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Given the association between high aerobic capacity and the prevention of metabolic diseases, elucidating the mechanisms by which high aerobic capacity regulates whole-body metabolic homeostasis is a major research challenge. Oxidative post-translational modifications (Ox-PTMs) of proteins can regulate cellular homeostasis in skeletal and cardiac muscles, but the relationship between Ox-PTMs and intrinsic components of oxidative energy metabolism is still unclear. Here, we evaluated the Ox-PTM profile in cardiac and skeletal muscles of rats bred for low (LCR) and high (HCR) intrinsic aerobic capacity. Redox proteomics screening revealed different cysteine (Cys) Ox-PTM profile between HCR and LCR rats. HCR showed a higher number of oxidized Cys residues in skeletal muscle compared to LCR, while the opposite was observed in the heart. Most proteins with differentially oxidized Cys residues in the skeletal muscle are important regulators of oxidative metabolism. The most oxidized protein in the skeletal muscle of HCR rats was malate dehydrogenase (MDH1). HCR showed higher MDH1 activity compared to LCR in skeletal, but not cardiac muscle. These novel findings indicate a clear association between Cys Ox-PTMs and aerobic capacity, leading to novel insights into the role of Ox-PTMs as an essential signal to maintain metabolic homeostasis. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

A significant body of evidence supports the protective role of exercise training (ET) in cardiovascular diseases, skeletal muscle dystrophies, several types of cancer, Alzheimer disease or even in the recovery of spinal cord injury. In spite of this, the molecular mechanisms underlying the beneficial effects of exercise training are not well understood and remain elusive. Several mechanisms have been proposed in the past, but more recently microRNAs (miRNAs), small non-coding RNA molecules involved in a variety of basic biological processes that negatively modulate gene expression, recognized as important regulatory molecules. In this review, we highlight recent advances on the miRNA involvement in the benefits of ET. Here, we assess the role of microRNAs expressed in the heart, in the skeletal muscle, detected in the circulation (serum and plasma), and in other conditions (e.g., spinal cord injury). Additionally, the long-term effects of diverse ET modalities (e.g., running, cycling, resistance training) in the cardiac miRNA profile are properly addressed.

Contains fulltext : 154705.pdf (Publisher’s version ) (Open Access) Pathological cardiac hypertrophy is characterized by a shift in metabolic substrate utilization from fatty acids to glucose, but the molecular events underlying the metabolic remodeling remain poorly understood. Here, we investigated the role of liver X receptors (LXRs), which are key regulators of glucose and lipid metabolism, in cardiac hypertrophic pathogenesis. Using a transgenic approach in mice, we show that overexpression of LXRalpha acts to protect the heart against hypertrophy, fibrosis, and dysfunction. Gene expression profiling studies revealed that genes regulating metabolic pathways were differentially expressed in hearts with elevated LXRalpha. Functionally, LXRalpha overexpression in isolated cardiomyocytes and murine hearts markedly enhanced the capacity for myocardial glucose uptake following hypertrophic stress. Conversely, this adaptive response was diminished in LXRalpha-deficient mice. Transcriptional changes induced by LXRalpha overexpression promoted energy-independent utilization of glucose via the hexosamine biosynthesis pathway, resulting in O-GlcNAc modification of GATA4 and Mef2c and the induction of cytoprotective natriuretic peptide expression. Our results identify LXRalpha as a key cardiac transcriptional regulator that helps orchestrate an adaptive metabolic response to chronic cardiac stress, and suggest that modulating LXRalpha may provide a unique opportunity for intervening in myocyte metabolism.

