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Introduction and objectives Telomeres are noncoding regions located at the end of chromosomes and their shortening has been associated with risk factors and cardiovascular disease. The aim of this study was to evaluate the association between ideal cardiovascular health (Life's simple 7) and the odds of having short telomeres in a subsample of participants older than 55 years from the Seguimiento Universidad de Navarra (SUN) study. Methods We included 886 participants older than 55 years (645 men and 241 women). Telomere length was measured using a real-time quantitative polymerase chain reaction. Cardiovascular health score was defined by the American Heart Association as a composite score of 7 key risk factors (smoking status, physical activity, diet, body mass index , blood pressure, total cholesterol, and fasting blood glucose) with 0 to 2 points for each factor. We categorized this score in tertiles as poor (0-9 points), intermediate (10-11 points) and ideal (12-14 points). The odds of having short telomeres was defined as telomere length below the 20th percentile. Results Individuals with higher ideal cardiovascular health had a lower prevalence of having short telomeres (adjusted OR, 0.60; 95%CI, 0.34-1.05; P trend = .052). This association was statistically significant in men (adjusted OR, 0.37; 95%CI, 0.17-0.83; P trend = .025) but not in women. Conclusions An inverse association between cardiovascular health score and short telomeres was found especially for men older than 55 years in the SUN population. The SUN project was registered at ClinicalTrials.gov (Identifier: NCT02669602).

Resumen Introduccion y objetivos Los telomeros son regiones no codificantes localizadas al final de los cromosomas de celulas eucariotas, y su acortamiento se ha visto relacionado con la enfermedad cardiovascular y sus factores de riesgo. El objetivo de este estudio es evaluar la asociacion entre el indice de salud cardiovascular ideal y el riesgo de telomero corto en una poblacion de sujetos de edad avanzada de la cohorte Seguimiento Universidad de Navarra (SUN). Metodos Se valoro a 886 adultos mayores de 55 anos (645 varones y 241 mujeres). La longitud telomerica se midio utilizando qPCR (quantitative protein chain reaction) en tiempo real y reaccion unica. El indice de salud cardiovascular «Life's simple 7» se definio segun la American Heart Association mediante la puntuacion de 7 items con valores de 0 a 2 para cada uno: tabaquismo, actividad fisica, dieta, indice de masa corporal, presion arterial, colesterol total y glucosa en sangre. La maxima puntuacion del indice corresponde a 14 puntos. Se categorizo en terciles: pobre (0-9 puntos), intermedio (10-11 puntos) e ideal (12-14 puntos). El riesgo de telomero corto se definio como una longitud telomerica por debajo del percentil 20. Resultados Sujetos con altos valores en el indice de salud cardiovascular ideal tenian menos riesgo de telomero corto (OR ajustada = 0,60; IC95%, 0,34-1,05; p de tendencia lineal = 0,052). Esta asociacion fue significativa en varones (OR ajustada = 0,37; IC95%, 0,17-0,83; p de tendencia lineal = 0,025), pero no en mujeres. Conclusiones En varones mayores de 55 anos, existe una asociacion inversa entre el indice de salud cardiovascular y el riesgo de tener telomeros cortos. El proyecto SUN fue registrado en ClinicalTrials.gov (Identificador: NCT02669602).

Background and aims In lifestyle intervention studies, we demonstrated that changes in telomere length (TL) were associated with changes in anthropometric indices. Therefore, our new hypothesis is that TL could be a predictor of changes in anthropometric or metabolic measures in children with abdominal obesity. The aim of the study was to evaluate the association between anthropometric and biochemical measurements with TL before and after an 8-week lifestyle intervention in children with abdominal obesity (7–16 years old). Methods and results We assessed anthropometric and biochemical outcomes at baseline and after 8-week lifestyle intervention in 106 children with abdominal obesity (11.30 ± 2.49 years old, 63% girls). TL was measured by monochrome multiplex real-time quantitative PCR. After the lifestyle intervention, anthropometric parameters and glucose metabolism indicators significantly improved in the participants. TL did not change after the intervention in participants. Significant negative correlations between baseline TL and anthropometric measures (BMI, body weight and waist circumference) were observed. Furthermore, baseline TL was a predictor for changes in blood glucose levels after the lifestyle intervention. Conclusions An inverse correlation between TL and obesity traits was observed in children with abdominal obesity. Interestingly, we found that baseline TL could predict changes in blood glucose levels. Clinical trial NCT03147261. Registered 10 May 2017.

Blood-Brain barrier (BBB) disruption is a hallmark of central nervous system (CNS) dysfunction, and oxidative stress is one of the molecular mechanisms that may underlie this process. NADPH oxidases (NOX) are involved in oxidative stress-mediated vascular dysfunction and participate in the pathophysiology of its target organs. The NADPH oxidase 5 (NOX5) isoform is absent in rodents, and although little is known about the role it may play in disrupting the BBB, it has recently been implicated in experimental stroke. Our aim was to investigate the role of NADPH oxidase 5 (NOX5) in promoting vascular alterations and to identify its impact on the cognitive status of aged mice. No differences were detected in the arterial blood pressure or body weight between knock-in mice expressing endothelial NOX5 and the control mice. The Morris water maze test showed memory impairments in the aged knock-in mice expressing NOX5 compared with their control littermates. For assessing the BBB integrity, we studied the protein expression of two tight junction (TJ) proteins: Zonula occludens-1 (ZO-1) and occludin. Compared to the control animals, Aged NOX5 mice exhibited reduced levels of both proteins, demonstrating an alteration of the BBB integrity. Our data indicate that vascular NOX5 may favor behavioral changes with aging through oxidative stress-mediated BBB breakdown.

The purpose of this study was to assess the effect of physical activity (PA) changes, measured by accelerometry, on telomere length (TL) in pediatric patients with abdominal obesity after a lifestyle intervention. One hundred and twenty-one children (7–16 years old) with abdominal obesity were randomized to the intervention (a moderately hypocaloric Mediterranean diet) or the usual care group (standard pediatric recommendations) for 22 months (a 2 month intensive phase and a subsequent 20 month follow-up). Both groups were encouraged to accumulate an extra 200 min/week of PA. TL was measured by MMqPCR. Data were analyzed in 102 subjects after 2 months and 64 subjects at the first 10 months of follow-up. Light PA level decreased in both groups after 12 months of intervention. At month 2, moderate to vigorous PA (MVPA) increased in the intervention group (+5.4 min/day, p = 0.035) and so did sedentary time in the usual care group (+49.7 min/day, p = 0.010). TL changes were positively associated (p < 0.050) with metabolic equivalents (METs), MVPA level, and number of steps, and were inversely associated with sedentary and light PA levels in the intervention group after the intensive phase. In conclusion, favourable changes in PA levels in the intensive phase of a lifestyle intervention could contribute to TL maintenance in a pediatric population with abdominal obesity. Novelty Changes in physical activity levels had a direct effect on telomere length, a biomarker of cellular aging and oxidative stress. PA advice based on The American College of Sports Medicine included in this intervention is easy to implement in primary care.

