RESUMO
OBJECTIVE: The gut hormone glucose-dependent insulinotropic polypeptide (GIP) stimulates beta cell function and improves glycemia through its incretin actions. GIP also regulates endothelial function and suppresses adipose tissue inflammation through control of macrophage activity. Activation of the GIP receptor (GIPR) attenuates experimental atherosclerosis and inflammation in mice, however whether loss of GIPR signaling impacts the development of atherosclerosis is uncertain. METHODS: Atherosclerosis and related metabolic phenotypes were studied in Apoe-/-:Gipr-/- mice and in Gipr+/+ and Gipr-/- mice treated with an adeno-associated virus expressing PCSK9 (AAV-PCSK9). Bone marrow transplantation (BMT) studies were carried out using donor marrow from Apoe-/-:Gipr-/-and Apoe-/-:Gipr+/+mice transplanted into Apoe-/-:Gipr-/- recipient mice. Experimental endpoints included the extent of aortic atherosclerosis and inflammation, body weight, glucose tolerance, and circulating lipid levels, the proportions and subsets of circulating leukocytes, and tissue gene expression profiles informing lipid and glucose metabolism, and inflammation. RESULTS: Body weight was lower, circulating myeloid cells were reduced, and glucose tolerance was not different, however, aortic atherosclerosis was increased in Apoe-/-:Gipr-/- mice and trended higher in Gipr-/- mice with atherosclerosis induced by AAV-PCSK9. Levels of mRNA transcripts for genes contributing to inflammation were increased in the aortae of Apoe-/-:Gipr-/- mice and expression of a subset of inflammation-related hepatic genes were increased in Gipr-/- mice treated with AAV-PCSK9. BMT experiments did not reveal marked atherosclerosis, failing to implicate bone marrow derived GIPR + cells in the control of atherosclerosis or aortic inflammation. CONCLUSIONS: Loss of the Gipr in mice results in increased aortic atherosclerosis and enhanced inflammation in aorta and liver, despite reduced weight gain and preserved glucose homeostasis. These findings extend concepts of GIPR in the suppression of inflammation-related pathophysiology beyond its classical incretin role in the control of metabolism.
Assuntos
Aterosclerose , Pró-Proteína Convertase 9 , Animais , Camundongos , Apolipoproteínas E/genética , Aterosclerose/genética , Glicemia , Peso Corporal , Polipeptídeo Inibidor Gástrico/metabolismo , Incretinas , Inflamação/metabolismo , Receptores Acoplados a Proteínas G , Receptores dos Hormônios Gastrointestinais , RNA MensageiroRESUMO
OBJECTIVE: Glucose-dependent insulinotropic polypeptide (GIP) and Glucagon-like peptide-1 (GLP-1) are incretin hormones that exert overlapping yet distinct actions on islet ß-cells. We recently observed that GIP, but not GLP-1, upregulated islet expression of Transcription Factor 7 (TCF7), a gene expressed in immune cells and associated with the risk of developing type 1 diabetes. TCF7 has also been associated with glucose homeostasis control in the liver. Herein we studied the relative metabolic importance of TCF7 expression in hepatocytes vs. islet ß-cells in mice. METHODS: Tcf7 expression was selectively inactivated in adult mouse hepatocytes using adenoviral Cre expression and targeted in ß-cells using two different lines of insulin promoter-Cre mice. Glucose homeostasis, plasma insulin and triglyceride responses, islet histology, hepatic and islet gene expression, and body weight gain were evaluated in mice fed regular chow or high fat diets. Tcf7 expression within pancreatic islets and immune cells was evaluated using published single cell RNA-seq (scRNA-seq) data, and in islet RNA from immunodeficient Rag2-/-Il2rg-/- mice. RESULTS: Reduction of hepatocyte Tcf7 expression did not impair glucose homeostasis, lipid tolerance or hepatic gene expression profiles linked to control of metabolic or immune pathways. Similarly, oral and intraperitoneal glucose tolerance, plasma insulin responses, islet histology, body weight gain, and insulin tolerance were not different in mice with targeted recombination of Tcf7 in insulin-positive ß-cells. Surprisingly, islet Tcf7 mRNA transcripts were not reduced in total islet RNA containing endocrine and associated non-endocrine cell types from Tcf7ßcell-/- mice, despite Cre-mediated recombination of islet genomic DNA. Furthermore, glucose tolerance was normal in whole body Tcf7-/- mice. Analysis of scRNA-seq datasets localized pancreatic Tcf7 expression to islet progenitors during development, and immune cells, but not within differentiated islet ß-cells or endocrine lineages within mature islets. Moreover, the expression of Tcf7 was extremely low in islet RNA from Rag2-/-Il2rg-/- mice and, consistent with expression within immune cells, Tcf7 was highly correlated with levels of Cd3g mRNA transcripts in RNA from wild type mouse islets. CONCLUSIONS: These findings demonstrate that Tcf7 expression is not a critical determinant of glucose homeostasis in mice. Moreover, the detection of Tcf7 expression within islet mRNA is attributable to the expression of Tcf7 RNA in islet-associated murine immune cells, and not in islet ß-cells.
