RESUMEN
Gut intraepithelial lymphocytes (IELs) are thought to calibrate glucagon-like peptide 1 (GLP-1) bioavailability, thereby regulating systemic glucose and lipid metabolism. Here, we show that the gut IEL GLP-1 receptor (GLP-1R) is not required for enteroendocrine L cell GLP-1 secretion and glucose homeostasis nor for the metabolic benefits of GLP-1R agonists (GLP-1RAs). Instead, the gut IEL GLP-1R is essential for the full effects of GLP-1RAs on gut microbiota. Moreover, independent of glucose control or weight loss, the anti-inflammatory actions of GLP-1RAs require the gut IEL GLP-1R to selectively restrain local and systemic T cell-induced, but not lipopolysaccharide-induced, inflammation. Such effects are mediated by the suppression of gut IEL effector functions linked to the dampening of proximal T cell receptor signaling in a protein-kinase-A-dependent manner. These data reposition key roles of the L cell-gut IEL GLP-1R axis, revealing mechanisms linking GLP-1R activation in gut IELs to modulation of microbiota composition and control of intestinal and systemic inflammation.
Asunto(s)
Microbioma Gastrointestinal , Linfocitos Intraepiteliales , Glucemia , Péptido 1 Similar al Glucagón/metabolismo , Receptor del Péptido 1 Similar al Glucagón , Glucosa/metabolismo , Humanos , Inflamación , Intestinos , Linfocitos Intraepiteliales/metabolismo , Receptores de Antígenos de Linfocitos TRESUMEN
OBJECTIVE: Glucagon-like peptide-1 is a nutrient-sensitive hormone secreted from enteroendocrine L cells within the small and large bowel. Although GLP-1 levels rise rapidly in response to food ingestion, the greatest density of L cells is localized to the distal small bowel and colon. Here, we assessed the importance of the distal gut in the acute L cell response to diverse secretagogues. METHODS: Circulating levels of glucose and plasma GLP-1 were measured in response to the administration of L cell secretagogues in wild-type mice and in mice with (1) genetic reduction of Gcg expression throughout the small bowel and large bowel (GcgGut-/-) and (2) selective reduction of Gcg expression in the distal gut (GcgDistalGut-/-). RESULTS: The acute GLP-1 response to olive oil or arginine administration was markedly diminished in GcgGut-/- but preserved in GcgDistalGut-/- mice. In contrast, the increase in plasma GLP-1 levels following the administration of the GPR119 agonist AR231453, or the melanocortin-4 receptor (MC4R) agonist LY2112688, was markedly diminished in the GcgDistalGut-/- mice. The GLP-1 response to LPS was also markedly attenuated in the GcgGut-/- mice and remained submaximal in the GcgDistalGut-/- mice. Doses of metformin sufficient to lower glucose and increase GLP-1 levels in the GcgGut+/+ mice retained their glucoregulatory activity, yet they failed to increase GLP-1 levels in the GcgGut-/- mice. Surprisingly, the actions of metformin to increase plasma GLP-1 levels were substantially attenuated in the GcgDistalGut-/- mice. CONCLUSION: These findings further establish the importance of the proximal gut for the acute response to nutrient-related GLP-1 secretagogues. In contrast, we identify essential contributions of the distal gut to (i) the rapid induction of circulating GLP-1 levels in response to pharmacological selective agonism of G-protein-coupled receptors, (ii) the increased GLP-1 levels following the activation of Toll-Like Receptors with LPS, and iii) the acute GLP-1 response to metformin. Collectively, these results reveal that distal gut Gcg + endocrine cells are rapid responders to structurally and functionally diverse GLP-1 secretagogues.
Asunto(s)
Células Enteroendocrinas/metabolismo , Péptido 1 Similar al Glucagón/metabolismo , Glucagón/metabolismo , Animales , Glucemia/análisis , Colon/metabolismo , Colon/fisiología , Células Enteroendocrinas/patología , Femenino , Expresión Génica/genética , Regulación de la Expresión Génica/genética , Glucagón/genética , Péptido 1 Similar al Glucagón/sangre , Péptido 1 Similar al Glucagón/genética , Receptor del Péptido 2 Similar al Glucagón/metabolismo , Glucosa/metabolismo , Insulina/metabolismo , Intestino Delgado/metabolismo , Intestino Delgado/fisiología , Masculino , Metformina/farmacología , Ratones , Ratones Noqueados , Proglucagón/metabolismoRESUMEN
A glucagon-like peptide-2 (GLP-2) analog is used in individuals with intestinal failure who are at risk for liver disease, yet the hepatic actions of GLP-2 are not understood. Treatment of high-fat diet-fed (HFD-fed) mice with GLP-2 did not modify the development of hepatosteatosis or hepatic inflammation. In contrast, Glp2r-/- mice exhibited increased hepatic lipid accumulation, deterioration in glucose tolerance, and upregulation of biomarkers of hepatic inflammation. Both mouse and human liver expressed the canonical GLP-2 receptor (GLP-2R), and hepatic Glp2r expression was upregulated in mice with hepatosteatosis. Cell fractionation localized the Glp2r to hepatic stellate cells (HSCs), and markers of HSC activation and fibrosis were increased in livers of Glp2r-/- mice. Moreover, GLP-2 directly modulated gene expression in isolated HSCs ex vivo. Taken together, these findings define an essential role for the GLP-2R in hepatic adaptation to nutrient excess and unveil a gut hormone-HSC axis, linking GLP-2R signaling to control of HSC activation.