Objective: Aortic valve (AV) calcification plays an important role in the progression of aortic stenosis (AS). MMP-10 (matrix metalloproteinase-10 or stromelysin-2) is involved in vascular calcification in atherosclerosis. We hypothesize that MMP-10 may play a pathophysiological role in calcific AS. Approach and Results: Blood samples (n=112 AS and n=349 controls) and AVs (n=88) from patients undergoing valve replacement were analyzed. Circulating MMP-10 was higher in patients with AS compared with controls ( P S =0.451; P S =0.5045, P S =0.5003, P P Conclusions: MMP-10, which is overexpressed in aortic valve from patients with AS, seems to play a central role in calcification in AS through Akt phosphorylation. MMP-10 could be a new therapeutic target for delaying the progression of aortic valve calcification in AS.

Telomere shortening and oxidative stress are involved in the pathogenesis of atherosclerosis. Different studies have shown that phagocytic NADPH oxidase is associated with this disease. This study aimed to investigate the association between phagocytic NADPH oxidase and telomere shortening in human atherosclerosis. To assess this potential association, telomere length and phagocytic NADPH oxidase activity were determined by PCR and chemiluminescence, respectively, in a population of asymptomatic subjects free of overt clinical atherosclerosis. We also measured serum 8-hydroxy-2-deoxyguanosine (8-OHdG) levels (an index of oxidative stress) and carotid intima-media thickness (IMT), a surrogate marker of atherosclerosis. After adjusting them for age and sex, telomere length inversely correlated (p &lt, 0.05) with NADPH oxidase-mediated superoxide production, with 8-OHdG values, and with carotid IMT. Interestingly, the asymptomatic subjects with plaques have a lower telomere length (p &lt, 0.05), and higher values of plasma 8-OHdG and superoxide production (p &lt, 0.05). These data were confirmed in a second population in which patients with coronary artery disease showed lower telomere length and higher 8-OHdG and superoxide production than the asymptomatic subjects. In both studies, NADPH oxidase-dependent superoxide production in phagocytic cells was only due to the specific expression of the Nox2 isoform. In conclusion, these findings suggest that phagocytic NADPH oxidase may be involved in oxidative stress-mediated telomere shortening, and that this axis may be critically involved in human atherosclerosis.

Background Telomere length (TL) is a marker of biological age that may be affected by dietary factors through oxidation and inflammation mechanisms. In addition, ultra-processed food (UPF) consumption has increased worldwide and it has been associated with the risk of developing several diseases. Objectives We aimed to evaluate the association between UPF consumption and the risk of having short telomeres in an elderly population of the Seguimiento Universidad de Navarra (SUN) Project. Methods This is a cross-sectional study of 886 participants (645 men and 241 women) aged 57-91 y recruited from the SUN Project (Spain, 1999-2018). TL was measured from saliva samples by real-time qPCR at baseline and UPF consumption was collected using a validated 136-item FFQ and classified according to the NOVA system. We evaluated the association between consumption of energy-adjusted UPF categorized into quartiles (low, medium-low, medium-high, and high consumption) and the risk of having short telomeres ( Results Those participants with the highest UPF consumption had almost twice the odds of having short telomeres compared with those with the lowest consumption (adjusted OR: 1.82; 95% CI: 1.05, 3.22; P-trend = 0.03). Conclusions A higher consumption of UPF (>3 servings/d) was associated with higher risk of having shorter telomeres in an elderly Spanish population of the SUN Project.This trial was registered at clinicaltrials.gov as NCT02669602.

BACKGROUND Dietary factors seem to influence telomere length. Moreover, associations between changes in adiposity indices and telomere length (TL) have been found in intervention studies. OBJECTIVE We evaluated changes in two diet quality indices and their association with TL in children with abdominal obesity in a 12-month lifestyle intervention. METHODS Eighty-seven participants (7-16 years old) were assigned to the intervention (moderate hypocaloric Mediterranean diet) or usual care group (standard paediatric recommendations) for a 2-month intensive phase and a subsequent 10-month follow-up. Diet quality was assessed using the Diet Quality Index for Adolescents (DQI-A) and the Healthy Lifestyle Diet Index (HLD-I). TL was measured by monochrome multiplex real-time quantitative PCR. The intra-class correlation coefficient for TL was 0.793 (95% CI 0.707, 0.857). RESULTS After a 12-month lifestyle intervention, a significant reduction in BMI-SDS (-0.57 and -0.49 for the intervention and usual care groups, respectively) and fat mass was observed in all subjects without differences between groups. Changes in DQI-A (+12.36% vs +5.53%, P = .005) and HLD-I (+4.43 vs +1.09, P

Shorter telomeres are associated with several age-related diseases, and lifestyle factors could influence this relationship. The aim of this study was to examine associations between salivary telomere length (TL) and diet quality using 5 evidence-based dietary indexes in an elderly (55 years old) Spanish population of the SUN project (n = 886).TL was measured using the quantitative real-time polymerase chain reaction. Age-adjusted TL variable through residuals methods was used for all analysis. Diet quality was assessed by the Prime Diet Quality Score (PDQS), Fat Quality Index (FQI), Mediterranean Diet Adherence Screener (MEDAS), Dietary Approaches to Stop Hypertension (DASH) index and the Alternative Healthy Eating Index (AHEI-2010).TL did differ according to sex, smoking status, and dyslipidemia in elderly subjects of the SUN study. In addition, subjects with dyslipidemia (compared to absence of dyslipidemia) had a significantly higher risk (27% vs. 18%, p = 0.015) of short telomeres (percentile 20th). Interestingly, a lower risk of having short telomeres was observed among participants in the top tertiles of the following diet quality score PDQS, MEDAS and DASH compared to the bottom tertiles in crude and adjusted models. Moreover, FQI and AHEI-2010 scores showed an inverse association with the risk of having short telomeres after adjustment for potential confounders (model adjusted for dyslipidemia interaction, p for trend = 0.025 and 0.021, respectively; and model additionally adjusted for sex and smoking status, p for trend = 0.033 and 0.029, respectively).Adherence to high quality diet is associated to longer salivary TL in our elderly Spanish population of the SUN study.

SummaryBackground Shorter telomeres have been associated with elevated risk for age-related diseases. However, little is known about the biomarker role of telomere length (TL) for predicting inflammation and glucose alterations. Objective The objective of this research is to evaluate the association between TL, inflammatory markers and glucose levels after a 2-month weight-loss programme in obese adolescents. Methods Telomere length was measured using a quantitative polymerase chain reaction in 66 obese adolescents aged 12–17 years (51% men) from the EVASYON programme. The adolescents were genotyped for the polymorphism -174G/C (rs1800795) in the IL-6gene, and anthropometric and biochemical markers as well as inflammatory cytokines were analysed. Results Multiple-adjusted models showed that longer telomeres at baseline were associated with a higher reduction in glucose (B = −4.08, 95% confidence interval: −6.66 to −1.50) and IL-6 (B = −1.03, 95% confidence interval: −2.01 to −0.05) serum levels after 2 months of the weight-loss treatment. The -174G/C polymorphism modulated the association between basal TL and changes in IL-6 (P interaction = 0.029). Thus, subjects with the GG + GC genotype and with longer telomeres showed a higher decrease in IL-6 levels than CC homozygotes. Conclusion Longer telomeres are associated with an improvement in glucose tolerance and inflammation after a weight-loss programme in obese adolescents. Moreover, the -174G/C polymorphism may influence the relationship between TL and IL-6 changes.

