Daiane Silvello

An analysis of the global expression of microRNAs in an experimental model of physiological left ventricular hypertrophy.

Background MicroRNAs (miRs) are a class of small non-coding RNAs that regulate gene expression. Studies of transgenic mouse models have indicated that deregulation of a single miR can induce pathological cardiac hypertrophy and cardiac failure. The roles of miRs in the genesis of physiological left ventricular hypertrophy (LVH), however, are not well understood. Objective To evaluate the global miR expression in an experimental model of exercise-induced LVH. Methods Male Balb/c mice were divided into sedentary (SED) and exercise (EXE) groups. Voluntary exercise was performed on an odometer-monitored metal wheels for 35 days. Various tests were performed after 7 and 35 days of training, including a transthoracic echocardiography, a maximal exercise test, a miR microarray (miRBase v.16) and qRT-PCR analysis. Results The ratio between the left ventricular weight and body weight was increased by 7% in the EXE group at day 7 (p<0.01) and by 11% at day 35 of training (p<0.001). After 7 days of training, the microarray identified 35 miRs that were differentially expressed between the two groups: 20 were up-regulated and 15 were down-regulated in the EXE group compared with the SED group (p = 0.01). At day 35 of training, 25 miRs were differentially expressed: 15 were up-regulated and 10 were decreased in the EXE animals compared with the SED animals (p<0.01). The qRT-PCR analysis demonstrated an increase in miR-150 levels after 35 days and a decrease in miR-26b, miR-27a and miR-143 after 7 days of voluntary exercise. Conclusions We have identified new miRs that can modulate physiological cardiac hypertrophy, particularly miR-26b, -150, -27a and -143. Our data also indicate that previously established regulatory gene pathways involved in pathological LVH are not changed in physiological LVH.

Genetic polymorphisms of the adrenergic system and implantable cardioverter-defibrillator therapies in patients with heart failure

AIMS We investigated whether the combination of beta(1)-Gly389Arg and GNB3 C825T, two genetic polymorphisms strictly related to adrenergic system modulation, could act as predictors of appropriate therapies in patients with heart failure (HF) using implantable cardioverter-defibrillators (ICDs). METHODS AND RESULTS Patients with HF and ICD implantation for primary and secondary prevention were studied. All ICD therapies were registered and classified as appropriate (secondary to ventricular tachycardia) or inappropriate (others). Genetic analysis was performed by polymerase chain reaction and restriction fragment length polymorphism methods. Seventy-three patients with mean left ventricular ejection fraction of 35 +/- 11% were evaluated. Overall, 35 ICD therapies occurred during follow-up in 31 (42.5%) patients. Twenty-four therapies (33%) were appropriate, and 11 (15%) were inappropriate. Individual analysis of each polymorphism only identified T825 carriers of GNB3 C825T as predictor of appropriate shocks. The combined presence of risk genotypes (Arg389 of the beta(1)-Gly389Arg and T825 of the GNB3 C825T) identified patients with higher risk of appropriate shocks. Patients with two at-risk genotypes had a survival rate free of appropriate shocks lower than those with none or only one of these markers (87 vs. 54%, respectively; log-rank statistic = 0.006). Using a Cox regression model, each at-risk genotype was associated with an increment of risk of appropriate ICD shocks (odds ratio = 3.9, 95% confidence interval of 1.3-12.0; P = 0.02). CONCLUSION Genetic polymorphisms of the adrenergic system may help to identify HF patients who are more likely to receive appropriate ICD therapies. Further studies are necessary to determine the clinical applicability of these polymorphisms as predictors of arrhythmias.

Circulating microRNAs in obese and lean heart failure patients: A case–control study with computational target prediction analysis

AIMS MicroRNAs (miRs) regulate processes involved in both cardiac remodeling and obesity. We investigated if the expression of selected miRs in patients with heart failure (HF) is influenced by the presence of obesity. METHODS In this case-control study, we compared plasma levels of miR-21, -130b, -221, -423-5p, and the -221/-130b ratio in 57 age- and gender-matched subjects: 40 HF patients (20 obese HF and 20 lean HF) and 17 lean healthy controls. Body composition was estimated by bioelectrical impedance analysis. MiRs were measured by quantitative reverse transcription-PCR. Bioinformatics analysis was performed based on miRs findings to predict their putative targets and investigate their biological function. RESULTS HF was associated with increased miR-423-5p levels in both lean and obese patients (P<0.05 vs. controls) without differences between HF groups. MiR-130b levels were reduced in obese HF patients compared with HF lean (P=0.036) and controls (P=0.025). MiR-221 levels were non-significantly increased in obese HF patients. MiR-21 levels were not different among the groups. MiR-221/-130b ratio was increased in obese HF patients, and was positively associated with body fat percentage (r=0.43; P=0.002), body mass index (r=0.44; P=0.002), and waist circumference (r=0.40; P=0.020). Computational prediction of target genes followed by functional enrichment analysis indicated a relevant role of miR-130b and miR-221 in modulating the expression of genes associated to cardiovascular and endocrine diseases, and suggested their influence in important signaling mechanisms and in numerous processes related to the circulatory and endocrine systems. CONCLUSIONS In HF patients, the presence of obesity is associated with a differential expression of selected miRs and the miR-221/-130b ratio had significant correlations with adiposity parameters. Computational target prediction analysis identified several interrelated pathways targeted by miR-130b and miR-221 with a known relationship with endocrine and cardiovascular diseases, representing potential mechanisms to be further validated.