Asunto(s)
Hígado Graso/metabolismo , Péptido 2 Similar al Glucagón/metabolismo , Receptor del Péptido 2 Similar al Glucagón/metabolismo , Células Estrelladas Hepáticas/metabolismo , Animales , Dieta Alta en Grasa/efectos adversos , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Transducción de Señal/fisiologíaRESUMEN
Glucagon-like peptide-2 (GLP-2), secreted from enteroendocrine cells, attenuates gut motility, enhances barrier function, and augments nutrient absorption, actions mediated by a single GLP-2 receptor (GLP-2R). Despite extensive analyses, the precise distribution and cellular localization of GLP-2R expression remains controversial, confounded by the lack of suitable GLP-2R antisera. Here, we reassessed murine Glp2r expression using regular and real-time quantitative PCR (qPCR), in situ hybridization (ISH), and a Glp2rLacZ reporter mouse. Glp2r mRNA expression was detected from the stomach to the rectum and most abundant in the jejunum. Glp2r transcripts were also detected in cerebral cortex, mesenteric lymph nodes, gallbladder, urinary bladder, and mesenteric fat. Surprisingly, Glp2r mRNA was found in testis by qPCR at levels similar to jejunum. However, the testis Glp2r transcripts, detected by different primer pairs and qPCR, lacked 5' mRNA coding sequences, and only a minute proportion of them corresponded to full-length Glp2r mRNA. Within the gut, Glp2r-driven LacZ expression was localized to enteric neurons and lamina propria stromal cells, findings confirmed by ISH analysis of the endogenous Glp2r mRNA. Unexpectedly, vascular Glp2rLacZ expression was localized to mesenteric veins and not arteries. Moreover, mesenteric fat Glp2rLacZ expression was detected within blood vessels and not adipocytes. Reporter LacZ expression was not detected in all tissues expressing an endogenous Glp2r transcript, such as gallbladder, urinary bladder, and mesenteric lymph nodes. Collectively, these findings extend our understanding of the cellular domains of Glp2r expression and highlight limitations inherent in application of commonly used technologies to infer analysis of gene expression.
Asunto(s)
Receptor del Péptido 2 Similar al Glucagón/genética , Animales , Tracto Gastrointestinal/metabolismo , Ratones , Ratones Endogámicos C57BL , ARN Mensajero/análisis , Reacción en Cadena en Tiempo Real de la Polimerasa , beta-Galactosidasa/análisisRESUMEN
OBJECTIVE: Therapeutic interventions that improve glucose homeostasis such as attenuation of glucagon receptor (Gcgr) signaling and bariatric surgery share common metabolic features conserved in mice and humans. These include increased circulating levels of bile acids (BA) and the proglucagon-derived peptides (PGDPs), GLP-1 and GLP-2. Whether BA acting through TGR5 (Gpbar1) increases PGDP levels in these scenarios has not been examined. Furthermore, although the importance of GLP-1 action has been interrogated in Gcgr-/- mice and after bariatric surgery, whether GLP-2 contributes to the metabolic benefits of these interventions is not known. METHODS: To assess whether BA acting through Gpbar1 mediates improved glucose homeostasis in Gcgr-/- mice we generated and characterized Gcgr-/-:Gpbar1-/- mice. The contribution of GLP-2 receptor (GLP-2R) signaling to intestinal and metabolic adaptation arising following loss of the Gcgr was studied in Gcgr-/-:Glp2r-/- mice. The role of the GLP-2R in the metabolic improvements evident after bariatric surgery was studied in high fat-fed Glp2r-/- mice subjected to vertical sleeve gastrectomy (VSG). RESULTS: Circulating levels of BA were markedly elevated yet similar in Gcgr-/-:Gpbar1+/+ vs. Gcgr-/-:Gpbar1-/- mice. Loss of GLP-2R lowered levels of BA in Gcgr-/- mice. Gcgr-/-:Glp2r-/- mice also exhibited shifts in the proportion of circulating BA species. Loss of Gpbar1 did not impact body weight, intestinal mass, or glucose homeostasis in Gcgr-/- mice. In contrast, small bowel growth was attenuated in Gcgr-/-:Glp2r-/- mice. The improvement in glucose tolerance, elevated circulating levels of GLP-1, and glucose-stimulated insulin levels were not different in Gcgr-/-:Glp2r+/+ vs. Gcgr-/-:Glp2r-/- mice. Similarly, loss of the GLP-2R did not attenuate the extent of weight loss and improvement in glucose control after VSG. CONCLUSIONS: These findings reveal that GLP-2R controls BA levels and relative proportions of BA species in Gcgr-/- mice. Nevertheless, the GLP-2R is not essential for i) control of body weight or glucose homeostasis in Gcgr-/- mice or ii) metabolic improvements arising after VSG in high fat-fed mice. Furthermore, despite elevations of circulating levels of BA, Gpbar1 does not mediate elevated levels of PGDPs or major metabolic phenotypes in Gcgr-/- mice. Collectively these findings refine our understanding of the relationship between Gpbar1, elevated levels of BA, PGDPs, and the GLP-2R in amelioration of metabolic derangements arising following loss of Gcgr signaling or after vertical sleeve gastrectomy.