Oxidative stress is one of the main mechanisms involved in the pathophysiology of vascular diseases. Among others, oxidative stress promotes endothelial dysfunction, and accelerated ageing and remodelling of vasculature. Lately, NADPH oxidases have been demonstrated to be involved in cardiovascular diseases. NADPH oxidase 5 has emerged as a new player in oxidative stress-mediated endothelial alterations, involved in the pathophysiology of hypertension, diabetes, atherosclerosis, myocardial infarction and stroke. This oxidase seems to mediate its detrimental effects by promoting inflammation. NADPH oxidase 5 has been studied in a lesser extent compared with the other members of the NADPH oxidase family due to its loss in the rodent genome, the main experimental research model. In addition, its potential as a therapeutic target remains unexplored given the lack of specific inhibitors. In this review the latest findings on NADPH oxidase 5 regulation, implications in vascular pathophysiology and therapeutic approaches will be updated.

BACKGROUND AND AIMS: Matrix metalloproteinases (MMPs) have been implicated in atherosclerosis and vascular calcification. Among them, we reported that MMP10 is present in human atheroma, associated with atherosclerosis. However, it remains unclear whether MMP10 is involved in atherogenesis and vascular calcification. METHODS: MMP10 was measured in serum from patients with subclinical atherosclerosis and analyzed in carotid endarterectomies by immunostaining. ApoE-deficient mice (Apoe-/-) were crossed to MMP10-deficient (Mmp10-/-) mice and followed up to 20 months. Plaque area and composition were assessed by histology and immunohistochemistry. Inflammatory markers were measured in atherosclerotic plaques by RT-qPCR, and leukocyte subpopulations were analyzed by flow cytometry. In vitro calcification assays were performed in aortic vascular smooth muscle cells (VSMC). RESULTS: MMP10 serum levels were associated with coronary calcification in subjects with subclinical atherosclerosis. Immunostaining revealed MMP10 expression in human atheromas, spatially associated with calcification areas, and complicated plaques released higher amounts of MMP10 than non-diseased segments. Interestingly, vascular MMP10 expression was confined to the atherosclerotic lesion in Apoe-/- mice, and Apoe-/-Mmp10-/- showed a substantial reduction in atherosclerotic lesion size, macrophage content and plaque calcification. Reduced local and systemic inflammatory markers could be demonstrated in Apoe-/-Mmp10-/- by gene expression and flow cytometry analysis. Calcium phosphate deposition and vascular calcification markers were downregulated in VSMC from Apoe-/-Mmp10-/- mice. CONCLUSIONS: Delayed plaque progression and altered cellular composition in the absence of MMP10 suggests that MMP10 plays a role in atherosclerosis, favoring inflammation, development and complication of the plaque. Funded through the "UTE project CIMA" (University of Navarra), Ministerio de Educacion y Ciencia (SAF2009-12039, SAF2016-79151-R), Ministerio de Sanidad y Consumo (PI14/01152 and PI15/01807), Sociedad Espanola de Cardiologia, Sociedad Espanola de Arteriosclerosis, Red de Investigacion Cardiovascular RIC (RD12/0042/0009), the gs5: Fondation pour la Recherche Medicale and the Fondation de France. Sí

Oxidative stress, a pathological situation created by an imbalance between reactive oxygen species production and antioxidant capacity, is a critical mechanism that underlies the pathophysiology of cardiovascular diseases and is involved in obesity, metabolic syndrome, and diabetes. The nicotinamide adenine dinucleotide phosphate (NADPH) oxidase enzyme family is the main source of reactive oxygen species and its expression and activity is enhanced and plays a critical role in the onset and/or development of cardiovascular diseases. The phagocytic NADPH oxidase, a member of the NADPH oxidase family present in lymphocytes, monocytes, and neutrophils, has been implicated in the pathophysiology of these vascular and metabolic disorders, by promoting detrimental prooxidant processes that could increase the morbidity and mortality in these diseases. A present challenge consists of the development of novel NADPH oxidase-selective drugs that could prevent and/or treat cardiovascular and metabolic diseases.

Vascular calcification is a common feature in atherosclerosis and associates with cardiovascular events. Oxidative stress may be involved in the pathogenesis of vascular calcification. Previous studies have shown that the phagocytic NADPH oxidase is associated with atherosclerosis. The objective of the present study was to investigate the association between phagocytic NADPH oxidase-mediated superoxide production and coronary artery calcium (CAC). NADPH oxidase-mediated superoxide production was determined by chemiluminescence and CAC by computed tomography in 159 asymptomatic men free of overt clinical atherosclerosis. Multivariate linear regression analyses were used to assess the relationship between CAC and NADPH oxidase-mediated superoxide production. Compared with individuals in the lowest score of CAC (= 0 Agatston units), those in the upper score (400 Agatston units) showed higher superoxide production (p 0.05). In correlation analysis, superoxide production positively (p 0.01) correlated with CAC, which in multivariate analysis remained significant after adjusting for age, HDL-cholesterol, triglycerides, body mass index, smoking, arterial hypertension and diabetes mellitus. In conclusion, in a population of men without clinically overt atherosclerotic disease, increased NADPH oxidase-mediated superoxide production associated with enhanced CAC. Albeit descriptive, these findings suggest a potential involvement of phagocytic NADPH oxidase-mediated oxidative stress in CAC.

Summary Background & aims A healthy lifestyle has been associated with longer telomeres, but whether Mediterranean Diet (MeDiet) affect telomere length (TL) has not been fully elucidated yet. Our aim was to assess the relationship between MeDiet and TL in high cardiovascular risk subjects in the context of a randomized nutritional intervention trial. Methods We assessed 520 participants (55–80 years, 55% women) from the PREDIMED-NAVARRA trial. Leukocyte TL was measured by qPCR at baseline and after 5 years of a dietary intervention program where subjects were randomly assigned to a low-fat control diet or to two MeDiets, one supplemented with extra virgin olive oil (MeDiet-EVOO) and the other with mixed nuts (MeDiet-nuts). A validated 14-item questionnaire was used to appraise baseline adherence of participants to the MeDiet. Results Better adherence to MeDiet (as appraised by the 14-item score) was associated with longer basal telomeres in women in the baseline cross-sectional analysis, whereas the opposite was observed in men (P interaction = 0.036). Female subjects who scored 10 points had longer basal telomeres (0.27, 95% CI: 0.03–0.52) than women scoring ≤6 points at the beginning of the study (−0.46, 95% CI: −0.85 to −0.7) (P = 0.003). However, allocation to the MeDiet-nuts group (−0.24, 95% CI: −0.38 to −0.01) was associated with a higher risk of telomere shortening after 5 years of intervention, whereas no differences were found for the MeDiet-EVOO group (0.14, 95% CI: 0.02–0.27), in comparison with the Control group (0.07, 95% CI: −0.08 to 0.23) (P = 0.003 and P = 0.537, respectively). Conclusion A greater baseline adherence to a Mediterranean dietary pattern was associated with longer telomeres only in women. No beneficial effect of the intervention with the MeDiet for the prevention of telomere shortening in comparison with a low-fat diet was observed.

