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Osipova et al.1 recently identified an inactivating gene mutation that contributed to the evolution of the hummingbird species by increasing flux of pathways for energy production that are necessary for the unique ability for hovering flight. Lessons from the natural selection for this mutation are applied to physiology and medicine.
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Aves , Vuelo Animal , Animales , Vuelo Animal/fisiología , Aves/genética , Aves/metabolismo , Metabolismo Energético/genética , Consumo de Oxígeno , Selección GenéticaRESUMEN
Metabolic dysfunction-associated steatotic liver disease is the most common form of liver disease and poses significant health risks to patients who progress to metabolic dysfunction-associated steatohepatitis. Fatty acid overload alters endoplasmic reticulum (ER) calcium stores and induces mitochondrial oxidative stress in hepatocytes, leading to hepatocellular inflammation and apoptosis. Obese mice have impaired liver sarco/ER Ca2+-ATPase (SERCA) function, which normally maintains intracellular calcium homeostasis by transporting Ca2+ ions from the cytoplasm to the ER. We hypothesized that restoration of SERCA activity would improve diet-induced steatohepatitis in mice by limiting ER stress and mitochondrial dysfunction. WT and melanocortin-4 receptor KO (Mc4r-/-) mice were placed on either chow or Western diet (WD) for 8 weeks. Half of the WD-fed mice were administered CDN1163 to activate SERCA, which reduced liver fibrosis and inflammation. SERCA activation also restored glucose tolerance and insulin sensitivity, improved histological markers of metabolic dysfunction-associated steatohepatitis, increased expression of antioxidant enzymes, and decreased expression of oxidative stress and ER stress genes. CDN1163 decreased hepatic citric acid cycle flux and liver pyruvate cycling, enhanced expression of mitochondrial respiratory genes, and shifted hepatocellular [NADH]/[NAD+] and [NADPH]/[NADP+] ratios to a less oxidized state, which was associated with elevated PUFA content of liver lipids. In sum, the data demonstrate that pharmacological SERCA activation limits metabolic dysfunction-associated steatotic liver disease progression and prevents metabolic dysfunction induced by WD feeding in mice.
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Hígado , ATPasas Transportadoras de Calcio del Retículo Sarcoplásmico , Animales , ATPasas Transportadoras de Calcio del Retículo Sarcoplásmico/metabolismo , Ratones , Hígado/metabolismo , Hígado/patología , Masculino , Hígado Graso/metabolismo , Hígado Graso/patología , Estrés del Retículo Endoplásmico , Ratones Endogámicos C57BL , Estrés Oxidativo/efectos de los fármacos , Dieta Occidental/efectos adversos , Ratones NoqueadosRESUMEN
Regular exercise elicits adaptations in glucose and lipid metabolism that allow the body to meet energy demands of subsequent exercise bouts more effectively and mitigate metabolic diseases including fatty liver. Energy discharged during the acute exercise bouts that comprise exercise training may be a catalyst for liver adaptations. During acute exercise, liver glycogenolysis and gluconeogenesis are accelerated to supply glucose to working muscle. Lower liver energy state imposed by gluconeogenesis and related pathways activates AMP-activated protein kinase (AMPK), which conserves ATP partly by promoting lipid oxidation. This study tested the hypothesis that AMPK is necessary for liver glucose and lipid adaptations to training. Liver-specific AMPKα1α2 knockout (AMPKα1α2fl/fl+AlbCre) mice and littermate controls (AMPKα1α2fl/fl) completed sedentary and exercise training protocols. Liver nutrient fluxes were quantified at rest or during acute exercise following training. Liver metabolites and molecular regulators of metabolism were assessed. Training increased liver glycogen in AMPKα1α2fl/fl mice, but not in AMPKα1α2fl/fl+AlbCre mice. The inability to increase glycogen led to lower glycogenolysis, glucose production, and circulating glucose during acute exercise in trained AMPKα1α2fl/fl+AlbCre mice. Deletion of AMPKα1α2 attenuated training-induced declines in liver diacylglycerides. In particular, training lowered the concentration of unsaturated and elongated fatty acids comprising diacylglycerides in AMPKα1α2fl/fl mice, but not in AMPKα1α2fl/fl+AlbCre mice. Training increased liver triacylglycerides and the desaturation and elongation of fatty acids in triacylglycerides of AMPKα1α2fl/fl+AlbCre mice. These lipid responses were independent of differences in tricarboxylic acid cycle fluxes. In conclusion, AMPK is required for liver training adaptations that are critical to glucose and lipid metabolism.NEW & NOTEWORTHY This study shows that the energy sensor and transducer, AMP-activated protein kinase (AMPK), is necessary for an exercise training-induced: 1) increase in liver glycogen that is necessary for accelerated glycogenolysis during exercise, 2) decrease in liver glycerolipids independent of tricarboxylic acid (TCA) cycle flux, and 3) decline in the desaturation and elongation of fatty acids comprising liver diacylglycerides. The mechanisms defined in these studies have implications for use of regular exercise or AMPK-activators in patients with fatty liver.
