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1.
Nature ; 579(7798): 279-283, 2020 03.
Artículo en Inglés | MEDLINE | ID: mdl-32132708

RESUMEN

Although it is well-established that reductions in the ratio of insulin to glucagon in the portal vein have a major role in the dysregulation of hepatic glucose metabolism in type-2 diabetes1-3, the mechanisms by which glucagon affects hepatic glucose production and mitochondrial oxidation are poorly understood. Here we show that glucagon stimulates hepatic gluconeogenesis by increasing the activity of hepatic adipose triglyceride lipase, intrahepatic lipolysis, hepatic acetyl-CoA content and pyruvate carboxylase flux, while also increasing mitochondrial fat oxidation-all of which are mediated by stimulation of the inositol triphosphate receptor 1 (INSP3R1). In rats and mice, chronic physiological increases in plasma glucagon concentrations increased mitochondrial oxidation of fat in the liver and reversed diet-induced hepatic steatosis and insulin resistance. However, these effects of chronic glucagon treatment-reversing hepatic steatosis and glucose intolerance-were abrogated in Insp3r1 (also known as Itpr1)-knockout mice. These results provide insights into glucagon biology and suggest that INSP3R1 may represent a target for therapies that aim to reverse nonalcoholic fatty liver disease and type-2 diabetes.


Asunto(s)
Glucagón/farmacología , Gluconeogénesis/efectos de los fármacos , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Hígado/efectos de los fármacos , Acetilcoenzima A/metabolismo , Tejido Adiposo/efectos de los fármacos , Animales , Diabetes Mellitus Tipo 2/fisiopatología , Activación Enzimática/efectos de los fármacos , Glucagón/sangre , Receptores de Inositol 1,4,5-Trifosfato/genética , Lipasa/metabolismo , Lipólisis/efectos de los fármacos , Lipólisis/genética , Ratones Noqueados , Mitocondrias/efectos de los fármacos , Enfermedad del Hígado Graso no Alcohólico/fisiopatología , Oxidación-Reducción/efectos de los fármacos
2.
Cell Rep ; 28(3): 759-772.e10, 2019 07 16.
Artículo en Inglés | MEDLINE | ID: mdl-31315053

RESUMEN

Mechanisms coordinating pancreatic ß cell metabolism with insulin secretion are essential for glucose homeostasis. One key mechanism of ß cell nutrient sensing uses the mitochondrial GTP (mtGTP) cycle. In this cycle, mtGTP synthesized by succinyl-CoA synthetase (SCS) is hydrolyzed via mitochondrial PEPCK (PEPCK-M) to make phosphoenolpyruvate, a high-energy metabolite that integrates TCA cycling and anaplerosis with glucose-stimulated insulin secretion (GSIS). Several strategies, including xenotopic overexpression of yeast mitochondrial GTP/GDP exchanger (GGC1) and human ATP and GTP-specific SCS isoforms, demonstrated the importance of the mtGTP cycle. These studies confirmed that mtGTP triggers and amplifies normal GSIS and rescues defects in GSIS both in vitro and in vivo. Increased mtGTP synthesis enhanced calcium oscillations during GSIS. mtGTP also augmented mitochondrial mass, increased insulin granule number, and membrane proximity without triggering de-differentiation or metabolic fragility. These data highlight the importance of the mtGTP signal in nutrient sensing, insulin secretion, mitochondrial maintenance, and ß cell health.

3.
Cell Rep ; 24(1): 47-55, 2018 07 03.
Artículo en Inglés | MEDLINE | ID: mdl-29972790

RESUMEN

Obesity is associated with colon cancer pathogenesis, but the underlying mechanism is actively debated. Here, we confirm that diet-induced obesity promotes tumor growth in two murine colon cancer models and show that this effect is reversed by an orally administered controlled-release mitochondrial protonophore (CRMP) that acts as a liver-specific uncoupler of oxidative phosphorylation. This agent lowered circulating insulin, and the reduction of tumor growth was abrogated by an insulin infusion raising plasma insulin to the level of high-fat-fed mice. We also demonstrate that hyperinsulinemia increases glucose uptake and oxidation in vivo in tumors and that CRMP reverses these effects. This study provides evidence that perturbations of whole-organism energy balance or hepatic energy metabolism can influence neoplastic growth. Furthermore, the data show that glucose uptake and utilization by cancers in vivo are not necessarily constitutively high but rather may vary according to the hormonal milieu.


