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1.
Mol Cell Endocrinol ; 521: 111107, 2021 02 05.
Artículo en Inglés | MEDLINE | ID: mdl-33309639

RESUMEN

Here we show that scratch family transcriptional repressor 1 (SCRT1), a zinc finger transcriptional regulator, is a novel regulator of beta cell function. SCRT1 was found to be expressed in beta cells in rodent and human islets. In human islets, expression of SCRT1 correlated with insulin secretion capacity and the expression of the insulin (INS) gene. Furthermore, SCRT1 mRNA expression was lower in beta cells from T2D patients. siRNA-mediated Scrt1 silencing in INS-1832/13 cells, mouse- and human islets resulted in impaired glucose-stimulated insulin secretion and decreased expression of the insulin gene. This is most likely due to binding of SCRT1 to E-boxes of the Ins1 gene as shown with ChIP. Scrt1 silencing also reduced the expression of several key beta cell transcription factors. Moreover, Scrt1 mRNA expression was reduced by glucose and SCRT1 protein was found to translocate between the nucleus and the cytosol in a glucose-dependent fashion in INS-1832/13 cells as well as in a rodent model of T2D. SCRT1 was also regulated by a GSK3ß-dependent SCRT1-serine phosphorylation. Taken together, SCRT1 is a novel beta cell transcription factor that regulates insulin secretion and is affected in T2D.


Asunto(s)
Diabetes Mellitus Tipo 2/metabolismo , Regulación de la Expresión Génica/genética , Glucosa/metabolismo , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Factores de Transcripción/metabolismo , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Línea Celular , Núcleo Celular/genética , Núcleo Celular/metabolismo , Inmunoprecipitación de Cromatina , Citoplasma/genética , Citoplasma/metabolismo , Modelos Animales de Enfermedad , Regulación de la Expresión Génica/efectos de los fármacos , Silenciador del Gen , Humanos , Inmunohistoquímica , Insulina/genética , Secreción de Insulina/efectos de los fármacos , Células Secretoras de Insulina/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , ARN Interferente Pequeño , RNA-Seq , Reacción en Cadena en Tiempo Real de la Polimerasa , Análisis de la Célula Individual , Factores de Transcripción/genética
2.
Mol Cell Endocrinol ; 511: 110835, 2020 07 01.
Artículo en Inglés | MEDLINE | ID: mdl-32371087

RESUMEN

It is not known how ghrelin affects insulin secretion in human islets from patients with type 2 diabetes (T2D) or whether islet ghrelin expression or circulating ghrelin levels are altered in T2D. Here we sought out to identify the effect of ghrelin on insulin secretion in human islets and the impact of T2D on circulating ghrelin levels and on islet ghrelin cells. The effect of ghrelin on insulin secretion was assessed in human T2D and non-T2D islets. Ghrelin expression was assessed with RNA-sequencing (n = 191) and immunohistochemistry (n = 21). Plasma ghrelin was measured with ELISA in 40 T2D and 40 non-T2D subjects. Ghrelin exerted a glucose-dependent insulin-suppressing effect in islets from both T2D and non-T2D donors. Compared with non-T2D donors, T2D donors had reduced ghrelin mRNA expression and 75% less islet ghrelin cells, and ghrelin mRNA expression correlated negatively with HbA1c. T2D subjects had 25% lower fasting plasma ghrelin levels than matched controls. Thus, ghrelin has direct insulin-suppressing effects in human islets and T2D patients have lower fasting ghrelin levels, likely as a result of reduced number of islet ghrelin cells. These findings support inhibition of ghrelin signaling as a potential therapeutic avenue for stimulation of insulin secretion in T2D patients.


Asunto(s)
Diabetes Mellitus Tipo 2/sangre , Ghrelina/sangre , Ghrelina/farmacología , Secreción de Insulina , Islotes Pancreáticos/patología , Recuento de Células , Ayuno/sangre , Glucosa/metabolismo , Humanos , Secreción de Insulina/efectos de los fármacos , Islotes Pancreáticos/efectos de los fármacos , Fenotipo , RNA-Seq , Donantes de Tejidos
3.
Mol Cell Endocrinol ; 476: 8-16, 2018 11 15.
Artículo en Inglés | MEDLINE | ID: mdl-29627317