Assuntos
Glicemia/metabolismo , Fator 1-alfa Nuclear de Hepatócito/metabolismo , Hepatócitos/metabolismo , Homeostase/genética , Células Secretoras de Insulina/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Animais , Células Cultivadas , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Feminino , Expressão Gênica , Fator 1-alfa Nuclear de Hepatócito/genética , Insulina/sangue , Insulina/genética , Subunidade gama Comum de Receptores de Interleucina/genética , Subunidade gama Comum de Receptores de Interleucina/metabolismo , Linfócitos/metabolismo , Masculino , Camundongos , Camundongos Knockout , Hepatopatia Gordurosa não Alcoólica/genética , Triglicerídeos/sangue , Aumento de Peso/genéticaRESUMO
Postprandial triglycerides (TGs) are elevated in people with type 2 diabetes (T2D). Glucose-lowering agents, such as glucagon-like peptide-1 (GLP-1) receptor agonists and dipeptidyl peptidase-4 (DPP-4) inhibitors, also reduce postprandial TG excursion. Although the glucose-lowering mechanisms of DPP-4 have been extensively studied, how the reduction of DPP-4 activity improves lipid tolerance remains unclear. Here, we demonstrate that gut-selective and systemic inhibition of DPP-4 activity reduces postprandial TG excursion in young mice. Genetic inactivation of Dpp4 simultaneously within endothelial cells and hematopoietic cells using Tie2-Cre reduced intestinal lipoprotein secretion under regular chow diet conditions. Bone marrow transplantation revealed a key role for hematopoietic cells in modulation of lipid responses arising from genetic reduction of DPP-4 activity. Unexpectedly, deletion of Dpp4 in enterocytes increased TG excursion in high-fat diet-fed (HFD-fed) mice. Moreover, chemical reduction of DPP-4 activity and increased levels of GLP-1 were uncoupled from TG excursion in older or HFD-fed mice, yet lipid tolerance remained improved in older Dpp4-/- and Dpp4EC-/- mice. Taken together, this study defines roles for specific DPP-4 compartments, age, and diet as modifiers of DPP-4 activity linked to control of gut lipid metabolism.
Assuntos
Dipeptidil Peptidase 4/metabolismo , Enterócitos/enzimologia , Triglicerídeos/metabolismo , Animais , Transplante de Medula Óssea , Dieta Hiperlipídica/efeitos adversos , Dipeptidil Peptidase 4/sangue , Dipeptidil Peptidase 4/genética , Dipeptidil Peptidase 4/isolamento & purificação , Inibidores da Dipeptidil Peptidase IV/farmacologia , Peptídeo 1 Semelhante ao Glucagon/sangue , Células-Tronco Hematopoéticas/enzimologia , Mucosa Intestinal/enzimologia , Mucosa Intestinal/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/fisiologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Camundongos Transgênicos , Período Pós-Prandial/efeitos dos fármacos , Período Pós-Prandial/fisiologia , Fosfato de Sitagliptina/farmacologiaRESUMO
Dipeptidyl peptidase-4 (DPP4) modulates inflammation by enzymatic cleavage of immunoregulatory peptides and through its soluble form (sDPP4) that directly engages immune cells. Here we examine whether reduction of DPP4 activity alters inflammation. Prolonged DPP4 inhibition increases plasma levels of sDPP4, and induces sDPP4 expression in lymphocyte-enriched organs in mice. Bone marrow transplantation experiments identify hematopoietic cells as the predominant source of plasma sDPP4 following catalytic DPP4 inhibition. Surprisingly, systemic DPP4 inhibition increases plasma levels of inflammatory markers in regular chow-fed but not in high fat-fed mice. Plasma levels of sDPP4 and biomarkers of inflammation are lower in metformin-treated subjects with type 2 diabetes (T2D) and cardiovascular disease, yet exhibit considerable inter-individual variation. Sitagliptin therapy for 12 months reduces DPP4 activity yet does not increase markers of inflammation or levels of sDPP4. Collectively our findings dissociate levels of DPP4 enzyme activity, sDPP4 and biomarkers of inflammation in mice and humans.
