Luis M. Jiménez

Simvastatin improves endothelial function in spontaneously hypertensive rats through a superoxide dismutase mediated antioxidant effect.

Background Hydroxymethylglutaryl coenzyme A (HMGCoA) reductase inhibitors have beneficial effects beyond their cholesterol-lowering properties. The antioxidant mechanism of HMGCoA reductase inhibitors is not completely understood. Objectives To elucidate the antioxidant effect of simvastatin. Methods We studied the influence of simvastatin treatment on the development of hypertension, modification of antioxidant systems, and reactivity of aortic rings in Wistar–Kyoto (WKY) and spontaneously hypertensive (SHR) rats. Results Simvastatin had no effect on blood pressure (BP). Simvastatin treatment (either 1 or 2 mg/kg body weight for 12 or 20 weeks) increased superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities in SHR rats compared with untreated control SHR rats. Carbachol-induced relaxation of aortic rings was impaired in control SHR rats and was restored by simvastatin treatment. Addition of SOD improved the response in control SHR rats and did not have any effect in treated SHR rats. Addition of diethyldithiocarbamic acid, a selective inhibitor of SOD, produced a mild non-significant impairment in carbachol-induced relaxation in control SHR rats, suggesting a deficient antioxidant system in these animals. However, in treated SHR and in WKY rats, impairment of the relaxation was marked, implying that SOD activity in these animals was important to maintain endothelial function. In aortic rings without endothelium from SHR rats, contraction induced by free radicals was substantially higher than in WKY rats. This effect was attenuated in 1-mg-treated rats and abolished in 2-mg-treated rats. Conclusions Simvastatin promotes intracellular antioxidant systems, fundamentally SOD, restoring endothelial function but not having any effect on blood pressure.

Improvement of age-related endothelial dysfunction by simvastatin: effect on NO and COX pathways

The effects of oral administration of the HMG‐CoA reductase inhibitor, simvastatin (SV), on age‐related endothelial dysfunction were investigated in the aorta of male Wistar rats. Adult (12–14 weeks) and old (60–80 weeks) rats were treated daily for 12 weeks with either vehicle or SV (1 mg kg−1). In old rats, SV treatment did not significantly affect systolic blood pressure and LDL‐cholesterol, but it reduced plasma cholesterol, triglycerides and oxidised LDL though it did not affect total antioxidant status. SV improved endothelium‐dependent relaxation to acetylcholine and A‐23187 in vessels from aged, but not adult, rats. This effect was linked to a greater NO vasodilatation via an increased expression of endothelial NO‐synthase. A mechanism sensitive to superoxide dismutase and catalase also accounts for enhanced endothelial vasodilatation. Finally, SV did not affect the release of prostacyclin, but it inhibited the generation of thromboxane (TX) A2 from COX‐2 isoform. The effect of the latter was sensitive to the Tp receptor antagonist, ICI‐192,605. The present study provides evidence that oral administration of SV improves endothelial dysfunction in the aorta from aged rats by mechanisms associated with enhanced NO vasodilatation, reduced release of TXA2 from cyclo‐oxygenase, and increased antioxidant properties of the vessel wall. These data underscore a new therapeutic perspective for SV in age‐related endothelial dysfunction.

Determination of Organic Contaminants in Landfill Leachates: A Review

Leachates derived from landfills constitute a potential risk of groundwater pollution because a variety of contaminants can be released by leaking from the contention system. Therefore, the leachate composition is of interest of their appropriate management. Although the leachate characterisation is usually carried out by global parameters (i.e. DOC, BOD, COD, AOX, etc), its characterisation at molecular level is of increasing interest and will be reviewed in the present article. Sample handling and determination techniques for a variety of organic contaminants is discussed and pitfalls as well as limitations of each analytical technique will be highlighted.

Genotype of the CYBA promoter -930A/G, polymorphism C677T of the MTHFR and APOE genotype in patients with hypertensive disorders of pregnancy: an observational study.

BACKGROUND AND OBJECTIVE Hypertensive disorders of pregnancy could be favoured by polymorphisms in genes affecting vascular physiology. The aim of our work was to study several variants in the genes regulating oxidative stress, plasma lipids metabolism and endothelial function (observational study). MATERIAL AND METHODS We studied the -930A/G polymorphism of the CYBA gene promoter, the apolipoprotein E (APOE) genotype and the methylene-tetrahydrofolate reductase (MTHFR) gene C677T polymorphism in 134 healthy pregnant women, 266 pregnant with non-proteinuric hypertension (NPH) and 184 patients with preeclampsia (PE). RESULTS The GG genotype of the CYBA gene promoter was present in 32.1% of the control population, 38.7% of patients with NPH (P=0.19) and 21.2% of the women with PE (P=0.03). A higher frequency of epsilon 3/epsilon 4 and epsilon 4/epsilon 4 genotypes of APOE was observed in patients with PE or NPH compared with controls (P<0.01). There were no significant differences detected in genotype or allele distribution of the MTHFR, C677T polymorphism. APOE epsilon 3/epsilon 4 and epsilon 4/epsilon 4 genotypes had a worse lipoprotein profile characterized by higher plasma values of total cholesterol (P<0.05) and triglycerides (P<0.005). Despite no differences in MTHFR C677T polymorphism distribution, higher levels of plasma homocysteine were observed in patients with PE than in patients with NPH or controls. CONCLUSIONS CYBA and APOE polymorphism showed a different distribution in the groups studied, while no differences were observed in MTHFR C677T polymorphism. APOE genotype was associated with changes in lipid and lipoprotein profiles in pregnant women.