RESUMEN

Impaired incretin effect is a culprit in Type 2 Diabetes. Cocaine- and amphetamine-regulated transcript (CART) is a regulatory peptide controlling pancreatic islet hormone secretion and beta-cell survival. Here we studied the potential expression of CART in enteroendocrine cells and examined the role of CART as a regulator of incretin secretion and expression. CART expression was found in glucose-dependent insulinotropic polypeptide (GIP)-producing K-cells and glucagon-like peptide-1 (GLP-1)-producing L-cells in human duodenum and jejunum and circulating CART levels were increased 60 min after a meal in humans. CART expression was increased by fatty acids and GIP, but unaffected by glucose in GLUTag and STC-1 cells. Exogenous CART had no effect on GIP and GLP-1 expression and secretion in GLUTag or STC-1 cells, but siRNA-mediated silencing of CART reduced GLP-1 expression and secretion. Furthermore, acute intravenous administration of CART increased GIP and GLP-1 secretion during an oral glucose-tolerance test in mice. We conclude that CART is a novel constituent of human K- and L-cells with stimulatory actions on incretin secretion and that interfering with the CART system may be a therapeutic avenue for T2D.


Asunto(s)
Polipéptido Inhibidor Gástrico/metabolismo , Péptido 1 Similar al Glucagón/metabolismo , Intestinos/química , Proteínas del Tejido Nervioso/metabolismo , Adulto , Animales , Ácidos Grasos/metabolismo , Femenino , Glucosa/metabolismo , Prueba de Tolerancia a la Glucosa , Humanos , Incretinas/metabolismo , Masculino , Ratones , Persona de Mediana Edad , Proteínas del Tejido Nervioso/sangre , Proteínas del Tejido Nervioso/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo
4.
Mol Cell Endocrinol ; 447: 52-60, 2017 05 15.
Artículo en Inglés | MEDLINE | ID: mdl-28237718

RESUMEN

Impaired beta-cell function is key to the development of type 2 diabetes. Cocaine- and amphetamine-regulated transcript (CART) is an islet peptide with insulinotropic and glucagonostatic properties. Here we studied the role of endogenous CART in beta-cell function. CART silencing in INS-1 (832/13) beta-cells reduced insulin secretion and production, ATP levels and beta-cell exocytosis. This was substantiated by reduced expression of several exocytosis genes, as well as reduced expression of genes important for insulin secretion and processing. In addition, CART silencing reduced the expression of a network of transcription factors essential for beta-cell function. Moreover, in RNAseq data from human islet donors, CARTPT expression levels correlated with insulin, exocytosis genes and key beta-cell transcription factors. Thus, endogenous beta-cell CART regulates insulin expression and secretion in INS-1 (832/13) cells, via actions on the exocytotic machinery and a network of beta-cell transcription factors. We conclude that CART is important for maintaining the beta-cell phenotype.


Asunto(s)
Regulación de la Expresión Génica , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Transcripción Genética , Animales , Apoptosis/genética , Exocitosis/genética , Técnicas de Silenciamiento del Gen , Silenciador del Gen , Humanos , Secreción de Insulina , Células Secretoras de Insulina/citología , Modelos Biológicos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Factores de Transcripción/metabolismo
5.
Peptides ; 71: 113-20, 2015 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-26206285

RESUMEN

Islet produced 5-hydroxy tryptamine (5-HT) is suggested to regulate islet hormone secretion in a paracrine and autocrine manner in rodents. Hitherto, no studies demonstrate a role for this amine in human islet function, nor is it known if 5-HT signaling is involved in the development of beta cell dysfunction in type 2 diabetes (T2D). To clarify this, we performed a complete transcriptional mapping of 5-HT receptors and processing enzymes in human islets and investigated differential expression of these genes in non-diabetic and T2D human islet donors. We show the expression of fourteen 5-HT receptors as well as processing enzymes involved in the biosynthesis of 5-HT at the mRNA level in human islets. Two 5-HT receptors (HTR1D and HTR2A) were over-expressed in T2D islet donors. Both receptors (5-HT1d and 5-HT2a) were localized to human alpha, beta and delta cells. 5-HT inhibited both insulin and glucagon secretion in non-diabetic islet donors. In islets isolated from T2D donors the amine significantly increased release of insulin in response to glucose. Our results suggest that 5-HT signaling participates in regulation of overall islet hormone secretion in non- diabetic individuals and over-expression of HTR1D and HTR2A may either contribute to islet dysfunction in T2D or arise as a consequence of an already dysfunctional islet.


