Guillermo T. Sáez

Effect of a traditional Mediterranean diet on lipoprotein oxidation: a randomized controlled trial.

BACKGROUND Despite the richness in antioxidants of the Mediterranean diet, to our knowledge, no randomized controlled trials have assessed its effect on in vivo lipoprotein oxidation. METHODS A total of 372 subjects at high cardiovascular risk (210 women and 162 men; age range, 55-80 years), who were recruited into a large, multicenter, randomized, controlled, parallel-group clinical trial (the Prevención con Dieta Mediterránea [PREDIMED] Study) directed at testing the efficacy of the traditional Mediterranean diet (TMD) on the primary prevention of coronary heart disease, were assigned to a low-fat diet (n = 121) or one of 2 TMDs (TMD + virgin olive oil or TMD + nuts). The TMD participants received nutritional education and either free virgin olive oil for all the family (1 L/wk) or free nuts (30 g/d). Diets were ad libitum. Changes in oxidative stress markers were evaluated at 3 months. RESULTS After the 3-month interventions, mean (95% confidence intervals) oxidized low-density lipoprotein (LDL) levels decreased in the TMD + virgin olive oil (-10.6 U/L [-14.2 to -6.1]) and TMD + nuts (-7.3 U/L [-11.2 to -3.3]) groups, without changes in the low-fat diet group (-2.9 U/L [-7.3 to 1.5]). Change in oxidized LDL levels in the TMD + virgin olive oil group reached significance vs that of the low-fat group (P = .02). Malondialdehyde changes in mononuclear cells paralleled those of oxidized LDL. No changes in serum glutathione peroxidase activity were observed. CONCLUSIONS Individuals at high cardiovascular risk who improved their diet toward a TMD pattern showed significant reductions in cellular lipid levels and LDL oxidation. Results provide further evidence to recommend the TMD as a useful tool against risk factors for CHD. Trial Registration isrctn.org Identifier: ISRCTN35739639.

Cohort Profile: Design and methods of the PREDIMED study

The Spanish Ministry of Health—Instituto de Salud Carlos III (ISCIII) funded the project for the period 2003–05 (RTIC G03/140). In 2006 a new funding modality was established by ISCIII through the CIBER (Centros de Investigacion Biomedica En Red). Fisiopatologia de la Obesidad y Nutricion (CIBERobn), which is providing funding for 7 of the original research groups, whereas the other 12 were funded by a new research network (RTIC RD 06/0045). Other official funds from Spanish government agencies have been obtained for subprojects related to intermediate outcomes (lipoproteins, inflammatory markers, vascular imaging, genomic and proteomic studies, etc.). Obviously, the donation by food companies of all the VOO and mixed nuts needed throughout the duration of the study is a substantial contribution.

The production of free radicals during the autoxidation of cysteine and their effect on isolated rat hepatocytes

Autoxidizing cysteine has been shown to produce thiyl and hydroxyl radicals. Hydrogen peroxide increased the yield of both radicals which was inhibited by catalase but stimulated by copper/zinc superoxide dismutase. This effect is due to increased hydrogen peroxide production by copper/zinc superoxide dismutase as a result of superoxide dismutation. The production of superoxide radicals could not be detected probably because of its low reactivity, however, measurement of oxygen uptake and reduction of ferricytochrome c by autoxidizing cysteine clearly implicate the involvement of super oxide radicals. The production of hydroxyl radicals is postulated to proceed through a fenton reaction, however, this may not necessarily be metal ion controlled. Autoxidizing cysteine disrupts the integrity of hepatocytes causing release of glutathione, adenosine triphosphate and lactate dehydrogenase indicating that it is of little use as a therapeutic agent.