Asunto(s)
Ácidos y Sales Biliares/metabolismo , Receptor del Péptido 2 Similar al Glucagón/metabolismo , Receptores de Glucagón/metabolismo , Animales , Glucemia/metabolismo , Peso Corporal/fisiología , Dieta Alta en Grasa , Gastrectomía/métodos , Glucagón , Péptido 1 Similar al Glucagón/metabolismo , Péptido 2 Similar al Glucagón/metabolismo , Receptor del Péptido 1 Similar al Glucagón/genética , Glucosa/metabolismo , Prueba de Tolerancia a la Glucosa , Homeostasis/fisiología , Insulina/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proglucagón/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Glucagón/genética , Transducción de Señal , Pérdida de Peso/fisiologíaRESUMEN
Glucagonlike peptide-1 receptor (GLP-1R) agonists, which are used to treat type 2 diabetes and obesity, reduce the rates of myocardial infarction and cardiovascular death. GLP-1R has been localized to the human sinoatrial node; however, its expression in ventricular tissue remains uncertain. Here we studied GLP-1R expression in the human heart using GLP-1R-directed antisera, quantitative polymerase chain reaction (PCR), reverse transcription PCR to detect full-length messenger RNA (mRNA) transcripts, and in situ hybridization (ISH). GLP1R mRNA transcripts, encompassing the entire open reading frame, were detected in all four cardiac chambers from 15 hearts at levels approximating those detected in human pancreas. In contrast, cardiac GLP2R expression was relatively lower, and cardiac GCGR expression was sporadic and not detected in the left ventricle. GLP1R mRNA transcripts were not detected in RNA from human cardiac fibroblasts, coronary artery endothelial, or vascular smooth muscle cells. Human Brunner glands and pancreatic islets exhibited GLP-1R immunopositivity and abundant expression of GLP1R mRNA transcripts by ISH. GLP1R transcripts were also detected by ISH in human cardiac sinoatrial node tissue. However, definitive cellular localization of GLP1R mRNA transcripts or immunoreactive GLP-1R protein within human cardiomyocytes or cardiac blood vessels remained elusive. Moreover, validated GLP-1R antisera lacked sufficient sensitivity to detect expression of the endogenous islet or cardiac GLP-1R by Western blotting. Hence, although human cardiac ventricles express the GLP1R, the identity of one or more ventricular cell type(s) that express a translated GLP1R protein requires further clarification with highly sensitive methods of detection.
Asunto(s)
Receptor del Péptido 1 Similar al Glucagón/metabolismo , Receptor del Péptido 2 Similar al Glucagón/metabolismo , Miocardio/metabolismo , Animales , Línea Celular , Cricetinae , Femenino , Fibroblastos/metabolismo , Receptor del Péptido 1 Similar al Glucagón/genética , Receptor del Péptido 2 Similar al Glucagón/genética , Humanos , Islotes Pancreáticos/metabolismo , Músculo Liso Vascular/metabolismo , Miocitos del Músculo Liso/metabolismoRESUMEN
Incretin hormones exert pleiotropic metabolic actions beyond the pancreas. Although the heart expresses both incretin receptors, the cardiac biology of GIP receptor (GIPR) action remains incompletely understood. Here we show that GIPR agonism did not impair the response to cardiac ischemia. In contrast, genetic elimination of the Gipr reduced myocardial infarction (MI)-induced ventricular injury and enhanced survival associated with reduced hormone sensitive lipase (HSL) phosphorylation; it also increased myocardial triacylglycerol (TAG) stores. Conversely, direct GIPR agonism in the isolated heart reduced myocardial TAG stores and increased fatty acid oxidation. The cardioprotective phenotype in Gipr-/- mice was partially reversed by pharmacological activation or genetic overexpression of HSL. Selective Gipr inactivation in cardiomyocytes phenocopied Gipr-/- mice, resulting in improved survival and reduced adverse remodeling following experimental MI. Hence, the cardiomyocyte GIPR regulates fatty acid metabolism and the adaptive response to ischemic cardiac injury. These findings have translational relevance for developing GIPR-based therapeutics.
Asunto(s)
Infarto del Miocardio/patología , Receptores de la Hormona Gastrointestinal/metabolismo , Adenilil Ciclasas/metabolismo , Animales , Activación Enzimática , Polipéptido Inhibidor Gástrico/metabolismo , Células HEK293 , Insuficiencia Cardíaca/patología , Humanos , Sistema de Señalización de MAP Quinasas , Ratones , Ratones Endogámicos C57BL , Infarto del Miocardio/diagnóstico por imagen , Infarto del Miocardio/fisiopatología , Infarto del Miocardio/prevención & control , Miocardio/metabolismo , Miocardio/patología , Miocitos Cardíacos/metabolismo , Fosforilación , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores de la Hormona Gastrointestinal/deficiencia , Receptores de la Hormona Gastrointestinal/genética , Transducción de Señal , Esterol Esterasa/metabolismo , Triglicéridos/metabolismo , Remodelación VentricularRESUMEN
OBJECTIVE: Glucagon-like peptides (GLPs) are secreted from enteroendocrine cells in response to nutrients and bile acids and control metabolism via actions on structurally-related yet distinct G protein coupled receptors. GLP-1 regulates gut motility, appetite, islet function, and glucose homeostasis, whereas GLP-2 enhances intestinal nutrient absorption. GLP-1R agonists are used to treat diabetes and obesity, and a GLP-2R agonist is approved to treat short bowel syndrome. Unexpectedly, reports of gallbladder disease have been associated with the use of both GLP-1R and GLP-2R agonists and after bariatric surgery, although the mechanisms remain unknown. METHODS: We investigated whether GLP-1 or GLP-2 acutely controls gallbladder (GB) volume and whether GLP-2 regulates GB muscle activity in mice. The expression of Tgr5, Glp2r, and Glp1r was assessed in mouse GB, and the effects of GLP-2 on hepatic bile acid (BA) flow, intestinal and liver BA uptake, and GB gene expression were determined. GLP-2 regulation of GB volume was assessed in wildtype, Glp2r-/- and Tgr5-/- mice. The effect of GLP-2 on GB smooth muscle (GBSM) calcium transients was characterized ex vivo. RESULTS: Acute administration of the GLP-1R agonist exendin-4 lowered glucose but had no effect on GB volume in mice. In contrast, GLP-2 rapidly enhanced GB filling in a dose-dependent manner, actions maintained in the presence of cholecystokinin, and mediated through the canonical GLP-2R. GLP-2 also rapidly induced immediate early gene expression in GB, consistent with detection of the endogenous Glp2r in GB RNA. The ability of GLP-2 to increase GB volume was not abrogated by systemic administration of hexamethonium, propranolol, a vasoactive peptide receptor antagonist or N-Nitroarginine methyl ester, and was maintained in Tgr5-/- mice. In contrast, lithocholic acid, a Tgr5 agonist, increased GB filling in Glp2r-/- but not in Tgr5-/- mice. GLP-2 had no effect on ileal uptake or hepatic clearance of taurocholic acid or on hepatic bile flow, yet reduced the frequency of spontaneous calcium transients in murine GBSM ex vivo, in a tetrodotoxin-sensitive manner. CONCLUSIONS: Our data extend endocrine concepts of regulation of GB filling beyond FXR-FGF15/19 and the direct effects of BA via Tgr5, to encompass a novel BA-Tgr5-L cell GLP-2 axis providing nutrient-mediated feedback from BA to terminate meal-related GB contraction. These findings have implications for conditions characterized by elevated circulating levels of GLP-2 such as after bariatric surgery and the development and use of agents that promote Tgr5 activation, L cell secretion, or GLP-2R agonism for the treatment of metabolic disease.