Effects of simvastatin on endothelial function after chronic inhibition of nitric oxide synthase by L-NAME.

Blood pressure, plasma NO2 and NO3 level, heart weight index, antioxidant enzyme activity, and vascular reactivity in rat intact aortic rings were assessed to investigate the effects of 8-week treatment with the hydroxy-methyl-glutaryl coenzyme A reductase inhibitor simvastatin (1 mg/kg per day) on endothelial dysfunction induced by chronic Nω-nitro-l-arginine methyl ester (l-NAME 70 mg/kg per day). Results were compared with those obtained in rats receiving l-NAME, simvastatin or control animals. Coadministration of simvastatin did not restore l-NAME–increased blood pressure but normalized heart weight index (P < 0.05), endothelium-dependent relaxation to acetylcholine (P < 0.001), and plasma NO2 and NO3 concentration (P < 0.001) without affecting relaxation to sodium nitroprusside. Endothelium-dependent relaxation in these animals was abolished by acute incubation with l-NAME, unaffected by thromboxane synthetase inhibitor and TXA2/PGH2 receptor antagonist, ridogrel, and decreased by indomethacin. Simvastatin treatment also increased plasma NO2+NO3 without affecting endothelial function, heart weight index, and blood pressure of control rats. The presence of superoxide dismutase (SOD) and catalase improved endothelial relaxation only in l-NAME–treated rats, but O2− generated by hypoxanthine and xanthine oxidase inhibited the relaxant effect in both l-NAME and simvastatin plus l-NAME–treated rats. SOD activity was increased in all groups receiving simvastatin. Long-term treatment with simvastatin restored l-NAME–induced endothelial dysfunction, probably by preventing nitric oxide decrease. Other effects of simvastatin, including release of compensating vasodilatory cyclo-oxygenase products and increased SOD activity, could also be involved.

Role of ketoconazole treatment in urinary-free cortisol-to-cortisone and tetrahydrocortisol-to-tetrahydrocortisone ratios in nonectopic Cushing's syndrome.

We hypothesized that in nonectopic Cushing syndrome there is an insufficient activity of type II (renal) 11β-hydroxysteroid dehydrogenase (11β-HSD2) that is related to cortisol excess, rather than to corticotropin (adrenocorticotropic hormone [ACTH]) levels. We measured plasma ACTH and urinary-free cortisol (UFF), urinary-free cortisone (UFE), tetrahydrocortisol (UTHF), and tetrahydrocortisone (UTHE) in 24-h urine samples of 24 healthy subjects and 15 patients diagnosed with nonectopic Cushing syndrome. Then, in the group of patients, a new 24-h urine sample was collected after treatment with 800 mg daily of ketoconazole. The UFF/UFE and UTHF/UTHE ratios were calculated as an estimation of 11β-HSD2 activity. The patients had an increase in both the UFF/UFE (19.95±10.3 vs 5.78±4.72 nmol/24 h; p<0.0001) and UTHF/UTHE ratios (5.36±5.23 vs 1.39±0.95 nmol/24 h; p<0.001). Both UFF/UFE and UTHF/UTHE ratios decreased after ketoconazole treatment (19.95±10.3 vs 12.2±6.9 nmol/24 h; p<0.005; and 5.36±5.23 vs 1.62 vs 1.21 nmol/24 h; p<0.001, respectively). The control subjects had a significant relationship between UFF and UFE (r=0.70, p<0.0001), and between UTHF and UTHE (r=0.75, p<0.0001) that did not exist in the patient group. After ketoconazole treatment, the decrease in cortisol excretion in the patient group allowed a positive and significant relation between UFF and UFE (r=0.64, p<0.01) and between UTHF and UTHE (r=0.56, p<0.05) to appear. There was not any significant relationship between either UFF/UFE or UTHF/UTHE ratios and plasma levels of ACTH.

Detection of c. -32T>G (IVS1-13T>G) mutation of Pompe disease by real-time PCR in dried blood spot specimen.

BACKGROUND Pompe disease, or acid maltase deficiency, is a genetic muscle disorder caused by mutations in the gene encoding the acid alpha-glucosidase (GAA) enzyme, which is essential for the degradation of glycogen to glucose in lysosomes. The wide clinical variability is resulted from genetic heterogeneity, and many different mutations of the GAA gene have been reported. Some of these mutations are associated with specific phenotypes, such as the c. -32T>G (IVS1-13T>G) mutation seen in late-onset Pompe disease. METHODS We used a real-time PCR, after genomic DNA extraction isolated from DBS (dried blood spots) and PCR amplification. RESULTS Our results successfully detected in controls and patients have been 100% concordant with sequencing results. CONCLUSIONS This assay combines simple sample processing and rapid analysis and it allows to detect the patients with a milder form and slower progression of this disease with a high reliability.