In vivo nutrigenomic effects of virgin olive oil polyphenols within the frame of the Mediterranean diet: a randomized controlled trial

The aim of the study was to assess whether benefits associated with the traditional Mediterranean diet (TMD) and virgin olive oil (VOO) consumption could be mediated through changes in the expression of atherosclerosis‐related genes. A randomized, parallel, controlled clinical trial in healthy volunteers (n=90) aged 20 to 50 yr was performed. Threemonth intervention groups were as follows: 1) TMD with VOO (TMD+VOO), 2) TMD with washed virgin olive oil (TMD+WOO), and 3) control with participants’ habitual diet. WOO was similar to VOO, but with a lower polyphenol content (55 vs. 328 mg/kg, respectively). TMD consumption decreased plasma oxidative and inflammatory status and the gene expression related with both inflammation [INF‐γ (INFy), Rho GTPase‐activating protein15 (ARHGAP15), and interleukin‐7 receptor (IL7R)] and oxidative stress [adrenergic ß2‐receptor (ADRB2) and polymerase (DNA‐directed) κ (POLK)] in peripheral blood mononuclear cells. All effects, with the exception of the decrease in POLK expression, were particularly observed when VOO, rich in polyphenols, was present in the TMD dietary pattern. Our results indicate a significant role of olive oil polyphenols in the down‐regulation of proatherogenic genes in the context of a TMD. In addition, the benefits associated with a TMD and olive oil polyphenol consumption on cardiovascular risk can be mediated through nutrigenomic effects.—Konstantinidou, V., Covas, M.‐I., Mun˜oz‐Aguayo, D., Khymenets, O., de la Torre, R., Saez, G., del Carmen Tormos, M., Toledo, E., Marti, A., Ruiz‐Gutiérrez, V., Ruiz Mendez, M. V., Fito, M. In vivo nutrigenomic effects of virgin olive oil polyphenols within the frame of the Mediterranean diet: a randomized controlled trial. FASEBJ. 24, 2546–2557 (2010). www.fasebj.org

Effect of the Mediterranean diet on blood pressure in the PREDIMED trial: results from a randomized controlled trial

BackgroundHypertension can be prevented by adopting healthy dietary patterns. Our aim was to assess the 4-year effect on blood pressure (BP) control of a randomized feeding trial promoting the traditional Mediterranean dietary pattern.MethodsThe PREDIMED primary prevention trial is a randomized, single-blinded, controlled trial conducted in Spanish primary healthcare centers. We recruited 7,447 men (aged 55 to 80 years) and women (aged 60 to 80 years) who had high risk for cardiovascular disease. Participants were assigned to a control group or to one of two Mediterranean diets. The control group received education on following a low-fat diet, while the groups on Mediterranean diets received nutritional education and also free foods; either extra virgin olive oil, or nuts. Trained personnel measured participants’ BP at baseline and once yearly during a 4-year follow-up. We used generalized estimating equations to assess the differences between groups during the follow-up.ResultsThe percentage of participants with controlled BP increased in all three intervention groups (P-value for within-group changes: P<0.001). Participants allocated to either of the two Mediterranean diet groups had significantly lower diastolic BP than the participants in the control group (−1.53 mmHg (95% confidence interval (CI) −2.01 to −1.04) for the Mediterranean diet supplemented with extra virgin olive oil, and −0.65 mmHg (95% CI -1.15 to −0.15) mmHg for the Mediterranean diet supplemented with nuts). No between-group differences in changes of systolic BP were seen.ConclusionsBoth the traditional Mediterranean diet and a low-fat diet exerted beneficial effects on BP and could be part of advice to patients for controlling BP. However, we found lower values of diastolic BP in the two groups promoting the Mediterranean diet with extra virgin olive oil or with nuts than in the control group.Trial registrationCurrent Controlled Trials ISRCTN35739639

Dietary intake and major food sources of polyphenols in a Spanish population at high cardiovascular risk: The PREDIMED study

