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BACKGROUND AND AIMS: Metabolic syndrome (MetS) is currently considered to raise the risk for type 2 diabetes and cardiovascular events. It has been suggested that part of this risk excess may be due to a cluster of additional factors associated with MetS. We aimed to investigate the role of inflammation on the ventricular-vascular coupling in patients with MetS. METHODS AND RESULTS: We enrolled a total of 227 hypertensive patients (106 with MetS and 121 without MetS) matched for age and gender. Aortic pulse wave velocity (aPWV), intima-media thickness (IMT) and high sensitivity C-reactive protein (CRP) increased according to the number of MetS components. Patients with MetS showed increased aPWV (11.5 ± 3.7 vs. 10.3 ± 2.5 m/s, P = 0.03) compared with controls. In a model adjusted for age, sex, heart rate and mean blood pressure, aPWV resulted increased in patients with CKD (beta 1.29 m/s, 95%CI 0.61-1.96 m/s, P < 0.001) and MetS (beta 0.89 m/s, 95%CI 0.28-1.51 m/s, P = 0.005). After additional adjustment for CRP and IMT, the slope of aPWV was respectively reduced by 16% and 62%, suggesting that inflammation and intima-media thickening could contribute to aortic stiffening in patients with MetS. In these patients, aPWV was also associated with left-ventricular mass index (beta 0.79 g/m2.7, 95%CI 0.05-1.52 g/m2.7, P = 0.05). CONCLUSION: MetS is characterized by an inflammation-dependent acceleration in cardiovascular ageing. This pattern of pathophysiological abnormalities may contribute to amplify the burden of cardiovascular risk in patients with MetS.
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Hemodinámica , Hipertensión/fisiopatología , Inflamación/fisiopatología , Síndrome Metabólico/fisiopatología , Función Ventricular Izquierda , Biomarcadores/sangre , Proteína C-Reactiva/análisis , Estudios de Casos y Controles , Estudios Transversales , Femenino , Humanos , Hipertensión/sangre , Hipertensión/diagnóstico , Inflamación/sangre , Inflamación/diagnóstico , Mediadores de Inflamación/sangre , Italia , Masculino , Síndrome Metabólico/sangre , Síndrome Metabólico/diagnóstico , Pronóstico , Medición de Riesgo , Factores de Riesgo , Rigidez Vascular , Remodelación VentricularRESUMEN
BACKGROUND AND AIMS: Familial hypercholesterolemia (FH) is underdiagnosed and public cholesterol screening may be useful to find new subjects. In this study, we aim to investigate the prevalence of FH patients in a hospital screening program and evaluate their atherosclerotic burden using intima-media thickness (IMT). METHODS AND RESULTS: We screened 1575 lipid profiles and included for genetic analysis adults with a low-density lipoprotein (LDL) cholesterol >190 mg/dL and triglycerides <200 mg/dL and first-degree child relatives with LDL cholesterol >160 mg/dL and triglycerides <200 mg/dL. The diagnosis of FH was presumed by Dutch Lipid Clinic Network (DLCN) criteria and confirmed by the presence of the genetic variant. Mean common carotid intima-media thickness (IMT) was assessed using consensus criteria. After confirming LDL cholesterol value and excluding secondary hypercholesterolemia, 56 subjects with a DLCN ≥4 performed genetic analysis. Of these, 26 had an FH genetic variant. The proportion of patients with a mutation having a DLCN score of 6-8 was 75%; in individuals with a DLCN score >8 it was 100%. Mean IMT was higher in FH patients compared to non FH (0.73 [0.61-0.83] vs 0.71 [0.60-0.75] mm, p < 0.01). Moreover, we detected two mutations not previously described. Finally, simple regression analysis showed a correlation of IMT with LDL cholesterol >190 mg/dL and corneal arcus (p < 0.01 and p < 0.001, respectively). CONCLUSIONS: A hospital screening was useful to detect FH subjects with increased atherosclerosis. Also, next-generation sequencing was able to detect new FH mutations.
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Enfermedades de las Arterias Carótidas/diagnóstico , Arteria Carótida Común/diagnóstico por imagen , Grosor Intima-Media Carotídeo , Análisis Mutacional de ADN/métodos , Hospitales , Hiperlipoproteinemia Tipo II/diagnóstico , Lípidos/sangre , Tamizaje Masivo/métodos , Mutación , Anciano , Biomarcadores/sangre , Enfermedades de las Arterias Carótidas/sangre , Enfermedades de las Arterias Carótidas/epidemiología , Enfermedades de las Arterias Carótidas/genética , Femenino , Predisposición Genética a la Enfermedad , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Hiperlipoproteinemia Tipo II/sangre , Hiperlipoproteinemia Tipo II/epidemiología , Hiperlipoproteinemia Tipo II/genética , Italia/epidemiología , Masculino , Persona de Mediana Edad , Fenotipo , Placa Aterosclerótica , Valor Predictivo de las Pruebas , Prevalencia , Evaluación de Programas y Proyectos de Salud , Factores de RiesgoRESUMEN
BACKGROUND AND AIMS: Modern diets are high in advanced glycation end-products (dAGEs), derived from processing methods, exerting a pivotal role in promoting atherosclerotic risk. In this cross-sectional study we investigate the relationship between dAGE intake, arterial stiffness, inflammatory profile and macronutrient composition, in subjects with type 2 diabetes without overt cardiovascular disease. METHODS AND RESULTS: Arterial stiffness, carboxy-methyl-lysine, endogenous secretory receptor for AGEs (esRAGE), high sensitivity C reactive protein (hs-CRP), S100A12 and macronutrient intake were evaluated in 85 subjects with type 2 diabetes. The subjects were stratified into two groups according to dAGE consumption: high and low dAGE intake (≥ or <15.000 kU/day, respectively). Subjects with high dAGE intake (n = 45) showed a higher augmentation, augmentation index and pulse wave velocity (PWV) compared with those subjects with low dAGE intake (18 ± 5.4 vs 12.2 ± 6.3 mmHg, P < 0.05; 38.3 ± 5.4 vs 29.3 ± 10%; 9.2 ± 1.4 m/sec vs 7.9 ± 1.7, P < 0.05, respectively). hs-CRP were higher in subjects with high dAGE intake [0.42 (0.18-0.54) vs 0.21 (0.14-0.52) mg/dL, P < 0.05] whereas esRAGE plasma levels were lower [0.16 (0.23-0.81) vs 0.2 (0.14-0.54) ng/dL, P < 0.05]. Simple regression analysis showed a correlation between dAGEs and fat intake. Multivariate analysis showed an independent association between augmentation, systolic blood pressure (BP) and dAGE consumption; BMI and esRAGE were the major determinants of PWV. CONCLUSIONS: Our data suggests that a chronic high dAGE diet could lead to a vascular dysfunction and inflammatory activation, contributing to the development of vascular complications in subjects with type 2 diabetes. Testing this hypothesis may represent a direction of future research.
