Veronica Gonzalez-Albert

Renin polymorphisms and haplotypes are associated with blood pressure levels and hypertension risk in postmenopausal women

Background Renin is a key protein of the renin–angiotensin system involved in the physiological control of blood pressure; the renin gene is therefore a candidate for essential hypertension in humans. We tested the association between polymorphisms and haplotypes of the renin gene and the risk of hypertension and blood pressure levels in two Spanish populations. Methods Two population-based studies from different regions of Spain were performed. Study A included 1502 individuals (748 women) 40–70 years old, and Study B included 670 women 45–70 years old. Fourteen polymorphisms of the renin gene were selected based on position, spacing, heterozygosity (> 10% for the minor allele frequency) and previous information, and were assessed by SNPlex. Results Genotype GG of the rs5707 polymorphism was significantly associated with blood pressure levels (P = 0.005) and with the risk of having hypertension (odds ratio, 6.16; 95% confidence interval, 1.19–31.8) in women 40–70 years old from study A, but not the men. This association was also present in the women of study B (P < 0.001 for blood pressure values; odds ratio, 2.11 and 95% confidence interval, 1.07–4.17 for hypertension). Two haplotypes defined by five selected polymorphisms were associated with increased risk of hypertension in these aged women. Conclusion Polymorphisms of the renin gene were associated with blood pressure levels and risk of hypertension in women over 40 years old. The interaction between the potential functional impact of this genetic background and the estrogen fall could explain the association in women of this age group.

Influence of microsomal triglyceride transfer protein promoter polymorphism −493 Gt on fasting plasma triglyceride values and interaction with treatment response to atorvastatin in subjects with heterozygous familial hypercholesterolaemia

Familial hypercholesterolaemia (FH) is an autosomal dominant disease characterized by elevated levels of low-density lipoprotein-cholesterol (LDL-C). Phenotypic expression is highly variable, being influenced by diet, age, gender, body mass index, apolipoprotein E genotype and type of LDL-receptor gene mutation. Microsomal triglyceride (TG) transfer protein (MTP) is a protein involved in lipid metabolism. Polymorphism MTP −493 GT has been shown to modulate lipid levels in several populations. To analyse the effect of this polymorphism in the lipid phenotype expression of FH and treatment response, we studied a sample of 222 Spanish FH patients, of whom 147 were studied before and after treatment with 20 mg of atorvastatin daily during 6 weeks. The variant was analysed by polymerase chain reaction amplification and single-strand confirmation polymorphism. Treatment reduced LDL-C, total cholesterol and TGs. Baseline fasting TGs and very-low-density lipoprotein cholesterol levels were lower in female T allele carriers (TG: 111±51 mg/dl GG, 89±35 mg/dl GT, 83±26 mg/dl TT, P=0.022; very-low-density lipoprotein cholesterol: 24±13 mg/dl GG, 16±5 mg/dl GT, 17±5 mg/dl TT, P=0.018). Triglyceride response to atorvastatin was modulated by this polymorphism in men (P=0.009), but not in women, although differences between genotypes were maintained after treatment. In conclusion, the MTP −493 GT polymorphism modulates pre- and post-treatment plasma TG values of FH in Spanish subjects in a gender-specific way. Other environmental and genetic factors likely also modulate this response.

Different Impacts of Cardiovascular Risk Factors on Oxidative Stress

The objective of the study was to evaluate oxidative stress (OS) status in subjects with different cardiovascular risk factors. With this in mind, we have studied three models of high cardiovascular risk: hypertension (HT) with and without metabolic syndrome, familial hypercholesterolemia (FH) and familial combined hyperlipidemia (FCH) with and without insulin resistance. Oxidative stress markers (oxidized/reduced glutathione ratio, 8-oxo-deoxyguanosine and malondialdehide) together with the activity of antioxidant enzyme triad (superoxide dismutase, catalase, glutathione peroxidase) and activation of both pro-oxidant enzyme (NAPDH oxidase components) and AGTR1 genes, as well as antioxidant enzyme genes (CuZn-SOD, CAT, GPX1, GSR, GSS and TXN) were measured in mononuclear cells of controls (n = 20) and patients (n = 90) by assessing mRNA levels. Activity of some of these antioxidant enzymes was also tested. An increase in OS and pro-oxidant gene mRNA values was observed in patients compared to controls. The hypertensive group showed not only the highest OS values, but also the highest pro-oxidant activation compared to those observed in the other groups. In addition, in HT a significantly reduced antioxidant activity and mRNA induction of antioxidant genes were found when compared to controls and the other groups. In FH and FCH, the activation of pro-oxidant enzymes was also higher and antioxidant ones lower than in the control group, although it did not reach the values obtained in hypertensives. The thioredoxin system was more activated in patients as compared to controls, and the highest levels were in hypertensives. The increased oxidative status in the presence of cardiovascular risk factors is a consequence of both the activation of pro-oxidant mechanisms and the reduction of the antioxidant ones. The altered response of the main cytoplasmic antioxidant systems largely contributes to OS despite the apparent attempt of the thioredoxin system to control it.

