Cristina Hernández

Metabolic Fingerprints of Proliferative Diabetic Retinopathy: An 1H-NMR–Based Metabonomic Approach Using Vitreous Humor

PURPOSEnTo explore the metabolic profile of vitreous fluid of patients with proliferative diabetic retinopathy (PDR) using 1H-NMR-based metabonomic analysis.nnnMETHODSn1H-NMR spectra were acquired from vitreous samples obtained during vitrectomy from 22 patients with type 1 diabetes with PDR and from 22 nondiabetic patients with macular hole (control group). Data analysis included a principal component analysis and partial least squares discriminant analysis (PLS-DA). In addition, 1H-(1)H and 1H-(13)C HMQC correlation spectra were acquired for the identification of metabolites. Furthermore, the main metabolites accounting for the differences in metabolic profile were assessed by current biochemical methods.nnnRESULTSnLactate was the most abundant metabolite, and it was present at higher levels in samples from PDR patients than from nondiabetic patients (P=0.02). Glucose was significantly higher in samples from PDR patients than nondiabetic patients (P=0.03). After removing the lactate peak at 1.35 ppm and with the use of PLS-DA, a model was obtained that was able to correctly classify 19 of 22 patients with PDR and 18 of 22 controls, resulting in a sensitivity of 86% and a specificity of 81%. The main metabolites involved in this specific pattern recognition were galactitol and ascorbic acid (AA); levels of both were significantly lower in PDR patients.nnnCONCLUSIONSn1H-NMR-based metabonomic analysis of vitreous fluid permits the obtainment of a metabolic signature of PDR. Apart from the higher abundance of lactate and glucose, significant deficits of galactitol and AA are the main metabolic fingerprints of vitreous fluid from PDR patients.

Gene expression of paired abdominal adipose AQP7 and liver AQP9 in patients with morbid obesity Relationship with glucose abnormalities

The trafficking of glycerol from adipose and hepatic tissue is mainly mediated by 2 aquaporin channel proteins: AQP7 and AQP9, respectively. In rodents, both aquaporins were found to act in a coordinated manner. The aim was to study the relationship between adipose AQP7 and hepatic AQP9 messenger RNA expression and the presence of glucose abnormalities simultaneously in morbid obesity. Adipose tissue (subcutaneous [SAT] and visceral [VAT]) and liver biopsies from the same patient were obtained during bariatric surgery in 30 (21 male and 9 female) morbidly obese subjects. Real-time quantification of AQP7 in SAT and VAT and hepatic AQP9 gene expression were performed. A 75-g oral glucose tolerance test was performed in all subjects. The homeostasis model assessment of insulin resistance and lipidic profile were also determined. Visceral adipose tissue AQP7 expression levels were significantly higher than SAT AQP7 (P = .009). Subcutaneous adipose tissue AQP7 positively correlated with both VAT AQP7 and hepatic AQP9 messenger RNA expression (r = 0.44, P = .013 and r = 0.45, P = .012, respectively). The correlation between SAT AQP7 and liver AQP9 was stronger in intolerant and type 2 diabetes mellitus subjects (r = 0.602, P = .011). We have found no differences in compartmental AQP7 adipose tissue distribution or AQP9 hepatic gene expression according to glucose tolerance classification. The present study provides, for the first time, evidence of coordinated regulation between adipose aquaglyceroporins, with a greater expression found in visceral fat, and between subcutaneous adipose AQP7 and hepatic AQP9 gene expression within the context of human morbid obesity.

Effects of high glucose concentration on the barrier function and the expression of tight junction proteins in human retinal pigment epithelial cells.

