the composition and/or the activity of gut bacteria, namely, bacteria from clostridial cluster XIVa.
Asunto(s)
Quitina/farmacología , Dieta Alta en Grasa/efectos adversos , Fibras de la Dieta/farmacología , Tracto Gastrointestinal/microbiología , Bacterias Grampositivas/efectos de los fármacos , Animales , Suplementos Dietéticos , Hígado Graso/inducido químicamente , Hígado Graso/prevención & control , Tracto Gastrointestinal/efectos de los fármacos , Péptido 1 Similar al Glucagón/metabolismo , Glucanos/farmacología , Glucosa/metabolismo , Intolerancia a la Glucosa , Hiperglucemia/tratamiento farmacológico , Metabolismo de los Lípidos , Lípidos/sangre , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Obesos , Proglucagón/metabolismo , Triglicéridos/metabolismo , Aumento de Peso/efectos de los fármacosRESUMEN
The distinct lack of cell lines derived from the adult brain is evident. Ciliary neurotrophic factor (CNTF) triggers neurogenesis in primary culture from adult mouse hypothalamus, as detected by bromodeoxyuridine and Ki67 immunostaining. Using SV-40 T-antigen, we immortalized dividing neurons and generated clonal cell lines expressing neuropeptides and receptors involved in neuroendocrine function. We hypothesized that proglucagon-derived peptides may be the mechanistic downstream effectors of CNTF due to documented neuroprotective and proliferative effects. Indeed, proglucagon gene expression was induced by CNTF, and exposure of primary cells to glucagon-like peptide-1 receptor (GLP-1) agonist, exendin-4, induced cell proliferation. Intracerebroventricular injection of CNTF into adult mice caused increased expression of proglucagon peptide in the hypothalamus. Using a specific GLP-1-receptor antagonist, we found that neurogenesis was significantly attenuated and primary culture from GLP-1-receptor-knockout mice lacked CNTF-mediated neuronal proliferation, thus linking the induction of neurogenesis in the hypothalamus to GLP-1-receptor signaling.
Asunto(s)
Factor Neurotrófico Ciliar/farmacología , Péptido 1 Similar al Glucagón/metabolismo , Hipotálamo/citología , Neurogénesis/fisiología , Neuronas/citología , Animales , Línea Celular , Proliferación Celular , Factor Neurotrófico Ciliar/metabolismo , Regulación de la Expresión Génica , Masculino , Ratones , Ratones Endogámicos C57BL , Proglucagón/genética , Proglucagón/metabolismo , Transducción de SeñalRESUMEN
AIM OF THE STUDY: Huang-lian-jie-du-decoction (HLJDD), a well-known Chinese herbal formula, has been used for diabetic treatment. The purpose of the study was to investigate whether HLJDD affected glucagon-like peptide (GLP)-1 (7-36) amide level in diabetic rats. MATERIALS AND METHODS: Streptozotocin (STZ)-induced diabetic rats were treated with HLJDD at low dose (2 g/kg/day) or high dose (4 g/kg/day). After 5-week treatment, GLP-1 (7-36) amide level and insulin level in portal vein and tissues stimulated by oral glucose load were measured by ELISA kits. The proglucagon gene expression in intestinal tracts and the proliferation of intestinal L cell and pancreatic beta cell were measured using RT-PCR and immunohistochemistry techniques, respectively. RESULTS: It was found that 5-week HLJDD treatment attenuated alteration of glucose level and insulin level in plasma and tissues of diabetic rats induced by STZ, accompanied by improvement of diabetic syndrome. 5-week HLJDD treatment increased GLP-1 (7-36) amide level in portal vein plasma and distal ileum. Further studies showed that 5-week HLJDD treatment increased the mRNA level of proglucagon gene in distal ileum, promoted pancreatic beta cell and intestinal L cell proliferation in a dose-dependent manner. CONCLUSION: All the results indicated that HLJDD exerted its anti-diabetic effects partly via modulating GLP-1 (7-36) amide level.