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Proteínas Quinasas Activadas por AMP , Hígado Graso , Humanos , Animales , Ratones , Proteínas Quinasas Activadas por AMP/metabolismo , Glucógeno Hepático , Hígado/metabolismo , Glucosa/metabolismo , Hígado Graso/metabolismo , Ácidos Grasos/metabolismoRESUMEN
The Mouse Metabolic Phenotyping Center (MMPC)Live Program was established in 2023 by the National Institute for Diabetes, Digestive and Kidney Diseases (NIDDK) at the National Institutes of Health (NIH) to advance biomedical research by providing the scientific community with standardized, high quality phenotyping services for mouse models of diabetes and obesity. Emerging as the next iteration of the MMPC Program which served the biomedical research community for 20 years (2001-2021), MMPCLive is designed as an outwardly-facing consortium of service cores that collaborate to provide reduced-cost consultation and metabolic, physiologic, and behavioral phenotyping tests on live mice for U.S. biomedical researchers. Four MMPCLive Centers located at universities around the country perform complex and often unique procedures in vivo on a fee for service basis, typically on mice shipped from the client or directly from a repository or vendor. Current areas of expertise include energy balance and body composition, insulin action and secretion, whole body carbohydrate and lipid metabolism, cardiovascular and renal function, food intake and behavior, microbiome and xenometabolism, and metabolic pathway kinetics. Additionally, an opportunity arose to reduce barriers to access and expand the diversity of the biomedical research workforce by establishing the VIBRANT Program. Directed at researchers historically underrepresented in the biomedical sciences, VIBRANT-eligible investigators have access to testing services, travel and career development awards, expert advice and experimental design consultation, and short internships to learn test technologies. Data derived from experiments run by the Centers belongs to the researchers submitting mice for testing which can be made publicly available and accessible from the MMPCLive database following publication. In addition to services, MMPCLive staff provide expertise and advice to researchers, develop and refine test protocols, engage in outreach activities, publish scientific and technical papers, and conduct educational workshops and training sessions to aid researchers in unraveling the heterogeneity of diabetes and obesity.
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Research conducted over the last 50 yr has provided insight into the mechanisms by which insulin stimulates glucose transport across the skeletal muscle cell membrane Transport alone, however, does not result in net glucose uptake as free glucose equilibrates across the cell membrane and is not metabolized. Glucose uptake requires that glucose is phosphorylated by hexokinases. Phosphorylated glucose cannot leave the cell and is the substrate for metabolism. It is indisputable that glucose phosphorylation is essential for glucose uptake. Major advances have been made in defining the regulation of the insulin-stimulated glucose transporter (GLUT4) in skeletal muscle. By contrast, the insulin-regulated hexokinase (hexokinase II) parallels Robert Frost's "The Road Not Taken." Here the case is made that an understanding of glucose phosphorylation by hexokinase II is necessary to define the regulation of skeletal muscle glucose uptake in health and insulin resistance. Results of studies from different physiological disciplines that have elegantly described how hexokinase II can be regulated are summarized to provide a framework for potential application to skeletal muscle. Mechanisms by which hexokinase II is regulated in skeletal muscle await rigorous examination.
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Hexoquinasa , Insulina , Transporte Biológico , Glucosa/metabolismo , Hexoquinasa/metabolismo , Humanos , Insulina/metabolismo , Músculo Esquelético/metabolismoRESUMEN
BACKGROUND: The mechanism of adverse limb events associated with peripheral artery disease remains incompletely understood. We investigated whether microvascular disease is associated with amputation in a large cohort of veterans to determine whether microvascular disease diagnosed in any location increases the risk of amputation alone and in concert with peripheral artery disease. METHODS: Participants in the Veterans Aging Cohort Study were recruited from April 1, 2003 through December 31, 2014. We excluded participants with known prior lower limb amputation. Using time-updated Cox proportional hazards regression, we analyzed the effect of prevalent microvascular disease (retinopathy, neuropathy, and nephropathy) and peripheral artery disease status on the risk of incident amputation events after adjusting for demographics and cardiovascular risk factors. RESULTS: Among 125 674 veterans without evidence of prior amputation at baseline, the rate of incident amputation over a median of 9.3 years of follow-up was 1.16 per 1000 person-years, yielding a total of 1185 amputations. In time-updated multivariable-adjusted analyses, compared with those without peripheral artery disease or microvascular disease, microvascular disease alone was associated with a 3.7-fold (95% CI, 3.0-4.6) increased risk of amputation; peripheral artery disease alone conferred a 13.9-fold (95% CI, 11.3-17.1) elevated risk of amputation; and the combination of peripheral artery disease and microvascular disease was associated with a 22.7-fold (95% CI, 18.3-28.1) increased risk of amputation. CONCLUSIONS: Independent of traditional risk factors, the presence of microvascular disease increases the risk of amputation alone and synergistically increases risk in patients with peripheral artery disease. Further research is needed to understand the mechanisms by which this occurs.