There is no information on the direct effect of high glucose concentrations on the barrier function of retinal pigment epithelium (RPE). The aim of this study was to explore the effect of high glucose concentrations on the permeability and the expression of tight junction proteins (occludin, zonula occludens-1 (ZO-1) and claudin-1) in a human RPE line (ARPE-19). For this purpose, ARPE-19 cells were cultured for 3 weeks in a medium containing 5.5 mM D-glucose (mimicking physiological conditions) and 25 mM D-glucose (mimicking hyperglycemia that occurs in diabetic patients). The permeability was evaluated by measuring transepithelial electrical resistance (TER) and apical-basolateral movements of dextran. The expression of tight junction proteins was evaluated by real-time PCR (RT-PCR) and Western blot. Cells grown at 25 mM of D-glucose showed a significant higher TER and a significant lower dextran diffusion than the ones maintained at 5.5 mM of D-glucose. Occludin and ZO-1 mRNA levels and protein content were similar in cultures maintained in 5.5 mM and 25 mM D-glucose. By contrast, high glucose concentrations induced a significant overexpression of claudin-1 (mRNA: 1.03 +/- 0.48 vs 2.29 +/- 0.7 RQ; p = 0.039, at 21 days. Protein levels: 0.92 +/- 0.12 vs 1.14 +/- 0.28 arbitrary units; p = 0.03, at 21 days). However, after blocking claudin-1 expression using siRNA no changes in TER and permeability were observed. We conclude that high glucose concentration results in a reduction of permeability in ARPE-19 cells. In addition, our results suggest that the overexpression of claudin-1 induced by high glucose concentrations is not involved in the mechanisms by which glucose increases the tight junction sealing function. Further studies addressed to unravel the complexity of permeability regulation in RPE are needed.

Differential effects of gemfibrozil and fenofibrate on reverse cholesterol transport from macrophages to feces in vivo

Gemfibrozil and fenofibrate, two of the fibrates most used in clinical practice, raise HDL cholesterol (HDLc) and are thought to reduce the risk of atherosclerotic cardiovascular disease. These drugs act as PPARα agonists and upregulate the expression of genes crucial in reverse cholesterol transport (RCT). In the present study, we determined the effects of these two fibrates on RCT from macrophages to feces in vivo in human apoA-I transgenic (hApoA-ITg) mice. [(3)H]cholesterol-labeled mouse macrophages were injected intraperitoneally into hApoA-ITg mice treated with intragastric doses of fenofibrate, gemfibrozil or a vehicle solution for 17days, and radioactivity was determined in plasma, liver and feces. Fenofibrate, but not gemfibrozil, enhanced [(3)H]cholesterol flux to plasma and feces of female hApoA-ITg mice. Fenofibrate significantly increased plasma HDLc, HDL phospholipids, hApoA-I levels and phospholipid transfer protein activity, whereas these parameters were not altered by gemfibrozil treatment. Unlike gemfibrozil, fenofibrate also induced the generation of larger HDL particles, which were more enriched in cholesteryl esters, together with higher potential to generate preβ-HDL formation and caused a significant increase in [(3)H]cholesterol efflux to plasma. Our findings demonstrate that fenofibrate promotes RCT from macrophages to feces in vivo and, thus, highlight a differential action of this fibrate on HDL.

High glucose concentration leads to differential expression of tight junction proteins in human retinal pigment epithelial cells

INTRODUCTIONnOne of the early features of diabetic retinopathy is the breakdown of the blood-retinal barrier (BRB) due to disruption of the tight junctions. Whereas impairment of the proteins involved in the disruption of the tight junctions of the internal BRB has been extensively studied, there is no information on the direct effect of high glucose concentration on the barrier function of the outer blood-retinal barrier (formed by the retinal pigment epithelium [RPE]). The aim of this study was to explore the effect of high glucose concentration on the expression of tight junction proteins (occludin, zonula occludens-1 [ZO-1] and claudin-1) in a human RPE line under two distinct glucose concentrations.nnnMATERIALS AND METHODSnAn RPE cell line (ARPE-19) were cultured for 3 weeks in a medium supplemented with 10% fetal calf serum containing 5.5 mmol D-glucose (mimicking physiological conditions) or 25 mmol Dglucose (mimicking the hyperglycemia that occurs in diabetic patients). Occludin, ZO-1 and claudin-1 were studied by real-time polymerase chain reaction and Western blot at 14 and 21 days.nnnRESULTSnOccludin and ZO-1 mRNA levels and protein content were similar in cultures maintained at 5.5 mmol and 25 mmol of D-glucose. In contrast, high glucose concentration (25 mmol) induced a clear upregulation in claudin-1 mRNA expression and protein content at 21 days (mRNA level: 1.03 vs 2.29; protein content: 0.92 vs 1.14).nnnCONCLUSIONSnHigh glucose concentration leads to differential expression of tight junction proteins in ARPE-19 cells. In addition, our results suggest that the upregulation of claudin-1by glucose is involved in the increase of tight junction sealing function. The functional consequences and clinical applicability of these findings require further investigation.