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Diabetes Mellitus Tipo 2/complicaciones , Angiopatías Diabéticas/etiología , Dieta/efectos adversos , Productos Finales de Glicación Avanzada/efectos adversos , Inflamación/etiología , Rigidez Vascular , Adulto , Anciano , Biomarcadores/sangre , Proteína C-Reactiva/análisis , Grosor Intima-Media Carotídeo , Estudios Transversales , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/diagnóstico , Angiopatías Diabéticas/sangre , Angiopatías Diabéticas/diagnóstico , Angiopatías Diabéticas/fisiopatología , Femenino , Productos Finales de Glicación Avanzada/administración & dosificación , Humanos , Inflamación/sangre , Inflamación/diagnóstico , Masculino , Persona de Mediana Edad , Análisis de la Onda del Pulso , Receptor para Productos Finales de Glicación Avanzada/sangre , Medición de Riesgo , Factores de RiesgoRESUMEN
BACKGROUND & AIMS: Nonalcoholic fatty liver disease (NAFLD) represents the most common chronic liver disease in industrialized countries. NAFLD has the potential to progress through the inflammatory phase of nonalcoholic steatohepatitis (NASH) to fibrosis, cirrhosis, and hepatocellular carcinoma. Identifying patients at risk for this transition is a relevant clinical challenge. The complexity of these phenotypes in vivo made necessary the development of in vitro models in order to dissect the molecular signalling affected in NAFLD and NASH, but also to identify potential circulating biomarkers. METHODS AND RESULTS: We profiled the expression of 754 cellular and medium-secreted human miRNAs in HepG2 cells after lipotoxic (Palmitate, model of NASH) or not-lipotoxic stimuli (Oleate-Palmitate, model of NAFLD). Results were validated through Single TaqMan assays. We performed computational analysis of miRNA targets and pathways. Oleate-palmitate treatment induced a variation of 2.8% and 10% of total miRNAs in cells and medium, respectively; palmitate treatment caused 10% and 19% intracellular and extracellular miRNA deregulation, respectively. We validated miR-126, miR-150, miR-223, miR-483-3p, miR-1226*, and miR-1290 deregulation. Through computational analysis, we observed that targets of both intracellular and extracellular DE miRNAs were involved in processes associated with the onset and progression of NAFLD and NASH, such as fatty acid metabolism, apoptosis and inflammation. CONCLUSIONS: These data would be useful to elucidate the role of miRNAs in the pathogenesis and progression of the NAFLD spectrum, but they also allow the identification of novel potential biomarkers for differential diagnosis to be tested in vivo.
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Hepatocitos/metabolismo , Hígado/metabolismo , MicroARNs/genética , Enfermedad del Hígado Graso no Alcohólico/genética , Antígenos CD36/genética , Antígenos CD36/metabolismo , Supervivencia Celular , Ceramidas/metabolismo , Coenzima A Ligasas/genética , Coenzima A Ligasas/metabolismo , Biología Computacional , Diglicéridos/metabolismo , Perfilación de la Expresión Génica/métodos , Regulación de la Expresión Génica , Redes Reguladoras de Genes , Marcadores Genéticos , Células Hep G2 , Hepatocitos/efectos de los fármacos , Hepatocitos/patología , Humanos , Proteínas Sustrato del Receptor de Insulina/genética , Proteínas Sustrato del Receptor de Insulina/metabolismo , Hígado/efectos de los fármacos , Hígado/patología , MicroARNs/metabolismo , Enfermedad del Hígado Graso no Alcohólico/inducido químicamente , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/patología , Ácido Oléico/toxicidad , Análisis de Secuencia por Matrices de Oligonucleótidos , Ácido Palmítico/toxicidad , Fosforilación , Mapas de Interacción de Proteínas , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal , Factores de TiempoRESUMEN
BACKGROUND AND AIMS: We aimed to investigate lipid abnormalities and liver steatosis in patients with HbA1c-defined prediabetes and type 2 diabetes compared to individuals with HbA1c-defined normoglycaemia. METHODS AND RESULTS: Ninety-one subjects with prediabetes according to HbA1c, i.e. from 5.7 to 6.4% (39-46 mmol/mol), 50 newly diagnosed patients with HbA1c-defined type 2 diabetes (HbA1c ≥6.5% [≥48 mmol/mol]), and 67 controls with HbA1c lower than 5.7% (<39 mmol/mol), were studied. Fasting blood samples for lipid profiles, fatty liver index (FLI), bioimpedance analysis, ultrasound scan of the liver, and BARD (body mass index, aspartate aminotransferase/alanine aminotransferase ratio, diabetes) score for evaluation of liver fibrosis, were performed in all subjects. In comparison to controls, subjects with prediabetes were characterised by: lower apolipoprotein AI and HDL cholesterol levels, higher blood pressure, triglycerides levels and apolipoprotein B/apolipoprotein AI ratio, higher FLI, increased prevalence of and more severe hepatic steatosis, similar BARD score, and higher total body fat mass. In comparison to subjects with diabetes, subjects with prediabetes exhibited: similar blood pressure and apolipoprotein B/apolipoprotein AI ratio, similar FLI, reduced prevalence of and less severe hepatic steatosis, lower BARD score, increased percent fat and lower total body muscle mass. In comparison to controls, subjects with diabetes showed: lower apolipoprotein AI and HDL cholesterol levels, higher blood pressure and triglycerides levels, higher FLI, increased prevalence of and more severe hepatic steatosis, higher BARD score, and higher total body muscle mass. Moreover, HbA1c was correlated with BMI, HOMA-IR, triglycerides, HDL cholesterol, AST, and ALT. CONCLUSIONS: Subjects with HbA1c-defined prediabetes and type 2 diabetes, respectively, are characterised by abnormalities in lipid profile and liver steatosis, thus exhibiting a severe risk profile for cardiovascular and liver diseases.