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Amputación Quirúrgica , Microcirculación , Enfermedad Arterial Periférica/cirugía , Adulto , Anciano , Amputación Quirúrgica/estadística & datos numéricos , Comorbilidad , Complicaciones de la Diabetes/epidemiología , Susceptibilidad a Enfermedades , Extremidades/irrigación sanguínea , Femenino , Estudios de Seguimiento , Humanos , Isquemia/etiología , Isquemia/cirugía , Enfermedades Renales/epidemiología , Masculino , Persona de Mediana Edad , Enfermedad Arterial Periférica/complicaciones , Enfermedad Arterial Periférica/fisiopatología , Enfermedades del Sistema Nervioso Periférico/epidemiología , Prevalencia , Utilización de Procedimientos y Técnicas , Modelos de Riesgos Proporcionales , Estudios Prospectivos , Enfermedades de la Retina/epidemiología , Factores de Riesgo , Veteranos/estadística & datos numéricosRESUMEN
Integrins are important for mechanosensation in tissue and play, together with nutrition, a role in regulating extracellular matrix (ECM) in skeletal muscle and tendon. Integrin receptors are dimers that consist of an α and ß subunit and bridge extracellular and intracellular signals. The present study investigates whether the deletion of the integrin receptor α1 subunit influences collagen and other matrix proteins in the musculotendinous tissue and whether it causes any compensatory changes in other integrin subunits in C57BL/6J mice. In addition, we study whether a high-fat diet (HFD) influences these responses in muscle or tendon. Mice on a HFD had a higher number of non-enzymatic cross-links in skeletal muscle ECM and increased gene expression of collagen and other extracellular matrix proteins. In contrast to gene expression, total collagen protein content was decreased by HFD in the muscle with no change in tendon. Integrin α1 subunit knockout resulted in a decrease of collagen type I and III, TGF-ß1 and IGF-1 gene expression in muscle of HFD mice but did not affect total collagen protein compared with wild-type (WT) littermates in either muscle or tendon. There was no compensatory increase in the genes that express other integrin subunits. In conclusion, HFD induced a significant increase in expression of ECM genes in muscle. On the protein level, HFD resulted in a lower collagen content in muscle. Tendons were unaffected by the diet. Deletion of the integrin α1 subunit did not affect collagen protein or gene expression in muscle or tendon.
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Tendón Calcáneo/metabolismo , Colágeno/metabolismo , Matriz Extracelular , Integrina alfa1/fisiología , Músculo Esquelético/metabolismo , Animales , Dieta Alta en Grasa , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Obesidad/metabolismoRESUMEN
The frequency of prediabetes is increasing as the prevalence of obesity rises worldwide. In prediabetes, hyperglycemia, insulin resistance, and inflammation and metabolic derangements associated with concomitant obesity cause endothelial vasodilator and fibrinolytic dysfunction, leading to increased risk of cardiovascular and renal disease. Importantly, the microvasculature affects insulin sensitivity by affecting the delivery of insulin and glucose to skeletal muscle; thus, endothelial dysfunction and extracellular matrix remodeling promote the progression from prediabetes to diabetes mellitus. Weight loss is the mainstay of treatment in prediabetes, but therapies that improved endothelial function and vasodilation may not only prevent cardiovascular disease but also slow progression to diabetes mellitus.