Asunto(s)
Vesícula Biliar/efectos de los fármacos , Fármacos Gastrointestinales/farmacología , Receptor del Péptido 2 Similar al Glucagón/metabolismo , Péptidos/farmacología , Receptores Acoplados a Proteínas G/metabolismo , Animales , Vesícula Biliar/metabolismo , Vesícula Biliar/fisiología , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Contracción Muscular , Receptores Acoplados a Proteínas G/genéticaRESUMEN
OBJECTIVE: Glucagon-like peptide-2 (GLP-2) is co-secreted with GLP-1 from gut endocrine cells, and both peptides act as growth factors to expand the surface area of the mucosal epithelium. Notably, GLP-2 also enhances glucose and lipid transport in enterocytes; however, its actions on control of amino acid (AA) transport remain unclear. Here we examined the mechanisms linking gain and loss of GLP-2 receptor (GLP-2R) signaling to control of intestinal amino acid absorption in mice. METHODS: Absorption, transport, and clearance of essential AAs, specifically lysine, were measured in vivo by Liquid Chromatography triple quadrupole Mass Spectrometry (LC-MS/MS) and ex vivo with Ussing chambers using intestinal preparations from Glp2r+/+ and Glp2r-/- mice. Immunoblotting determined jejunal levels of protein components of signaling pathways (PI3K-AKT, and mTORC1-pS6-p4E-BP1) following administration of GLP-2, protein gavage, and rapamycin to fasted Glp2r+/+ and Glp2r-/- mice. Expression of AA transporters from full thickness jejunum and 4F2hc from brush border membrane vesicles (BBMVs) was measured by real-time PCR and immunoblotting, respectively. RESULTS: Acute administration of GLP-2 increased basal AA absorption in vivo and augmented basal lysine transport ex vivo. GLP-2-stimulated lysine transport was attenuated by co-incubation with wortmannin, rapamycin, or tetrodotoxin ex vivo. Phosphorylation of mTORC1 effector proteins S6 and 4E-BP1 was significantly increased in wild-type mice in response to GLP-2 alone, or when co-administered with protein gavage, and abolished following oral gavage of rapamycin. In contrast, activation of GLP-1R signaling did not enhance S6 phosphorylation. Disruption of GLP-2 action in Glp2r-/- mice reduced lysine transport ex vivo and attenuated the phosphorylation of S6 and 4E-BP1 in response to oral protein. Moreover, the expression of cationic AA transporter slc7a9 in response to refeeding, and the abundance of 4F2hc in BBMVs following protein gavage, was significantly attenuated in Glp2r-/- mice. CONCLUSIONS: These findings reveal an important role for GLP-2R signaling in the physiological and pharmacological control of enteral amino acid sensing and assimilation, defining an enteroendocrine cell-enterocyte axis for optimal energy absorption.
Asunto(s)
Sistemas de Transporte de Aminoácidos/metabolismo , Péptido 2 Similar al Glucagón/metabolismo , Absorción Intestinal/efectos de los fármacos , Aminoácidos/metabolismo , Animales , Células Enteroendocrinas/metabolismo , Péptido 1 Similar al Glucagón/metabolismo , Péptido 1 Similar al Glucagón/fisiología , Péptido 2 Similar al Glucagón/fisiología , Receptor del Péptido 2 Similar al Glucagón/metabolismo , Glucosa/metabolismo , Mucosa Intestinal/metabolismo , Intestino Delgado/metabolismo , Yeyuno/metabolismo , Ratones , Ratones Endogámicos C57BL , Péptidos/metabolismo , Receptores de Glucagón/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Pharmacological inhibition of the dipeptidyl peptidase-4 (DPP4) enzyme potentiates incretin action and is widely used to treat type 2 diabetes. Nevertheless, the precise cells and tissues critical for incretin degradation and glucose homeostasis remain unknown. Here, we use mouse genetics and pharmacologic DPP4 inhibition to identify DPP4+ cell types essential for incretin action. Although enterocyte DPP4 accounted for substantial intestinal DPP4 activity, ablation of enterocyte DPP4 in Dpp4Gut-/- mice did not produce alterations in plasma DPP4 activity, incretin hormone levels, and glucose tolerance. In contrast, endothelial cell (EC)-derived DPP4 contributed substantially to levels of soluble plasma DPP4 activity, incretin degradation, and glucose control. Surprisingly, DPP4+ cells of bone marrow origin mediated the selective degradation of fasting GIP, but not GLP-1. Collectively, these findings identify distinct roles for DPP4 in the EC versus the bone marrow compartment for selective incretin degradation and DPP4i-mediated glucoregulation.