Background: Telomeres are nucleoprotein structures that protect the ends of eukaryote chromosomes. Shorter telomere length (TL) is associated with some age-related human disorders, but its relationship with obesity or adiposity parameters remains unclear. Objective: The aim of this study was to assess the relationship between TL and changes in adiposity indices after a 5-year nutritional intervention. Design and subjects: TL was measured by quantitative real-time PCR in 521 subjects (55-80 years, 55% women). Participants were randomly selected from the PREDIMED-NAVARRA centre after they completed a 5-year intervention programme. Anthropometric parameters were directly measured by trained personnel at baseline and on a yearly basis thereafter. TL at baseline and changes in TL after a 5-year intervention were assessed. Results: Higher baseline TL significantly predicted a greater decrease in body weight (B=−1.09 kg, 95% confidence interval (CI): −2.01 to −0.16), body mass index (BMI) (B=−0.47 kg m−2, 95% CI: −0.83 to −0.11), waist circumference (B=−1.15 cm, 95% CI: −2.28 to −0.01) and waist to height ratio (B=−0.008, 95% CI: −0.010 to −0.001) in multiple-adjusted models. In addition, changes in TL during the 5-year intervention were inversely associated with changes in the four anthropometric variables. The reduction in adiposity indices during the intervention, associated with increasing TL, was even higher among subjects with the longest telomeres at baseline. Logistic regression analysis showed that the risk of remaining obese after 5 years was lower in those participants who initially had the longest telomeres and increased their TL after intervention (odds ratio=0.27, 95% CI: 0.03-2.03). Conclusions: Our research suggests that TL is inversely associated with changes in obesity parameters. The assessment of TL can provide further insights for biological pathways leading to adiposity. We show for the first time an improvement of obesity indices when an increase in TL is observed after a 5-year Mediterranean diet intervention.

Introduction and objectives: Advanced kidney disease is a major health problem due to its association with high cardiovascular morbidity and mortality. Early recognition of advanced kidney disease is the mainstay to avoid its progression. Since metabolic syndrome and insulin resistance are risk factors for both cardiovascular and advanced kidney disease, we investigated the relationship of early kidney disease (EKD) with metabolic syndrome and insulin resistance, and their association with surrogate markers of arteriosclerosis. Methods: We studied 1,498 subjects. Insulin resistance was defined as HOMA 3.7 mmol (mU)/L 2 and EKD as stages 1 and 2 of the NKF-KDOQI. Carotid intima-media thickness was used as a surrogate marker of arteriosclerosis. Results: The presence of one trait of metabolic syndrome was associated with an odds ratio (OR) for EKD of 2.3 (95% confidence interval [CI], 1.18-4.48) that increased to 6.72 (95% CI, 3.56-13.69) in subjects with the syndrome. All the traits of the syndrome except low level of high-density lipoproteins showed an increased OR for EKD. Increasing HOMA was also directly correlated with higher OR for EKD, being as high as 3.89 (95% CI, 1.99-7.59) for subjects in the fourth quartile. Subjects with the syndrome plus EKD showed an increased intima-media thickness compared with those without kidney disease. Conclusions: Insulin resistance and all metabolic syndrome traits except low level of high-density lipoproteins were significantly associated with an increased OR for EKD. Both metabolic syndrome and EKD were independently and additively related to the presence of surrogate markers of arteriosclerosis.

OBJECTIVES: Hyperleptinemia and oxidative stress play a major role in the development of cardiovascular diseases in obesity. This study aimed to investigate whether there is a relationship between plasma levels of leptin and phagocytic nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity, and its potential relevance in the vascular remodeling in obese patients. METHODS: The study was performed in 164 obese and 94 normal-weight individuals (controls). NADPH oxidase activity was evaluated by luminescence in phagocytic cells. Levels of leptin were quantified by ELISA in plasma samples. Carotid intima-media thickness (cIMT) was measured by ultrasonography. In addition, we performed in-vitro experiments in human peripheral blood mononuclear cells and murine macrophages. RESULTS: Phagocytic NADPH oxidase activity and leptin levels were enhanced (P < 0.05) in obese patients compared with controls. NADPH oxidase activity positively correlated with leptin in obese patients. This association remained significant in a multivariate analysis. cIMT was higher (P < 0.05) in obese patients compared with controls. In addition, cIMT also correlated positively with leptin and NADPH oxidase activity in obese patients. In-vitro studies showed that leptin induced NADPH oxidase activation. Inhibition of the leptin-induced NADPH oxidase activity by wortmannin and bisindolyl maleimide suggested a direct involvement of the phosphatidylinositol 3-kinase and protein kinase C pathways, respectively. Finally, leptin-induced NADPH oxidase activation promoted macrophage proliferation. CONCLUSIONS: These findings show that phagocytic NADPH oxidase activity is increased in obesity and is related to preclinical atherosclerosis in this condition. We also suggest that hyperleptinemia may contribute to phagocytic NADPH oxidase overactivity in obesity.

AIMS: The purpose of this study was to analyse the expression of CD74 in human atherosclerotic plaques and peripheral blood mononuclear cells (PBMC) as well as its potential participation in proinflammatory responses in cultured human vascular smooth muscle cells (VSMC). METHODS AND RESULTS: CD74 expression was analysed in human atherosclerotic plaques (immunohistochemistry), PBMC (real-time PCR), and human aortic VSMC (real-time PCR and western blotting). Nuclear factor-kappaB (NF-kappaB) activation was assessed by southwestern histochemistry and electrophoretic mobility shift assay. Monocyte chemoattractant protein-1 (MCP-1) levels were studied by both real-time PCR and enzyme-linked immunosorbent assay. CD74 immunostaining was increased in the inflammatory vs. the fibrous region of atherosclerotic plaques (n = 70, 18.2 +/- 1.3 vs. 7.8 +/- 0.6% positive staining/mm2, P < 0.001). CD74 colocalized with the transcription factor NF-kappaB in both VSMC and macrophages. In cultured VSMC, CD74 expression was induced by interferon gamma (IFNgamma). Incubation with an agonistic anti-CD74 antibody or with IFNgamma elicited MCP-1 expression, which was prevented by AKT and gamma-secretase inhibitors. Moreover, CD74 small-interfering RNA decreased NF-kappaB activation and MCP-1 production induced by IFNgamma in VSMC. Finally, CD74 mRNA levels in PBMC from patients with carotid stenosis were higher than in healthy subjects (n = 20, 3 +/- 0.5 vs. 2 +/- 0.5 AU, P < 0.001). Additionally, a linear trend between CD74 mRNA expression tertiles and intima-media thickness (IMT) was observed in PBMC from asymptomatic subjects (n = 185, P < 0.001). CONCLUSION: CD74 levels are increased in plaques and PBMC from patients with carotid stenosis and are associated with IMT in subjects free from clinical cardiovascular diseases. CD74 could be a novel therapeutic target to decrease the inflammatory response in atherosclerosis.