Asunto(s)
Diabetes Mellitus Tipo 2/metabolismo , Glucagón/metabolismo , Insulina/metabolismo , Islotes Pancreáticos/metabolismo , Receptor de Serotonina 5-HT1D/biosíntesis , Receptor de Serotonina 5-HT2A/biosíntesis , Diabetes Mellitus Tipo 2/patología , Femenino , Regulación de la Expresión Génica , Humanos , Secreción de Insulina , Islotes Pancreáticos/patología , Masculino , Transducción de Señal
6.
Regul Pept ; 182: 35-40, 2013 Mar 10.
Artículo en Inglés | MEDLINE | ID: mdl-23318496

RESUMEN

Cocaine- and amphetamine-regulated transcript (CART) is a regulatory peptide expressed in the nervous system and in endocrine cells, e.g. in pancreatic islets. CART deficient mice exhibit islet dysfunction, impaired insulin secretion and increased body weight. A mutation in the CART gene in humans is associated with reduced metabolic rate, obesity and diabetes. Furthermore, CART is upregulated in islets of type-2 diabetic rats and regulates islet hormone secretion in vitro. While the function of CART in the nervous system has been extensively studied, there is no information on its expression or function in white adipose tissue. CART mRNA and protein were found to be expressed in both subcutaneous and visceral white adipose tissue from rat and man. Stimulating rat primary adipocytes with CART significantly potentiated isoprenaline-induced lipolysis, and hormone sensitive lipase activation (phosphorylation of Ser 563). On the other hand, CART significantly potentiated the inhibitory effect of insulin on isoprenaline-induced lipolysis. CART inhibited insulin-induced glucose uptake and lipogenesis, which was associated with inhibition of PKB phosphorylation. In conclusion, CART is a novel constituent of human and rat adipocytes and affects several biological processes central in both lipid- and glucose homeostasis. Depending on the surrounding conditions, the effects of CART are insulin-like or insulin-antagonistic.


Asunto(s)
Adipocitos/metabolismo , Glucosa/metabolismo , Homeostasis/genética , Metabolismo de los Lípidos , Proteínas del Tejido Nervioso/genética , ARN Mensajero/metabolismo , Adipocitos/efectos de los fármacos , Animales , Secuencia de Bases , Cartilla de ADN , Isoproterenol/farmacología , Masculino , Ratas , Ratas Sprague-Dawley , Reacción en Cadena en Tiempo Real de la Polimerasa
7.
Diabetes Obes Metab ; 12(11): 976-82, 2010 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-20880344

RESUMEN

AIM: Clinical studies have suggested a role for dietary glycaemic index (GI) in body weight regulation and diabetes risk. Here, we investigated the long-term metabolic effects of low and high glycaemic diets using the C57BL/6J mouse model. METHODS: Female C57BL/6J mice were fed low or high glycaemic starch in either low-fat or medium-fat diets for 22 weeks. Oral and intravenous glucose tolerance tests were performed to investigate the effect of the experimental diets on glucose tolerance and insulin resistance. RESULTS: In this study, a high glycaemic diet resulted in impaired oral glucose tolerance compared to a low glycaemic diet. This effect was more pronounced in the group fed a medium-fat diet, suggesting that a lower dietary fat content ameliorates the negative effect of a high glycaemic diet. No effect on body weight or body fat content was observed in either a low-fat diet or a medium-fat diet. Static incubation of isolated islets did not show any differences in basal (3.3 mM glucose) or glucose-stimulated (8.6 and 16.7 mM glucose) insulin secretion between mice fed a low or high glycaemic diet. CONCLUSION: Together, our data suggest that the impaired glucose tolerance seen after a high glycaemic diet is not explained by altered ß-cell function.


Asunto(s)
Dieta para Diabéticos , Resistencia a la Insulina/fisiología , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Animales , Peso Corporal/fisiología , Dieta con Restricción de Grasas , Femenino , Prueba de Tolerancia a la Glucosa/métodos , Índice Glucémico , Secreción de Insulina , Ratones , Ratones Endogámicos C57BL
8.
Diabetologia ; 52(2): 271-80, 2009 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-19023560

RESUMEN

AIMS/HYPOTHESIS: The enzyme hormone-sensitive lipase (HSL) is produced and is active in pancreatic beta cells. Because lipids are known to play a crucial role in normal control of insulin release and in the deterioration of beta cell function, as observed in type 2 diabetes, actions of HSL in beta cells may be critical. This notion has been addressed in different lines of HSL knockout mice with contradictory results. METHODS: To resolve this, we created a transgenic mouse lacking HSL specifically in beta cells, and characterised this model with regard to glucose metabolism and insulin secretion, using both in vivo and in vitro methods. RESULTS: We found that fasting basal plasma glucose levels were significantly elevated in mice lacking HSL in beta cells. An IVGTT at 12 weeks revealed a blunting of the initial insulin response to glucose with delayed elimination of the sugar. Additionally, arginine-stimulated insulin secretion was markedly diminished in vivo. Investigation of the exocytotic response in single HSL-deficient beta cells showed an impaired response to depolarisation of the plasma membrane. Beta cell mass and islet insulin content were increased, suggesting a compensatory mechanism, by which beta cells lacking HSL strive to maintain normoglycaemia. CONCLUSIONS/INTERPRETATION: Based on these results, we suggest that HSL, which is located in close proximity of the secretory granules, may serve as provider of a lipid-derived signal essential for normal insulin secretion.