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Vasos Sanguíneos/fisiopatología , Microcirculación , Estado Prediabético/fisiopatología , Inhibidores de la Enzima Convertidora de Angiotensina/uso terapéutico , Animales , Enfermedades Cardiovasculares/etiología , Enfermedades Cardiovasculares/fisiopatología , Terapia Combinada , Diabetes Mellitus Tipo 2/fisiopatología , Dieta Reductora , Progresión de la Enfermedad , Endotelio Vascular/fisiopatología , Matriz Extracelular/metabolismo , Ácidos Grasos no Esterificados/metabolismo , Fibrinólisis , Glucosa/metabolismo , Humanos , Hiperglucemia/etiología , Hiperglucemia/fisiopatología , Hipoglucemiantes/uso terapéutico , Inflamación/fisiopatología , Resistencia a la Insulina , Estilo de Vida , Síndrome Metabólico/fisiopatología , Ratones , MicroARNs/genética , Músculo Esquelético/metabolismo , Obesidad/fisiopatología , Estado Prediabético/epidemiología , Estado Prediabético/patología , Estado Prediabético/terapia , Riesgo , Pérdida de PesoRESUMEN
Aberrant extracellular matrix (ECM) remodelling in muscle, liver and adipose tissue is a key characteristic of obesity and insulin resistance. Despite its emerging importance, the effective ECM targets remain largely undefined due to limitations of current approaches. Here, we developed a novel ECM-specific mass spectrometry-based proteomics technique to characterise the global view of the ECM changes in the skeletal muscle and liver of mice after high fat (HF) diet feeding. We identified distinct signatures of HF-induced protein changes between skeletal muscle and liver where the ECM remodelling was more prominent in the muscle than liver. In particular, most muscle collagen isoforms were increased by HF diet feeding whereas the liver collagens were differentially but moderately affected highlighting a different role of the ECM remodelling in different tissues of obesity. Moreover, we identified a novel association between collagen 24α1 and insulin resistance in the skeletal muscle. Using quantitative gene expression analysis, we extended this association to the white adipose tissue. Importantly, collagen 24α1 mRNA was increased in the visceral adipose tissue, but not the subcutaneous adipose tissue of obese diabetic subjects compared to lean controls, implying a potential pathogenic role of collagen 24α1 in obesity and type 2 diabetes.
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Colágeno/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Resistencia a la Insulina , Grasa Intraabdominal/metabolismo , Músculo Esquelético/metabolismo , Obesidad/metabolismo , Grasa Subcutánea/metabolismo , Animales , Colágeno/genética , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Matriz Extracelular/metabolismo , Expresión Génica , Humanos , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Obesidad/etiología , ARN Mensajero/genéticaRESUMEN
Glycine N-methyltransferase (GNMT) is the most abundant liver methyltransferase regulating the availability of the biological methyl donor, S-adenosylmethionine (SAM). Moreover, GNMT has been identified to be down-regulated in hepatocellular carcinoma (HCC). Despite its role in regulating SAM levels and association of its down-regulation with liver tumorigenesis, the impact of reduced GNMT on metabolic reprogramming before the manifestation of HCC has not been investigated in detail. Herein, we used 2H/13C metabolic flux analysis in conscious, unrestrained mice to test the hypothesis that the absence of GNMT causes metabolic reprogramming. GNMT-null (KO) mice displayed a reduction in blood glucose that was associated with a decline in both hepatic glycogenolysis and gluconeogenesis. The reduced gluconeogenesis was due to a decrease in liver gluconeogenic precursors, citric acid cycle fluxes, and anaplerosis and cataplerosis. A concurrent elevation in both hepatic SAM and metabolites of SAM utilization pathways was observed in the KO mice. Specifically, the increase in metabolites of SAM utilization pathways indicated that hepatic polyamine synthesis and catabolism, transsulfuration, and de novo lipogenesis pathways were increased in the KO mice. Of note, these pathways utilize substrates that could otherwise be used for gluconeogenesis. Also, this metabolic reprogramming occurs before the well-documented appearance of HCC in GNMT-null mice. Together, these results indicate that GNMT deletion promotes a metabolic shift whereby nutrients are channeled away from glucose formation toward pathways that utilize the elevated SAM.
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Carbono/metabolismo , Eliminación de Gen , Gluconeogénesis , Glicina N-Metiltransferasa/genética , Metionina/metabolismo , Animales , Ciclo del Ácido Cítrico , Metabolismo Energético , Hígado Graso/genética , Hígado Graso/metabolismo , Glucosa/metabolismo , Glicina N-Metiltransferasa/metabolismo , Hígado/metabolismo , Masculino , Análisis de Flujos Metabólicos , Ratones , Ratones Noqueados , S-Adenosilmetionina/metabolismoRESUMEN
Integrin-linked kinase (ILK) is a critical intracellular signaling node for integrin receptors. Its role in liver development is complex, as ILK deletion at E10.5 (before hepatocyte differentiation) results in biochemical and morphological differences that resolve as mice age. Nevertheless, mice with ILK depleted specifically in hepatocytes are protected from the hepatic insulin resistance during obesity. Despite the potential importance of hepatocyte ILK to metabolic health, it is unknown how ILK controls hepatic metabolism or glucoregulation. The present study tested the role of ILK in hepatic metabolism and glucoregulation by deleting it specifically in hepatocytes, using a cre-lox system that begins expression at E15.5 (after initiation of hepatocyte differentiation). These mice develop the most severe morphological and glucoregulatory abnormalities at 6 wk, but these gradually resolve with age. After identifying when the deletion of ILK caused a severe metabolic phenotype, in depth studies were performed at this time point to define the metabolic programs that coordinate control of glucoregulation that are regulated by ILK. We show that 6-wk-old ILK-deficient mice have higher glucose tolerance and decreased net glycogen synthesis. Additionally, ILK was shown to be necessary for transcription of mitochondrial-related genes, oxidative metabolism, and maintenance of cellular energy status. Thus, ILK is required for maintaining hepatic transcriptional and metabolic programs that sustain oxidative metabolism, which are required for hepatic maintenance of glucose homeostasis.