Asunto(s)
Dipeptidil Peptidasa 4/metabolismo , Polipéptido Inhibidor Gástrico/metabolismo , Péptido 1 Similar al Glucagón/metabolismo , Glucosa/metabolismo , Homeostasis , Incretinas/metabolismo , Animales , Trasplante de Médula Ósea , Dieta Alta en Grasa , Dipeptidil Peptidasa 4/sangre , Nutrición Enteral , Conducta Alimentaria/efectos de los fármacos , Prueba de Tolerancia a la Glucosa , Homeostasis/efectos de los fármacos , Resistencia a la Insulina , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/metabolismo , Intestinos/efectos de los fármacos , Intestinos/enzimología , Masculino , Ratones , Modelos Biológicos , Fosfato de Sitagliptina/farmacologíaRESUMEN
Glucagon-like peptide-1 (GLP-1) secreted from enteroendocrine L cells promotes nutrient disposal via the incretin effect. However, the majority of L cells are localized to the distal gut, suggesting additional biological roles for GLP-1. Here, we demonstrate that GLP-1 receptor (GLP-1R) signaling controls mucosal expansion of the small bowel (SB) and colon. These actions did not require the epidermal growth factor (EGF) or intestinal epithelial insulin-like growth factor (IGF1) receptors but were absent in Glp1r(-/-) mice. Polyp number and size were increased in SB of exendin-4-treated Apc(Min/+) mice, whereas polyp number was reduced in SB and colon of Glp1r(-/-):Apc(Min/+) mice. Exendin-4 increased fibroblast growth factor 7 (Fgf7) expression in colonic polyps of Apc(Min/+) mice and failed to increase intestinal growth in mice lacking Fgf7. Exogenous exendin-4 and Fgf7 regulated an overlapping set of genes important for intestinal growth. Thus, gain and loss of GLP-1R signaling regulates gut growth and intestinal tumorigenesis.
Asunto(s)
Colon/metabolismo , Factor 7 de Crecimiento de Fibroblastos/metabolismo , Receptor del Péptido 1 Similar al Glucagón/agonistas , Receptor del Péptido 1 Similar al Glucagón/metabolismo , Mucosa Intestinal/metabolismo , Intestino Delgado/metabolismo , Animales , Carcinogénesis/metabolismo , Proliferación Celular/fisiología , Colon/fisiología , Colon/fisiopatología , Factor de Crecimiento Epidérmico/metabolismo , Exenatida , Femenino , Incretinas/metabolismo , Mucosa Intestinal/fisiología , Mucosa Intestinal/fisiopatología , Intestino Delgado/patología , Intestino Delgado/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Péptidos/metabolismo , Receptor IGF Tipo 1/metabolismo , Transducción de Señal/fisiología , Ponzoñas/metabolismoRESUMEN
Obesity and diabetes are characterized by increased inflammation reflecting disordered control of innate immunity. We reveal a local intestinal intraepithelial lymphocyte (IEL)-GLP-1 receptor (GLP-1R) signaling network that controls mucosal immune responses. Glp1r expression was enriched in intestinal IEL preparations and copurified with markers of Tαß and Tγδ IELs, the two main subsets of intestinal IELs. Exendin-4 increased cAMP accumulation in purified IELs and reduced the production of cytokines from activated IELs but not from splenocytes ex vivo. These actions were mimicked by forskolin, absent in IELs from Glp1r(-/-) mice, and attenuated by the GLP-1R agonist exendin (9-39) consistent with a GLP-1R-dependent mechanism of action. Furthermore, Glp1r(-/-) mice exhibited dysregulated intestinal gene expression, an abnormal representation of microbial species in feces, and enhanced sensitivity to intestinal injury following administration of dextran sodium sulfate. Bone marrow transplantation using wild-type C57BL/6 donors normalized expression of multiple genes regulating immune function and epithelial integrity in Glp1r(-/-) recipient mice, whereas acute exendin-4 administration robustly induced the expression of genes encoding cytokines and chemokines in normal and injured intestine. Taken together, these findings define a local enteroendocrine-IEL axis linking energy availability, host microbial responses, and mucosal integrity to the control of innate immunity.
Asunto(s)
Mucosa Intestinal/inmunología , Linfocitos/inmunología , Receptores de Glucagón/agonistas , Receptores de Glucagón/fisiología , Animales , Citocinas/análisis , Exenatida , Femenino , Péptido 1 Similar al Glucagón/metabolismo , Péptido 1 Similar al Glucagón/farmacología , Receptor del Péptido 1 Similar al Glucagón , Inmunidad Innata , Masculino , Ratones , Ratones Endogámicos C57BL , Péptidos/farmacología , Transducción de Señal , Ponzoñas/farmacologíaRESUMEN
Glucagon-like peptide-2 (GLP-2) is a 33-amino-acid proglucagon-derived peptide secreted from enteroendocrine L cells. GLP-2 circulates at low basal levels in the fasting period, and plasma levels rise rapidly after food ingestion. Renal clearance and enzymatic inactivation control the elimination of bioactive GLP-2. GLP-2 increases mesenteric blood flow and activates proabsorptive pathways in the gut, facilitating nutrient absorption. GLP-2 also enhances gut barrier function and induces proliferative and cytoprotective pathways in the small bowel. The actions of GLP-2 are transduced via a single G protein-coupled receptor (GLP-2R), expressed predominantly within the gastrointestinal tract. Disruption of GLP-2R signaling increases susceptibility to gut injury and impairs the adaptive mucosal response to refeeding. Sustained augmentation of GLP-2R signaling reduces the requirement for parenteral nutrition in human subjects with short-bowel syndrome. Hence GLP-2 integrates nutrient-derived signals to optimize mucosal integrity and energy absorption.