Background Galectin‐3 (Gal‐3) participates in different mechanisms involved in atherothrombosis, such as inflammation, proliferation, or macrophage chemotaxis. Thus, there have been committed intensive efforts to elucidate the function of Gal‐3 in cardiovascular ( CV ) diseases. The role of Gal‐3 as a circulating biomarker has been demonstrated in patients with heart failure, but its importance as a biomarker in atherothrombosis is still unknown. Methods and Results Because Gal‐3 is involved in monocyte‐to‐macrophage transition, we used fresh isolated monocytes and the in vitro model of macrophage differentiation of THP ‐1 cells stimulated with phorbol myristate acetate ( PMA) . Gal‐3 release is increased by PMA in human monocytes and macrophages, a process involving exosomes and regulated by reactive oxygen species/ NADPH oxidase activity. In asymptomatic subjects (n=199), Gal‐3 plasma levels are correlated with NADPH oxidase activity in peripheral blood mononuclear cells ( r =0.476; P r =0.438; P mL ; P P Conclusions Gal‐3 extracellular levels could reflect key underlying mechanisms involved in atherosclerosis etiology, development, and plaque rupture, such as inflammation, infiltration of circulating cells and oxidative stress. Moreover, circulating Gal‐3 concentrations are associated with clinical outcomes in patients with atherothrombosis.

Left ventricular hypertrophy (LVH) is an independent marker of mortality in hypertension. Although the mechanisms contributing to LVH are complex, inflammation and oxidative stress may favor its development. We analyzed the association of the phagocytic NADPH oxidase–mediated superoxide anion release and LVH in patients with essential hypertension and the role of cardiotrophin-1 (CT-1) and interleukin-6 (IL-6), cytokines implicated in cardiac growth. Blood pressure, echocardiography data, and serum CT-1 and IL-6 levels were obtained in 140 subjects: 18 normotensives without LVH, 42 hypertensives without LVH, and 80 hypertensives with LVH. The NADPH oxidase–dependent superoxide production was assessed by chemiluminescence in peripheral blood mononuclear cells. Peripheral blood mononuclear cells were stimulated with CT-1 in vitro. Superoxide anion production by peripheral blood mononuclear cells associated with LVH and correlated with the left ventricular mass index. Serum CT-1 and IL-6 levels, which associated with the left ventricular mass index, correlated with superoxide production. Serum CT-1 and IL-6 levels were correlated. CT-1 stimulated NADPH oxidase superoxide production in peripheral blood mononuclear cells, which resulted in an increased release of IL-6. Our results show that superoxide anion production by the phagocytic NADPH oxidase associates with hypertensive heart disease, being significantly enhanced in hypertensive patients with LVH. This may be attributable to the activation of the NADPH oxidase by CT-1 and the subsequent release of IL-6. The phagocytic NADPH oxidase may be a therapeutic target in hypertensive heart disease

Hypertension is associated with increased plasma inflammatory markers such as cytokines and increased vascular cyclooxygenase-2 (COX-2) expression. The ability of peroxisome proliferator-activated receptor-γ (PPARγ) agonists to reduce oxidative stress seems to contribute to their anti-inflammatory properties. This study analyzes the effect of pioglitazone, a PPARγ agonist, on interleukin-1β-induced COX-2 expression and the role of reactive oxygen species (ROS) on this effect.Vascular smooth muscle cells from hypertensive rats stimulated with interleukin-1β (10 ng/ml, 24 h) were used. Interleukin-1β increased: 1) COX-2 protein and mRNA levels; 2) protein and mRNA levels of the NADPH oxidase subunit NOX-1, NADPH oxidase activity and ROS production; and 3) phosphorylation of inhibitor of nuclear factor kappa B (IκB) kinase (IKK) nuclear expression of the p65 nuclear factor kappa B (NF-κB) subunit and cell proliferation, all of which were reduced by apocynin (30 μmol/l). Interleukin-1β-induced COX-2 expression was reduced by apocynin, tempol (10 μmol/l), catalase (1000 U/ml) and lactacystin (5 μmol/l). Moreover, H2O2 (50 μmol/l, 90 min) induced COX-2 expression, which was reduced by lactacystin. Pioglitazone (10 μmol/l) reduced the effects of interleukin-1β on: 1) COX-2 protein and mRNA levels; 2) NOX-1 protein and mRNA levels, NADPH oxidase activity and ROS production; and 3) p-IKK, p65 expressions and cell proliferation. Pioglitazone also reduced the H2O2-induced COX-2 expression and increased Cu/Zn and Mn-superoxide dismutase protein expression. PPARγ small interfering RNA (5 nmol/l) further increased interleukin-1β-induced COX-2 and NOX-1 mRNA levels. In addition, pioglitazone increased the interleukin-1β-induced PPARγ mRNA levels.PPARγ activation with pioglitazone reduces interleukin-1β-induced COX-2 expression by interference with the redox-sensitive transcription factor NF-κB.

Advanced kidney disease is a major health problem due to its association with high cardiovascular morbidity and mortality. Early recognition of advanced kidney disease is the mainstay to avoid its progression. Since metabolic syndrome and insulin resistance are risk factors for both cardiovascular and advanced kidney disease, we investigated the relationship of early kidney disease (EKD) with metabolic syndrome and insulin resistance, and their association with surrogate markers of arteriosclerosis. METHODS: We studied 1498 subjects. Insulin resistance was defined as HOMA >/=3.7 mmol (muU)/L(2) and EKD as stages 1 and 2 of the NKF-KDOQI. Carotid intima-media thickness was used as a surrogate marker of arteriosclerosis. RESULTS: The presence of one trait of metabolic syndrome was associated with an odds ratio (OR) for EKD of 2.3 (95% confidence interval [CI], 1.18-4.48) that increased to 6.72 (95% CI, 3.56-13.69) in subjects with the syndrome. All the traits of the syndrome except low level of high-density lipoproteins showed an increased OR for EKD. Increasing HOMA was also directly correlated with higher OR for EKD, being as high as 3.89 (95% CI, 1.99-7.59) for subjects in the fourth quartile. Subjects with the syndrome plus EKD showed an increased intima-media thickness compared with those without kidney disease. CONCLUSIONS: Insulin resistance and all metabolic syndrome traits except low level of high-density lipoproteins were significantly associated with an increased OR for EKD. Both metabolic syndrome and EKD were independently and additively related to the presence of surrogate markers of arteriosclerosis.