Asunto(s)
Neoplasias del Colon/patología , Hígado/metabolismo , Fosforilación Oxidativa , Proteína de la Poliposis Adenomatosa del Colon/metabolismo , Animales , Glucemia/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Neoplasias del Colon/sangre , Pólipos del Colon/patología , Modelos Animales de Enfermedad , Hiperinsulinismo/metabolismo , Insulina/metabolismo , Hígado/efectos de los fármacos , Masculino , Metformina/farmacología , Ratones Endogámicos C57BL , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Oxidación-Reducción , Fosforilación Oxidativa/efectos de los fármacos , Protones
4.
Diabetologia ; 61(6): 1435-1446, 2018 06.
Artículo en Inglés | MEDLINE | ID: mdl-29497783

RESUMEN

AIMS/HYPOTHESIS: Targeting regulators of adipose tissue lipoprotein lipase could enhance adipose lipid clearance, prevent ectopic lipid accumulation and consequently ameliorate insulin resistance and type 2 diabetes. Angiopoietin-like 8 (ANGPTL8) is an insulin-regulated lipoprotein lipase inhibitor strongly expressed in murine adipose tissue. However, Angptl8 knockout mice do not have improved insulin resistance. We hypothesised that pharmacological inhibition, using a second-generation antisense oligonucleotide (ASO) against Angptl8 in adult high-fat-fed rodents, would prevent ectopic lipid accumulation and insulin resistance by promoting adipose lipid uptake. METHODS: ANGPTL8 expression was assessed by quantitative PCR in omental adipose tissue of bariatric surgery patients. High-fat-fed Sprague Dawley rats and C57BL/6 mice were treated with ASO against Angptl8 and insulin sensitivity was assessed by hyperinsulinaemic-euglycaemic clamps in rats and glucose tolerance tests in mice. Factors mediating lipid-induced hepatic insulin resistance were assessed, including lipid content, protein kinase Cε (PKCε) activation and insulin-stimulated Akt phosphorylation. Rat adipose lipid uptake was assessed by mixed meal tolerance tests. Murine energy balance was assessed by indirect calorimetry. RESULTS: Omental fat ANGPTL8 mRNA expression is higher in obese individuals with fatty liver and insulin resistance compared with BMI-matched insulin-sensitive individuals. Angptl8 ASO prevented hepatic steatosis, PKCε activation and hepatic insulin resistance in high-fat-fed rats. Postprandial triacylglycerol uptake in white adipose tissue was increased in Angptl8 ASO-treated rats. Angptl8 ASO protected high-fat-fed mice from glucose intolerance. Although there was no change in net energy balance, Angptl8 ASO increased fat mass in high-fat-fed mice. CONCLUSIONS/INTERPRETATION: Disinhibition of adipose tissue lipoprotein lipase is a novel therapeutic modality to enhance adipose lipid uptake and treat non-alcoholic fatty liver disease and insulin resistance. In line with this, adipose ANGPTL8 is a candidate therapeutic target for these conditions.


Asunto(s)
Tejido Adiposo/metabolismo , Proteínas Similares a la Angiopoyetina/genética , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Oligonucleótidos Antisentido/genética , Hormonas Peptídicas/genética , Animales , Composición Corporal , Calorimetría Indirecta , Dieta Alta en Grasa , Prueba de Tolerancia a la Glucosa , Resistencia a la Insulina , Metabolismo de los Lípidos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratas , Ratas Sprague-Dawley
5.
J Clin Invest ; 126(11): 4361-4371, 2016 11 01.
Artículo en Inglés | MEDLINE | ID: mdl-27760050

RESUMEN

Nonalcoholic fatty liver disease (NAFLD) is a risk factor for type 2 diabetes (T2D), but whether NAFLD plays a causal role in the pathogenesis of T2D is uncertain. One proposed mechanism linking NAFLD to hepatic insulin resistance involves diacylglycerol-mediated (DAG-mediated) activation of protein kinase C-ε (PKCε) and the consequent inhibition of insulin receptor (INSR) kinase activity. However, the molecular mechanism underlying PKCε inhibition of INSR kinase activity is unknown. Here, we used mass spectrometry to identify the phosphorylation site Thr1160 as a PKCε substrate in the functionally critical INSR kinase activation loop. We hypothesized that Thr1160 phosphorylation impairs INSR kinase activity by destabilizing the active configuration of the INSR kinase, and our results confirmed this prediction by demonstrating severely impaired INSR kinase activity in phosphomimetic T1160E mutants. Conversely, the INSR T1160A mutant was not inhibited by PKCε in vitro. Furthermore, mice with a threonine-to-alanine mutation at the homologous residue Thr1150 (InsrT1150A mice) were protected from high fat diet-induced hepatic insulin resistance. InsrT1150A mice also displayed increased insulin signaling, suppression of hepatic glucose production, and increased hepatic glycogen synthesis compared with WT controls during hyperinsulinemic clamp studies. These data reveal a critical pathophysiological role for INSR Thr1160 phosphorylation and provide further mechanistic links between PKCε and INSR in mediating NAFLD-induced hepatic insulin resistance.