Asunto(s)
Antiinflamatorios no Esteroideos/farmacología , Diabetes Mellitus Experimental/metabolismo , Medicamentos Herbarios Chinos/farmacología , Péptido 1 Similar al Glucagón/metabolismo , Animales , Glucemia/metabolismo , Peso Corporal/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Diabetes Mellitus Experimental/sangre , Ingestión de Alimentos/efectos de los fármacos , Inmunohistoquímica , Indicadores y Reactivos , Insulina/sangre , Células Secretoras de Insulina/efectos de los fármacos , Mucosa Intestinal/metabolismo , Masculino , Páncreas/efectos de los fármacos , Páncreas/metabolismo , Extractos Vegetales/farmacología , Proglucagón/biosíntesis , Proglucagón/genética , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Ratas , Ratas Sprague-Dawley , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
To determine whether portal plasma flow (PPF) and net portal appearance of amino acids (AA) could be affected by 2-hydroxy-4-methylthiobutyrate (HMB), six barrows (35-day-old, 8.6+/-1.4 kg), implanted with arterial, portal and mesenteric catheters, were fed a DL-methionine (as the control) or HMB-supplemented diet once hourly and infused intramesenterically with 1% p-amino hippurate. PPF was numerically 9% higher (P=0.09) in HMB-fed pigs than in controls over a 4-6 h period. Compared with controls, pigs fed the HMB diet had increased (P<0.05) net portal balance and/or appearance of leucine, isoleucine, histidine, arginine and alanine, but had decreased (P<0.05) portal appearance of glutamate over a 6-h period. The concentration of acetate in the lumen of the distal small intestine was higher (P=0.01) in HMB-fed pigs than in controls (25.14 vs. 7.64 mmol/kg). mRNA levels for proglucagon and endothelial nitric-oxide synthase (eNOS) in stomach and proximal small intestine, and mRNA levels for GLP-2 receptor (GLP-2R) in stomach were higher (P<0.05) in HMB-fed pigs compared with those in controls. Collectively, HMB supplementation increased concentrations of short-chain fatty acids in intestinal lumen, expression of proglucagon, GLP-2R, and eNOS genes, and net portal absorption of AA. These novel findings from the study with pigs may also have important implications for intestinal nutrition and health in humans.
Asunto(s)
Aminoácidos/sangre , Dieta , Metionina/análogos & derivados , Sistema Porta/efectos de los fármacos , Aminoácidos/metabolismo , Animales , Ácidos Grasos/sangre , Ácidos Grasos/metabolismo , Receptor del Péptido 2 Similar al Glucagón , Mucosa Intestinal/metabolismo , Intestinos/irrigación sanguínea , Metionina/administración & dosificación , Óxido Nítrico Sintasa de Tipo III/análisis , Sistema Porta/metabolismo , Proglucagón/análisis , ARN Mensajero/análisis , Receptores de Glucagón/análisis , Flujo Sanguíneo Regional/efectos de los fármacos , PorcinosRESUMEN
Recent data reported that inulin-type fructans extracted from chicory roots regulate appetite and lipid/glucose metabolism, namely, by promoting glucagon-like peptide-1 (GLP-1) production in the colon. The Agave genus growing in different regions of Mexico also contains important amounts of original fructans, with interesting nutritional and technological properties, but only few data report their physiological effect when added in the diet. Therefore, we decided to evaluate in parallel the effect of supplementation with 10 % agave or chicory fructans on glucose and lipid metabolism in mice. Male C57Bl/6J mice were fed a standard (STD) diet or diet supplemented with Raftilose P95 (RAF), fructans from Agave tequilana Gto. (TEQ) or fructans from Dasylirion spp. (DAS) for 5 weeks. The body weight gain and food intake in mice fed fructans-containing diets were significantly lower than the ones of mice fed the STD diet, TEQ leading to the lowest value. Serum glucose and cholesterol were similarly lower in all fructans-fed groups than in the STD group and correlated to body weight gain. Only RAF led to a significant decrease in serum TAG. As previously shown for RAF, the supplementation with agave fructans (TEQ and DAS) induced a higher concentration of GLP-1 and its precursor, proglucagon mRNA, in the different colonic segments, thus suggesting that fermentable fructans from different botanical origin and chemical structure are able to promote the production of satietogenic/incretin peptides in the lower part of the gut, with promising effects on glucose metabolism, body weight and fat mass development.