Objective To analyze whether genetic variants of PPARA are associated with the development of stage C heart failure. Methods We analyzed the distribution of the rs1800206, rs4253778 and rs135551 polymorphisms in genomic DNA extracted from peripheral blood cells of 534 patients in different heart failure stages and 63 healthy individuals. The mRNA expression of the peroxisome proliferator-activated receptor (PPAR)α target genes long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) and medium-chain acyl-CoA dehydrogenase (MCAD) was measured in myocardial biopsies of a subgroup of stage B and C patients. Functional studies were performed in HL-1 cardiomyocytes. Results The V162 allele of the rs1800206 polymorphism was more frequent in stage C patients than in stage A and B patients and healthy individuals. Patients with the V162 allele exhibited decreased myocardial LCHAD and MCAD mRNA expression as compared to L162 homozygote patients. In addition, stage C patients exhibited lower myocardial LCHAD and MCAD mRNA expression than stage B patients. Cardiomyocytes transfected with the V162 allele presented decreased PPARα transcriptional activity, LCHAD mRNA expression and ATP production compared to cardiomyocytes transfected with the L162 variant. Conclusions These findings suggest that the V162 allele of the human PPARA gene can be a new risk factor in the development of stage C heart failure, likely via depressed cardiac PPARα activity.

Objective Experimental and clinical evidence supports a role of cardiotrophin-1 (CT-1) in the development of hypertensive left ventricular hypertrophy (LVH). The goal of this study was to investigate the relationship between human CT-1 genetic background and LVH in essential hypertension. Methods A total of 900 individuals were genotyped for the 1742(C/G) polymorphism of the human CT-1 gene. Serum CT-1 levels were assessed by ELISA in 681 individuals. Left ventricular parameters were determined by two-dimensional echocardiography in 297 individuals. Results The prevalence of the GG genotype of the 1742(C/G) polymorphism was reduced in essential hypertension (8.4% in normotensive individuals, 4.9% in hypertensive patients, P = 0.046 versus CC/CG individuals) and in LVH (11.5% in nonhypertrophic normotensive individuals, 12.2% in nonhypertrophic hypertensive patients, 2.6% in hypertensive patients with LVH, P = 0.008 versus CC/CG individuals). Apart from this, GG individuals presented lower serum concentration of CT-1 (GG, 147.1 ± 10.5 fmol/ml; CC/CG, 187.1 ± 4.8 fmol/ml; P = 0.036) and left ventricular mass index (GG, 91 ± 6 g/m; CC/CG, 119 ± 3 g/m; P = 0.002). Multivariate analyses showed that the 1742(C/G) polymorphism was a significant determinant of both left ventricular mass index and serum CT-1, after adjusting for confounding factors. Finally, in-vitro studies supported the functionality of the 1742(C/G) polymorphism. Conclusion Our results indicate that the 1742(C/G) polymorphism of the human CT-1 gene is associated with LVH in hypertension and that the GG genotype may have a protective role. It is suggested that CT-1 is one of the mediators of this association.

Oxidative stress plays a key role in the pathophysiology of several major cardiovascular diseases, including atherosclerosis, hypertension, heart failure, stroke, and diabetes. Reactive oxygen species (ROS) induce cardiovascular alterations by modulating cell contraction/dilatation, migration, growth/apoptosis, and extracellular matrix protein turnover, which contribute to vascular and cardiac remodeling. Of the several sources of ROS within the cardiovascular system, the family of the multisubunit NADPH oxidases appears to be a predominant contributor generating superoxide anions. Recent data suggest a significant role of the genetic background in NADPH oxidase regulation. Common genetic polymorphisms within CYBA, the gene that encodes the p22phox subunit of the NADPH oxidase, have been characterized in the context of cardiovascular diseases. This chapter aims to present the current state of research into these polymorphisms in their relationship to cardiovascular diseases.

OBJECTIVE: Metabolic syndrome (MetS) is associated with insulin resistance and increases the cardiovascular risk. Oxidative stress constitutes a potential mechanism that links insulin resistance and cardiovascular disease. The aim of this study was to analyze the relationship of NADPH oxidase activation with insulin resistance, and the effect of this interaction on the cardiovascular risk in MetS patients. METHODS: NADPH oxidase-dependent superoxide production and expression was evaluated by luminescence and western blot, respectively, in peripheral blood mononuclear cells obtained from 125 patients with MetS. Insulin resistance was defined by the homeostasis model assessment index. Matrix metalloproteinase-9 was quantified by enzyme-linked immunosorbent assay in plasma samples. To ascertain the mechanisms involved in vivo, we performed in-vitro experiments in cultured macrophages. RESULTS: Fifty-six percent of patients with MetS showed insulin resistance. Plasma matrix metalloproteinase-9 levels were higher (P < 0.05) in insulin-resistant patients than in patients with insulin sensitivity. NADPH oxidase-dependent superoxide production was augmented (P < 0.05) in insulin-resistant patients with respect to insulin-sensitive patients. The interaction between insulin resistance and abnormally high NADPH oxidase-mediated superoxide production was associated with the highest matrix metalloproteinase-9 values. Increased NADPH oxidase-dependent superoxide production was significantly associated with higher NADPH oxidase p22phox expression in insulin-resistant than in insulin-sensitive patients. Interestingly, insulin upregulated p22phox in peripheral blood mononuclear cells and in murine macrophages. CONCLUSION: Insulin resistance is associated with phagocytic NADPH oxidase activation. This association results in the highest cardiovascular risk in MetS patients.

Endothelial dysfunction is a marker of atherosclerosis and contributes to the atherogenic process and the development of atherothrombotic complications. Oxidative stress has been implicated in the development of endothelial dysfunction through alterations of the nitric oxide metabolism. A number of evidence suggests a role for phagocytic-cell-mediated oxidative stress in diminished nitric oxide availability that is present in patients with atherosclerotic risk factors such as arterial hypertension. Thus, the combination of an excessive production of reactive oxygen species, namely superoxide anion, with an impaired antioxidant defense capacity leading to oxidative stress may facilitate the development and progression of atherosclerosis. Findings from recent clinical studies suggest that this mechanism can be operative in patients with cerebrovascular disease. This view may increase our capabilities to understand the pathophysiology of cerebrovascular disease, as well as to stimulate the design of new therapeutic strategies aimed to prevent and control the atherosclerotic process in patients presenting this condition.

OBJECTIVES Cardiotrophin-1 (CT-1) induces hypertrophic growth and contractile dysfunction in cardiomyocytes. This cross-sectional study was aimed to analyze CT-1 associations with echocardiographically assessed left ventricular systolic properties taking into account the influence of left ventricular growth [i.e. left ventricular hypertrophy (LVH) and inappropriate left ventricular mass (iLVM)] in asymptomatic hypertensive patients. METHODS Serum CT-1 was measured by ELISA in 278 asymptomatic hypertensive patients with a left ventricular ejection fraction more than 50% and in 25 age and sex-matched normotensive patients. RESULTS Serum CT-1 was increased in hypertensive patients as compared to normotensive patients. CT-1 was directly correlated with parameters of left ventricular mass (LVM) and inversely correlated with parameters assessing myocardial systolic function and left ventricular chamber contractility in hypertensive patients, these associations being independent of a number of potential confounding factors. Interestingly, the associations of CT-1 with myocardial systolic function were independent of LVM even in patients with LVH or iLVM. In addition, there was a significant increment of serum CT-1 in hypertensive patients with LVH or iLVM, especially in those in whom LVH or iLVM were accompanied by impaired myocardial systolic function, as compared to the remaining hypertensive patients and normotensive patients. Plasma amino-terminal pro-brain natriuretic peptide was not correlated with any of the assessed left ventricular systolic parameters in either group of patients. CONCLUSION These findings suggest that serum CT-1 is associated with myocardial systolic dysfunction in asymptomatic hypertensive patients, independently of LVM, even in those patients with pathologic left ventricular growth.