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Glucemia/metabolismo , Hepatocitos/metabolismo , Resistencia a la Insulina , Insulina/metabolismo , Hígado/metabolismo , Obesidad/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Factores de Edad , Animales , Diferenciación Celular , Respiración de la Célula , Metabolismo Energético , Técnicas de Inactivación de Genes , Glucosa/metabolismo , Prueba de Tolerancia a la Glucosa , Homeostasis , Inflamación , Hígado/embriología , Hígado/patología , Cirrosis Hepática , Ratones , Proteínas Serina-Treonina Quinasas/metabolismoRESUMEN
These studies test, using intravital microscopy (IVM), the hypotheses that perfusion effects on insulin-stimulated muscle glucose uptake (MGU) are 1) capillary recruitment independent and 2) mediated through the dispersion of glucose rather than insulin. For experiment 1, capillary perfusion was visualized before and after intravenous insulin. No capillary recruitment was observed. For experiment 2, mice were treated with vasoactive compounds (sodium nitroprusside, hyaluronidase, and lipopolysaccharide), and dispersion of fluorophores approximating insulin size (10-kDa dextran) and glucose (2-NBDG) was measured using IVM. Subsequently, insulin and 2[14C]deoxyglucose were injected and muscle phospho-2[14C]deoxyglucose (2[C14]DG) accumulation was used as an index of MGU. Flow velocity and 2-NBDG dispersion, but not perfused surface area or 10-kDa dextran dispersion, predicted phospho-2[14C]DG accumulation. For experiment 3, microspheres of the same size and number as are used for contrast-enhanced ultrasound (CEU) studies of capillary recruitment were visualized using IVM. Due to their low concentration, microspheres were present in only a small fraction of blood-perfused capillaries. Microsphere-perfused blood volume correlated to flow velocity. These findings suggest that 1) flow velocity rather than capillary recruitment controls microvascular contributions to MGU, 2) glucose dispersion is more predictive of MGU than dispersion of insulin-sized molecules, and 3) CEU measures regional flow velocity rather than capillary recruitment.
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Velocidad del Flujo Sanguíneo/fisiología , Glucosa/metabolismo , Microcirculación/fisiología , Músculo Esquelético/irrigación sanguínea , Músculo Esquelético/metabolismo , 4-Cloro-7-nitrobenzofurazano/análogos & derivados , 4-Cloro-7-nitrobenzofurazano/metabolismo , Animales , Velocidad del Flujo Sanguíneo/efectos de los fármacos , Radioisótopos de Carbono , Desoxiglucosa/análogos & derivados , Desoxiglucosa/metabolismo , Dextranos/metabolismo , Hipoglucemiantes/farmacología , Insulina/farmacología , Microscopía Intravital , Ratones , Microcirculación/efectos de los fármacos , Microesferas , Músculo Esquelético/anatomía & histología , Músculo Esquelético/diagnóstico por imagen , UltrasonografíaRESUMEN
Sepsis costs the healthcare system $23 billion annually and has a mortality rate between 10 and 40%. An early indication of sepsis is the onset of hyperglycemia, which is the result of sepsis-induced insulin resistance in skeletal muscle. Previous investigations have focused on events in the myocyte (e.g., insulin signaling and glucose transport and subsequent metabolism) as the causes for this insulin-resistant state. However, the delivery of insulin to the skeletal muscle is also an important determinant of insulin action. Skeletal muscle microvascular blood flow, which delivers the insulin to the muscle, is known to be decreased during sepsis. Here we test whether the reduced capillary blood flow to skeletal muscle belies the sepsis-induced insulin resistance by reducing insulin delivery to the myocyte. We hypothesize that decreased capillary flow and consequent decrease in insulin delivery is an early event that precedes gross cardiovascular alterations seen with sepsis. This hypothesis was examined in mice treated with either lipopolysaccharide (LPS) or polymicrobial sepsis followed by intravital microscopy of the skeletal muscle microcirculation. We calculated insulin delivery to the myocyte using two independent methods and found that LPS and sepsis rapidly reduce insulin delivery to the skeletal muscle by ~50%; this was driven by decreases in capillary flow velocity and the number of perfused capillaries. Furthermore, the changes in skeletal muscle microcirculation occur before changes in both cardiac output and arterial blood pressure. These data suggest that a rapid reduction in skeletal muscle insulin delivery contributes to the induction of insulin resistance during sepsis.