Asunto(s)
Hiperglucemia/etiología , Células Secretoras de Insulina/enzimología , Insulina/metabolismo , Esterol Esterasa/deficiencia , Esterol Esterasa/genética , Tejido Adiposo/enzimología , Animales , Área Bajo la Curva , Glucemia/metabolismo , Exocitosis/genética , Exones , Prueba de Tolerancia a la Glucosa , Hiperglucemia/sangre , Secreción de Insulina , Células Secretoras de Insulina/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mutación , Reacción en Cadena de la Polimerasa , ARN Mensajero/genética , Vesículas Secretoras/enzimología
9.
Exp Diabetes Res ; 2008: 697035, 2008.
Artículo en Inglés | MEDLINE | ID: mdl-18497871

RESUMEN

Human islet amyloid polypeptide (hIAPP), a pancreatic islet protein of 37 amino acids, is the main component of islet amyloid, seen at autopsy in patients with type 2 diabetes mellitus (DM2). To investigate the roles of hIAPP and islet amyloid in DM2, we generated transgenic mice expressing hIAPP in their islet beta cells. In this study, we found that after a long-term, high-fat diet challenge islet amyloid was observed in only 4 of 19 hIAPP transgenic mice. hIAPP transgenic females exhibited severe glucose intolerance, which was associated with a downregulation of GLUT-2 mRNA expression. In isolated islets from hIAPP males cultured for 3 weeks on high-glucose medium, the percentage of amyloid containing islets increased from 5.5% to 70%. This ex vivo system will allow a more rapid, convenient, and specific study of factors influencing islet amyloidosis as well as of therapeutic strategies to interfere with this pathological process.


Asunto(s)
Amiloide/metabolismo , Amiloidosis/etiología , Diabetes Mellitus Tipo 2/metabolismo , Islotes Pancreáticos/metabolismo , Amiloide/sangre , Amiloide/genética , Amiloidosis/genética , Amiloidosis/metabolismo , Amiloidosis/patología , Animales , Peso Corporal , Supervivencia Celular , Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/patología , Grasas de la Dieta/efectos adversos , Modelos Animales de Enfermedad , Regulación hacia Abajo , Femenino , Intolerancia a la Glucosa/metabolismo , Prueba de Tolerancia a la Glucosa , Transportador de Glucosa de Tipo 2/genética , Transportador de Glucosa de Tipo 2/metabolismo , Humanos , Células Secretoras de Insulina/metabolismo , Polipéptido Amiloide de los Islotes Pancreáticos , Islotes Pancreáticos/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , ARN Mensajero/metabolismo , Factores de Tiempo , Técnicas de Cultivo de Tejidos
10.
Eur J Pediatr Surg ; 17(3): 184-9, 2007 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-17638157

RESUMEN

AIMS: Cocaine- and amphetamine-regulated transcript (CART)-peptide is found in the brain and participates in the control of feeding behavior. It is also expressed in the peripheral nervous system and is suggested to have neuromodulatory and/or neurotrophic effects in rat intestine. The aims of this study were to investigate the presence of CART-peptide in the normal ganglionic as well as aganglionic intestine from patients with Hirschsprung's disease and the peptide's possible coexistence with other neurotransmitters. METHODS: Intestinal specimens from nine patients with Hirschsprung's disease were examined using immunohistochemistry. A double immunostaining technique was used in order to elucidate the presence of CART-peptide in NOS and VIP-containing enteric neurons. RESULTS: In ganglionic intestine, CART-peptide was found in numerous nerve fibers, predominantly within the smooth muscle layers and in myenteric nerve cell bodies. A high degree of co-localization of CART with NOS and VIP was seen. Only very few CART immunoreactive nerve fibers and no nerve cell bodies were found in the aganglionic intestine. CONCLUSIONS: This is the first report on the presence of CART-peptide in the human intestine. In the ganglionic intestine CART was detected mainly in myenteric neurons, while only very few CART-IR nerve fibers were found in the aganglionic intestine. This, together with the coexistence of CART with NOS and VIP, indicates an intrinsic origin of the CART-containing neurons and suggests that CART may influence NO and VIP-induced effects.