Objective Leptin attenuates the angiotensin II-induced increase of cytosolic calcium ([Ca 2+ ] i ) and vasoconstriction in the aorta of normotensive Wistar rats. To determine whether these effects may be altered in hypertension, we assessed the effect of leptin on angiotensin II-induced vascular response in the aorta of 10-week-old spontaneously hypertensive rats (SHR). Methods Contractile responses to angiotensin II (100 nmol/l) in the presence of different concentrations of leptin (0.1,1, 10, 100 nmol/l) were evaluated in isolated aortic rings by the organ bath system. [Ca 2+ ] i was measured in vascular smooth muscle cells (VSMCs) using Fura-2 fluorescence. The expression of the short (OB-Ra) and long (OB-Rb) isoforms of the leptin receptor in VSMCs was evaluated by real-time reverse transcriptase-polymerase chain reaction and western-blot analysis. Results Circulating leptin concentrations were increased in SHR. Serum metabolic parameters, including glucose, insulin, total cholesterol and triglyceride levels, were also elevated in SHR. Leptin did not modify the angiotensin II-induced vasoconstriction in SHR either in intact or endothelium-denuded aortic rings. In addition, leptin was not able either to diminish the angiotensin II-induced the peak rise of [Ca 2+ ] i or to accelerate the recovery rate to basal calcium levels in VSMCs from SHR. However, OB-Ra and OB-Rb mRNA and protein expression were increased in SHR VSMCs. Conclusions The lack of effect of leptin on angiotensin II-induced contraction in the aorta of SHR is due to an impaired handling of [Ca 2+ ] i in VSMCs. Hyperleptinemia and overexpression of OB-R in VSMCs could be compensatory mechanisms against VSMC leptin resistance in genetically hypertensive rats.

OBJECTIVE: Oxidative stress is implicated in hypertension. The reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidases are the main source of superoxide in phagocytic and vascular cells. The C242T polymorphism of CYBA, the human gene that encodes p22phox, has been found to be functionally associated with vascular NADPH oxidase activity in atherosclerotic patients. We investigated the association of the C242T polymorphism with hypertension and its potential impact on NADPH oxidase activity. We also analysed the interaction of C242T polymorphism with the -930A/G CYBA variant. DESIGN: Case-control study in a random sample of 623 subjects (326 hypertensive patients and 297 normotensive controls) from the general population. METHODS: CYBA polymorphisms were determined by restriction fragment length polymorphism (RFLP) or allelic discrimination. NADPH oxidase activity and p22phox expression were quantified in phagocytic cells by chemiluminescence and by northern and western blots, respectively. RESULTS: The prevalence of the CC genotype and the C allele frequency were significantly higher (P < 0.05) in hypertensives than in normotensives. CC genotype remained associated with hypertension after adjusting for potential confounders in a logistic regression analysis. Increased phagocytic NADPH oxidase activity was observed in CC hypertensives compared with CT and TT hypertensives (P < 0.05). Enhanced plasma levels of von Willebrand factor were found in CC hypertensives compared with TT hypertensives (P < 0.05). The C242T polymorphism was not in linkage disequilibrium with the -930A/G CYBA promoter variation, which also associates with hypertension. CONCLUSION: The C242T CYBA polymorphism is associated with essential hypertension. Furthermore, hypertensives carrying the CC genotype of this polymorphism exhibit features of NADPH oxidase-mediated oxidative stress and endothelial damage.

Background. Oxidative stress has been implicated in the pathogenesis of atherosclerosis that develops in patients with advanced chronic kidney disease (CKD). This study was designed to investigate whether a relationship exists between phagocytic nicotinamide adenine dinucleotide phosphate (NADPH) oxidase–dependent superoxide anion (•O2 −) production and subclinical atherosclerosis in patients with early CKD. Methods. Superoxide production was assayed by chemiluminescence under baseline and stimulated conditions on mononuclear cells obtained from asymptomatic patients with stage 1 to 2 CKD (N = 22) and healthy controls (N = 21). Ultrasonographic determination of carotid intima-media thickness (IMT) was used to assess the presence of atherosclerosis. Results. Although there were no differences in baseline •O2 − production between controls and patients, the •O2 − production in phorbol myristate acetate–stimulated mononuclear cells was increased (P < 0.05) in patients compared with controls. The phorbol myristate acetate–induced •O2 − production was completely abolished by apocynin, a specific inhibitor of NADPH oxidase. A direct correlation (r = 0.441, P < 0.05) was found between plasma insulin levels and NADPH oxidase–mediated •O2 − production in patients. Carotid IMT was higher (P < 0.005) in patients than in controls. CarotidIMTvalues above the upper normal limit in controls were found in 70% and 40% of patients with increased or normal NADPH oxidase–mediated •O2 − production, respectively. Conclusion. Generation of •O2 − that is mainly dependent on NADPH oxidase is abnormally enhanced in patients with early CKD. It is suggested that this alteration could be related to the development of subclinical atherosclerosis in these patients.

Objective— Oxidative stress plays a critical role in the pathogenesis of atherosclerosis. The NADPH oxidase constitutes the main source of superoxide in phagocytic and vascular cells. This study aimed to investigate the levels of NADPH oxidase–mediated superoxide production in phagocytic cells and the association between phagocytic superoxide production and carotid intima-media thickness (IMT), a surrogate marker of asymptomatic atherosclerosis. Methods and Results— NADPH oxidase–mediated superoxide production was determined by a chemiluminescence assay using lucigenin and associated with IMT for 184 asymptomatic subjects free of overt clinical atherosclerotic disease. Compared with individuals in the lowest tertile of superoxide production, those in the upper tertile (>20 counts/sec) showed significantly higher IMT ( P P Conclusions— In a population sample of adults without clinically overt atherosclerotic disease, increased NADPH oxidase activity was associated with enhanced carotid IMT, suggesting a relationship between phagocytic NADPH oxidase–mediated oxidative stress and the development of atherosclerosis.