Although aberrant reactivation of embryonic gene programs is intricately linked to pathological heart disease, the transcription factors driving these gene programs remain ill-defined. Here we report that increased calcineurin/Nfat signalling and decreased miR-25 expression integrate to re-express the basic helix-loop-helix (bHLH) transcription factor dHAND (also known as Hand2) in the diseased human and mouse myocardium. In line, mutant mice overexpressing Hand2 in otherwise healthy heart muscle cells developed a phenotype of pathological hypertrophy. Conversely, conditional gene-targeted Hand2 mice demonstrated a marked resistance to pressure-overload-induced hypertrophy, fibrosis, ventricular dysfunction and induction of a fetal gene program. Furthermore, in vivo inhibition of miR-25 by a specific antagomir evoked spontaneous cardiac dysfunction and sensitized the murine myocardium to heart failure in a Hand2-dependent manner. Our results reveal that signalling cascades integrate with microRNAs to induce the expression of the bHLH transcription factor Hand2 in the postnatal mammalian myocardium with impact on embryonic gene programs in heart failure.

Background Respiratory muscle weakness contributes to exercise intolerance in patients with heart failure with a preserved ejection fraction ( HF p EF )—a condition characterized by multiple comorbidities with few proven treatments. We aimed, therefore, to provide novel insight into the underlying diaphragmatic alterations that occur in HF p EF by using an obese cardiometabolic rat model and further assessed whether exercise training performed only after the development of overt HF p EF could reverse impairments. Methods and Results Obese ZSF 1 rats (n=12) were compared with their lean controls (n=8) at 20 weeks, with 3 additional groups of obese ZSF 1 rats compared at 28 weeks following 8 weeks of either sedentary behavior (n=13), high‐intensity interval training (n=11), or moderate‐continuous training (n=11). Obese rats developed an obvious HF p EF phenotype at 20 and 28 weeks. In the diaphragm at 20 weeks, HF p EF induced a shift towards an oxidative phenotype and a fiber hypertrophy paralleled by a lower protein expression in Mu RF 1 and MuRF2, yet mitochondrial and contractile functional impairments were observed. At 28 weeks, neither the exercise training regimen of high‐intensity interval training or moderate‐continuous training reversed any of the diaphragm alterations induced by HF p EF . Conclusions This study, using a well‐characterized rat model of HF p EF underpinned by multiple comorbidities and exercise intolerance (ie, one that closely resembles the patient phenotype), provides evidence that diaphragm alterations and dysfunction induced in overt HF p EF are not reversed following 8 weeks of aerobic exercise training. As such, whether alternative therapeutic interventions are required to treat respiratory muscle weakness in HF p EF warrants further investigation.

OBJECTIVE: Increased glucose 6-phosphate dehydrogenase activity has been demonstrated in heart failure. This study sought to assess myocardial glucose 6-phosphate dehydrogenase activity in retraining of the subpulmonary ventricle of adult goats. METHODS: Eighteen adult goats were divided into three groups: traditional (fixed banding), sham, and intermittent (adjustable banding, daily 12-hour systolic overload). Systolic overload (70% of systemic pressure) was maintained during a 4-week period. Right ventricle, pulmonary artery and aortic pressures were measured throughout the study. All animals were submitted to echocardiographic and hemodynamic evaluations throughout the protocol. After the study period, the animals were killed for morphological and glucose 6-phosphate dehydrogenase activity assessment. RESULTS: A 55.7% and 36.7% increase occurred in the intermittent and traditional right ventricle masses, respectively, when compared with the sham group (P

Coronary heart disease is the most common cause of death, and the number of individuals at risk is increasing. To better manage this pandemic, improved tool for risk prediction, including more accurate biomarkers are needed. The objective of this study was to assess the utility of circulating microRNAs (miRs) to predict future fatal acute myocardial infarction (AMI) in healthy participants.We performed a prospective nested case-control study with 10-year observation period and fatal AMI as endpoint. In total, 179 miRs were quantified by real-time polymerase chain reaction in serum of 112 healthy participants (40-70 years) that either (1) suffered from fatal AMI within 10 years [n = 56], or (2) remained healthy [n = 56, risk factor-matched controls]. Candidate miRs were validated in a separate cohort of healthy individuals (n = 100).Twelve miRs were differently expressed in cases and controls in the derivation cohort (p < 0.05). Among these, 10 miRs differed significantly between cases and controls in the validation cohort (p < 0.05). We identified gender dimorphisms, as miR-424-5p and miR-26a-5p were associated exclusively with risk in men and women, respectively. The best model for predicting future AMI consisted of miR-106a-5p, miR-424-5p, let-7g-5p, miR-144-3p and miR-660-5p, providing 77.6% correct classification for both genders, and 74.1% and 81.8% for men and women, respectively. Adding these 5 miRs to the Framingham Risk Score, increased the AUC from 0.72 to 0.91 (p < 0.001).In conclusion, we identified several miRs associated with future AMI, revealed gender-specific associations, and proposed a panel of 5 miRs to enhance AMI risk prediction in healthy individuals.