BACKGROUND AND AIMS Epidemiological data have shown an inverse association between the consumption of polyphenol-rich foods and the risk of cardiovascular disease or overall mortality. A comprehensive estimation of individual polyphenol intake in nutritional cohorts is needed to gain a better understanding of this association. The aim of this study was to estimate the quantitative intake of polyphenols and the major dietary sources in the PREDIMED (PREvención con DIeta MEDiterránea) cohort using individual food consumption records. METHODS AND RESULTS The PREDIMED study is a large, parallel-group, multicentre, randomised, controlled 5-year feeding trial aimed at assessing the effects of the Mediterranean diet on the primary prevention of cardiovascular disease. A total of 7200 participants, aged 55-80 years, completed a validated 1-year food frequency questionnaire (FFQ) at baseline. Polyphenol consumption was calculated by matching food consumption data from the FFQ with the recently developed Phenol-Explorer database on polyphenol content in foods. The mean total polyphenol intake was 820 ± 323 mg day⁻¹ (443 ± 218 mg day⁻¹ of flavonoids and 304 ± 156 mg day⁻¹ of phenolic acids). Hydroxycinnamic acids were the phenolic group with the highest consumption and 5-caffeoylquinic acid was the most abundantly ingested individual polyphenol. The consumption of olives and olive oil was a differentiating factor in the phenolic profile of this Spanish population compared with other countries. CONCLUSION In Mediterranean countries, such as Spain, the main dietary source of polyphenols is coffee and fruits, but the most important differentiating factor with respect to other countries is the consumption of polyphenols from olives and olive oil.

Effects of dietary fibre intake on risk factors for cardiovascular disease in subjects at high risk

Background: Epidemiological studies and feeding trials with supplements suggest that fibre intake is associated with a reduction in cardiovascular risk. However, the effects of changes in dietary fibre on risk factor levels have not been evaluated in free-living individuals. Thus, the effects of changes in dietary fibre intake on cardiovascular risk factors were assessed over 3 months in free-living high-risk subjects. Methods: 772 high-risk subjects (age 69±5 years) were assigned to a low-fat diet or two Mediterranean-style diets. All participants received behavioural and nutritional education, including recommendations for increasing the consumption of vegetables, fruits, and legumes. Changes in food and nutrient intake, body weight, blood pressure, lipid profiles, glucose control and inflammatory markers were evaluated. Results: Most participants increased consumption of vegetable products, but the increase in dietary fibre exhibited wide between-subject variability (6–65 g/day). Body weight, waist circumference, and mean systolic and diastolic blood pressure decreased across quintiles of fibre intake (p<0.005; all). Reductions in fasting glucose and total cholesterol levels, and increments in HDL cholesterol were highest among participants in the upper 20% of fibre intake (p = 0.04 and 0.02 respectively). Plasma concentrations of C-reactive protein, but not those of inflammatory cytokines, decreased in parallel with increasing dietary fibre (p = 0.04). Significant reductions in LDL cholesterol were observed only among participants with the greatest increases in soluble fibre intake (p = 0.04). Conclusions: Increasing dietary fibre intake with natural foods is associated with reductions in classical and novel cardiovascular risk factors in a high-risk cohort.

Genetic alterations and oxidative metabolism in sporadic colorectal tumors from a Spanish community

Deletions of loci on chromosomes 5q, 17p, 18q, and 22q, together with the incidence of p53 mutations and amplification of the double minute‐2 gene were investigated in the sporadic colorectal tumors of 44 patients from a Spanish community. Chromosome deletions were analyzed by means of loss of heterozygosity analysis using a restriction fragment length polymorphism assay. Allelic losses were also detected by polymerase chain reaction (PCR)‐single‐stranded conformation polymorphism (SSCP) analysis of a polymorphic site in intron 2 of the p53 gene. The percentages of genetic deletions on the screened chromosomes were 39.3% (5q), 58.3% (17p), 40.9% (18q), and 40% (22q). Mutations in p53 exons 2–9 were examined by PCR‐SSCP analysis and direct sequencing of the mutated region. Twenty of 44 tumor samples (45.45%) showed mutations at various exons except for exons 2, 3, and 9, the most frequent changes being G → T transversion and C → T transition. Because oxygen‐free radicals play a role in the carcinogenesis process, we evaluated the oxidative status of the colorectal tumors. Antioxidant activities, lipid peroxidation, and DNA‐damaged product concentrations in colon tumors and normal mucosa were compared. In tumor tissues, superoxide dismutase and catalase decreased fourfold and twofold, respectively, whereas glutathione peroxidase and reduced glutathione increased threefold. Malondialdehyde and 8‐hydroxy‐2′‐deoxyguanosine (8‐OHdG) levels were twofold higher in colorectal tumors than in normal mucosa. Seven of 10 DNA tumor samples (70%) showing higher values of 8‐OHdG also had genetic alterations at different chromosomal loci. In these samples, the p53 gene was deleted or mutated in 71.4% of cases. We concluded that the observed changes in the oxidative metabolism of the tumor cells and the consecutive increase in DNA damage may potentiate the genomic instability of different chromosomal regions, leading to further cell malignancy and tumor expansion. Mol. Carcinog. 18:232–243, 1997.