Asunto(s)
Colon/inervación , Sistema Nervioso Entérico/metabolismo , Enfermedad de Hirschsprung/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Anticuerpos Antiidiotipos/análisis , Biomarcadores/metabolismo , Preescolar , Femenino , Estudios de Seguimiento , Enfermedad de Hirschsprung/patología , Humanos , Inmunoglobulina G/inmunología , Inmunohistoquímica , Lactante , Masculino , Músculo Liso/inervación , Músculo Liso/metabolismo , Músculo Liso/patología , Fibras Nerviosas/metabolismo , Fibras Nerviosas/patología , Proteínas del Tejido Nervioso/inmunología , Neurotransmisores , Óxido Nítrico Sintasa/metabolismo , Pronóstico , Estudios Retrospectivos , Índice de Severidad de la Enfermedad , Péptido Intestinal Vasoactivo/metabolismo
11.
Diabetologia ; 50(7): 1453-62, 2007 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-17479245

RESUMEN

AIMS/HYPOTHESIS: Increased glucagon secretion predicts deterioration of glucose tolerance, and high glucagon levels contribute to hyperglycaemia in type 2 diabetes. Inhibition of glucagon action may therefore be a potential novel target to reduce hyperglycaemia. Here, we investigated whether chronic treatment with a glucagon receptor antagonist (GRA) improves islet dysfunction in female mice on a high-fat diet (HFD). MATERIALS AND METHODS: After 8 weeks of HFD, mice were treated with a small molecule GRA (300 mg/kg, gavage once daily) for up to 30 days. Insulin secretion was studied after oral and intravenous administration of glucose and glucagon secretion after intravenous arginine. Islet morphology was examined and insulin secretion and glucose oxidation were measured in isolated islets. RESULTS: Fasting plasma glucose levels were reduced by GRA (6.0 +/- 0.2 vs 7.4 +/- 0.5 mmol/l; p = 0.017). The acute insulin response to intravenous glucose was augmented (1,300 +/- 110 vs 790 +/- 64 pmol/l; p < 0.001). The early insulin response to oral glucose was reduced in mice on HFD + GRA (1,890 +/- 160 vs 3,040 +/- 420 pmol/l; p = 0.012), but glucose excursions were improved. Intravenous arginine significantly increased the acute glucagon response (129 +/- 12 vs 36 +/- 6 ng/l in controls; p < 0.01), notably without affecting plasma glucose. GRA caused a modest increase in alpha cell mass, while beta cell mass was similar to that in mice on HFD + vehicle. Isolated islets displayed improved glucose-stimulated insulin secretion after GRA treatment (0.061 +/- 0.007 vs 0.030 +/- 0.004 pmol islet(-1) h(-1) at 16.7 mmol/l glucose; p < 0.001), without affecting islet glucose oxidation. CONCLUSIONS/INTERPRETATION: Chronic glucagon receptor antagonism in HFD-fed mice improves islet sensitivity to glucose and increases insulin secretion, suggesting improvement of key defects underlying impaired glucose tolerance and type 2 diabetes.


Asunto(s)
Alimentación Animal , Islotes Pancreáticos/metabolismo , Receptores de Glucagón/antagonistas & inhibidores , Animales , Glucemia/metabolismo , Dieta , Femenino , Glucagón/metabolismo , Prueba de Tolerancia a la Glucosa , Insulina/metabolismo , Resistencia a la Insulina , Ratones , Ratones Endogámicos C57BL , Oxígeno/metabolismo , Receptores de Glucagón/metabolismo , Factores de Tiempo
12.
Diabetologia ; 50(1): 74-83, 2007 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-17093947

RESUMEN

AIM/HYPOTHESIS: Hyperinsulinaemia maintains euglycaemia in insulin-resistant states. The precise cellular mechanisms by which the beta cells adapt are still unresolved. A peripherally derived cue, such as increased circulating fatty acids, may instruct the beta cell to initiate an adaptive programme to maintain glucose homeostasis. When this fails, type 2 diabetes ensues. Because mitochondria play a key role in beta cell pathophysiology, we tested the hypothesis that mitochondrial metabolism is critical for beta cell adaptation to insulin resistance. METHODS: C57BL/6J mice were given high-fat (HF) diet for 12 weeks. We then analysed islet hormone secretion, metabolism in vivo and in vitro, and beta cell morphology. RESULTS: HF diet resulted in insulin resistance and glucose intolerance but not frank diabetes. Basal insulin secretion was elevated in isolated islets from HF mice with almost no additional response provoked by high glucose. In contrast, a strong secretory response was seen when islets from HF mice were stimulated with fuels that require mitochondrial metabolism, such as glutamate, glutamine, alpha-ketoisocaproic acid and succinate. Moreover, while glucose oxidation was impaired in islets from HF mice, oxidation of glutamine and palmitate was enhanced. Ultrastructural analysis of islets in HF mice revealed an accumulation of lipid droplets in beta cells and a twofold increase in mitochondrial area. CONCLUSIONS/INTERPRETATION: We propose that beta cells exposed to increased lipid flux in insulin resistance respond by increasing mitochondrial volume. This expansion is associated with enhanced mitochondrial metabolism as a means of beta cell compensation.