Asunto(s)
Péptido 2 Similar al Glucagón/farmacología , Péptido 2 Similar al Glucagón/fisiología , Animales , Enfermedades Gastrointestinales/fisiopatología , Motilidad Gastrointestinal/fisiología , Tracto Gastrointestinal/irrigación sanguínea , Tracto Gastrointestinal/fisiología , Receptor del Péptido 2 Similar al Glucagón , Humanos , Absorción Intestinal , Isquemia/fisiopatología , Traumatismos por Radiación/fisiopatología , Receptores de Glucagón/biosíntesis , Receptores de Glucagón/efectos de los fármacos , Receptores de Glucagón/metabolismo , Síndrome del Intestino Corto/fisiopatología , Transducción de SeñalRESUMEN
Glucagon-like peptide-1 receptor (GLP-1R) agonists reduce lipid accumulation in peripheral tissues, attenuating atherosclerosis and hepatic steatosis in preclinical studies. We examined whether GLP-1R activation decreases atherosclerosis progression in high-fat diet-fed male ApoE(-/-) mice after administration of streptozotocin and treatment with the long-acting GLP-1R agonist taspoglutide administered once monthly vs. metformin in the drinking water for 12 wk. Taspoglutide did not reduce plaque area or lipid content in the aortic arch or abdominal aorta, and no significant change in aortic macrophage accumulation was detected after taspoglutide or metformin. In contrast, hepatic triglyceride levels were significantly reduced in livers from taspoglutide-treated mice. Both peripheral and intracerebroventricular administration of exendin-4 rapidly decreased plasma triglyceride levels in fasted mice, and taspoglutide therapy in ApoE(-/-) mice modulated the expression of hepatic genes controlling fatty acid uptake and oxidation. We were unable to detect expression of the entire Glp1r coding sequence in macrophages isolated from ApoE(-/-), C57BL/6, and IL10(-/-) mice. Similarly, Glp1r mRNA transcripts were not detected in RNA from isolated murine hepatocytes. Using Western blotting and tissue extracts from Glp1r(+/+) and Glp1r(-/-) mice, and cells transfected with a tagged murine GLP-1R cDNA, we could not validate the sensitivity and specificity of three different GLP-1R antisera commonly used for the detection of GLP-1R protein. Taken together, these findings illustrate divergent actions of GLP-1R agonists on atherosclerosis progression and accumulation of ectopic lipid in ApoE(-/-) mice and highlight the importance of indirect GLP-1R actions for the control of hepatic lipid accumulation.
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Aterosclerosis/metabolismo , Hígado/metabolismo , Receptores de Glucagón/metabolismo , Animales , Apolipoproteínas E/deficiencia , Apolipoproteínas E/genética , Western Blotting , Composición Corporal/efectos de los fármacos , Receptor del Péptido 1 Similar al Glucagón , Prueba de Tolerancia a la Glucosa , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Interleucina-10/deficiencia , Interleucina-10/genética , Hígado/efectos de los fármacos , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , Ratones , Ratones Noqueados , Péptidos/farmacología , Receptores de Glucagón/agonistas , Receptores de Glucagón/genéticaRESUMEN
The enteroendocrine and enteric nervous systems convey signals through an overlapping network of regulatory peptides that act either as circulating hormones or as localized neurotransmitters within the gastrointestinal tract. Because recent studies invoke an important role for vasoactive intestinal peptide (VIP) as a downstream mediator of glucagon-like peptide-2 (GLP-2) action in the gut, we examined the importance of the VIP-GLP-2 interaction through analysis of Vip(-/-) mice. Unexpectedly, we detected abnormal villous architecture, expansion of the crypt compartment, increased crypt cell proliferation, enhanced Igf1 and Kgf gene expression, and reduced expression of Paneth cell products in the Vip(-/-) small bowel. These abnormalities were not reproduced by antagonizing VIP action in wild-type mice, and VIP administration did not reverse the intestinal phenotype of Vip(-/-) mice. Exogenous administration of GLP-2 induced the expression of ErbB ligands and immediate-early genes to similar levels in Vip(+/+) vs. Vip(-/-) mice. Moreover, GLP-2 significantly increased crypt cell proliferation and small bowel growth to comparable levels in Vip(+/+) vs. Vip(-/-) mice. Unexpectedly, exogenous GLP-2 administration had no therapeutic effect in mice with dextran sulfate-induced colitis; the severity of colonic injury and weight loss was modestly reduced in female but not male Vip(-/-) mice. Taken together, these findings extend our understanding of the complex intestinal phenotype arising from loss of the Vip gene. Furthermore, although VIP action may be important for the antiinflammatory actions of GLP-2, the Vip gene is not required for induction of a gene expression program linked to small bowel growth after enhancement of GLP-2 receptor signaling.