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Capilares/metabolismo , Hiperglucemia/metabolismo , Resistencia a la Insulina , Insulina/metabolismo , Microcirculación , Músculo Esquelético/metabolismo , Sepsis/metabolismo , Animales , Permeabilidad Capilar , Modelos Animales de Enfermedad , Ecocardiografía , Lipopolisacáridos , Ratones , Microvasos/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/irrigación sanguíneaRESUMEN
Extracellular matrix hyaluronan is increased in skeletal muscle of high-fat-fed insulin-resistant mice, and reduction of hyaluronan by PEGPH20 hyaluronidase ameliorates diet-induced insulin resistance (IR). CD44, the main hyaluronan receptor, is positively correlated with type 2 diabetes. This study determines the role of CD44 in skeletal muscle IR. Global CD44-deficient (cd44-/-) mice and wild-type littermates (cd44+/+) were fed a chow diet or 60% high-fat diet for 16 wk. High-fat-fed cd44-/- mice were also treated with PEGPH20 to evaluate its CD44-dependent action. Insulin sensitivity was measured by hyperinsulinemic-euglycemic clamp (ICv). High-fat feeding increased muscle CD44 protein expression. In the absence of differences in body weight and composition, despite lower clamp insulin during ICv, the cd44-/- mice had sustained glucose infusion rate (GIR) regardless of diet. High-fat diet-induced muscle IR as evidenced by decreased muscle glucose uptake (Rg) was exhibited in cd44+/+ mice but absent in cd44-/- mice. Moreover, gastrocnemius Rg remained unchanged between genotypes on chow diet but was increased in high-fat-fed cd44-/- compared with cd44+/+ when normalized to clamp insulin concentrations. Ameliorated muscle IR in high-fat-fed cd44-/- mice was associated with increased vascularization. In contrast to previously observed increases in wild-type mice, PEGPH20 treatment in high-fat-fed cd44-/- mice did not change GIR or muscle Rg during ICv, suggesting a CD44-dependent action. In conclusion, genetic CD44 deletion improves muscle IR, and the beneficial effects of PEGPH20 are CD44-dependent. These results suggest a critical role of CD44 in promoting hyaluronan-mediated muscle IR, therefore representing a potential therapeutic target for diabetes.
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Dieta Alta en Grasa , Glucosa/metabolismo , Receptores de Hialuranos/genética , Ácido Hialurónico/metabolismo , Resistencia a la Insulina/genética , Músculo Esquelético/metabolismo , Animales , Peso Corporal , Técnica de Clampeo de la Glucosa , Receptores de Hialuranos/metabolismo , Hialuronoglucosaminidasa/farmacología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Músculo Esquelético/irrigación sanguínea , Músculo Esquelético/efectos de los fármacosRESUMEN
Pathologies including diabetes and conditions such as exercise place an unusual demand on liver energy metabolism, and this demand induces a state of energy discharge. Hepatic AMP-activated protein kinase (AMPK) has been proposed to inhibit anabolic processes such as gluconeogenesis in response to cellular energy stress. However, both AMPK activation and glucose release from the liver are increased during exercise. Here, we sought to test the role of hepatic AMPK in the regulation of in vivo glucose-producing and citric acid cycle-related fluxes during an acute bout of muscular work. We used 2H/13C metabolic flux analysis to quantify intermediary metabolism fluxes in both sedentary and treadmill-running mice. Additionally, liver-specific AMPK α1 and α2 subunit KO and WT mice were utilized. Exercise caused an increase in endogenous glucose production, glycogenolysis, and gluconeogenesis from phosphoenolpyruvate. Citric acid cycle fluxes, pyruvate cycling, anaplerosis, and cataplerosis were also elevated during this exercise. Sedentary nutrient fluxes in the postabsorptive state were comparable for the WT and KO mice. However, the increment in the endogenous rate of glucose appearance during exercise was blunted in the KO mice because of a diminished glycogenolytic flux. This lower rate of glycogenolysis was associated with lower hepatic glycogen content before the onset of exercise and prompted a reduction in arterial glucose during exercise. These results indicate that liver AMPKα1α2 is required for maintaining glucose homeostasis during an acute bout of exercise.