Asunto(s)
Grasas de la Dieta/efectos adversos , Resistencia a la Insulina , Hígado/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Receptor de Insulina/metabolismo , Transducción de Señal/efectos de los fármacos , Sustitución de Aminoácidos , Animales , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/patología , Grasas de la Dieta/farmacología , Glucógeno/biosíntesis , Glucógeno/genética , Hígado/patología , Ratones , Ratones Mutantes , Mutación Missense , Enfermedad del Hígado Graso no Alcohólico/inducido químicamente , Enfermedad del Hígado Graso no Alcohólico/genética , Fosforilación , Proteína Quinasa C-epsilon/genética , Proteína Quinasa C-epsilon/metabolismo , Receptor de Insulina/genética
6.
Nat Commun ; 7: 12639, 2016 08 31.
Artículo en Inglés | MEDLINE | ID: mdl-27577745

RESUMEN

Insulin resistance is a key driver of type 2 diabetes (T2D) and is characterized by defective insulin receptor (INSR) signalling. Although surface INSR downregulation is a well-established contributor to insulin resistance, the underlying molecular mechanisms remain obscure. Here we show that the E3 ubiquitin ligase MARCH1 impairs cellular insulin action by degrading cell surface INSR. Using a large-scale RNA interference screen, we identify MARCH1 as a negative regulator of INSR signalling. March1 loss-of-function enhances, and March1 overexpression impairs, hepatic insulin sensitivity in mice. MARCH1 ubiquitinates INSR to decrease cell surface INSR levels, but unlike other INSR ubiquitin ligases, MARCH1 acts in the basal state rather than after insulin stimulation. Thus, MARCH1 may help set the basal gain of insulin signalling. MARCH1 expression is increased in white adipose tissue of obese humans, suggesting that MARCH1 contributes to the pathophysiology of T2D and could be a new therapeutic target.


Asunto(s)
Antígenos CD/metabolismo , Diabetes Mellitus Tipo 2/patología , Resistencia a la Insulina/fisiología , Insulina/metabolismo , Obesidad/patología , Receptor de Insulina/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Tejido Adiposo Blanco/patología , Adolescente , Animales , Antígenos CD/genética , Biopsia , Línea Celular , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/terapia , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Regulación hacia Abajo , Femenino , Técnicas de Silenciamiento del Gen , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mutagénesis Sitio-Dirigida , Obesidad/sangre , Obesidad/etiología , Obesidad/terapia , Oligonucleótidos Antisentido/administración & dosificación , Oligonucleótidos Antisentido/genética , Fosforilación , ARN Interferente Pequeño/metabolismo , Receptor de Insulina/genética , Transducción de Señal/fisiología , Ubiquitina-Proteína Ligasas/genética , Ubiquitinación , Regulación hacia Arriba
7.
Bone ; 81: 104-111, 2015 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-26164475

RESUMEN

Parathyroid hormone-related protein (PTHrP) is widely expressed in the fibrous outer layer of the periosteum (PO), and the PTH/PTHrP type I receptor (PTHR1) is expressed in the inner PO cambial layer. The cambial layer gives rise to the PO osteoblasts (OBs) and osteoclasts (OCs) that model/remodel the cortical bone surface during development as well as during fracture healing. PTHrP has been implicated in the regulation of PO modeling during development, but nothing is known as regards a role of PTHrP in this location during fracture healing. We propose that PTHrP in the fibrous layer of the PO may be a key regulatory factor in remodeling bone formation during fracture repair. We first assessed whether PTHrP expression in the fibrous PO is associated with PO osteoblast induction in the subjacent cambial PO using a tibial fracture model in PTHrP-lacZ mice. Our results revealed that both PTHrP expression and osteoblast induction in PO were induced 3 days post-fracture. We then investigated a potential functional role of PO PTHrP during fracture repair by performing tibial fracture surgery in 10-week-old CD1 control and PTHrP conditional knockout (PTHrP cKO) mice that lack PO PTHrP. We found that callus size and formation as well as woven bone mineralization in PTHrP cKO mice were impaired compared to that in CD1 mice. Concordant with these findings, functional enzyme staining revealed impaired OB formation and OC activity in the cKO mice. We conclude that deleting PO PTHrP impairs cartilaginous callus formation, maturation and ossification as well as remodeling during fracture healing. These data are the initial genetic evidence suggesting that PO PTHrP may induce osteoblastic activity and regulate fracture healing on the cortical bone surface.