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Diabetes Mellitus Tipo 2/complicaciones , Dislipidemias/epidemiología , Enfermedad del Hígado Graso no Alcohólico/epidemiología , Estado Prediabético/complicaciones , Adulto , Apolipoproteína A-I/sangre , Composición Corporal , HDL-Colesterol/sangre , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/diagnóstico , Diabetes Mellitus Tipo 2/metabolismo , Angiopatías Diabéticas/epidemiología , Dislipidemias/complicaciones , Femenino , Hemoglobina Glucada/análisis , Humanos , Hipertensión/complicaciones , Hipertensión/epidemiología , Resistencia a la Insulina , Italia/epidemiología , Lípidos/sangre , Hígado/diagnóstico por imagen , Masculino , Persona de Mediana Edad , Enfermedad del Hígado Graso no Alcohólico/complicaciones , Enfermedad del Hígado Graso no Alcohólico/diagnóstico por imagen , Enfermedad del Hígado Graso no Alcohólico/fisiopatología , Estado Prediabético/sangre , Estado Prediabético/diagnóstico , Estado Prediabético/metabolismo , Prevalencia , Factores de Riesgo , Índice de Severidad de la Enfermedad , UltrasonografíaRESUMEN
OBJECTIVE: We investigated the separate impact of metabolic syndrome (MS) and altered glucose tolerance on early markers of vascular injuries. METHODS: Intima-media thickness (IMT) and pulse wave analysis (PWA), were evaluated in 132 overweight or obese subjects, with (MS(+)) or without (MS(-)) MS; subjects were further classified as normotolerant (NT) or with altered glucose tolerance (AGT) according to a 2 h oral glucose tolerance test (OGTT). RESULTS: In MS(+) patients, IMT was higher than in the MS(-) group, and PWA revealed higher augmentation pressure (Aug, the contribution that wave reflection makes to systolic arterial pressure) and lower subendocardial viability ratio (SEVR, an estimate of myocardial perfusion). When analyzed according to glucose tolerance, IMT was higher in MS(+)NT subjects and AGT patients with and without MS, vs. MS(-)NT subjects. Logistic regression modeling showed that both AGT and MS were independently associated with increased IMT. However, only MS remained associated with IMT after adjustment for age. SEVR was reduced only in MS(+) patients, independently of glucose tolerance. In both groups, Aug and AugI were higher in the AGT group, but the correlation with 2 h-plasma glucose disappeared when corrected for age. CONCLUSION: Both MS and AGT altered IMT, but the effect of AGT disappears when age is added to the multiple regression model. In contrast, arterial stiffness was affected differently in the two categories: in subjects with MS, the subendocardial viability ratio (an estimate of myocardial perfusion) was impaired, while in subjects with AGT, both Aug and AugI were increased. These data suggest that applying the definition of MS might help to better characterize cardiovascular risk in subjects with altered glucose tolerance or obesity.
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Enfermedades de las Arterias Carótidas/epidemiología , Trastornos del Metabolismo de la Glucosa/epidemiología , Síndrome Metabólico/epidemiología , Adulto , Factores de Edad , Biomarcadores/sangre , Arterias Carótidas/diagnóstico por imagen , Arterias Carótidas/fisiopatología , Enfermedades de las Arterias Carótidas/sangre , Enfermedades de las Arterias Carótidas/diagnóstico , Enfermedades de las Arterias Carótidas/fisiopatología , Grosor Intima-Media Carotídeo , Distribución de Chi-Cuadrado , Diagnóstico Precoz , Femenino , Trastornos del Metabolismo de la Glucosa/sangre , Trastornos del Metabolismo de la Glucosa/diagnóstico , Prueba de Tolerancia a la Glucosa , Hemodinámica , Humanos , Italia/epidemiología , Modelos Logísticos , Masculino , Síndrome Metabólico/sangre , Síndrome Metabólico/diagnóstico , Persona de Mediana Edad , Análisis Multivariante , Obesidad/epidemiología , Oportunidad Relativa , Valor Predictivo de las Pruebas , Análisis de la Onda del Pulso , Medición de Riesgo , Factores de Riesgo , Rigidez VascularRESUMEN
BACKGROUND: In rat pancreatic islets, chronic exposure to high free fatty acid (FFA) levels impairs insulin secretion and ß cell mass. The mechanisms underlying this defect are not completely understood. Since islets have intrinsically low anti-oxidant enzyme defense, oxidative stress might be responsible for ß cell damage. AIM: In this study, we investigated if FFA could induce oxidative stress in rat pancreatic islets and if metformin might reverse adverse effects. MATERIAL AND METHODS: We cultured rat pancreatic islets in the presence or absence of FFA (oleate/palmitate 2:1, 2 mM) for 72 h. In some experiments, we used metformin (2.5 µg/ml) during the last 24 h. RESULTS: In our model, glucosestimu lated insulin release was markedly reduced (p<0.005) after chronic FFA exposure, and the ATP/ADP ratio was altered (p<0.05). We observed a significant increase of reactive oxygen species (ROS) (p<0.001), malondialdehyde a lipid peroxidation product (p<0.01) and nitric oxide (NO) levels in the culture media (p<0.001). Inducible NO synthase (iNOS) and heat shock protein-70 (HSP-70) protein expression were also increased (p<0.001 and p<0.01, respectively). When metformin was present during the last 24 h of culture, insulin secretion was restored, and the ATP/ADP ratio was normalized. ROS production, NO production, lipid peroxidation, iNOS and HSP-70 protein expression levels had decreased. CONCLUSIONS: These data indicate that, in rat pancreatic islets, chronic exposure to high FFA induces oxidative stress and that metformin, by reducing this effect, may have a direct beneficial effect on insulin secretion impaired by lipotoxicity.