Assuntos
Doenças Cardiovasculares/imunologia , Diabetes Mellitus Tipo 2/tratamento farmacológico , Dipeptidil Peptidase 4/sangue , Inibidores da Dipeptidil Peptidase IV/efeitos adversos , Inflamação/imunologia , Idoso , Animais , Biomarcadores/análise , Biomarcadores/metabolismo , Doenças Cardiovasculares/sangue , Doenças Cardiovasculares/prevenção & controle , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/imunologia , Dieta Aterogênica/efeitos adversos , Dieta Hiperlipídica/efeitos adversos , Dipeptidil Peptidase 4/imunologia , Dipeptidil Peptidase 4/metabolismo , Inibidores da Dipeptidil Peptidase IV/administração & dosagem , Modelos Animais de Doenças , Feminino , Receptor do Peptídeo Semelhante ao Glucagon 1/genética , Humanos , Inflamação/sangue , Inflamação/diagnóstico , Inflamação/tratamento farmacológico , Mediadores da Inflamação/análise , Mediadores da Inflamação/metabolismo , Masculino , Metformina/administração & dosagem , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/sangue , Isoformas de Proteínas/metabolismo , Fosfato de Sitagliptina/administração & dosagem , Fosfato de Sitagliptina/efeitos adversosRESUMO
OBJECTIVE: Glucose-dependent insulinotropic polypeptide (GIP) is secreted from the gut in response to nutrient ingestion and promotes meal-dependent insulin secretion and lipid metabolism. Loss or attenuation of GIP receptor (GIPR) action leads to resistance to diet-induced obesity through incompletely understood mechanisms. The GIPR is expressed in white adipose tissue; however, its putative role in brown adipose tissue (BAT) has not been explored. METHODS: We investigated the role of the GIPR in BAT cells in vitro and in BAT-specific (GiprBAT-/-) knockout mice with selective elimination of the Gipr within the Myf5+ expression domain. We analyzed body weight, adiposity, glucose homeostasis, insulin and lipid tolerance, energy expenditure, food intake, body temperature, and iBAT oxygen consumption ex vivo. High-fat diet (HFD)-fed GiprBAT-/- mice were studied at room temperature (21 °C), 4 °C, and 30 °C ambient temperatures. RESULTS: The mouse Gipr gene is expressed in BAT, and GIP directly increased Il6 mRNA and IL-6 secretion in BAT cells. Additionally, levels of thermogenic, lipid and inflammation mRNA transcripts were altered in BAT cells transfected with Gipr siRNA. Body weight gain, energy expenditure, and glucose and insulin tolerance were normal in HFD-fed GiprBAT-/- mice housed at room temperature. However, GiprBAT-/- mice exhibited higher body temperatures during an acute cold challenge and a lower respiratory exchange ratio and impaired lipid tolerance at 21 °C. In contrast, body weight was lower and iBAT oxygen consumption was higher in HFD-fed mice housed at 4 °C but not at 30 °C. CONCLUSIONS: The BAT GIPR is linked to the control of metabolic gene expression, fuel utilization, and oxygen consumption. However, the selective loss of the GIPR within BAT is insufficient to recapitulate the findings of decreased weight gain and resistance to obesity arising in experimental models with systemic disruption of GIP action.
Assuntos
Tecido Adiposo Marrom/metabolismo , Receptores dos Hormônios Gastrointestinais/metabolismo , Animais , Linhagem Celular , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Glucagon-like peptide 1 (GLP-1) receptors are widely distributed throughout the nervous system, enabling physiological and pharmacological control of glucose and energy homeostasis. Here we elucidated the importance of Glp1r expression within cellular domains targeted by expression of Wnt1-Cre2 or Phox2b-Cre. Widespread loss of neural Glp1r in Glp1rΔWnt1-/- mice had no effect on basal food intake, gastric emptying, and glucose homeostasis. However, the glucoregulatory actions of GLP-1R agonists, but not gut-selective DPP-4 inhibition, were preserved in Glp1rΔWnt1-/- mice. Unexpectedly, selective reduction of Glp1r expression within neurons targeted by Phox2b-Cre impaired glucose homeostasis and gastric emptying and attenuated the extent of weight loss achieved with sustained GLP-1R agonism. Collectively, these studies identify discrete neural domains of Glp1r expression mediating GLP-1-regulated control of metabolism and the gut-brain axis and reveal the unexpected importance of neuronal Phox2b+ cells expressing GLP-1R for physiological regulation of gastric emptying, islet hormone responses, and glucose homeostasis.