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Regulación de la Expresión Génica/efectos de los fármacos , Péptido 2 Similar al Glucagón/farmacología , Transducción de Señal/efectos de los fármacos , Péptido Intestinal Vasoactivo/metabolismo , Proteína de la Poliposis Adenomatosa del Colon/genética , Proteína de la Poliposis Adenomatosa del Colon/metabolismo , Animales , Western Blotting , Proliferación Celular/efectos de los fármacos , Colon/efectos de los fármacos , Colon/metabolismo , Colon/patología , Sistema Nervioso Entérico/efectos de los fármacos , Sistema Nervioso Entérico/metabolismo , Femenino , Receptor del Péptido 2 Similar al Glucagón , Factor I del Crecimiento Similar a la Insulina/genética , Factor I del Crecimiento Similar a la Insulina/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Péptidos y Proteínas de Señalización Intercelular/farmacología , Mucosa Intestinal/metabolismo , Intestino Delgado/efectos de los fármacos , Intestino Delgado/metabolismo , Intestinos/efectos de los fármacos , Yeyuno/efectos de los fármacos , Yeyuno/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Tamaño de los Órganos/efectos de los fármacos , Células de Paneth/efectos de los fármacos , Células de Paneth/metabolismo , Receptores de Glucagón/genética , Receptores de Glucagón/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores Sexuales , Péptido Intestinal Vasoactivo/genéticaRESUMEN
Exogenous glucagon-like peptide-2 receptor (GLP-2R) activation elicits proliferative and cytoprotective responses in the gastrointestinal mucosa and ameliorates experimental small and large bowel gut injury. Nevertheless, the essential physiological role(s) of the endogenous GLP-2R remain poorly understood. We studied the importance of the GLP-2R for gut growth, epithelial cell lineage allocation, the response to mucosal injury, and host-bacterial interactions in Glp2r(-/-) and littermate control Glp2r(+/+) mice. Glp2r(-/-) mice exhibit normal somatic growth and preserved small and large bowel responses to IGF-I and keratinocyte growth factor. However, Glp2r(-/-) mice failed to up-regulate intestinal epithelial c-fos expression in response to acute GLP-2 administration and do not exhibit changes in small bowel conductance or small or large bowel growth after administration of GLP-2R agonists. The crypt and villus compartment and the numbers and localization of Paneth, enteroendocrine, and goblet cells were comparable in Glp2r(+/+) vs. Glp2r(-/-) mice. Although the severity and extent of colonic mucosal injury in response to 3% oral dextran sulfate was similar across Glp2r genotypes, Glp2r(-/-) mice exhibited significantly increased morbidity and mortality and increased bacterial translocation after induction of enteritis with indomethacin and enhanced mucosal injury in response to irinotecan. Moreover, bacterial colonization of the small bowel was significantly increased, expression of Paneth cell antimicrobial gene products was reduced, and mucosal bactericidal activity was impaired in Glp2r(-/-) mice. Although the Glp2r is dispensable for gut development and the response to colonic injury, Glp2r(-/-) mice exhibit enhanced sensitivity to small bowel injury, and abnormal host-bacterial interactions in the small bowel.
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Enteritis/patología , Predisposición Genética a la Enfermedad , Intestinos/patología , Células de Paneth/citología , Receptores de Glucagón/genética , Receptores de Glucagón/fisiología , Animales , Peso Corporal , Exones , Femenino , Factor 7 de Crecimiento de Fibroblastos/metabolismo , Receptor del Péptido 2 Similar al Glucagón , Humanos , Factor I del Crecimiento Similar a la Insulina/metabolismo , Mucosa Intestinal/metabolismo , Intestino Delgado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Permeabilidad , ARN Ribosómico 16S/metabolismoRESUMEN
BACKGROUND & AIMS: The small bowel mucosa is sensitive to nutrients and undergoes rapid adaptation to nutrient deprivation and refeeding through changes in apoptosis and cell proliferation, respectively. Although glucagon-like peptide-2 (GLP-2) exerts trophic effects on the gut and levels increase with refeeding, mechanisms linking GLP-2 to mucosal adaptation to refeeding remain unclear. METHODS: Fasting and refeeding were studied in wild-type (WT) and Glp2r(-/-) mice and in WT mice treated with the pan ErbB inhibitor CI-1033. Experimental end points included intestinal weights, histomorphometry, gene and protein expression, and crypt cell proliferation. RESULTS: Fasting was associated with significant reductions in small bowel mass, decreased crypt plus villus height, and reduced crypt cell proliferation. Refeeding increased plasma levels of GLP-2, reversed small bowel atrophy, increased villus height and cell number, and stimulated jejunal crypt cell proliferation. In contrast, refeeding failed to increase small bowel weight, crypt cell proliferation, or villus cell number in Glp2r(-/-) mice. Levels of mRNA transcripts for egf, kgf, and igfr were lower in fasted Glp2r(-/-) mice. Epidermal growth factor but not insulin-like growth factor-1 restored the intestinal adaptive response to refeeding in Glp2r(-/-) mice. Furthermore, CI-1033 prevented adaptive crypt cell proliferation, Akt activation, and induction of ErbB ligand gene expression after refeeding. Up-regulation of ErbB ligand expression and intestinal Akt phosphorylation were significantly diminished in refed Glp2r(-/-) mice. CONCLUSIONS: These findings identify Glp2r and ErbB pathways as essential components of the signaling network regulating the adaptive mucosal response to refeeding in the mouse intestine.