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Adenosina Difosfato/metabolismo , Adenosina Trifosfato/metabolismo , Ácidos Grasos no Esterificados/efectos adversos , Glucosa/farmacología , Insulina/metabolismo , Islotes Pancreáticos/efectos de los fármacos , Metformina/farmacología , Estrés Oxidativo/efectos de los fármacos , Animales , Apoptosis/efectos de los fármacos , Western Blotting , Supervivencia Celular/efectos de los fármacos , Proteínas HSP70 de Choque Térmico/metabolismo , Hipoglucemiantes/farmacología , Secreción de Insulina , Islotes Pancreáticos/metabolismo , Masculino , Malondialdehído/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Nitritos/metabolismo , Ratas , Ratas WistarRESUMEN
BACKGROUND AND AIMS: Metabolic syndrome (MS) has been mainly related to insulin resistance, but the role of changes in insulin secretion has not been thoroughly investigated. METHODS AND RESULTS: Using an oral glucose tolerance test (OGTT) we studied beta-cell function and insulin sensitivity in subjects with normal fasting glucose with and without MS, and their relationship to fatty liver which was evaluated by abdominal-ultrasonography. In MS early phase insulin secretion, as measured by insulinogenic index (IG(30)), was increased (p<0.05) independently from insulin sensitivity. Furthermore IG(30) was progressively higher as the number of factors needed for the diagnosis of MS increased (p<0.01). Insulin and C-peptide AUC were also increased (p<0.01 and p<0.05, respectively) but, in contrast to IG(30), these differences disappeared when ISI was used as a covariate. After OGTT, 51% of the subjects with MS had altered post-load glucose tolerance compared to 24.9% without MS (p<0.01). In both groups, the altered glucose tolerance was associated with a similar IG(30) reduction. In normo-tolerant subjects with MS the IG(30) was higher (+54.1%, p<0.01), and this elevation occurred irrespective of ISI; however, the beta-cell compensatory capacity for insulin resistance (disposition index) was impaired (p<0.001). Fatty liver was more frequent (p<0.001) and more severe (p<0.01) in MS, and it was significantly related to total AUC-insulin (p<0.001), independently from ISI. CONCLUSION: These findings indicate that the prevalence of altered tolerance is more frequent in subjects with normal fasting glucose and MS. The hyperinsulinemia might not only be an adaptive response to insulin resistance, but a primary defect of beta-cell function contributing to glucose intolerance.
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Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Síndrome Metabólico/metabolismo , Síndrome Metabólico/fisiopatología , Estado Prediabético/metabolismo , Estado Prediabético/fisiopatología , Adulto , Algoritmos , Glucemia/análisis , Proteína C-Reactiva/análisis , Diabetes Mellitus Tipo 2/epidemiología , Diabetes Mellitus Tipo 2/etiología , Hígado Graso/diagnóstico por imagen , Hígado Graso/epidemiología , Hígado Graso/etiología , Femenino , Intolerancia a la Glucosa/epidemiología , Intolerancia a la Glucosa/etiología , Humanos , Hiperinsulinismo/epidemiología , Hiperinsulinismo/etiología , Insulina/sangre , Resistencia a la Insulina , Secreción de Insulina , Cinética , Masculino , Síndrome Metabólico/sangre , Persona de Mediana Edad , Estado Prediabético/sangre , Prevalencia , Índice de Severidad de la Enfermedad , UltrasonografíaRESUMEN
Glycemic control in elderly persons with type 2 diabetes mellitus (T2DM) is challenging because they are more likely to have other age-associated medical conditions and to experience hypoglycemia during intensive therapy. A best therapeutic strategy for these patients has not yet been defined. We investigated the efficacy and safety of adding once-daily insulin glargine to patients' current oral antidiabetic drugs (OAD) regimen, compared to increasing the OAD doses. The study enrolled patients aged 65 years or more, with poor glycemic control. Patients were randomized to two groups and entered a 3-week titration period in which their actual therapy was adjusted to meet the study's glycemic goals, by either adding insulin glargine to current therapy (group A, 27 patients) or increasing current OAD dosages (group B, 28 patients). Thereafter, therapies were continued unchanged for a 24-week observation period. The mean therapeutic dosage of insulin glargine in group A was 14.9 IU/day (SD = 5.0 IU/day). During the observation period, mean levels of glycosylated hemoglobin (HbA1c) reduced by 1.5% in group A and 0.6% in group B (P = 0.381). An HbA1c level <7.0% was achieved by five patients in each group. Mean fasting blood glucose levels reduced by 29 and 15% in groups A and B, respectively (P = 0.029). Group A had fewer total hypoglycemic events (23 vs. 79, P = 0.030) and fewer patients experiencing any such event (9 vs. 17, P = 0.045). Neither a serious hypoglycemic event nor other adverse event occurred. These results suggest that, compared to increasing OAD dosage, the addition of insulin glargine to current OAD therapy is as effective but safer in terms of the risk for hypoglycemia in elderly patients with T2DM.