Enhanced reduction in oxidative stress and altered glutathione and thioredoxin system response to unsaturated fatty acid load in familial hypercholesterolemia

OBJECTIVES Familial hypercholesterolemia (FH) is characterized by increased oxidative stress (OS) levels. In the postprandial state, lipids and lipoproteins modulate OS status through their impact on pro-oxidant and antioxidant mechanisms. The objective of this study was to evaluate in patients with FH the response to an unsaturated oral fat load test (OFLT) by analyzing the mRNA levels of genes involved in the glutathione and thioredoxin antioxidant systems. DESIGN AND METHODS We analyzed 14 FH patients and 20 normolipidemic and normoglycemic controls. In both groups, mRNA values of antioxidant enzyme genes (glutathione and thioredoxin systems) were determined at baseline and at 2, 4, 6, and 8h after OFLT by real time PCR. RESULTS In the fasting state the mRNA levels of antioxidant enzymes GPX4 and the GSR, GSS, and GCLC enzymes (involved in glutathione regeneration and synthesis) and thioredoxin (TXN), were significantly increased in the FH group compared to the healthy controls. Some genes (GPX1 and GPX4) were increased at 4h in both groups, but values for the rest of the antioxidant enzyme mRNAs were decreased in FH patients after 4h from unsaturated OFLT and were increased in controls. CONCLUSIONS We concluded that an OFLT with predominantly unsaturated fat has a different effect on postprandial antioxidant enzyme mRNA levels in controls than in FH patients. Increased antioxidant enzyme mRNA is not the main way to reduce postprandial oxidative stress in FH. This difference could determine the influence of dietary patterns in these patients.

Are IL18RAP gene polymorphisms associated with body mass regulation? A cross-sectional study

Objectives To investigate the association between IL18RAP and body mass index (BMI) and obesity and to verify the effect of a polymorphism in the microRNA136 (MIR136) IL18RAP binding region. Design We analysed samples from two Spanish cross-sectional studies, VALCAR (Spanish Mediterranean coast) and Hortega (Spanish centre). These studies aimed at analysing cardiovascular risk and development of cardiovascular disease in the general population. Both populations correspond to regions with different characteristics. Setting Five IL18RAP single nucleotide polymorphisms were selected using the SYSNPs web tool and analysed by oligonucleotide ligation assay (SNPlex). For the MIR136 functional study, cells were transfected with plasmids containing different rs7559479 polymorphism alleles and analysed by luciferase reporter assays. Participants 1970 individuals (Caucasian, both genders): VALCAR (468) and Hortega (1502). Results rs2293225, rs2272127 and rs7559479 showed the following associations: rs7559479 G allele correlated with a higher obesity risk (P=0.01; OR=1.82; 95% CI 1.15 to 2.87 for the VALCAR group; P=0.033; OR=1.35; 95% CI 1.03 to 1.79 for the Hortega population) and higher body mass index (BMI) values (P=0.0045; P=0.1 for VALCAR and Hortega, respectively); a significant association with obesity (P=0.0024, OR=1.44, 95% CI 1.14 to 1.82) and increased BMI values (P=0.008) was found when considering both populations together. rs2293225 T allele was associated with lower obesity risk (P=0.036; OR=0.60; 95% CI 0.35 to 0.96) and lower BMI values (P=0.0038; OR=1.41) while the rs2272127 G allele was associated with lower obesity risk (P=0.028; OR=0.66; 95% CI 0.44 to 0.97) only in the VALCAR population. A reporter assay showed that the presence of the A allele in rs7559479 was associated with increased MIR136 binding to IL18RAP. Conclusions Our results suggest that polymorphisms in IL18RAP influence susceptibility to obesity. We demonstrated that the A allele in rs7559479 increases MIR136 binding, which regulates IL-18 system activity.

Correlation of zinc with oxidative stress biomarkers.

Hypertension and smoking are related with oxidative stress (OS), which in turn reports on cellular aging. Zinc is an essential element involved in an individual’s physiology. The aim of this study was to evaluate the relation of zinc levels in serum and urine with OS and cellular aging and its effect on the development of hypertension. In a Spanish sample with 1500 individuals, subjects aged 20–59 years were selected, whose zinc intake levels fell within the recommended limits. These individuals were classified according to their smoking habits and hypertensive condition. A positive correlation was found (Pearson’s C = 0.639; p = 0.01) between Zn serum/urine quotient and oxidized glutathione levels (GSSG). Finally, risk of hypertension significantly increased when the GSSG levels exceeded the 75 percentile; OR = 2.80 (95%CI = 1.09–7.18) and AOR = 3.06 (95%CI = 0.96–9.71). Low zinc levels in serum were related with OS and cellular aging and were, in turn, to be a risk factor for hypertension.