FUNDAMENTO: A transposicao corrigida das grandes arterias frequentemente evolui com disfuncao ventricular direita. O preparo ventricular para a correcao anatomica em pacientes adultos apresenta resultados desapontadores. OBJETIVO: Analisar a hipertrofia do ventriculo direito (VD) induzida por dois tipos de bandagem pulmonar (BP), convencional e intermitente em animais adultos. METODOS: Dezenove cabras adultas foram divididas em tres grupos: Convencional (seis animais), Intermitente (seis animais) e Controle (sete animais). O grupo Convencional foi submetido a BP fixa com fita cardiaca, enquanto no grupo Intermitente foi usado dispositivo de BP ajustavel, que gerava sobrecarga sistolica por 12 horas, alternada com 12 horas de descanso do VD. As pressoes de VD, tronco pulmonar e aorta foram medidas durante todo o estudo. Ecocardiograma foi realizado semanalmente. Apos quatro semanas, os animais foram eutanasiados para avaliacao morfologica dos ventriculos. O grupo Controle foi submetido a eutanasia para analise em condicoes basais. RESULTADOS: A sobrecarga pressorica foi menor no grupo Intermitente (p=0,001), comparada ao grupo Convencional. Houve aumento na espessura do VD do grupo Intermitente, medida pelo ecocardiograma, comparado ao seu momento basal (p

1. Clinical and experimental evidence highlights the importance of the renin-angiotensin system in renovascular hypertension. Furthermore, genetic factors affecting angiotensin-converting enzyme (ACE) could influence the development of renovascular hypertension. 2. To test the effect of small gene perturbations on the development of renovascular hypertension, mice harbouring two or three copies of the Ace gene were submitted to 4 weeks of two-kidney, one-clip (2K1C) hypertension. Blood pressure (BP), cardiac hypertrophy, baroreflex sensitivity and blood pressure and heart rate variability were assessed and compared between the different groups. 3. The increase in BP induced by 2K1C was higher in mice with three copies of the Ace gene compared with mice with only two copies (46 vs 23 mmHg, respectively). Moreover, there was a 3.8-fold increase in the slope of the left ventricle mass/BP relationship in mice with three copies of the Ace gene. Micewith three copies of the Ace gene exhibited greater increases in cardiac and serum ACE activity than mice with only two copies of the gene. Both baroreflex bradycardia and tachycardia were significantly depressed in mice with three copies of the Ace gene after induction of 2K1C hypertension. The variance in basal systolic BP was greater in mice with three copies of the Ace gene after 2K1C hypertension compared with those with only two copies of the gene (106 vs 54%, respectively). In addition, the low-frequency component of the pulse interval was higher mice with three copies of the Ace gene after 2K1C hypertension compared with those with only two (168 vs 86%, respectively). Finally, in mice with three copies of the Ace gene, renovascular hypertension induced a 6.1-fold increase in the sympathovagal balance compared with a 3.2-fold increase in mice with only two copies of the gene. 4. Collectively, these data provide direct evidence that small genetic disturbances in ACE levels per se have an influence on haemodynamic, cardiac mass and autonomic nervous system responses in mice under pathological perturbation.