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Glucemia/metabolismo , Hipoglucemiantes/administración & dosificación , Hipoglucemiantes/uso terapéutico , Insulina/análogos & derivados , Administración Oral , Anciano , Índice de Masa Corporal , Carbamatos/uso terapéutico , Quimioterapia Combinada , Femenino , Gliclazida/uso terapéutico , Hemoglobina Glucada/metabolismo , Humanos , Insulina/uso terapéutico , Insulina Glargina , Insulina de Acción Prolongada , Masculino , Metformina/uso terapéutico , Pioglitazona , Piperidinas/uso terapéutico , Proyectos de Investigación , Rosiglitazona , Tiazolidinedionas/uso terapéuticoRESUMEN
BACKGROUND: Long-chain polyunsaturated fatty acid omega-3 levels are decreased in the hepatic tissue of patients with nonalcoholic fatty liver disease. Polyunsaturated fatty acids are negative regulators of hepatic lipogenesis and attenuate the inflammatory response in mice. AIM: To investigate whether polyunsaturated fatty acid may be effective in the treatment of nonalcoholic fatty liver disease. METHODS: Forty patients with nonalcoholic fatty liver disease were randomized into two groups for treatment of 6 months duration. Group DP (n=20) received an AHA recommended diet and polyunsaturated fatty acid 2g/day; Group D (n=20) received only the AHA regular diet. Outcome measurements were fatty liver assessed by abdominal ultrasound, liver aminotransferase and tumour necrosis factor-alpha serum levels, and insulin resistance assessed by HOMA(IR). RESULTS: After 6 months of treatment, the DP group displayed a decrease in alanine aminotransferase levels (p<0.01), as well as in triglyceride levels (p<0.01), serum tumour necrosis factor-alpha levels (p<0.05) and in HOMA(IR) (p<0.05). In the D group, no significant modification was observed. In the DP group, complete fatty liver regression was observed in 33.4% of the patients, and an overall reduction in 50%. In contrast, no patient achieved complete regression in the D group, whereas some amount of reduction occurred in 27.7% of the patients; the remaining 72.2% did not change. CONCLUSION: Our results indicate that alanine aminotransferase, triglyceride and serum tumour necrosis factor-alpha levels, as well as fatty liver improved after polyunsaturated fatty acid administration.
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Ácidos Grasos Omega-3/uso terapéutico , Hígado Graso/dietoterapia , Hígado Graso/diagnóstico por imagen , Hígado Graso/metabolismo , Femenino , Estudios de Seguimiento , Humanos , Resistencia a la Insulina/fisiología , Lipogénesis/efectos de los fármacos , Hígado/diagnóstico por imagen , Hígado/metabolismo , Masculino , Persona de Mediana Edad , Transaminasas/metabolismo , Resultado del Tratamiento , Factor de Necrosis Tumoral alfa/sangre , UltrasonografíaRESUMEN
BACKGROUNDS AND AIMS: Non-alcoholic-steatohepatitis (NASH) is closely related to insulin resistance, but it is unknown whether insulin resistance may be localized in hepatocytes. This study investigates insulin signalling in liver tissue from NASH, and the molecular mechanisms by which insulin-resistance could lead to liver damage (apoptosis). Moreover, to investigate the mechanisms of lipid overload we studied key enzymes in hepatocytes lipid metabolism. METHODS AND RESULTS: In liver specimens from 11 patients with NASH and 7 histological normal livers, we measured total and phosphorylated Akt (active form), Bax and Bcl-2 by Western-blot analysis. In addition, we studied AMP-activated protein Kinase and Carnitine-Palmitoyl-Transferase-1 gene expression, key regulators of non-esterified fatty acid synthesis and oxidation, by reverse transcription polymerase chain reaction. In NASH, phosphorylated Akt was impaired (104.3+/-10.6 vs 152.6+/-22.4 AU, p<0.002) and correlated with necroinflammatory score (r=-0.62; p<0.05). Bax/Bcl-2 ratio was increased in NASH. Moreover, we observed a decrease of AMP-activated protein Kinase (10.74+/-6 vs 144.7+/-41.6 AU, p<0.0001) and Carnitine-Palmitoyl-Transferase-1 gene expression (38.7+/-14.6 vs 192.1+/-26.2 AU, p<0.0001), and both were correlated with steatosis score (r=-0.56, p<0.05, r=-0.87, p<0.05 respectively). CONCLUSIONS: Akt, a key molecule of insulin signalling and cell apoptosis is impaired in NASH, suggesting an important role of hepatic insulin resistance in liver failure. Moreover, decreased non-esterified fatty acid oxidation may cause hepatic lipid overload.
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Apoptosis/genética , Hígado Graso/genética , Resistencia a la Insulina/genética , Lípidos/fisiología , Adulto , Glucemia/metabolismo , Índice de Masa Corporal , Carnitina O-Palmitoiltransferasa/genética , Hígado Graso/patología , Femenino , Humanos , Inflamación/genética , Inflamación/patología , Hígado/fisiología , Masculino , Persona de Mediana Edad , Valores de ReferenciaRESUMEN
AIMS/HYPOTHESIS: Little information is available on the insulin release properties of pancreatic islets isolated from type 2 diabetic subjects. Since mitochondria represent the site where important metabolites that regulate insulin secretion are generated, we studied insulin release as well as mitochondrial function and morphology directly in pancreatic islets isolated from type 2 diabetic patients. METHODS: Islets were prepared by collagenase digestion and density gradient purification, and insulin secretion in response to glucose and arginine was assessed by the batch incubation method. Adenine nucleotides, mitochondrial membrane potential, the expression of UCP-2, complex I and complex V of the respiratory chain, and nitrotyrosine levels were evaluated and correlated with insulin secretion. RESULTS: Compared to control islets, diabetic islets showed reduced insulin secretion in response to glucose, and this defect was associated with lower ATP levels, a lower ATP/ADP ratio and impaired hyperpolarization of the mitochondrial membrane. Increased protein expression of UCP-2, complex I and complex V of the respiratory chain, and a higher level of nitrotyrosine were also found in type 2 diabetic islets. Morphology studies showed that control and diabetic beta cells had a similar number of mitochondria; however, mitochondrial density volume was significantly higher in type 2 diabetic beta cells. CONCLUSIONS/INTERPRETATION: In pancreatic beta cells from type 2 diabetic subjects, the impaired secretory response to glucose is associated with a marked alteration of mitochondrial function and morphology. In particular, UCP-2 expression is increased (probably due to a condition of fuel overload), which leads to lower ATP, decreased ATP/ADP ratio, with consequent reduction of insulin release.