Asunto(s)
Grasas de la Dieta/metabolismo , Resistencia a la Insulina/fisiología , Islotes Pancreáticos/metabolismo , Mitocondrias/metabolismo , Adaptación Fisiológica , Animales , Glucemia/metabolismo , Peso Corporal/fisiología , Femenino , Glutamina/farmacología , Insulina/sangre , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patología , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/patología , Cetoácidos/farmacología , Ratones , Ratones Endogámicos C57BL , Palmitatos/farmacología , Succinatos/farmacología
13.
Regul Pept ; 129(1-3): 203-11, 2005 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-15927717

RESUMEN

CART peptides are anorexigenic and are widely expressed in the central and peripheral nervous systems, as well as in endocrine cells in the pituitary, adrenal medulla and the pancreatic islets. To study the role of CART in islet function, we used CART null mutant mice (CART KO mice) and examined insulin secretion in vivo and in vitro, and expression of islet hormones and markers of beta-cell function using immunocytochemistry. We also studied CART expression in the normal pancreas. In addition, body weight development and food intake were documented. We found that in the normal mouse pancreas, CART was expressed in numerous pancreatic nerve fibers, both in the exocrine and endocrine portion of the gland. CART was also expressed in nerve cell bodies in the ganglia. Double immunostaining revealed expression in parasympathetic (vasoactive intestinal polypeptide (VIP)-containing) and in fewer sensory fibers (calcitonin gene-related peptide (CGRP)-containing). Although the expression of islet hormones appeared normal, CART KO islets displayed age dependent reduction of pancreatic duodenal homeobox 1 (PDX-1) and glucose transporter-2 (GLUT-2) immunoreactivity, indicating beta-cell dysfunction. Consistent with this, CART KO mice displayed impaired glucose-stimulated insulin secretion both in vivo after an intravenous glucose challenge and in vitro following incubation of isolated islets in the presence of glucose. The impaired insulin secretion in vivo was associated with impaired glucose elimination, and was apparent already in young mice with no difference in body weight. In addition, CART KO mice displayed increased body weight at the age of 40 weeks, without any difference in food intake. We conclude that CART is required for maintaining normal islet function in mice.


Asunto(s)
Intolerancia a la Glucosa/metabolismo , Insulina/metabolismo , Islotes Pancreáticos/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Aumento de Peso , Animales , Intolerancia a la Glucosa/genética , Intolerancia a la Glucosa/patología , Secreción de Insulina , Islotes Pancreáticos/patología , Ratones , Ratones Noqueados , Proteínas del Tejido Nervioso/deficiencia , Aumento de Peso/genética
15.
J Histochem Cytochem ; 52(3): 301-10, 2004 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-14966197

RESUMEN

Ghrelin is produced mainly by endocrine cells in the stomach and is an endogenous ligand for the growth hormone secretagogue receptor (GHS-R). It also influences feeding behavior, metabolic regulation, and energy balance. It affects islet hormone secretion, and expression of ghrelin and GHS-R in the pancreas has been reported. In human islets, ghrelin expression is highest pre- and neonatally. We examined ghrelin and GHS-R in rat islets during development with immunocytochemistry and in situ hybridization. We also studied the effect of ghrelin on insulin secretion from INS-1 (832/13) cells and the expression of GHS-R in these cells. We found ghrelin expression in rat islet endocrine cells from mid-gestation to 1 month postnatally. Islet expression of GHS-R mRNA was detected from late fetal stages to adult. The onset of islet ghrelin expression preceded that of gastric ghrelin. Islet ghrelin cells constitute a separate and novel islet cell population throughout development. However, during a short perinatal period a minor subpopulation of the ghrelin cells co-expressed glucagon or pancreatic polypeptide. Markers for cell lineage, proliferation, and duct cells revealed that the ghrelin cells proliferate, originate from duct cells, and share lineage with glucagon cells. Ghrelin dose-dependently inhibited glucose-stimulated insulin secretion from INS-1 (832/13) cells, and GHS-R was detected in the cells. We conclude that ghrelin is expressed in a novel developmentally regulated endocrine islet cell type in the rat pancreas and that ghrelin inhibits glucose-stimulated insulin secretion via a direct effect on the beta-cell.