The purpose of the present study was to determine the range of the influence of the baroreflex on blood pressure in chronic renal hypertensive rats. Supramaximal electrical stimulation of the aortic depressor nerve and section of the baroreceptor nerves (sinoaortic denervation) were used to obtain a global analysis of the baroreceptor-sympathetic reflex in normotensive control and in chronic (2 months) 1-kidney, 1-clip hypertensive rats. The fall in blood pressure produced by electrical baroreceptor stimulation was greater in renal hypertensive rats than in normotensive controls (right nerve: -47 +/- 8 vs -23 +/- 4 mmHg; left nerve: -51 +/- 7 vs -30 +/- 4 mmHg; and both right and left nerves: -50 +/- 8 vs -30 +/- 4 mmHg; P0.05). Furthermore, the increase in blood pressure level produced by baroreceptor denervation in chronic renal hypertensive rats was similar to that observed in control animals 2-5 h (control: 163 +/- 5 vs 121 +/- 1 mmHg; 1K-1C: 203 +/- 7 vs 170 +/- 5 mmHg; P0.05) and 24 h (control: 149 +/- 3 vs 121 +/- 1 mmHg; 1K-1C: 198 +/- 8 vs 170 +/- 5 mmHg; P0.05) after sinoaortic denervation. Taken together, these data indicate that the central and peripheral components of the baroreflex are acting efficiently at higher arterial pressure in renal hypertensive rats when the aortic nerve is maximally stimulated or the activity is abolished.

Objective Null genotypes of glutathione S-transferase (GSTs) exhibit absence of enzymatic activity and are hypothesized to modulate an increased risk of developing cardiovascular disease. The aim of this study was to identify the potential association between GSTM1 and GSTT1 deleted polymorphisms with cardiovascular risk factors and coronary atherosclerosis in two independent urban populations. Methods and results Genotype distribution of GSTM1 and GSTT1 deleted polymorphism were examined in a sample of 1577 individuals from the general population and a replication sample of 871 individuals submitted to coronary angiography. Triglycerides, HDL-cholesterol and the triglycerides/HDL ratio were significantly associated with a double-deleted genotype in individuals from the general population. These findings were replicated in a second, independent, population of individuals submitted to coronary angiography. In addition, coronary artery disease severity was also associated with GSTs genotypes and the risk conferred from GSTs genotype was mainly due to triglycerides/HDL ratio information. Conclusions The data suggest that the presence of a double deletion genotypes of the GSTM1 and GSTT1 genes is associated with hypertriglyceridemia and low HDL-cholesterol levels in humans. These novel findings may provide a new unexplored link between lipid metabolism and GST homeostasis.

Objectives: Adjustable pulmonary trunk (PT) bandingdevice may induce a more physiologic ventricle retrainingfor the two-stage Jatene operation. This experimental studyevaluates the acute hypertrophy (96 hours) of the rightventricle (RV) submitted to an intermittent pressureoverload. Methods: Five groups of seven young goats were distributedaccording to RV intermittent systolic overload duration (0,24, 48, 72 and 96 hours). The zero-hour group served as acontrol group. Echocardiographic and hemodynamicevaluations were performed daily. After completing thetraining program for each group, the animals were sacrificedfor water content and cardiac masses evaluation. Results: There was a significant increase in RV free wallthickness starting with the 48-hour group (p