Asunto(s)
Diabetes Mellitus Tipo 2/patología , Islotes Pancreáticos/patología , Mitocondrias/patología , Nucleótidos de Adenina/metabolismo , Arginina/farmacología , Separación Celular , Glucosa/farmacología , Humanos , Insulina/metabolismo , Secreción de Insulina , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/metabolismo , Valores de ReferenciaRESUMEN
Exposure of rat pancreatic islets to 20 mM leucine for 24 h reduced insulin release in response to glucose (16.7 and 22.2 mM). Insulin release was normal when the same islets were stimulated with leucine (40 mM) or glyburide (1 microM). To investigate the mechanisms responsible for the different effect of these secretagogues, we studied several steps of glucose-induced insulin secretion. Glucose utilization and oxidation rates in leucine-precultured islets were similar to those of control islets. Also, the ATP-sensitive K(+) channel-independent pathway of glucose-stimulated insulin release, studied in the presence of 30 mM K(+) and 250 microM diazoxide, was normal. In contrast, the ATP-to-ADP ratio after stimulation with 22.2 mM glucose was reduced in leucine-exposed islets with respect to control islets. The decrease of the ATP-to-ADP ratio was due to an increase of ADP levels. In conclusion, prolonged exposure of pancreatic islets to high leucine levels selectively impairs glucose-induced insulin release. This secretory abnormality is associated with (and might be due to) a reduced ATP-to-ADP ratio. The abnormal plasma amino acid levels often present in obesity and diabetes may, therefore, affect pancreatic islet insulin secretion in these patients.
Asunto(s)
Adenosina Difosfato/análisis , Adenosina Trifosfato/análisis , Glucosa/farmacología , Insulina/metabolismo , Islotes Pancreáticos/efectos de los fármacos , Leucina/administración & dosificación , Adenosina Trifosfato/farmacología , Animales , Células Cultivadas , Glucosa/metabolismo , Secreción de Insulina , Islotes Pancreáticos/química , Islotes Pancreáticos/metabolismo , Masculino , Oxidación-Reducción , Canales de Potasio/fisiología , Ratas , Ratas WistarRESUMEN
Because metformin affects glucose and free fatty acid (FFA) metabolism in peripheral insulin target tissues, we investigated the effect of this drug in restoring a normal secretory pattern in rat pancreatic islets whose function has been impaired by chronic exposure to elevated FFA or glucose concentrations. We cultured rat pancreatic islets with or without FFA (2 mmol/l oleate/palmitate 2:1) or high glucose (16.7 mmol/l) concentrations in the presence or absence of metformin (0.25-12.5 microg/ml) and then measured insulin release, glucose utilization, glucose, and FFA oxidation. When compared with control islets, islets exposed to high FFA or glucose concentrations showed an increased basal and a decreased glucose-induced insulin release. In islets cultured for an additional 24 h with FFA or glucose in the presence of metformin (2.5 microg/ml), both basal and glucose-induced insulin secretions were restored. Both glucose utilization and glucose oxidation were altered in islets pre-exposed to high FFA or glucose concentrations. In particular, regarding control islets, glucose utilization was increased at 2.8 mmol/l glucose and decreased at 16.7 mmol/l glucose; glucose oxidation was similar to control islets at 2.8 mmol/l glucose but decreased at 16.7 mmol/l glucose. In contrast, oleate oxidation was increased in islets pre-exposed to FFA. All of these abnormalities were reversed in islets cultured for an additional 24 h with high FFA or glucose concentrations in the presence of metformin (2.5 microg/ml). In conclusion, our data show that metformin is able to restore the intracellular abnormalities of glucose and FFA metabolism and to restore a normal secretory pattern in rat pancreatic islets whose secretory function has been impaired by chronic exposure to elevated FFA or glucose levels. These data raise the possibility that, in diabetic patients, metformin (in addition to its peripheral effects) may have a direct beneficial effect on the beta-cell secretory function.
Asunto(s)
Ácidos Grasos no Esterificados/farmacología , Glucosa/farmacología , Hipoglucemiantes/farmacología , Insulina/metabolismo , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/metabolismo , Metformina/farmacología , Animales , Relación Dosis-Respuesta a Droga , Ácidos Grasos no Esterificados/metabolismo , Glucosa/metabolismo , Técnicas In Vitro , Secreción de Insulina , Masculino , Oxidación-Reducción , Ratas , Ratas Wistar , Factores de Tiempo , Triglicéridos/metabolismoRESUMEN
Several data suggest that, during the natural history of Type 2 diabetes (T2DM), whereas the degree of insulin resistance is quite stable in a single individual, insulin secretion is often progressively decreasing. The decline in beta-cell function seems, at least in part, acquired. This review focuses on data showing that chronically elevated levels of glucose and fatty acids may impair beta-cell function and, at a later stage, also affect beta-cell survival. Results obtained both in vitro and in vivo demonstrate that beta-cell function is impaired after chronic exposure to glucose and fatty acids, with many similarities between the two conditions. Basal insulin release is increased as a result of an increase of the low-Km component of glucose phosphorylation. Glucose-induced insulin release is blunted, probably due to an impairment of glucose oxidation. If the metabolic abnormality persists in vivo or the culture is prolonged in vitro, a significant increase in the rate of beta-cell death was observed, eventually leading to severe destruction of the islet architecture. An attractive hypothesis, currently under investigation, is that genetic susceptibility to diabetes may confer an increased sensitivity to the adverse effect of high glucose and non-esterified fatty acid levels on beta-cell survival.