Assuntos
Encéfalo/metabolismo , Ingestão de Alimentos , Esvaziamento Gástrico , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Glucose/metabolismo , Incretinas/metabolismo , Animais , Encéfalo/citologia , Encéfalo/efeitos dos fármacos , Encéfalo/fisiologia , Peptídeo 1 Semelhante ao Glucagon/análogos & derivados , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Receptor do Peptídeo Semelhante ao Glucagon 1/genética , Peptídeos Semelhantes ao Glucagon/análogos & derivados , Peptídeos Semelhantes ao Glucagon/farmacologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Homeostase , Fragmentos Fc das Imunoglobulinas/farmacologia , Incretinas/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Neurônios/fisiologia , Proteínas Recombinantes de Fusão/farmacologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteína Wnt1/genética , Proteína Wnt1/metabolismoRESUMO
Dipeptidyl peptidase-4 (DPP-4) controls glucose homeostasis through enzymatic termination of incretin action. We report that plasma DPP-4 activity correlates with body weight and fat mass, but not glucose control, in mice. Genetic disruption of adipocyte Dpp4 expression reduced plasma DPP-4 activity in older mice but did not perturb incretin levels or glucose homeostasis. Knockdown of hepatocyte Dpp4 completely abrogated the obesity-associated increase in plasma DPP-4 activity, reduced liver cytokine expression, and partially attenuated inflammation in adipose tissue without changes in incretin levels or glucose homeostasis. In contrast, circulating levels of soluble DPP4 (sDPP4) were dissociated from inflammation in mice with endothelial-selective or global genetic inactivation of Dpp4. Remarkably, inhibition of DPP-4 enzymatic activity upregulated circulating levels of sDPP4 originating from endothelial or hematopoietic cells without inducing systemic or localized inflammation. Collectively, these findings reveal unexpected complexity in regulation of soluble versus enzymatic DPP-4 and control of inflammation and glucose homeostasis.
Assuntos
Dipeptidil Peptidase 4/fisiologia , Glucose/metabolismo , Hepatócitos/metabolismo , Incretinas/metabolismo , Inflamação/imunologia , Obesidade/metabolismo , Células 3T3-L1 , Animais , Citocinas/metabolismo , Hepatócitos/citologia , Camundongos , Camundongos Endogâmicos C57BLRESUMO
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.
Assuntos
Infarto do Miocárdio/patologia , Receptores dos Hormônios Gastrointestinais/metabolismo , Adenilil Ciclases/metabolismo , Animais , Ativação Enzimática , Polipeptídeo Inibidor Gástrico/metabolismo , Células HEK293 , Insuficiência Cardíaca/patologia , Humanos , Sistema de Sinalização das MAP Quinases , Camundongos , Camundongos Endogâmicos C57BL , Infarto do Miocárdio/diagnóstico por imagem , Infarto do Miocárdio/fisiopatologia , Infarto do Miocárdio/prevenção & controle , Miocárdio/metabolismo , Miocárdio/patologia , Miócitos Cardíacos/metabolismo , Fosforilação , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores dos Hormônios Gastrointestinais/deficiência , Receptores dos Hormônios Gastrointestinais/genética , Transdução de Sinais , Esterol Esterase/metabolismo , Triglicerídeos/metabolismo , Remodelação VentricularRESUMO
OBJECTIVES: Glucagon-like peptide-1 (GLP-1) is secreted from enteroendocrine cells and exerts a broad number of metabolic actions through activation of a single GLP-1 receptor (GLP-1R). The cardiovascular actions of GLP-1 have garnered increasing attention as GLP-1R agonists are used to treat human subjects with diabetes and obesity that may be at increased risk for development of heart disease. Here we studied mechanisms linking GLP-1R activation to control of heart rate (HR) in mice. METHODS: The actions of GLP-1R agonists were examined on the control of HR in wild type mice (WT) and in mice with cardiomyocyte-selective disruption of the GLP-1R (Glp1rCM-/-). Complimentary studies examined the effects of GLP-1R agonists in mice co-administered propranolol or atropine. The direct effects of GLP-1R agonism on HR and ventricular developed pressure were examined in isolated perfused mouse hearts ex vivo, and atrial depolarization was quantified in mouse hearts following direct application of liraglutide to perfused atrial preparations ex vivo. RESULTS: Doses of liraglutide and lixisenatide that were equipotent for acute glucose control rapidly increased HR in WT and Glp1rCM-/- mice in vivo. The actions of liraglutide to increase HR were more sustained relative to lixisenatide, and diminished in Glp1rCM-/- mice. The acute chronotropic actions of GLP-1R agonists were attenuated by propranolol but not atropine. Neither native GLP-1 nor lixisenatide increased HR or developed pressure in perfused hearts ex vivo. Moreover, liraglutide had no direct effect on sinoatrial node firing rate in mouse atrial preparations ex vivo. Despite co-localization of HCN4 and GLP-1R in primate hearts, HCN4-directed Cre expression did not attenuate levels of Glp1r mRNA transcripts, but did reduce atrial Gcgr expression in the mouse heart. CONCLUSIONS: GLP-1R agonists increase HR through multiple mechanisms, including regulation of autonomic nervous system function, and activation of the atrial GLP-1R. Surprisingly, the isolated atrial GLP-1R does not transduce a direct chronotropic effect following exposure to GLP-1R agonists in the intact heart, or isolated atrium, ex vivo. Hence, cardiac GLP-1R circuits controlling HR require neural inputs and do not function in a heart-autonomous manner.