SLEEP IS A COMPLEX AND HETEROGENEOUS STATE THAT ACCOUNTS FOR A SIGNIFICANT FRACTION OF THE BEHAVIORAL PATTERN EXHIBITED DURING A 24-hour period.1 The 2 states of natural sleep, namely slow-wave or synchronized sleep and rapid eye movement (REM) or desynchronized sleep depend on distinct neural mechanisms and affect the regulation of the cardiovascular system differently.2 Hemodynamic control during the sleep phases is remarkably different among species. In cats, the most detailed species studied, a slight decrease in blood pressure, cardiac output, and heart rate during synchronized sleep and desynchronized sleep phases has been previously described.3–8 The fall in blood pressure becomes more intense during desynchronized sleep, mainly due to a decrease in total vascular resistance (caused by renal and splanchnic vasodilatation) and an intense bradycardia (that contributes to decrease cardiac output). In rats, blood pressure remains unchanged and stable during synchronized sleep and increases during desynchronized sleep.9–11 In humans, Khatri and Freis12 described a small reduction in blood pressure and cardiac output during synchronized sleep in relation to the awake state. The decline in cardiac output was associated with a reduction in heart rate rather than in stroke volume, and total vascular resistance remains essentially unchanged. Conversely, blood pressure, cardiac output, total vascular resistance, heart rate, and respiratory rate have been found to be increased during the desynchronized sleep phase.12 These hemodynamic adjustments, which occur during synchronized and desynchronized sleep phases, could be a consequence of a baroreflex buffering the blood pressure levels. Indeed, arterial baroreflex is one of the most powerful and rapid mechanisms for blood pressure control, buffering the short-term pressure fluctuations that accompany the 24-hour period.13 Several techniques can be used to evaluate the relative role of baroreceptors on hemodynamic control.14 Sinoaortic denervation (SAD) is a well-established experimental technique,9–11,15–17 described in rats in our laboratory,18 that involves the differentiation and resection of aortic and carotid baroreceptors. During the sleep phases, we have also reported that rats that undergo SAD have a paradoxic blood pressure response, with increased blood pressure levels during synchronized sleep and marked hypotension during desynchronized sleep.9–11 However, the hemodynamic behavior during the sleep phases after baroreceptor deafferentation has not been completely explored in the rat. To our knowledge, no previous study in rats has analyzed the changes in cardiac output and vascular resistance during sleep. In the present study, our objective was to determine the relative role of arterial baroreflex on hemodynamic parameters during the synchronized and desynchronized sleep phases of natural sleep in rats, especially the changes observed in cardiac output and total and regional vascular resistance.

The influence of genetic factors on complex phenotypes is context dependent, posing a challenge to quantify the role of single gene variants on this process. Moreover, redundancy and reserve capacity among control systems prevent most physiological stimuli to destabilize these processes. To test whether small gene perturbation can disrupt this equilibrium under pathological conditions, mice harboring one, two, or three copies of the angiotensin converting enzyme ( Ace) gene were submitted to 3 and 6 wk of pressure overload (PO). Direct systolic blood pressure (SBP), as an index of cardiac afterload, and left ventricle mass index (LVMI) were measured. LVMI under normotension was the same regardless of the Ace genotype, but the slopes of the LVMI/SBP curves increased in the three- vs. one-copy group by ∼50% upon 3- or 6-wk PO. Angiotensin II AT1 receptor blocker treatment produced a significant pressure independent decrease in the LVMI/SBP ratio. Unlike the one-copy group, PO resulted in a significant reduction in angiotensinogen and an increase in Ace mRNA expression accompanied by an increase in cardiac angiotensin II levels in the three-copy group. Similarly, the human ACE D gene variant influenced cardiac mass, estimated by Sokolov-Lyon index, in a sample of 1,507 individuals from an urban population only in individuals in the 4th quartile of the blood pressure distribution. Collectively, these data provide direct evidence that ACE gene dosage per se does not influence cardiac mass but upon a pathological stimulus, such as elevation in blood pressure, it modulates cardiac mass in both mice and humans.

Exercise training improves arterial baroreflex control in heart failure (HF) rabbits. However, the mechanisms involved in the amelioration of baroreflex control are unknown. We tested the hypothesis that exercise training would increase the afferent aortic depressor nerve activity (AODN) sensitivity in ischemic-induced HF rats. Twenty ischemic-induced HF rats were divided into trained ( n = 11) and untrained ( n = 9) groups. Nine normal control rats were also studied. Power spectral analysis of pulse interval, systolic blood pressure, renal sympathetic nerve activity (RSNA), and AODN were analyzed by means of autoregressive parametric spectral and cross-spectral algorithms. Spontaneous baroreflex sensitivity of heart rate (HR) and RSNA were analyzed during spontaneous variation of systolic blood pressure. Left ventricular end-diastolic pressure was higher in HF rats compared with that in the normal control group ( P = 0.0001). Trained HF rats had a peak oxygen uptake higher than untrained rats and similar to normal controls ( P = 0.01). Trained HF rats had lower low-frequency [1.8 ± 0.2 vs. 14.6 ± 3 normalized units (nu), P = 0.0003] and higher high-frequency (97.9 ± 0.2 vs. 85.0 ± 3 nu, P = 0.0005) components of pulse interval than untrained rats. Trained HF rats had higher spontaneous baroreceptor sensitivity of HR (1.19 ± 0.2 vs. 0.51 ± 0.1 ms/mmHg, P = 0.003) and RSNA [2.69 ± 0.4 vs. 1.29 ± 0.3 arbitrary units (au)/mmHg, P = 0.04] than untrained rats. In HF rats, exercise training increased spontaneous AODN sensitivity toward normal levels (trained HF rats, 1,791 ± 215; untrained HF rats, 1,150 ± 158; and normal control rats, 2,064 ± 327 au/mmHg, P = 0.05). In conclusion, exercise training improves AODN sensitivity in HF rats.