Asunto(s)
Diabetes Mellitus Tipo 2/fisiopatología , Ácidos Grasos no Esterificados/fisiología , Hiperglucemia/fisiopatología , Insulina/metabolismo , Islotes Pancreáticos/fisiología , Animales , Humanos , Resistencia a la Insulina , Secreción de Insulina , Islotes Pancreáticos/metabolismo , Islotes Pancreáticos/patología , RatasRESUMEN
An increased sensitivity to glucose was observed in islets pre-exposed for 1 h to glibenclamide (0.1 micromol 1(-1)), but not to tolbutamide (100 micromol l(-1)), as indicated by a shift to the left of the dose-response curve (EC(50) at 5.8+/-0.3 mmol l(-1) glucose vs 10.6+/-0.8 in control islets; n=11, P<0.005). According to this secretory pattern also glucose utilization at 2.5 and 5.0 mmol l(-1) glucose was higher in islets exposed to glibenclamide. Since binding to mitochondria results in an increased enzyme activity, we measured hexokinase (HK) and glucokinase (GK) activity both in a cytosolic and in a mitochondrion-enriched fractions. Cytosolic hexokinase activity was similar in islets exposed to glibenclamide and in control islets but mitochondrial hexokinase activity was significantly increased after exposure to glibenclamide (124+/-7 vs 51+/-9 nmol microgram prot(-1) 90 min(-1), P<0.01), with no change in the enzyme protein content. In contrast, glucokinase activity in the two groups of islets was similar. When in islets < exposed to glibenclamide hexokinase binding to mitochondria was inhibited by the addition of 20 nmol l(-1) dicyclohexylcarbodiimide (DCC), no increase of glucose sensitivity was observed (EC(50) 10.9+/-1.3 mmol l(-1) glucose, n=3, similar to that of control islets). These data indicate that a 1 h exposure to glibenclamide causes the beta cell to become more sensitive to glucose. This increased sensitivity is associated with (and may be due to) an increased hexokinase activity, in particular the mitochondrial-bound, more active, form. This mechanism may contribute to the hypoglycemic action of this drug.
Asunto(s)
Glucosa/farmacología , Gliburida/farmacología , Hipoglucemiantes/farmacología , Islotes Pancreáticos/efectos de los fármacos , Animales , Separación Celular , Células Cultivadas , Diciclohexilcarbodiimida/farmacología , Glucosa/metabolismo , Hexoquinasa/metabolismo , Insulina/metabolismo , Secreción de Insulina , Islotes Pancreáticos/metabolismo , Cetoácidos/farmacología , Masculino , Fosforilación , Ratas , Ratas Wistar , Fracciones Subcelulares/efectos de los fármacos , Fracciones Subcelulares/metabolismoRESUMEN
When rat pancreatic islets are incubated in 5.5 or 16.7 mmol/l glucose for 3 h, an increased sensitivity is observed in islets pre-exposed to high glucose, as indicated by a shift to the left of the glucose dose-response curve (EC50 7.1 +/- 0.9 and 11.5 +/- 1.2 in high- and low-glucose-exposed islets, respectively; n = 5, P < 0.05). To investigate the mechanism(s) responsible for this effect, we measured hexokinase and glucokinase activity both in the cytosolic fraction and in a mitochondrion-enriched fraction, since binding to the outer mitochondrial membrane has been reported to result in an increased enzyme activity. In islets cultured at 16.7 mmol/l glucose, the cytosolic hexokinase activity was similar to control islets, but mitochondrial enzyme activity was significantly increased (124 +/- 7 vs. 51 +/- 9 nmol x microg(-1) x 90 min(-1), P < 0.01). As a consequence, the cytosolic:mitochondrial fraction ratio was altered in comparison with control islets. In contrast, glucokinase activity in the two groups of islets was similar in the cytosolic fraction and undetectable in the mitochondrial fraction. Hexokinase I quantitation by Western blot confirmed the enzyme translocation from the free cytosolic to the mitochondria-bound form in islets cultured at 16.7 mmol/l glucose. Glucose-induced alterations were reversible after 1 h exposure to 5.5 mmol/l glucose. Moreover, in islets exposed to 16.7 mmol/l glucose, inhibition of hexokinase binding to mitochondria by the addition of 20 nmol/l dicyclohexylcarbodiimide resulted in no increase of glucose sensitivity (EC50 10.9 +/- 0.4, n = 3, similar to that of control islets). These data indicate that after chronic exposure to high glucose, the beta-cell becomes more sensitive to glucose before eventually getting desensitized. This increased sensitivity is associated with (and may be due to) an increased hexokinase activity secondary to a subcellular shift of the enzyme from the free cytosolic to the mitochondria-bound, more active form.
Asunto(s)
Glucosa/farmacología , Hexoquinasa/metabolismo , Insulina/metabolismo , Islotes Pancreáticos/metabolismo , Mitocondrias/enzimología , Animales , Western Blotting , Células Cultivadas , Citosol/enzimología , Citosol/metabolismo , Relación Dosis-Respuesta a Droga , Electroforesis en Gel de Poliacrilamida , Glucoquinasa/metabolismo , Glucosa/metabolismo , Hexoquinasa/análisis , Hexoquinasa/inmunología , Secreción de Insulina , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/enzimología , Masculino , Mitocondrias/metabolismo , Fosforilación , Ratas , Ratas WistarRESUMEN
The prolonged exposure of pancreatic islets and isolated beta cells to elevated glucose concentrations induces a state of unresponsiveness to glucose (desensitization). However, an increased sensitivity to glucose (detected by a shift to the left of the dose-response curve of glucose-induced insulin release) has been also reported after chronic exposure to glucose, making the overall response less comprehensible. In vitro models have many theoretical and practical advantages in better understanding the effects of the prolonged glucose stimulation; moreover, they are also suitable for studying the mechanisms responsible of the observed alterations. We have performed a time-course study of the effect of the exposure to glucose at high concentration on the secretory behaviour of beta cells. Rat pancreatic islets exposed for 30 min to high glucose (300 mg/dl) showed increased basal insulin secretion (175 +/- 29 vs 44 +/- 8 pg/islet (per 30 min; n = 5, P < 0.002) was the only difference from control islets (exposed to 100 mg/dl). After 3 h exposure to high glucose, also increased sensitivity to glucose was observed, as indicated by a shift to the left of the glucose dose-response curve (EC50 123 +/- 10 and 177 +/- 11 mg/dl, respectively; n = 5, P < 0.05). After 6 h exposure to high glucose, besides the two alterations already described, also a decrease in glucose-induced insulin release was observed (688 +/- 104 vs 1184 +/- 34 pg/islet per 30 min; n = 5, P < 0.01). We studied the mechanism responsible for these alterations and we found that the "supersensitivity" to glucose may be related to alterations in the "glucose-sensing" mechanism of beta cells, in particular in glucose phosphorylation. In contrast, in islets desensitized to glucose our data suggest that ion flux and consequent membrane potential changes play a key role in determining the secretory defect. Since a normal response to glyburide was observed, a proximal signal defect for closure of potassium channels is more likely than an intrinsic defect in the channel. In conclusion, our data show what the prolonged stimulation of beta cells with glucose at high concentration induces a series of distinct secretory abnormalities, with a pattern of response that leads first to increased sensitivity and then to decreased responsiveness to glucose.