Immune-unreactive urinary albumin as a predictor of cardiovascular events: the Hortega Study

BACKGROUND We aimed to determine if immune-unreactive albumin excretion (IURAE) is associated with cardiovascular (CV) events in a representative sample of a general population from Spain. METHODS We included 1297 subjects (mean age ± standard error 48.0 ± 0.2 years, 48% females), who participated in the Hortega Follow-Up Study. The primary endpoint was incidence of fatal and non-fatal CV events. Urinary albumin excretion (UAE) was measured in spot voided urine, frozen at -80°C, by immunonephelometry [immune-reactive albumin excretion (IRAE)] and by high-performance liquid chromatography (HPLC) [total albumin excretion (AE)]. IURAE was calculated as the difference between HPLC measurements and IRAE. We estimated fully adjusted hazard ratios (HRs) of CV incidence by Cox regression for IRAE, IURAE and total AE. RESULTS After an average at-risk follow-up of 13 years, we observed 172 CV events. urinary albumin to creatinine ratio (UACR) of ≥30 mg/g assessed by IRAE, IURAE or total AE concentrations was observed in 74, 273 and 417 participants, respectively. Among discordant pairs, there were 49 events in those classified as micro- and macroalbuminuric by IURAE, but normoalbuminuric by IRAE. Only the IRAE was a significant independent factor for the incidence of CV events [HR (95% confidence interval) 1.15 (1.04-1.27)]. The association of UAE with CV events was mainly driven by heart failure (HF) [HR 1.33 (1.15-1.55) for IRAE; HR 1.38 (1.06-1.79) for IURAE; HR 1.62 (1.22-2.13) for total AE]. Those subjects who were micro- and macroalbuminuric by both IRAE and IURAE had a significant increase in risk for any CV event, and especially for HF. CONCLUSIONS IRAE, IURAE and AE were associated with an increased risk for CV events, but IRAE offered better prognostic assessment.

mRNA expression profiles obtained from microdissected pancreatic cancer cells can predict patient survival

Background Pancreatic ductal adenocarcinoma (PDAC) is one of the most devastating malignancies in developed countries because of its very poor prognosis and high mortality rates. By the time PDAC is usually diagnosed only 20-25% of patients are candidates for surgery, and the rate of survival for this cancer is low even when a patient with PDAC does undergo surgery. Lymph node invasion is an extremely bad prognosis factor for this disease. Methods We analyzed the mRNA expression profile in 30 PDAC samples from patients with resectable local disease (stages I and II). Neoplastic cells were isolated by laser-microdissection in order to avoid sample ‘contamination’ by non-tumor cells. Due to important differences in the prognoses of PDAC patients with and without lymph node involvement (stage IIB and stages I-IIA, respectively), we also analyzed the association between the mRNA expression profiles from these groups of patients and their survival. Results We identified expression profiles associated with patient survival in the whole patient cohort and in each group (stage IIB samples or stage I-IIA samples). Our results indicate that survival-associated genes are different in the groups with and without affected lymph nodes. Survival curves indicate that these expression profiles can help physicians to improve the prognostic classification of patients based on these profiles.

DIFFERENT IMPACT OF CARDIOVASCULAR RISK FACTORS ON OXIDATIVE STRESS: PP.21.343

Subjects and Methods: We have studied three models of high cardiovascular risk: hypertension (HT) with and without metabolic syndrome, familial hypercholesterolemia (FH) and familial combined hyperlipidemia (FCH) with and without insulin resistance. Activation of genes, assessed by mRNA level, of pro-oxidant enzyme (NAPDH oxidase components) and AGTR1 as well as of antioxidant systems (CuZn-SOD2, CAT, GPX1, GSR, GSS and TXN) were measured in mononuclear cells of controls (n = 20) and patients (n = 90) by using Ready-To-Go You Prime First-Strand beads (Amersham Pharmacia Biotech). Primers for PCR were designed by Primer3 software. The PCR reaction was done using SYBR Green PCR Master Mix and ABI PRISM 7000 Sequence Detection System (Applied Biosystems, Foster City, CA). Activity of some of these antioxidant enzymes was also tested. Results: An increase in OS and pro-oxidant gene mRNA values were observed in patients as compared to controls. Hypertensive group showed the highest OS values as well as the highest pro-oxidant activation as compared to that observed in the other groups. In addition, in hypertensive subjects a significantly reduced antioxidant activity and mRNA activation compared to controls and the others groups was found except for thioredoxin system that showed high level in HT. In HT and FCH the activation of pro-oxidant enzymes was also higher and the antioxidant lower than it was in the control group, although did not reach the values obtained in hypertensives. Thioredoxin system was activated in patients although the highest levels were in hypertension. Conclusion: The increased oxidative status in the presence of cardiovascular risk factors is consequence of both activation of prooxidant mechanisms and reduction of the antioxidant ones. The altered response of the main cytoplasmatic antioxidant systems largely contributes to OS despite the attempt of the thioredoxin system to control it.