Assuntos
Receptor do Peptídeo Semelhante ao Glucagon 1/fisiologia , Frequência Cardíaca/fisiologia , Animais , Sistema Nervoso Autônomo/fisiologia , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Frequência Cardíaca/efeitos dos fármacos , Liraglutida/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Peptídeos/farmacologiaRESUMO
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.
Assuntos
Colo/metabolismo , Fator 7 de Crescimento de Fibroblastos/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Mucosa Intestinal/metabolismo , Intestino Delgado/metabolismo , Animais , Carcinogênese/metabolismo , Proliferação de Células/fisiologia , Colo/fisiologia , Colo/fisiopatologia , Fator de Crescimento Epidérmico/metabolismo , Exenatida , Feminino , Incretinas/metabolismo , Mucosa Intestinal/fisiologia , Mucosa Intestinal/fisiopatologia , Intestino Delgado/patologia , Intestino Delgado/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Peptídeos/metabolismo , Receptor IGF Tipo 1/metabolismo , Transdução de Sinais/fisiologia , Peçonhas/metabolismoRESUMO
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.
Assuntos
Mucosa Intestinal/imunologia , Linfócitos/imunologia , Receptores de Glucagon/agonistas , Receptores de Glucagon/fisiologia , Animais , Citocinas/análise , Exenatida , Feminino , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Receptor do Peptídeo Semelhante ao Glucagon 1 , Imunidade Inata , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Peptídeos/farmacologia , Transdução de Sinais , Peçonhas/farmacologiaRESUMO
Glucagon-like peptide-1 (GLP-1) controls glucose homeostasis by regulating secretion of insulin and glucagon through a single GLP-1 receptor (GLP-1R). GLP-1R agonists also increase pancreatic weight in some preclinical studies through poorly understood mechanisms. Here we demonstrate that the increase in pancreatic weight following activation of GLP-1R signaling in mice reflects an increase in acinar cell mass, without changes in ductal compartments or ß-cell mass. GLP-1R agonists did not increase pancreatic DNA content or the number of Ki67(+) cells in the exocrine compartment; however, pancreatic protein content was increased in mice treated with exendin-4 or liraglutide. The increased pancreatic mass and protein content was independent of cholecystokinin receptors, associated with a rapid increase in S6 phosphorylation, and mediated through the GLP-1R. Rapamycin abrogated the GLP-1R-dependent increase in pancreatic mass but had no effect on the robust induction of Reg3α and Reg3ß gene expression. Mass spectrometry analysis identified GLP-1R-dependent upregulation of Reg family members, as well as proteins important for translation and export, including Fam129a, eIF4a1, Wars, and Dmbt1. Hence, pharmacological GLP-1R activation induces protein synthesis, leading to increased pancreatic mass, independent of changes in DNA content or cell proliferation in mice.