Effects of sardine-enriched diet on metabolic control, inflammation and gut microbiota in drug-naïve patients with type 2 diabetes: a pilot randomized trial

BackgroundNutrition therapy is the cornerstone of treating diabetes mellitus. The inclusion of fish (particularly oily fish) at least two times per week is recommended by current international dietary guidelines for type 2 diabetes. In contrast to a large number of human studies examining the effects of oily fish on different cardiovascular risk factors, little research on this topic is available in patients with type 2 diabetes. The aims of this pilot study were to investigate the effects of a sardine-enriched diet on metabolic control, adiponectin, inflammatory markers, erythrocyte membrane fatty acid (EMFA) composition, and gut microbiota in drug-naïve patients with type 2 diabetes.Methods35 drug-naïve patients with type 2 diabetes were randomized to follow either a type 2 diabetes standard diet (control group: CG), or a standard diet enriched with 100xa0g of sardines 5xa0days a week (sardine group: SG) for 6xa0months. Anthropometric, dietary information, fasting glycated hemoglobin, glucose, insulin, adiponectin, inflammatory markers, EMFA and specific bacterial strains were determined before and after intervention.ResultsThere were no significant differences in glycemic control between groups at the end of the study. Both groups decreased plasma insulin (SG: −35.3xa0%, Pu2009=u20090.01, CG: −22.6xa0%, Pu2009=u20090.02) and homeostasis model of assessment - insulin resistance (HOMA-IR) (SG: −39.2xa0%, Pu2009=u20090.007, CG: −21.8xa0%, Pu2009=u20090.04) at 6-months from baseline. However only SG increased adiponectin in plasma compared to baseline level (+40.7xa0%, Pu2009=u20090.04). The omega-3 index increased 2.6xa0% in the SG compared to 0.6xa0% in the CG (Pu2009=u20090.001). Both dietary interventions decreased phylum Firmicutes (SG and CG: Pu2009=u20090.04) and increased E. coli concentrations (SG: Pu2009=u20090.01, CG: Pu2009=u20090.03) at the end of the study from baseline, whereas SG decreased Firmicutes/Bacteroidetes ratio (Pu2009=u20090.04) and increased Bacteroides-Prevotella (Pu2009=u20090.004) compared to baseline.ConclusionsAlthough enriching diet with 100xa0g of sardines 5xa0days a week during 6xa0months to a type 2 diabetes standard diet seems to have neutral effects on glycemic control in drug-naïve patients with type 2 diabetes, this nutritional intervention could have beneficial effects on cardiovascular risk. Furthermore, both dietary interventions decreased HOMA-IR and altered gut microbiota composition of drug-naïve patients with type 2 diabetes.Trial registrationTrial number and name of the registry: NCT02294526, ClinicalTrials.gov

Impact of Glucose-Lowering Agents on the Risk of Cancer in Type 2 Diabetic Patients. The Barcelona Case-Control Study

Background The aim of the present study is to evaluate the impact of glucose-lowering agents in the risk of cancer in a large type 2 diabetic population. Methods A nested case-control study was conducted within a defined cohort (275,164 type 2 diabetic patients attending 16 Primary Health Care Centers of Barcelona). Cases (nu200a=u200a1,040) comprised those subjects with any cancer diagnosed between 2008 and 2010, registered at the Cancer Registry of Hospital Vall dHebron (Barcelona). Three control subjects for each case (nu200a=u200a3,120) were matched by age, sex, diabetes duration, and geographical area. The treatments analyzed (within 3 years prior to cancer diagnosis) were: insulin glargine, insulin detemir, human insulin, fast-acting insulin and analogues, metformin, sulfonylureas, repaglinide, thiazolidinediones, dipeptidyl peptidase-4 inhibitors, and alpha glucosidase inhibitors. Conditional logistic regressions were used to calculate the risk of cancer associated with the use of each drug adjusted by age, BMI, dose and duration of treatment, alcohol use, smoking habit, and diabetes duration. Results No differences were observed between case and control subjects for the proportion, dose or duration of exposure to each treatment. None of the types of insulin and oral agents analyzed showed a significant increase in the risk of cancer. Moreover, no cancer risk was observed when glargine was used alone or in combination with metformin. Conclusions Our results suggest that diabetes treatment does not influence the risk of cancer associated with type 2 diabetes. Therefore, an eventual increase of cancer should not be a reason for biasing the selection of any glucose-lowering treatment in type 2 diabetic population.