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Adaptación Fisiológica , Receptores ErbB/fisiología , Intestino Delgado/fisiología , Receptores de Glucagón/fisiología , Animales , Proliferación Celular , Ingestión de Alimentos , Factor de Crecimiento Epidérmico/farmacología , Receptor del Péptido 2 Similar al Glucagón , Factor I del Crecimiento Similar a la Insulina/farmacología , Ratones , Ratones Endogámicos C57BL , Fenotipo , Transducción de SeñalRESUMEN
BACKGROUND & AIMS: Glucagon-like peptide-2 (GLP-2) is a 33-amino acid peptide hormone secreted by enteroendocrine cells in response to nutrient ingestion. GLP-2 stimulates crypt cell proliferation leading to expansion of the mucosal epithelium; however, the mechanisms transducing the trophic effects of GLP-2 are incompletely understood. METHODS: We examined the gene expression profiles and growth-promoting actions of GLP-2 in normal mice in the presence or absence of an inhibitor of ErbB receptor signaling, in Glp2r(-/-) mice and in Egfr(wa2) mice harboring a hypomorphic point mutation in the epidermal growth factor receptor. RESULTS: Exogenous GLP-2 administration rapidly induced the expression of a subset of ErbB ligands including amphiregulin, epiregulin, and heparin binding (HB)-epidermal growth factor, in association with induction of immediate early gene expression in the small and large bowel. These actions of GLP-2 required a functional GLP-2 receptor because they were eliminated in Glp2r(-/-) mice. In contrast, insulin-like growth factor-I and keratinocyte growth factor, previously identified mediators of GLP-2 action, had no effect on the expression of these ErbB ligands. The GLP-2-mediated induction of ErbB ligand expression was not metalloproteinase inhibitor sensitive but was significantly diminished in Egfr(wa2) mice and completed abrogated in wild-type mice treated with the pan-ErbB inhibitor CI-1033. Furthermore, the stimulatory actions of GLP-2 on crypt cell proliferation and bowel growth were eliminated in the presence of CI-1033. CONCLUSIONS: These findings identify the ErbB signaling network as a target for GLP-2 action leading to stimulation of growth factor-dependent signal transduction and bowel growth in vivo.
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Proliferación Celular , Colon/citología , Péptido 2 Similar al Glucagón/farmacología , Yeyuno/citología , Proteínas Oncogénicas v-erbB/metabolismo , Transducción de Señal , Anfirregulina , Animales , Proliferación Celular/efectos de los fármacos , Colon/efectos de los fármacos , Colon/metabolismo , Familia de Proteínas EGF , Factor de Crecimiento Epidérmico/metabolismo , Epirregulina , Receptores ErbB/genética , Femenino , Glicoproteínas/metabolismo , Factor de Crecimiento Similar a EGF de Unión a Heparina , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Yeyuno/efectos de los fármacos , Yeyuno/metabolismo , Masculino , Ratones , Ratones Noqueados , Proteínas Oncogénicas v-erbB/antagonistas & inhibidores , Mutación PuntualRESUMEN
Glucagon-like peptide-2 (GLP-2) secreted from enteroendocrine cells exerts proabsorptive, regenerative, and cytoprotective actions in the normal and injured gut epithelium. Hence, sustained GLP-2 receptor (GLP-2R) activation represents a strategy under investigation for the prevention and treatment of chemotherapy-induced mucositis. Nevertheless, the consequences of increased GLP-2R signaling for the growth and survival of intestinal tumor cells remain poorly understood. We studied the proliferative and cytoprotective actions of GLP-2 in human colon cancer cells stably transfected with the GLP-2R and in nude mice harboring GLP-2R(+) human colon cancer cells. The importance of the GLP-2R for tumor growth was also examined in Apc(Min/+) mice chronically treated with exogenous GLP-2 and in Apc(Min/+):Glp2r(-/-) mice. GLP-2 increased cyclic AMP accumulation and produced cell-specific activation of growth and survival pathways in DLD-1, SW480, and HT29 cells. However, GLP-2 did not stimulate cell growth or attenuate cycloheximide-, LY294002-, indomethacin-, or chemotherapy-induced cytotoxicity in vitro. Moreover, chronic GLP-2 administration had no effect on the growth of human colon cancer cell xenografts in nude mice in vivo. Daily GLP-2 treatment for 7 weeks increased growth of normal gut mucosa but did not increase the number or size of polyps in Apc(Min/+) mice, and genetic disruption of the Glp2r gene in Apc(Min/+) mice did not modify polyp size or number. Taken together, although GLP-2R activation engages signaling pathways promoting cell proliferation and cytoprotection in the normal gut epithelium, sustained direct or indirect modulation of GLP-2R signaling does not modify intestinal tumor cell growth or survival.
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Proliferación Celular/efectos de los fármacos , Péptido 2 Similar al Glucagón/farmacología , Neoplasias Intestinales/patología , Animales , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Femenino , Genes APC , Receptor del Péptido 2 Similar al Glucagón , Células HT29 , Humanos , Neoplasias Intestinales/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Desnudos , Receptores de Glucagón/genética , Receptores de Glucagón/metabolismo , Células Tumorales CultivadasRESUMEN
BACKGROUND & AIMS: Glucagon action in the liver is essential for control of glucose homeostasis and the counterregulatory response to hypoglycemia. Because receptors for the related peptides glucagon-like peptide-1 and glucagon-like peptide-2 regulate beta-cell and enterocyte apoptosis, respectively, we examined whether glucagon receptor (Gcgr) signaling modulates hepatocyte survival. METHODS: The importance of the Gcgr for hepatocyte cell survival was examined using Gcgr+/+ and Gcgr-/- mice in vivo, and murine hepatocyte cultures in vitro. RESULTS: Gcgr-/- mice showed enhanced susceptibility to experimental liver injury induced by either Fas Ligord activation or a methionine- and choline-deficient diet. Restoration of hepatic Gcgr expression in Gcgr-/- mice attenuated the development of hepatocellular injury. Furthermore, exogenous glucagon administration reduced Jo2-induced apoptosis in wild-type mice and decreased caspase activation in fibroblasts expressing a heterologous Gcgr and in primary murine hepatocyte cultures. The anti-apoptotic actions of glucagon were independent of protein kinase A, phosphatidylinositol-3K, and mitogen-activated protein kinase, and were mimicked by the exchange protein directly activated by the cyclic AMP agonist 8-(4-chloro-phenylthio)-2'-O-methyladenosine-3', 5'-cyclic monophosphate-cAMP. CONCLUSIONS: These findings extend the essential actions of the Gcgr beyond the metabolic control of glucose homeostasis to encompass the regulation of hepatocyte survival.