Assuntos
Pâncreas/efeitos dos fármacos , Pâncreas/metabolismo , Receptores de Glucagon/agonistas , Receptores de Glucagon/metabolismo , Animais , Antígenos de Neoplasias/metabolismo , Biomarcadores Tumorais/metabolismo , Exenatida , Peptídeo 1 Semelhante ao Glucagon/análogos & derivados , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Receptor do Peptídeo Semelhante ao Glucagon 1 , Imuno-Histoquímica , Lectinas Tipo C/metabolismo , Liraglutida , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Associadas a Pancreatite , Peptídeos/farmacologia , Biossíntese de Proteínas/efeitos dos fármacos , Proteínas/metabolismo , Peçonhas/farmacologiaRESUMO
Glucagon-like peptide-1 (GLP-1) secretion is classically regulated by ingested nutrients. To identify novel molecular targets controlling incretin secretion, we analyzed enteroendocrine cell pathways important for hormone biosynthesis and secretion. We demonstrate that progesterone increases GLP-1 secretion and extracellular signal-related kinase 1/2 (ERK1/2) phosphorylation in enteroendocrine GLUTag cells via mechanisms sensitive to the mitogen-activated protein kinase inhibitor U0126. The stimulatory effects of progesterone (P4) or the synthetic progestin R5020 on ERK1/2 phosphorylation were independent of the classical progesterone receptor antagonist RU486. Furthermore, a cell-impermeable BSA-progesterone conjugate rapidly increased ERK1/2 phosphorylation and GLP-1 secretion. Knockdown of the membrane progesterone receptors Paqr5 or Paqr7 in GLUTag cells eliminated the stimulatory effect of R5020 and progesterone on GLP-1 secretion. Enteral progesterone administration increased plasma levels of GLP-1, glucose-dependent insulinotropic polypeptide (GIP), and insulin, and improved oral glucose tolerance in an RU486-insensitve manner in mice: however, systemic progesterone exposure did not improve glucose homeostasis. Unexpectedly, the glucoregulatory actions of enteral progesterone did not require classical incretin receptor signaling and were preserved in Glp1r(-/-) and Glp1r(-/-):Gipr(-/-) mice. Intestine-restricted activation of membrane progesterone receptors may represent a novel approach for stimulation of incretin hormone secretion and control of glucose homeostasis.
Assuntos
Células Enteroendócrinas/fisiologia , Glucose/metabolismo , Incretinas/metabolismo , Receptores de Progesterona/fisiologia , Animais , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Polipeptídeo Inibidor Gástrico/sangue , Glucagon/biossíntese , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Homeostase , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação , Promegestona/farmacologiaRESUMO
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.
Assuntos
Aterosclerose/metabolismo , Fígado/metabolismo , Receptores de Glucagon/metabolismo , Animais , Apolipoproteínas E/deficiência , Apolipoproteínas E/genética , Western Blotting , Composição Corporal/efeitos dos fármacos , Receptor do Peptídeo Semelhante ao Glucagon 1 , Teste de Tolerância a Glucose , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Interleucina-10/deficiência , Interleucina-10/genética , Fígado/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Knockout , Peptídeos/farmacologia , Receptores de Glucagon/agonistas , Receptores de Glucagon/genéticaRESUMO
BACKGROUND & AIMS: Glucagon-like peptide-1 (GLP-1) regulates glucose homeostasis through multiple mechanisms including direct actions on the endocrine pancreas and indirect activation of central nervous system circuits regulating gastric emptying, satiety, and body weight. Because native GLP-1 is rapidly degraded, there is considerable interest in development of more potent GLP-1 receptor (GLP-1R) agonists with sustained activity; however, the extent to which much larger GLP-1R agonists will mimic some or all of the actions of smaller peptides remains uncertain. METHODS: We studied the actions of CJC-1134-PC, a recombinant human serum albumin-exendin-4 conjugated protein, at the GLP-1R using heterologous cells expressing the GLP-1R in vitro and both wild-type and Glp1r(-/-) mice in vivo. RESULTS: CJC-1134-PC activated GLP-1R-dependent signaling in baby hamster kidney-GLP-1R cells and acutely lowered blood glucose in wild-type but not in Glp1r(-/-) mice. Moreover, acute administration of CJC-1134-PC rapidly activated c-Fos expression in multiple regions of the central nervous system, acutely inhibited gastric emptying, and produced sustained inhibition of food intake in a GLP-1R-dependent manner. Furthermore, chronic daily treatment of high-fat diet-fed wild-type mice with CJC-1134-PC for 4 weeks led to improved glucose tolerance, increased levels of glucose-stimulated insulin, decreased HbA1c, and weight loss associated with decreased hepatic triglyceride content. CONCLUSIONS: These findings illustrate that a high-molecular-weight exendin-4-albumin conjugate retains the ability to mimic a full spectrum of GLP-1R-dependent actions, including activation of central nervous system circuits regulating gastric emptying, food intake, and body weight.