Asunto(s)
Insulina/metabolismo , Islotes Pancreáticos/metabolismo , Hormonas Peptídicas/biosíntesis , Animales , Animales Recién Nacidos , Línea Celular , Ghrelina , Glucosa/farmacología , Inmunohistoquímica , Hibridación in Situ , Secreción de Insulina , Islotes Pancreáticos/embriología , Islotes Pancreáticos/crecimiento & desarrollo , Ratas , Ratas Sprague-Dawley , Receptores Acoplados a Proteínas G/biosíntesis , Receptores de Ghrelina
16.
J Histochem Cytochem ; 52(2): 169-77, 2004 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-14729868

RESUMEN

Cocaine- and amphetamine-regulated transcript (CART) is an anorexigenic peptide widely expressed in the central and peripheral, including the enteric, nervous systems. CART is also expressed in pituitary endocrine cells, adrenomedullary cells, islet somatostatin cells, and in rat antral gastrin cells. We used immunocytochemistry (IHC) and in situ hybridization (ISH) to study CART expression in developing rat pancreas. We also examined co-expression of CART and islet hormones and developmental markers and the effect of CART on proliferation using clonal insulin cells (INS-1 832/13). A major portion of each of the islet cell types, except the ghrelin cells, expressed CART during a period before and around birth. Two weeks postnatally, CART expression was restricted to somatostatin cells. Pre- and early postnatally, many of the CART-expressing cells co-expressed cytokeratin 20 (CK20), a marker of duct cells and islet precursor cells, the trophic hormone gastrin, and a smaller subpopulation also harbored the proliferation marker Ki67. CART was also expressed in pancreatic nerve fibers, both sensory and autonomic, and in ganglion nerve cell bodies. Although highly expressed in the developing islets, CART did not affect proliferation of INS-1 cells. We have demonstrated that CART is expressed in several islet cell types during rat development but is restricted to somatostatin cells and neurons in the adult rat.


Asunto(s)
Islotes Pancreáticos/metabolismo , Proteínas del Tejido Nervioso/biosíntesis , Animales , Animales Recién Nacidos , División Celular/efectos de los fármacos , Células Cultivadas , Inmunohistoquímica , Hibridación in Situ , Islotes Pancreáticos/embriología , Islotes Pancreáticos/crecimiento & desarrollo , Proteínas del Tejido Nervioso/farmacología , Páncreas/embriología , Páncreas/crecimiento & desarrollo , Páncreas/inervación , Ratas , Ratas Sprague-Dawley
17.
Neurogastroenterol Motil ; 15(5): 545-57, 2003 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-14507354

RESUMEN

Cocaine- and amphetamine-regulated transcript (CART) peptide, originally isolated from brain, is also expressed in the peripheral nervous system. The distribution, origin and projections of CART-expressing enteric neurones by immunocytochemistry and in situ hybridization in rat gastrointestinal (GI) tract were studied. Possible motor functions of CART were studied in vitro using longitudinal muscle strips from stomach, ileum and colon. Cocaine- and amphetamine-regulated transcript peptide was found in numerous myenteric neurones throughout the GI tract while CART-expressing submucous neurones were scarce. Cocaine- and amphetamine-regulated transcript was also expressed in the antral gastrin cells. Myenteric CART-expressing neurones in both small and large intestine issued short descending projections. In atrophic ileum, CART mRNA-expressing neurones increased in number while neurones containing CART peptide decreased. In hypertrophied ileum, no change in CART peptide or CART mRNA containing myenteric neurones was detected. Cocaine- and amphetamine-regulated transcript 55-102 (10(-9)-10(-7) mol L-1) did not induce any contractile or relaxatory responses in the muscle strips, neither did it affect responses induced by vasoactive intestinal peptide, pituitary adenylate cyclase-activating peptide or neuronal stimulation. In colonic, but not in ileal, strips addition of CART attenuated nitric oxide (NO) donor-induced relaxations. Although CART does not seem to play a pivotal role in classic neurotransmission to the longitudinal muscle, it may serve a modulatory role in NO transmission. It may, moreover, be involved in intestinal adaptation, and an additional hormonal role is also possible.


Asunto(s)
Tracto Gastrointestinal/química , Tracto Gastrointestinal/fisiología , Proteínas del Tejido Nervioso/análisis , Proteínas del Tejido Nervioso/fisiología , Animales , Relación Dosis-Respuesta a Droga , Femenino , Tracto Gastrointestinal/efectos de los fármacos , Relajación Muscular/efectos de los fármacos , Relajación Muscular/fisiología , Proteínas del Tejido Nervioso/farmacología , Ratas , Ratas Sprague-Dawley
18.
Regul Pept ; 107(1-3): 63-9, 2002 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-12137967