Effect of atorvastatin on lipoprotein (a) and interleukin-10: a randomized placebo-controlled trial.

AIMnThis study aimed to determine the effect of atorvastatin therapy on plasma lipoprotein (a) [Lp(a)] and biomarkers of inflammation in hypercholesterolaemic patients free of cardiovascular disease.nnnMETHODSnIn this three-month randomized double-blind placebo-controlled trial, 63 hypercholesterolaemic patients were randomly treated with either placebo or atorvastatin (10 or 40 mg/day) for 12 weeks. Lp(a) and biomarkers of inflammation (C-reactive protein [CRP], interleukin [IL]-6 and -10, and tumour necrosis factor-alpha receptors [TNF-Rs]) were measured at study entry, and at four and 12 weeks of follow-up.nnnRESULTSnAt the end of the study, patients allocated to atorvastatin (10 or 40 mg/day) presented with significantly lower Lp(a) levels than those taking placebo (10 [1-41]mg/dL versus 6 [1-38]mg/dL [P = 0.02] and 21 [1-138]mg/dL versus 15 [1-103]mg/dL [P = 0.04], respectively]. In multivariate analyses, the relative changes in Lp(a) were independently related to baseline Lp(a) levels and CRP changes. No significant changes in CRP, IL-6 and TNF-Rs were observed. In contrast, IL-10 (pg/mL) increased significantly in patients taking atorvastatin (2.14 [0.49-43]pg/mL versus 4.54 [0.51-37.5]pg/mL; P = 0.01), and was even more increased with the 40-mg dose than with 10mg.nnnCONCLUSIONnOur results suggest that 12-week atorvastatin is effective in reducing Lp(a) in dyslipidaemic patients free of CVD. Furthermore, this is also the first evidence that the drug increases IL-10 in a dose-dependent manner.

Gene expression profiling in hearts of diabetic mice uncovers a potential role of estrogen-related receptor γ in diabetic cardiomyopathy

Diabetic cardiomyopathy is characterized by an abnormal oxidative metabolism, but the underlying mechanisms remain to be defined. To uncover potential mechanisms involved in the pathophysiology of diabetic cardiomyopathy, we performed a gene expression profiling study in hearts of diabetic db/db mice. Diabetic hearts showed a gene expression pattern characterized by the up-regulation of genes involved in lipid oxidation, together with an abnormal expression of genes related to the cardiac contractile function. A screening for potential regulators of the genes differentially expressed in diabetic mice found that estrogen-related receptor γ (ERRγ) was increased in heart of db/db mice. Overexpression of ERRγ in cultured cardiomyocytes was sufficient to promote the expression of genes involved in lipid oxidation, increase palmitate oxidation and induce cardiomyocyte hypertrophy. Our findings strongly support a role for ERRγ in the metabolic alterations that underlie the development of diabetic cardiomyopathy.

Rendimiento diagnóstico de la determinación plasmática de tiroglobulina y del rastreo corporal con 131INa según el tipo de recurrencia en el carcinoma diferenciado de tiroides

Fundamento No existe un consenso claro sobre cual es el mejor protocolo de seguimiento en elcarcinoma diferenciado de tiroides (CDT). El objetivo del estudio es analizar la utilidad y el significadopronostico de la determinacion plasmatica de tiroglobulina (Tg) y el rastreo corporalcon 131 INa (RCT) en las recidivas de CDT tanto de forma global como segun el tipo de recurrencia(recidiva local o metastasis a distancia). Pacientes y metodos Se incluyen 34 pacientes con recurrencia de CDT recogidos durante un periodode 15 anos y con un seguimiento minimo de 5 anos, con anticuerpos antitiroglobulinanegativos y sin metastasis en el momento del diagnostico. En todos los casos se practico la tiroidectomiatotal o casi total y ablacion de restos con 131INa. El seguimiento posterior incluyola determinacion plasmatica de Tg y el RCT realizados en situacion de hipotiroidismo. Resultados La Tg estaba elevada en el 64,7% de los pacientes y el RCT en el 82,3%. En la recidivalocal aislada la sensibilidad del RCT fue superior a la de la Tg (93,7 frente a 43,7%; p Conclusiones La sensibilidad de la Tg y el RCT es diferente segun el tipo de recurrencia delCDT. Por tanto, ambas pruebas son complementarias y no puede prescindirse de ninguna deellas en el seguimiento del CDT.