Low-fat dairy products and blood pressure : follow-up of 2290 older persons at high cardiovascular risk participating in the PREDIMED study

High blood pressure (BP) has been ranked as the most important risk factor worldwide regarding attributable deaths. Dietary habits are major determinants of BP. Among them, frequent intake of low-fat dairy products may protect against hypertension. Our aim was to assess the relationship between low-fat dairy product intake and BP levels and their changes after 12-month follow-up in a cohort of asymptomatic older persons at high cardiovascular risk recruited into a large-scale trial assessing the effects of Mediterranean diets on cardiovascular outcomes. Data from 2290 participants, including 1845 with hypertension, were available for analyses. Dairy products were not a specific part of the intervention; thus, data were analysed as an observational cohort. Dietary information was collected with validated semi-quantitative FFQ and trained personnel measured BP. To assess BP changes, we undertook cross-sectional analyses at baseline and at the end of follow-up and longitudinal analyses. A statistically significant inverse association between low-fat dairy product intake and systolic BP was observed for the 12-month longitudinal analysis. In the longitudinal analysis, the adjusted systolic and diastolic BP were significantly lower in the highest quintile of low-fat dairy product intake (-4.2 (95% CI -6.9, -1.4) and -1.8 (95% CI -3.2, -0.4) mmHg respectively), whereas the point estimates for the difference in diastolic BP indicated a modest non-significant inverse association. Intake of low-fat dairy products was inversely associated with BP in an older population at high cardiovascular risk, suggesting a possible protective effect against hypertension.

Mitochondrial dysfunction, persistent oxidative damage, and catalase inhibition in immune cells of naïve and treated Crohn's disease

Background: Oxidative stress is considered a potential etiological factor for Crohns disease (CD). We characterized the reactive oxygen species (ROS) generated in immune peripheral cells of CD patients, as well as their antioxidant enzyme status and the presence of oxidative damage. In addition, mitochondrial function (&Dgr;&ggr;m) was analyzed to detect the possible origin of ROS. Methods: Cells were obtained from patients at the onset of disease, prior to any treatment. Experiments were repeated when patients were in clinical remission. A set of experiments was carried out in a group of CD patients in persistent morphological remission. Controls were healthy volunteers who were not receiving any treatment at the time. The generation of superoxide, hydrogen peroxide (H2O2) and nitric oxide, &Dgr;&ggr;m, superoxide dismutase (SOD) and catalase (CAT) activities, and concentrations of malondyaldehyde (MDA) and 8‐oxo‐deoxyguanosine (8‐oxo‐dG) were measured. Results: SOD activity and H2O2 production were significantly higher during active CD but returned to control levels in remission. &Dgr;&ggr;m was inhibited during active CD and, although it returned to control levels, its recovery took longer than clinical remission. CAT activity was permanently inhibited during CD, independent of the disease activity. MDA and 8‐oxo‐dG were permanently elevated. Conclusions: Oxidative stress during active CD depends on H2O2 production. The inhibition of &Dgr;&ggr;m suggests that this organelle is a source of ROS. CAT is permanently inhibited in CD, the biological significance of which is under study. The persistent oxidative damage detected may have implications for the evolution of the disease. Inflamm Bowel Dis 2010