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Proteínas Quinasas Activadas por AMP/metabolismo , Glucogenólisis , Hígado/enzimología , Condicionamiento Físico Animal , Proteínas Quinasas Activadas por AMP/deficiencia , Animales , Metabolismo Energético , Gluconeogénesis , Glucosa/metabolismo , Homeostasis , Marcaje Isotópico , Ratones , Ratones NoqueadosRESUMEN
OBJECTIVE: Changes in microvascular perfusion have been reported in many diseases, yet the functional significance of altered perfusion is often difficult to determine. This is partly because commonly used techniques for perfusion measurement often rely on either indirect or by-hand approaches. METHODS: We developed and validated a fully automated software technique to measure microvascular perfusion in videos acquired by fluorescence microscopy in the mouse gastrocnemius. Acute perfusion responses were recorded following intravenous injections with phenylephrine, SNP, or saline. RESULTS: Software-measured capillary flow velocity closely correlated with by-hand measured flow velocity (R2 = 0.91, P < 0.0001). Software estimates of capillary hematocrit also generally agreed with by-hand measurements (R2 = 0.64, P < 0.0001). Detection limits range from 0 to 2000 µm/s, as compared to an average flow velocity of 326 ± 102 µm/s (mean ± SD) at rest. SNP injection transiently increased capillary flow velocity and hematocrit and made capillary perfusion more steady and homogenous. Phenylephrine injection had the opposite effect in all metrics. Saline injection transiently decreased capillary flow velocity and hematocrit without influencing flow distribution or stability. All perfusion metrics were temporally stable without intervention. CONCLUSIONS: These results demonstrate a novel and sensitive technique for reproducible, user-independent quantification of microvascular perfusion.
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Automatización , Microscopía por Video , Microvasos/fisiología , Perfusión , Programas Informáticos , Animales , Velocidad del Flujo Sanguíneo , Hematócrito , Ratones , Microcirculación , Microscopía Fluorescente , Fenilefrina/farmacología , Reproducibilidad de los Resultados , Solución Salina/farmacologíaRESUMEN
AIMS/HYPOTHESIS: Insulin resistance is frequently associated with hypertension and type 2 diabetes. The cytochrome P450 (CYP) arachidonic acid epoxygenases (CYP2C, CYP2J) and their epoxyeicosatrienoic acid (EET) products lower blood pressure and may also improve glucose homeostasis. However, the direct contribution of endogenous EET production on insulin sensitivity has not been previously investigated. In this study, we tested the hypothesis that endogenous CYP2C-derived EETs alter insulin sensitivity by analysing mice lacking CYP2C44, a major EET producing enzyme, and by testing the association of plasma EETs with insulin sensitivity in humans. METHODS: We assessed insulin sensitivity in wild-type (WT) and Cyp2c44 -/- mice using hyperinsulinaemic-euglycaemic clamps and isolated skeletal muscle. Insulin secretory function was assessed using hyperglycaemic clamps and isolated islets. Vascular function was tested in isolated perfused mesenteric vessels. Insulin sensitivity and secretion were assessed in humans using frequently sampled intravenous glucose tolerance tests and plasma EETs were measured by mass spectrometry. RESULTS: Cyp2c44 -/- mice showed decreased glucose tolerance (639 ± 39.5 vs 808 ± 37.7 mmol/l × min for glucose tolerance tests, p = 0.004) and insulin sensitivity compared with WT controls (hyperinsulinaemic clamp glucose infusion rate average during terminal 30 min 0.22 ± 0.02 vs 0.33 ± 0.01 mmol kg-1 min-1 in WT and Cyp2c44 -/- mice respectively, p = 0.003). Although glucose uptake was diminished in Cyp2c44 -/- mice in vivo (gastrocnemius Rg 16.4 ± 2.0 vs 6.2 ± 1.7 µmol 100 g-1 min-1, p < 0.01) insulin-stimulated glucose uptake was unchanged ex vivo in isolated skeletal muscle. Capillary density was similar but vascular KATP-induced relaxation was impaired in isolated Cyp2c44 -/- vessels (maximal response 39.3 ± 6.5% of control, p < 0.001), suggesting that impaired vascular reactivity produces impaired insulin sensitivity in vivo. Similarly, plasma EETs positively correlated with insulin sensitivity in human participants. CONCLUSIONS/INTERPRETATION: CYP2C-derived EETs contribute to insulin sensitivity in mice and in humans. Interventions to increase circulating EETs in humans could provide a novel approach to improve insulin sensitivity and treat hypertension.