Assuntos
Sistema Nervoso Central/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Glucose/metabolismo , Homeostase/fisiologia , Hipoglicemiantes/farmacologia , Peptídeos/farmacologia , Peçonhas/farmacologia , Animais , Índice de Massa Corporal , Sistema Nervoso Central/fisiologia , Cricetinae , Exenatida , Esvaziamento Gástrico/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Quinases do Centro Germinativo , Receptor do Peptídeo Semelhante ao Glucagon 1 , Teste de Tolerância a Glucose , Proteínas de Homeodomínio/biossíntese , Proteínas de Homeodomínio/genética , Homeostase/efeitos dos fármacos , Insulina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Serina-Treonina Quinases/biossíntese , Proteínas Serina-Treonina Quinases/genética , RNA Mensageiro/genética , Receptores de Glucagon/biossíntese , Receptores de Glucagon/genética , Proteínas Recombinantes , Transativadores/biossíntese , Transativadores/genéticaRESUMO
Glucagon-like peptide-1 (GLP-1) regulates energy intake, gastrointestinal motility, and nutrient disposal. The relative importance of the islet beta-cell for GLP-1 actions remains unclear. We determined the role of the islet beta-cell and the pancreatic duodenal homeobox-1 (Pdx1) transcription factor for GLP-1 receptor (GLP-1R)-dependent actions through analysis of mice with beta-cell-specific inactivation of the Pdx1 gene (beta-cell(Pdx1-/-) mice). The GLP-1R agonist exendin-4 (Ex-4) reduced glycemic excursion following intraperitoneal (i.p.) glucose challenge in control littermates (beta-cell(Pdx1+/+) mice) but not in beta-cell(Pdx1-/-) mice. Similarly, Ex-4 failed to increase levels of plasma insulin, pancreatic insulin content, and pancreatic insulin mRNA transcripts in beta-cell(Pdx1-/-) mice. Furthermore, Ex-4 significantly increased beta-cell proliferation and reduced beta-cell apoptosis in beta-cell(Pdx1+/+) mice but not in beta-cell(Pdx1-/-) mice. Moreover, Ex-4 increased the levels of insulin and amylin mRNA transcripts and augmented glucose-stimulated insulin secretion in islets from beta-cell(Pdx1+/+) mice but not in beta-cell(Pdx1-/-) islets. Surprisingly, Ex-4 failed to reduce levels of plasma glucagon in beta-cell(Pdx1-/-) mice. These findings demonstrate that Pdx1 expression is essential for integrating GLP-1R-dependent signals regulating alpha-cell glucagon secretion and for the growth, differentiated function, and survival of islet beta-cells.
Assuntos
Glucagon/fisiologia , Proteínas de Homeodomínio/genética , Ilhotas Pancreáticas/fisiologia , Fragmentos de Peptídeos/fisiologia , Precursores de Proteínas/fisiologia , Receptores de Glucagon/fisiologia , Transativadores/genética , Animais , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Divisão Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Exenatida , Feminino , Peptídeo 1 Semelhante ao Glucagon , Receptor do Peptídeo Semelhante ao Glucagon 1 , Teste de Tolerância a Glucose , Homeostase , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/efeitos dos fármacos , Camundongos , Camundongos Knockout , Camundongos Mutantes , Peptídeos/farmacologia , Transativadores/deficiência , Peçonhas/farmacologiaRESUMO
Pdx-1 plays a key role in the development of the pancreas and the control of islet gene transcription and has also been proposed as a dominant regulator of the alpha- vs. beta-cell phenotype via extinction of proglucagon expression. To ascertain the relationship between Pdx-1 and proglucagon gene expression, we examined the effect of enhanced pdx-1 expression on proglucagon gene expression in murine islet alphaTC-1 and GLUTag enteroendocrine cells. Although adenoviral transduction increased the levels of pdx-1 mRNA transcripts and nuclear Pdx-1 protein, overexpression of pdx-1 did not repress endogenous proglucagon gene expression in alphaTC-1 or GLUTag cells or murine islets. Immunohistochemical analysis of cells transduced with Ad-pdx-1 demonstrated multiple individual islet or enteroendocrine cells exhibiting both nuclear Pdx-1 and cytoplasmic glucagon-like peptide-1 immunopositivity. The failure of pdx-1 to inhibit endogenous proglucagon gene expression was not attributable to defects in Pdx-1 nuclear translocation or DNA binding as demonstrated using Western blotting and EMSA analyses. Furthermore, Ad-pdx-1 transduction did not repress proglucagon promoter activity in alphaTC-1 or GLUTag cells. Taken together, these findings demonstrate that pdx-1 alone is not sufficient for specification of the hormonal phenotype or extinction of proglucagon gene expression in islet or enteroendocrine cells.