Asunto(s)
Remodelación Ósea/fisiología , Curación de Fractura/fisiología , Osteoblastos/metabolismo , Proteína Relacionada con la Hormona Paratiroidea/metabolismo , Periostio/metabolismo , Animales , Inmunohistoquímica , Captura por Microdisección con Láser , Masculino , Ratones , Ratones Noqueados , Osteogénesis/fisiología , Microtomografía por Rayos X
8.
J Anat ; 225(1): 71-82, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24762197

RESUMEN

The modeling of long bone surfaces during linear growth is a key developmental process, but its regulation is poorly understood. We report here that parathyroid hormone-related peptide (PTHrP) expressed in the fibrous layer of the periosteum (PO) drives the osteoclastic (OC) resorption that models the metaphyseal-diaphyseal junction (MDJ) in the proximal tibia and fibula during linear growth. PTHrP was conditionally deleted (cKO) in the PO via Scleraxis gene targeting (Scx-Cre). In the lateral tibia, cKO of PTHrP led to a failure of modeling, such that the normal concave MDJ was replaced by a mound-like deformity. This was accompanied by a failure to induce receptor activator of NF-kB ligand (RANKL) and a 75% reduction in OC number (P ≤ 0.001) on the cortical surface. The MDJ also displayed a curious threefold increase in endocortical osteoblast mineral apposition rate (P ≤ 0.001) and a thickened cortex, suggesting some form of coupling of endocortical bone formation to events on the PO surface. Because it fuses distally, the fibula is modeled only proximally and does so at an extraordinary rate, with an anteromedial cortex in CD-1 mice that was so moth-eaten that a clear PO surface could not be identified. The cKO fibula displayed a remarkable phenotype, with a misshapen club-like metaphysis and an enlargement in the 3D size of the entire bone, manifest as a 40-45% increase in the PO circumference at the MDJ (P ≤ 0.001) as well as the mid-diaphysis (P ≤ 0.001). These tibial and fibular phenotypes were reproduced in a Scx-Cre-driven RANKL cKO mouse. We conclude that PTHrP in the fibrous PO mediates the modeling of the MDJ of long bones during linear growth, and that in a highly susceptible system such as the fibula this surface modeling defines the size and shape of the entire bone.


Asunto(s)
Desarrollo Óseo/fisiología , Peroné/crecimiento & desarrollo , Proteína Relacionada con la Hormona Paratiroidea/fisiología , Periostio/fisiología , Tibia/crecimiento & desarrollo , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Eliminación de Gen , Ratones , Ratones Noqueados , Ligando RANK/genética
9.
J Bone Miner Res ; 28(3): 598-607, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-23109045

RESUMEN

The sites that receive ligament and tendon insertions (entheses) on the cortical surfaces of long bones are poorly understood, particularly regarding modeling and regulation. Entheses are classified as either fibrocartilaginous or fibrous based on their structures. Fibrous entheses typically insert into the metaphysis or diaphysis of a long bone, bear a periosteal component, and are modeled during long-bone growth. This modeling forms a root system by which the insertions attach to the cortical surface. In the case of the medial collateral ligament, modeling drives actual migration of the ligament along the cortical surface in order to accommodate linear growth, whereas in other sites modeling may excavate a deep cortical root system (eg, the teres major insertion) or a shallow root system with a large footprint (eg, the latissimus dorsi insertion). We report here that conditionally deleting parathyroid hormone-related protein (PTHrP) in fibrous entheses via Scleraxis-Cre targeting causes modeling to fail in these three iterations of osteoclast-driven enthesis excavation or migration. These iterations appear to represent formes frustes of a common modeling strategy, presumably differing from each other as a consequence of differences in biomechanical control. In sites in which PTHrP is not induced, either physiologically or because of conditional deletion, modeling does not take place and fibrocartilage is induced. These findings represent the initial genetic evidence that PTHrP regulates periosteal/intramembranous bone cell activity on cortical bone surfaces and indicate that PTHrP serves as a load-induced modeling tool in fibrous insertion sites during linear growth.


Asunto(s)
Desarrollo Óseo/fisiología , Fibrocartílago/crecimiento & desarrollo , Modelos Biológicos , Proteína Relacionada con la Hormona Paratiroidea/fisiología , Animales , Ratones
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