Polymorphisms of endothelial nitric oxide synthase gene in systolic heart failure: An haplotype analysis

BACKGROUND Endothelial nitric oxide synthase (eNOS) gene polymorphisms have been associated with the pathogenesis of cardiovascular diseases, but few studies have evaluated the role of eNOS haplotypes on the risk and prognosis of heart failure (HF). This prospective study was designed to analyze the impact of three eNOS polymorphisms (T-786C, VNTR4a/b and Glu298Asp) and their haplotypes on the susceptibility and clinical outcomes in HF outpatients with systolic dysfunction. METHODS AND RESULTS We conducted a case-control and a cohort study in which 316 HF patients and 360 healthy controls were recruited from a tertiary care university hospital. DNA was extracted from peripheral blood and eNOS polymorphisms were detected by PCR or PCR-RFLP. Patients were predominantly men, had a mean left ventricular ejection fraction of 31% and were followed-up for a median of 41months; there were 96 deaths, including 58 HF-related deaths. Genotype distribution of the eNOS T-786C, VNTR 4a/b and Glu298Asp was similar between HF patients and controls. Haplotype frequencies differed between HF patients and controls only in African-Brazilians (p=0.043). African-Brazilian patients that carried the haplotype -786C/4b/Asp298 had a better prognosis than patients that carried other haplotypes (log rank p value=0.016 for all-cause mortality). In a Cox proportional hazard model adjusted for clinical variables of risk, the -786C/4b/Asp298 haplotype remained as an independent genetic predictor of survival (adjusted HR=0.11; 95% CI=0.01-0.83; p=0.03). CONCLUSIONS The -786C/4b/Asp298 eNOS haplotype had a significant impact on HF susceptibility and prognosis, particularly in African-Brazilian patients.

Impact of β-2 Thr164Ile and combined β-adrenergic receptor polymorphisms on prognosis in a cohort of heart failure outpatients

Genetic polymorphisms of adrenergic receptors (ARs) have been associated with the development, progression, and prognosis of patients with heart failure (HF), with few data for the Brazilian population. We evaluated the role of the β2-AR Thr164Ile polymorphism at codon 164 on prognosis in a prospective study on 315 adult Brazilian HF patients, predominantly middleaged Caucasian men in functional class I-II, with severe left ventricular systolic dysfunction. Genomic DNA was extracted from peripheral blood and β2-AR164 genotypes were detected by PCR followed by restriction fragment length analysis. During a median follow-up of 3 years, 95 deaths occurred and 57 (60%) were HF-related. Unexpectedly, Ile164 carriers (N = 12) had no HF-related events (log-rank P value = 0.13). Analysis using genotype combination with β1-AR polymorphisms at codons 49 and 389 identified patients with favorable genotypes (Thr164Ile of β2-AR, Gly49Gly of β1-AR and/or Gly389Gly of β1-AR), who had lower HF-related mortality (P = 0.01). In a Cox proportional hazard model adjusted for other clinical characteristics, having any of the favorable genotypes remained as independent predictor of all-cause (hazard ratio (HR): 0.41, 95%CI: 0.17-0.95) and HF-related mortality (HR: 0.12, 95%CI: 0.02-0.90). These data show that the β2-AR Thr164Ile polymorphism had an impact on prognosis in a Brazilian cohort of HF patients. When combined with common β1-AR polymorphisms, a group of patients with a combination of favorable genotypes could be identified.

Encapsulated mononuclear stem cells: paracrine action for the treatment of acute myocardial infarction

Cell therapy is considered as a treatment option for acute myocardial infarction (AMI). Released molecules by cells paracrine action may promote tissue regeneration. Therefore we used bone-marrow mononuclear cells (BMMNCs) from GFP+ Wistar rats encapsulated in sodium alginate for AMI treatment. Animals were randomly allocated into groups – empty (EC); BMMNC capsules; or sham. AMI was induced by occlusion of left anterior artery and capsules were delivered intrathoracically. Troponin I was measured 24h after AMI and echocardiography was performed at 48h and 7d after AMI. On day 7 animals were euthanized and their hearts were harvested. Tissue levels of TNF-α, IL-6, IL-10, cleaved caspase-3, and catalase were measured. Technical procedures were performed by blinded operators. There was no difference in either heart morphofunctional parameters or biochemical analysis between AMI groups. We conclude that the paracrine effects of BMMNCs lacks efficacy to modulate events associated with AMI in the rat.