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Agave/química , Suplementos Dietéticos , Fructanos/farmacología , Obesidad/metabolismo , Animales , Glucemia/metabolismo , Ciego/patología , Colesterol/sangre , Colon/metabolismo , Defecación/efectos de los fármacos , Ingestión de Alimentos/efectos de los fármacos , Epidídimo/patología , Péptido 1 Similar al Glucagón/metabolismo , Liliaceae/química , Metabolismo de los Lípidos/efectos de los fármacos , Hígado/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Obesidad/fisiopatología , Tamaño de los Órganos/efectos de los fármacos , Proglucagón/biosíntesis , Proglucagón/genética , ARN Mensajero/genética , Triglicéridos/sangre , Aumento de Peso/efectos de los fármacosRESUMEN
We have evaluated the influence of oligofructose (OFS), a fermentable dietary fibre, on glucose homeostasis, insulin production and intestinal glucagon-like peptide-1 (GLP-1) in streptozotocin-treated diabetic rats. Male Wistar rats received either i.v. streptozotocin (STZ; 40 mg/kg) or vehicle (CT); one week later, they were fed for 6 weeks with either the standard diet (STZ-CT), or with a diet containing 10% oligofructose (STZ-OFS); both diets were available ad libitum. In a second set of experiments (duration 4 weeks), a supplemental group of food-restricted rats (STZ-Res) receiving a similar intake as CT rats, was added. OFS improved glucose tolerance and reduced food intake as compared with STZ-CT rats in both the post-prandial state and after an oral glucose tolerance test. After 6 weeks, portal and pancreatic insulin concentrations were doubled in STZ-OFS rats. Food restriction improved these parameters when compared with STZ-CT rats, but to a lesser extent than in the STZ-OFS group. We have shown that OFS treatment increased portal and colonic GLP-1(7-36) amide levels and doubled colonic proglucagon and prohormone convertase 1 mRNA levels; both OFS and food restriction lowered ileal GLP-1(7-36) amide levels as compared with levels in STZ-CT rats. We propose that OFS, through its fermentation in the colon, promotes the expression and secretion of colonic peptides, namely GLP-1(7-36) amide, with beneficial consequences on glycaemia, insulin secretion and hyperphagia in diabetic rats.
Asunto(s)
Glucemia/análisis , Colon/metabolismo , Diabetes Mellitus Experimental/tratamiento farmacológico , Oligosacáridos/uso terapéutico , Fragmentos de Péptidos/sangre , Actinas/genética , Animales , Colon/química , Diabetes Mellitus Experimental/metabolismo , Ingestión de Alimentos/efectos de los fármacos , Fermentación , Privación de Alimentos , Glucagón/sangre , Glucagón/genética , Péptido 1 Similar al Glucagón , Péptidos Similares al Glucagón , Prueba de Tolerancia a la Glucosa , Insulina/análisis , Insulina/sangre , Islotes Pancreáticos/química , Masculino , Fragmentos de Péptidos/análisis , Proglucagón , Proproteína Convertasa 1/genética , Precursores de Proteínas/sangre , Precursores de Proteínas/genética , ARN Mensajero/análisis , Ratas , Ratas Wistar , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
BACKGROUND & AIMS: Our understanding of the intestinotropic actions of glucagon-like peptide-2 (GLP-2)(1-33) is based on pharmacologic studies involving exogenous administration. However, the physiologic role of GLP-2 in mucosal growth and adaptation to nutritional stimulation remains poorly understood. METHODS: The properties of GLP-2(3-33), a GLP-2(1-33) metabolite, were determined in baby-hamster kidney cells transfected with the mouse GLP-2 receptor complementary DNA and in isolated murine intestinal muscle strips. To investigate the role of endogenous GLP-2(1-33) in gut adaptation, GLP-2(3-33) was administered to mice that were re-fed for 24 hours after 24 hours of fasting, and the small intestine was analyzed. GLP-2(3-33) also was injected into rats for analysis of circulating GLP-2(1-33) levels. RESULTS: GLP-2(3-33) antagonized the actions of GLP-2(1-33) in vitro and ex vivo. Fasting mice exhibited small intestinal atrophy (37% +/- 1% decrease in small intestinal weight, 19% +/- 2% decrease in crypt-villus height, and 99% +/- 35% increase in villus apoptosis, P < .05-.01). Adaptive growth in re-fed mice restored all these parameters, as well as crypt-cell proliferation, to normal control levels (P < .05 vs. fasting); these adaptive changes were prevented partially or completely by co-administration of GLP-2(3-33) to refeeding mice (by 32% +/- 19% to 103% +/- 15%, P < .05-.01 vs re-fed mice). Exogenous GLP-2(3-33) did not affect endogenous GLP-2(1-33) levels. CONCLUSIONS: These data show that endogenous GLP-2 regulates the intestinotropic response in re-fed mice through modulation of crypt-cell proliferation and villus apoptosis. GLP-2 is therefore a physiologic regulator of the dynamic adaptation of the gut mucosal epithelium in response to luminal nutrients.