RESUMEN

OBJECTIVES: Ghrelin, an endogenous ligand of the growth hormone secretagogue receptor (GHS-R), was recently identified in the stomach. Ghrelin is produced in a population of endocrine cells in the gastric mucosa, but expression in intestine, hypothalamus and testis has also been reported. Recent data indicate that ghrelin affects insulin secretion and plays a direct role in metabolic regulation and energy balance. On the basis of these findings, we decided to examine whether ghrelin is expressed in human pancreas. Specimens from fetal to adult human pancreas and stomach were studied by immunocytochemistry, for ghrelin and islet hormones, and in situ hybridisation, for ghrelin mRNA. RESULTS: We identified ghrelin expression in a separate population of islet cells in human fetal, neonatal, and adult pancreas. Pancreatic ghrelin cells were numerous from midgestation to early postnatally (10% of all endocrine cells). The cells were few, but regularly seen in adults as single cells at the islet periphery, in exocrine tissue, in ducts, and in pancreatic ganglia. Ghrelin cells did not express any of the known islet hormones. In fetuses, at midgestation, ghrelin cells in the pancreas clearly outnumbered those in the stomach. CONCLUSIONS: Ghrelin is expressed in a quite prominent endocrine cell population in human fetal pancreas, and ghrelin expression in the pancreas precedes by far that in the stomach. Pancreatic ghrelin cells remain in adult islets at lower numbers. Ghrelin is not co-expressed with any known islet hormone, and the ghrelin cells may therefore constitute a new islet cell type.


Asunto(s)
Islotes Pancreáticos/citología , Páncreas/metabolismo , Hormonas Peptídicas/biosíntesis , Adulto , Recuento de Células , Mucosa Gástrica/metabolismo , Ghrelina , Humanos , Inmunohistoquímica , Hibridación in Situ , Recién Nacido , Islotes Pancreáticos/embriología , Islotes Pancreáticos/metabolismo , Microscopía Fluorescente , Páncreas/embriología , Páncreas/crecimiento & desarrollo , Hormonas Pancreáticas/biosíntesis , ARN Mensajero/biosíntesis
19.
Eur J Neurosci ; 15(1): 120-32, 2002 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-11860512

RESUMEN

In an attempt to define clinically relevant models of akinesia and dyskinesia in 6-hydroxydopamine (6-OHDA)-lesioned rats, we have examined the effects of drugs with high (L-DOPA) vs. low (bromocriptine) dyskinesiogenic potential in Parkinson's disease on three types of motor performance, namely: (i) abnormal involuntary movements (AIMs) (ii) rotational behaviour, and (iii) spontaneous forelimb use (cylinder test). Rats with unilateral 6-OHDA lesions received single daily i.p. injections of L-DOPA or bromocriptine at therapeutic doses. During 3 weeks of treatment, L-DOPA but not bromocriptine induced increasingly severe AIMs affecting the limb, trunk and orofacial region. Rotational behaviour was induced to a much higher extent by bromocriptine than L-DOPA. In the cylinder test, the two drugs initially improved the performance of the parkinsonian limb to a similar extent. However, L-DOPA-treated animals showed declining levels of performance in this test because the drug-induced AIMs interfered with physiological limb use, and gradually replaced all normal motor activities. L-DOPA-induced axial, limb and orolingual AIM scores were significantly reduced by the acute administration of compounds that have antidyskinetic efficacy in parkinsonian patients and/or nonhuman primates (-91%, yohimbine 10 mg/kg; -19%, naloxone 4-8 mg/kg; -37%, 5-methoxy 5-N,N-dimethyl-tryptamine 2 mg/kg; -30%, clozapine 8 mg/kg; -50%, amantadine 40 mg/kg). L-DOPA-induced rotation was, however, not affected. The present results demonstrate that 6-OHDA-lesioned rats do exhibit motor deficits that share essential functional similarities with parkinsonian akinesia or dyskinesia. Such deficits can be quantified using novel and relatively simple testing procedures, whereas rotometry cannot discriminate between dyskinetic and antiakinetic effects of antiparkinsonian treatments.


Asunto(s)
Conducta Animal/efectos de los fármacos , Discinesia Inducida por Medicamentos/psicología , Enfermedad de Parkinson Secundaria/psicología , Animales , Antidiscinéticos/farmacología , Antiparkinsonianos/farmacología , Bromocriptina/farmacología , Discinesia Inducida por Medicamentos/tratamiento farmacológico , Femenino , Marcha/efectos de los fármacos , Levodopa/farmacología , Actividad Motora/efectos de los fármacos , Oxidopamina/farmacología , Enfermedad de Parkinson Secundaria/inducido químicamente , Ratas , Ratas Sprague-Dawley , Rotación , Conducta Estereotipada/efectos de los fármacos , Simpaticolíticos/farmacología
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