Asunto(s)
Diabetes Mellitus Experimental/fisiopatología , Glucosa/farmacología , Insulina/metabolismo , Islotes Pancreáticos/fisiología , Animales , Técnicas de Cultivo , Relación Dosis-Respuesta a Droga , Secreción de Insulina , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/metabolismo , Masculino , Ratas , Ratas WistarRESUMEN
The present study was done to achieve a better understanding of the role of ionic flux alterations in glucose-induced desensitization of pancreatic beta-cells. Moreover, we investigated the reversibility of glucose-induced desensitization after different times of exposure to high glucose to ascertain the time necessary for desensitization reversal and to determine whether it depends on the length of high glucose exposure. Glucose desensitization was obtained by incubating rat pancreatic islets for 6 h in CMRL medium containing 16.7 mmol/l glucose. At the end of this period, insulin release, 86Rb efflux, and 45Ca uptake were measured in parallel experiments. In islets cultured at 16.7 mmol/l glucose, maximal glucose-induced insulin release was reduced (848 +/- 97 pg x islet-1 x 30 min-1) in comparison to islets incubated at 5.5 mmol/l glucose (1,436 +/- 144, n = 7, P < 0.01). In contrast, insulin content was similar in the two groups, being 41.0 +/- 2.7 and 47.8 +/- 2.2 ng/islet in islets exposed to 16.7 or 5.5 mmol/l glucose, respectively (P = 0.167). The effect of glucose on both 86Rb efflux and 45Ca uptake was also significantly reduced in 16.7 mmol/l glucose-cultured islets. 86Rb efflux was inhibited only 19 +/- 4% in islets cultured at high glucose with respect to 56 +/- 7% in control islets (n = 5, P < 0.001). 45Ca uptake was 10.5 +/- 1.7 pmol/islet (mean +/- SE, n = 9) in islets cultured at high glucose with respect to 19.7 +/- 2.4 pmol/islet in control islets (P < 0.001). In contrast, the effect of glyburide (10 micromol/l) on insulin release, 86Rb efflux, and 45Ca uptake was similar in islets exposed to 5.5 or 16.7 mmol/l glucose. All the abnormalities observed in islets cultured at 16.7 mmol/l glucose were promptly and simultaneously reversible after islets were transferred in culture medium at 5.5 mmol/l glucose; both insulin secretion and glucose effects on 86Rb efflux and 45Ca uptake returned to values similar to control islets within 5 min. Also, islets exposed to high glucose for a longer period (24 h) recovered from both secretory and ionic abnormalities after 5 min of incubation in CMRL medium at 5.5 mmol/l glucose. Reversal from glucose desensitization was slower (45 - 60 min) when islets were incubated at 5.5 mmol/l glucose in Krebs-Ringer HEPES buffer instead of CMRL medium. The present data suggest that ion flux and consequent membrane-potential changes play a key role in the mechanism leading to glucose-induced desensitization of pancreatic beta-cells. Because a normal response to glyburide was observed in islets exposed to high glucose, a proximal signal defect for closure of K+ channels rather than an intrinsic defect in the channel is likely.
Asunto(s)
Calcio/metabolismo , Glucosa/farmacología , Insulina/metabolismo , Islotes Pancreáticos/fisiología , Rubidio/metabolismo , Animales , Transporte Biológico/efectos de los fármacos , Células Cultivadas , Medios de Cultivo , Relación Dosis-Respuesta a Droga , Secreción de Insulina , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/metabolismo , Cinética , Masculino , Ratas , Ratas Wistar , Factores de TiempoRESUMEN
In vitro exposure of rat pancreatic beta cells to interleukin-1 beta (IL-1 beta) inhibits glucose-stimulated insulin release (2140 +/- 239 and 323 +/- 80 pg.islet-1.h-1 at glucose levels of 16.7 mmol/l in control and IL-1 beta-exposed islets, respectively, n = 7, p < 0.001). Cholera toxin (2 micrograms/ml) or pertussis toxin (0.5 microgram/ml) potentiated, as expected, glucose-induced insulin release in control islets, but, in addition, when added together with IL-1 beta, were able to prevent the IL-1 beta mediated inhibition of glucose-stimulated insulin secretion (2087 +/- 301 and 1662 +/- 173 pg.islet-1.h-1, respectively, p < 0.05 vs islets exposed to IL-1 beta alone). To investigate the mechanism by which the toxins prevent the IL-1 beta effect, we then measured nitrite levels, glucose oxidation and Ca2+ uptake. Nitrite levels in the culture medium were 4.2 +/- 1.4 and 24.0 +/- 5 pmol.islet-1.24 h-1 in control islets and in IL-1 beta-exposed islets, respectively (n = 6, p = 0.05). In islets exposed to IL-1 beta and cholera or pertussis toxins, nitrite levels were 9.1 +/- 3 and 12.4 +/- 6 pmol.islet-1.24 h-1, respectively (n = 6, NS vs control islets). Glucose oxidation at 16.7 mmol/l glucose was 31.1 +/- 2.9 pmol.islet-1.120 min-1 in control islets and 16.8 +/- 2.7 pmol.islet-1.120 min-1 in IL-1 beta-treated islets (p < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)