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Adaptación Fisiológica/fisiología , Ingestión de Alimentos/fisiología , Mucosa Intestinal/fisiología , Fragmentos de Péptidos/fisiología , Adaptación Fisiológica/efectos de los fármacos , Secuencia de Aminoácidos , Animales , Apoptosis/fisiología , Peso Corporal , División Celular/fisiología , Clonación Molecular , Ingestión de Líquidos/fisiología , Femenino , Expresión Génica , Glucagón/genética , Péptido 2 Similar al Glucagón , Receptor del Péptido 1 Similar al Glucagón , Péptidos Similares al Glucagón , Mucosa Intestinal/citología , Mucosa Intestinal/enzimología , Intestino Delgado/citología , Intestino Delgado/enzimología , Intestino Delgado/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Datos de Secuencia Molecular , Tamaño de los Órganos , Fragmentos de Péptidos/sangre , Fragmentos de Péptidos/farmacología , Proglucagón , Precursores de Proteínas/genética , Ratas , Ratas Wistar , Receptores de Glucagón/genética , Receptores de Glucagón/metabolismoRESUMEN
Glucagon-like peptide-1 (GLP-1) is a peptide hormone from the gut that stimulates insulin secretion and protects beta-cells, inhibits glucagon secretion and gastric emptying, and reduces appetite and food intake. In agreement with these actions, it has been shown to be highly effective in the treatment of Type 2 diabetes, causing marked improvements in glycaemic profile, insulin sensitivity and beta-cell performance, as well as weight reduction. The hormone is metabolised rapidly by the enzyme dipeptidyl peptidase IV (DPP-IV) and, therefore, cannot be easily used clinically. Instead, resistant analogues of the hormone (or agonists of the GLP-1 receptor) are in development, along with DPP-IV inhibitors, which have been demonstrated to protect the endogenous hormone and enhance its activity. Agonists include both albumin-bound analogues of GLP-1 and exendin-4, a lizard peptide. Clinical studies with exendin have been carried out for > 6 months and have indicated efficacy in patients inadequately treated with oral antidiabetic agents. Orally active DPP-IV inhibitors, suitable for once-daily administration, have demonstrated similar efficacy. Diabetes therapy, based on GLP-1 receptor activation, therefore, appears very promising.
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Inhibidores de la Adenosina Desaminasa , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Glucagón/análogos & derivados , Glucagón/fisiología , Glicoproteínas/antagonistas & inhibidores , Hipoglucemiantes/uso terapéutico , Fragmentos de Péptidos/fisiología , Precursores de Proteínas/fisiología , Receptores de Glucagón/agonistas , Adenosina Desaminasa/fisiología , Vías Aferentes/fisiología , Animales , Apetito/efectos de los fármacos , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Tipo 2/fisiopatología , Dipeptidil Peptidasa 4/fisiología , Quimioterapia Combinada , Exenatida , Glucagón/agonistas , Glucagón/metabolismo , Glucagón/farmacología , Glucagón/uso terapéutico , Péptido 1 Similar al Glucagón , Receptor del Péptido 1 Similar al Glucagón , Glicoproteínas/fisiología , Humanos , Hipoglucemiantes/administración & dosificación , Hipoglucemiantes/farmacología , Hipotálamo/efectos de los fármacos , Hipotálamo/fisiopatología , Insulina/biosíntesis , Insulina/genética , Insulina/metabolismo , Secreción de Insulina , Mucosa Intestinal/inervación , Mucosa Intestinal/metabolismo , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/metabolismo , Liraglutida , Lagartos , Maleimidas/uso terapéutico , Ratones , Ratones Noqueados , Ratones Obesos , Fragmentos de Péptidos/agonistas , Fragmentos de Péptidos/metabolismo , Fragmentos de Péptidos/farmacología , Péptidos/farmacología , Péptidos/uso terapéutico , Proglucagón , Precursores de Proteínas/agonistas , Precursores de Proteínas/metabolismo , Precursores de Proteínas/farmacología , Ratas , Ratas Zucker , Receptores de Glucagón/deficiencia , Receptores de Glucagón/fisiología , Ponzoñas/farmacología , Ponzoñas/uso terapéuticoRESUMEN
BACKGROUND/PURPOSE: Glucocorticosteroids alter the function of the intestine. Budesonide (Bud) increases the jejunal D-glucose uptake, and this effect is prevented through a polyunsaturated fatty acid (PUFA) diet. This study was undertaken to assess the possible signalling effect of budesonide, prednisone (Pred), or dexamethasone (Dex) in animals with a 50% intestinal resection and fed chow or a diet enriched with saturated (SFA) or polyunsaturated fatty acids. METHODS: Northern blots were performed. RESULTS: Steroids reduced the jejunal but not the ileal expression of proglucagon. Ornithine decarboxylase (ODC) expression was reduced in the jejunum. CONCLUSIONS: c-jun, ODC, and proglucagon may be involved in the adaptive response that occurs with steroids and variations in dietary lipids after intestinal resection.