Asunto(s)
Sistema Enzimático del Citocromo P-450/metabolismo , Eicosanoides/metabolismo , Islotes Pancreáticos/metabolismo , Animales , Sistema Enzimático del Citocromo P-450/genética , Familia 2 del Citocromo P450/genética , Familia 2 del Citocromo P450/metabolismo , Resistencia a la Insulina/genética , Resistencia a la Insulina/fisiología , Masculino , Arterias Mesentéricas/metabolismo , RatonesRESUMEN
Hepatic insulin resistance is associated with increased collagen. Integrin α1ß1 is a collagen-binding receptor expressed on hepatocytes. Here, we show that expression of the α1 subunit is increased in hepatocytes isolated from high fat (HF)-fed mice. To determine whether the integrin α1 subunit protects against impairments in hepatic glucose metabolism, we analyzed glucose tolerance and insulin sensitivity in HF-fed integrin α1-null (itga1(-/-)) and wild-type (itga1(+/+)) littermates. Using the insulin clamp, we found that insulin-stimulated hepatic glucose production was suppressed by â¼50% in HF-fed itga1(+/+) mice. In contrast, it was not suppressed in HF-fed itga1(-/-) mice, indicating severe hepatic insulin resistance. This was associated with decreased hepatic insulin signaling in HF-fed itga1(-/-) mice. Interestingly, hepatic triglyceride and diglyceride contents were normalized to chow-fed levels in HF-fed itga1(-/-) mice. This indicates that hepatic steatosis is dissociated from insulin resistance in HF-fed itga1(-/-) mice. The decrease in hepatic lipid accumulation in HF-fed itga1(-/-) mice was associated with altered free fatty acid metabolism. These studies establish a role for integrin signaling in facilitating hepatic insulin action while promoting lipid accumulation in mice challenged with a HF diet.
Asunto(s)
Hígado Graso/metabolismo , Glucosa/metabolismo , Resistencia a la Insulina/genética , Integrina alfa1/biosíntesis , Animales , Dieta Alta en Grasa , Hígado Graso/patología , Hepatocitos/metabolismo , Humanos , Insulina/metabolismo , Integrina alfa1/genética , Hígado/metabolismo , Hígado/patología , Ratones , Ratones Noqueados , Triglicéridos/metabolismoAsunto(s)
American Heart Association/organización & administración , Enfermedades de la Aorta , Investigación Biomédica/organización & administración , Enfermedad Arterial Periférica , Investigadores/organización & administración , Comités Consultivos/organización & administración , American Heart Association/economía , Animales , Enfermedades de la Aorta/diagnóstico , Enfermedades de la Aorta/mortalidad , Enfermedades de la Aorta/fisiopatología , Enfermedades de la Aorta/terapia , Investigación Biomédica/economía , Humanos , Enfermedad Arterial Periférica/diagnóstico , Enfermedad Arterial Periférica/mortalidad , Enfermedad Arterial Periférica/fisiopatología , Enfermedad Arterial Periférica/terapia , Investigadores/economía , Apoyo a la Investigación como Asunto/organización & administración , Estados UnidosRESUMEN
Metabolic stress, as well as several antidiabetic agents, increases hepatic nucleotide monophosphate (NMP) levels, activates AMP-activated protein kinase (AMPK), and suppresses glucose production. We tested the necessity of hepatic AMPK for the in vivo effects of an acute elevation in NMP on metabolism. 5-Aminoimidazole-4-carboxamide 1-ß-D-ribofuranoside (AICAR; 8 mg·kg(-1)·min(-1))-euglycemic clamps were performed to elicit an increase in NMP in wild type (α1α2(lox/lox)) and liver-specific AMPK knock-out mice (α1α2(lox/lox) + Albcre) in the presence of fixed glucose. Glucose kinetics were equivalent in 5-h fasted α1α2(lox/lox) and α1α2(lox/lox) + Albcre mice. AMPK was not required for AICAR-mediated suppression of glucose production and increased glucose disappearance. These results demonstrate that AMPK is unnecessary for normal 5-h fasting glucose kinetics and AICAR-mediated inhibition of glucose production. Moreover, plasma fatty acids and triglycerides also decreased independently of hepatic AMPK during AICAR administration. Although the glucoregulatory effects of AICAR were shown to be independent of AMPK, these studies provide in vivo support for the AMPK energy sensor paradigm. AICAR reduced hepatic energy charge by â¼20% in α1α2(lox/lox), which was exacerbated by â¼2-fold in α1α2(lox/lox) + Albcre. This corresponded to a â¼6-fold rise in AMP/ATP in α1α2(lox/lox) + Albcre. Consistent with the effects on adenine nucleotides, maximal mitochondrial respiration was â¼30% lower in α1α2(lox/lox) + Albcre than α1α2(lox/lox) livers. Mitochondrial oxidative phosphorylation efficiency was reduced by 25%. In summary, these results demonstrate that the NMP capacity to inhibit glucose production in vivo is independent of liver AMPK. In contrast, AMPK promotes mitochondrial function and protects against a more precipitous fall in ATP during AICAR administration.