Objective: Oxidative stress has been implicated in the pathogenesis of hypertension and its complications through alterations in nitric oxide (NO) metabolism. This study was designed to investigate whether a relationship exists between phagocytic nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-dependent superoxide anion (•O2-) production and NO generation in patients with essential hypertension. Methods: Superoxide production was assayed by chemiluminescence under baseline and stimulated conditions on mononuclear cells obtained from hypertensives (n = 51) and normotensives (n = 43). NO production was evaluated by determining serum NO metabolites, nitrate plus nitrite (NOx). Results: Although there were no differences in baseline •O2- production between normotensives and hypertensives, the •O2- production in phorbol myristate acetate (PMA)-stimulated mononuclear cells was increased (P < 0.05) in hypertensives compared with normotensives. The PMA-induced •O2- production was completely abolished by apocynin, a specific inhibitor of NADPH oxidase. Moreover, stimulation of •O2- production by angiotensin II and endothelin-1 was higher (P < 0.05) in cells from hypertensives than in cells from normotensives. In addition, diminished (P < 0.001) serum NOx was detected in hypertensives compared with normotensives. Interestingly, an inverse correlation (r = 0.493, P < 0.01) was found between •O2- production and NOx in hypertensives. Conclusions: Generation of •O2- mainly dependent on NADPH oxidase is abnormally enhanced in stimulated mononuclear cells from hypertensives. It is suggested that this alteration could be involved in the diminished NO production observed in these patients.

Hypertensive heart disease is a progressive condition in which the compensatory left ventricular hypertrophy that maintains cardiac output leads to myocardial remodeling, characterized by fibrosis, insufficient vascularization, and alterations in cardiomyocytes, including contractile disturbances, changes in gene expression, and decrease in the number of cells. Structural abnormalities in the myocardial wall accelerate the development of diastolic and systolic dysfunction, resulting in heart failure. Many observations point to the apoptotic cell death of cardiomyocytes as a relevant factor in the transition from compensatory hypertrophy to pump failure in experimental and human hypertension. Potential inducers of cardiomyocyte apoptosis in overloaded hearts include extrinsic factors, such as mechanical forces, neurohormonal activation, oxidative stress, hypoxia, and cytokines. Some lines of evidence indicate that angiotensin II and the overstretching of cardiomyocytes are originally involved in the triggering of apoptosis in hypertension, whereas other factors are being investigated. Furthermore, intracellular changes, such as downregulation of survival proteins or activation of death proteins, seem to play an important role. The assumption that the apoptosis of cardiomyocytes worsens hypertensive heart disease prognosis brings forth new approaches to avoid or slow the transition to pump failure. In this respect, experimental data indicate that currently used antihypertensive drugs interfere with cardiomyocyte apoptosis. Moreover, the knowledge of intracellular apoptotic processes in cardiomyocytes provides novel therapeutic strategies to be added to the multimodal approach in the prevention of heart failure.

Abstract —Torasemide is a loop diuretic that is effective at low once-daily doses in the treatment of arterial hypertension. Because its antihypertensive mechanism of action may not be based entirely on the elimination of salt and water from the body, a vasodilator effect of this drug can be considered. In the present study, the ability of different concentrations of torasemide to modify angiotensin II (Ang II)–induced vascular responses was examined, with the use of an organ bath system, in endothelium-denuded aortic rings from spontaneously hypertensive rats. Ang II–induced increases of intracellular free calcium concentration ([Ca 2+ ] i ) were also examined by image analysis in cultured vascular smooth muscle cells (VSMCs) from spontaneously hypertensive rats. A dose-response curve to Ang II was plotted for cumulative concentrations (from 10 −9 to 10 −6 mol/L) in endothelium-denuded aortic rings (pD 2 =7.5±0.3). Isometric contraction induced by a submaximal concentration of Ang II (10 −7 mol/L) was reduced in a dose-dependent way by torasemide (IC 50 =0.5±0.04 μmol/L). Incubation of VSMCs with different concentrations of Ang II (from 10 −10 to 10 −6 mol/L) resulted in a dose-dependent rise of [Ca 2+ ] i (pD 2 =7.5±0.3). The stimulatory effect of [Ca 2+ ] i induced by a submaximal concentration of Ang II (10 −7 mol/L) was blocked by torasemide (IC 50 =0.5±0.3 nmol/L). Our findings suggest that torasemide blocks the vasoconstrictor action of Ang II in vitro. This action can be related to the ability of torasemide to block the increase of [Ca 2+ ] i induced by Ang II in VSMCs. It is proposed that these actions might be involved in the antihypertensive effect of torasemide observed in vivo.

OBJECTIVE: Previous studies have shown that as well as left ventricular hypertrophy, myocardial fibrosis develops early in rats with spontaneous hypertension (SHR). The present study was designed to investigate whether chronic treatment with the angiotensin II type 1 (AT1) receptor antagonist losartan modifies collagen type I metabolism and reverses left ventricular fibrosis in young SHR with left ventricular hypertrophy. DESIGN: The study was performed in 30-week-old normotensive Wistar-Kyoto (WKY) rats, untreated SHR and SHR treated with losartan (20 mg/mg per day, orally) for 14 weeks before they were killed. METHODS: Ventricular pro-alpha 1 (I) collagen messenger RNA was analyzed by Northern blot. Serum levels of the carboxy-terminal propeptide of procollagen type I (PIP) and the pyridoline cross-linked telopeptide domain of collagen type I (CITP) were determined by specific radioimmunoassays as markers of collagen type I synthesis and degradation, respectively. Collagen volume fraction was determined in the left ventricle by quantitative morphometry. RESULTS: Compared with WKY rats, SHR exhibited increased (P < 0.05) mean arterial pressure, pro-alpha 1 (I) collagen messenger RNA, PIP and left ventricular collagen volume fraction, and similar CITP values. After the treatment period, mean arterial pressure was higher (P < 0.05) in losartan-treated SHR than in WKY rats. Compared with untreated SHR, treated SHR showed no left ventricular hypertrophy and diminished (P < 0.05) values of mean arterial pressure, PIP and left ventricular collagen volume fraction. No changes in pro-alpha 1 (I) collagen messenger RNA and CITP values were observed with treatment in SHR. No significant differences in the left ventricular collagen volume fraction were observed between treated SHR with normal blood pressure and treated SHR with abnormally high blood pressure at the end of the treatment period. CONCLUSIONS: These results suggest that chronic AT1 blockade with losartan decreases the post-transcriptional synthesis of fibril-forming collagen type I molecules in young SHR. This effect may be involved in the ability of this drug to reverse left ventricular fibrosis in young rats with genetic hypertension. Apart from its antihypertensive action, other mechanisms may mediate the antifibrotic effect of losartan in this animal model.

Oxidative stress plays a key role in the pathophysiology of coronary artery disease, and constitutes a common mechanism behind the risk factors associated with this disease such as atherosclerosis, hypertension, diabetes and the metabolic syndrome. Oxidative stress is defined as an imbalance between the production of reactive oxygen and nitrogen species and the detoxification by the appropriate cellular systems. Reactive oxygen species induce cardiovascular dysfunction by modulating cell contraction/dilation, migration, growth/apoptosis and extracellular matrix protein turnover, which contribute to vascular and cardiac remodeling. In the last decade, the NADPH oxidase family has emerged as one of the most relevant sources of reactive oxygen species within the cardiovascular system. Recent data suggest a significant role of the genetic background in NADPH oxidase regulation. Common genetic polymorphisms within the promoter and exonic sequences of CYBA, the gene that encodes the p22phox subunit of the NADPH oxidase, have been characterized in the context of cardiovascular diseases. This review aims to present the current state of research into these polymorphisms with regards to their relationship to coronary artery disease.