Assuntos
Glândulas Endócrinas/metabolismo , Glucagon/antagonistas & inibidores , Glucagon/genética , Proteínas de Homeodomínio/fisiologia , Mucosa Intestinal/metabolismo , Ilhotas Pancreáticas/metabolismo , Precursores de Proteínas/antagonistas & inibidores , Precursores de Proteínas/genética , Transativadores/fisiologia , Transcrição Gênica/fisiologia , Animais , Linhagem Celular , Glândulas Endócrinas/citologia , Glucagon/metabolismo , Peptídeo 1 Semelhante ao Glucagon , Proteínas de Homeodomínio/metabolismo , Intestinos/citologia , Ilhotas Pancreáticas/citologia , Camundongos , Fragmentos de Peptídeos/metabolismo , Proglucagon , Precursores de Proteínas/metabolismo , Transativadores/metabolismo , Transdução GenéticaRESUMO
Despite interest in understanding glucagon-like peptide-1 (GLP-1) production, the factors important for GLP-1 biosynthesis remain poorly understood. We examined control of human proglucagon gene expression in NCI-H716 cells, a cell line that secretes GLP-1 in a regulated manner. Insulin, phorbol myristate acetate, or forskolin, known regulators of rodent proglucagon gene expression, had no effect, whereas sodium butyrate decreased levels of NCI-H716 proglucagon mRNA transcripts. The inhibitory effect of sodium butyrate was mimicked by trichostatin A but was not detected with sodium acetate or isobutyrate. The actions of butyrate were not diminished by the ERK1/2 inhibitor PD98059, p38 inhibitor SB203580, or soluble guanylate cyclase inhibitor LY83583 or following treatment of cells with KT5823, a selective inhibitor of cGMP-dependent protein kinase. NCI-H716 cells expressed multiple proglucagon gene transcription factors including isl-1, pax-6, pax-2, cdx-2/3, pax-4, hepatocyte nuclear factor (HNF)-3 alpha, HNF-3beta, HNF-3 gamma, and Nkx2.2. Nevertheless, the butyrate-dependent inhibition of proglucagon gene expression was not associated with coordinate changes in transcription factor expression and both the human and rat transfected proglucagon promoters were transcriptionally inactive in NCI-H716 cells. Hence, NCI-H716 cells may not be a physiologically optimal model for studies of human enteroendocrine proglucagon gene transcription.
Assuntos
Adenocarcinoma/genética , Neoplasias do Ceco/genética , Regulação da Expressão Gênica/fisiologia , Glucagon/genética , Intestinos/fisiopatologia , Precursores de Proteínas/genética , Adenocarcinoma/metabolismo , Animais , Butiratos/farmacologia , Neoplasias do Ceco/metabolismo , Expressão Gênica/efeitos dos fármacos , Glucagon/metabolismo , Peptídeo 1 Semelhante ao Glucagon , Proteína Homeobox Nkx-2.2 , Proteínas de Homeodomínio , Humanos , Proteínas Nucleares , Fragmentos de Peptídeos/metabolismo , Proglucagon , Precursores de Proteínas/metabolismo , Ratos , Fatores de Transcrição/metabolismo , Células Tumorais CultivadasRESUMO
In mammalian cells, the human adenovirus type 5 early region 1A (E1A) oncoprotein functions as a thyroid hormone (TH)-dependent activator of the thyroid hormone receptor (TR). Interestingly, in the cellular context of the yeast Saccharomyces cerevisiae, E1A acts as a TR-specific constitutive coactivator that is down-regulated by TH. TH reduces the interaction of E1A with the TR in yeast but not HeLa cells. The N-terminal 82 amino acids of E1A are sufficient for coactivation in yeast and residues 4-29 are essential. In yeast, expression of the nuclear receptor corepressor (N-CoR) could down-regulate constitutive transcriptional activation of the TR by E1A, whereas expression of the glucocorticoid receptor interacting protein 1 (GRIP-1) coactivator reconstituted the E1A-induced pattern of enhanced TH-dependent gene activation by TR observed in mammalian cells. We further show that the mating type switching gene (SWI)/sucrose nonfermenting (SNF) gene chromatin remodeling complex is required for both TH/GRIP-1- and E1A-dependent coactivator function, whereas the general control nonrepressed protein (GCN5)/alteration/deficiency in activation protein (ADA2) components of the SPT, ADA, GCN5, acetylation (SAGA) transcriptional adaptor complex are required for TH/GRIP-1, but not E1A-dependent activation of the TR. Taken together, these studies demonstrate that the novel TR-specific coactivator function of E1A in yeast depends on the SWI/SNF chromatin remodeling complex and can be further influenced by changes in the cellular complement of transcriptional coregulatory proteins.