Glucosa tetrasacárido como biomarcador diagnóstico de la enfermedad de Pompe: estudio en 35 pacientes

Background and objectives Pompe disease is a disorder originating from an acid alpha-glycosidase (AAG) enzyme deficiency. This disease produces an accumulation of lysosomal glycogen in different tissues, whereby the skeletal and heart muscles are especially involved. The established diagnosis is achieved through the identification of the AAG deficiency. There are also other secondary diagnostic biomarkers, such as tetra-saccharide glucose (Glc4), which shows high levels in the urine of these patients. In this study it is highlighted the usefulness of Glc4 as a diagnostic biomarker for Pompe disease in its different forms of presentation, using a high-performance liquid chromatography with ultraviolet detection (HPLC/UV) adapted to the study.BACKGROUND AND OBJECTIVES Pompe disease is a disorder originating from an acid alpha-glycosidase (AAG) enzyme deficiency. This disease produces an accumulation of lysosomal glycogen in different tissues, whereby the skeletal and heart muscles are especially involved. The established diagnosis is achieved through the identification of the AAG deficiency. There are also other secondary diagnostic biomarkers, such as tetra-saccharide glucose (Glc4), which shows high levels in the urine of these patients. In this study it is highlighted the usefulness of Glc4 as a diagnostic biomarker for Pompe disease in its different forms of presentation, using a high-performance liquid chromatography with ultraviolet detection (HPLC/UV) adapted to the study. PATIENTS AND METHODS A total of 75 individuals have been analyzed: 40 healthy controls and 35 patients diagnosed with Pompe disease. Twenty-four hour samples of urine were collected from all of the patients and their Glc4 levels were determined by means of HPLC/UV. RESULTS The evaluation of the urinary Glc4 shows a high discrimination ability between healthy/sick individuals. In addition, the results obtained have allowed to establish the most appropriate level of decision or cut-off point for the identification of sick people. CONCLUSIONS Glc4 urinary levels are found to be high in patients suffering from Pompe disease and even though increased levels are also found in other conditions, the existence of a AAG deficiency together with a compatible clinical symptoms, prove very helpful for a correct diagnosis of this serious disease.

Alteraciones del metabolismo glucídico en la hipertensión arterial esencial. Papel de la sobrecarga oral con glucosa

BACKGROUND AND OBJECTIVE: There is a high prevalence of diabetes in essential hypertensive patients, hence increasing the cardiovascular risk of these subjects. Our aim was to test the importance of a routine assessment of an oral glucose tolerance test (OGTT) in a sample of recently diagnosed essential hypertensives. PATIENTS AND METHOD: We studied 270 recently diagnosed untreated hypertensives (56% males). We measured fasting lipids and plasma glucose and insulin at baseline and after OGTT. RESULTS: 38.5% of the studied subjects had any abnormality of glucose metabolism, glucose intolerance (22.2%) and type 2 diabetes (12.2%) being the most common. An impaired fasting glucose was present in 4.1% of subjects. Patients with intolerance or type 2 diabetes had a worse plasma lipid profile, and those with impaired fasting glucose, a higher HOMA index. CONCLUSIONS: The high incidence of glucidic abnormalities found in hypertensive patients signals a need to perform an OGTT test in them. This can lead to the implementation of strategies aimed at halting vascular impairment in hypertensives with type 2 diabetes and delay the development of type 2 diabetes in patients with glucose intolerance.

Evaluation of a Multiplex ELISA for Autoantibody Profiling in Patients with Autoimmune Connective Tissue Diseases

The performance of immunoassays for the detection of autoantibodies is of critical importance in the diagnosis and assessment of patients with autoimmune connective tissue diseases (ACTD). Our objective was to compare the features of two multiplexed assays—INNO-LIA ANA and Gennova-PictArray ENA ELISA—for measurement of multiple autoantibodies and their utility as a clinical tool in ACTD diagnosis. The antigens included SS-A/Ro (60 and 52), SSB/La, Sm, Sm/RNP, CENP-B, Jo-1, and Scl-70. Stored sera from 85 ACTD patients and 80 controls consisting of patients with vasculitis, rheumatoid arthritis and infectious diseases, as well as healthy subjects were analyzed jointly with clinical and laboratory data. Agreement between the two methods varied between 58 and 99% (Cohens kappa: 0.21–0.71) mostly for SSA and SSB. The frequency of specific autoantibodies measured using the two methods was more variable for SSA, SSB, and RNP/Sm. There were a higher number of ambiguous results when using INNO-LIA. The optimized cut-off values of the Gennova-PictArray resulted in over 99% specificities in samples obtained from the control group. Sensitivity patterns were more accurate in Gennova-PictArray than in INNO-LIA, as suggested in previously reported studies. A third method could be applied to determine which of the two methods is more accurate.