We tested a preemptive combined cell/gene therapy strategy of skeletal myoblasts transfected with Ad(5)RSVVEGF-165 in an ischemia/reperfusion rat model to increase collateral blood flow to nonischemic heart tissue.Lewis rats were injected with placebo (Control), 10(6) skeletal myoblasts (SkM), or 10(6) skeletal myoblasts transfected with Ad(5)RSVVEGF-165 (SkM(+)) into the left ventricle 1week before ischemia. Left ventricle end-diastolic pressure, scar area, and capillary density were assessed 4weeks later.Local expression of human vascular endothelial growth factor was accompanied by an increase in capillary density in the SkM(+) group compared with that in the SkM and Control groups (700+/-40 vs. 289+/-18 and 318+/-59capillaries/mm(2), respectively; p0.05). After 3weeks, the myocardial scar area was reduced in SkM(+) vs. Control (5.3+/-0.4% and 14.8+/-1.6%, p0.05), while injected cells alone (SkM) did not cause improvement compared with Control (11.8+/-2.1% vs. 14.8+/-1.6%, p0.05). The decrease in the scar area in SkM(+) was accompanied by an increase in the capillary density compared with that in SkM and Control 30days after cell injection (1005+/-108 vs. 524+/-16 and 528+/-26capillaries/mm(2), respectively; p0.05). The scar areas were discrete (5.3-14.8%) and left ventricle end-diastolic pressure in all groups were comparable (p0.05).The combined cell/gene therapy strategy of genetically modified myoblast cells expressing angiogenic factors injected into the myocardium induced capillary formation and prevented the extension and development of cardiac damage associated with ischemia/reperfusion in rats.

SUMMARY 1 In chronic hypertension, the baroreceptors reset to hypertensive levels with a decrease in gain sensitivity, but only a few studies have evaluated baroreceptor resetting during chronic hypotension and, under these conditions, no consistent information is available concerning changes in baroreceptor gain sensitivity. Therefore, in the present study, the aortic baroreceptor function curve and the baroreflex control of heart rate (HR) were evaluated in chronic hypotension produced by myocardial infarction (MI) with no heart failure. 2 Aortic baroreceptor function curves were studied in anaesthetized three groups of rats: (i) MI-7, six rats 7 days after MI; (ii) MI-30, nine rats 30 days after MI; and (iii) five control animals (SHAM). The pressure–nerve activity relationship was measured during rapid changes in blood pressure by integrating the whole-nerve activity of the baroreceptors in a computerized beat-to-beat analysis. 3 Both long-term periods (7 or 30 days) of hypotension were accompanied by complete resetting of the baroreceptor in rats (the leftward displacement of the baroreceptor curve matched the decrease in blood pressure). Moreover, the resetting of the baroreceptor function curve was not accompanied by changes in gain sensitivity (1.47, 1.64 and 1.67%/mmHg for SHAM, MI-7 and MI-30 groups, respectively) and the baroreflex control of HR was normal comparing SHAM and MI-30 groups (bradycardic 1.62 ± 0.18 vs 1.99 ± 0.52 b.p.m./mmHg, respectively; tachycardic 3.6 ± 0.5 vs 4.1 ± 0.4 b.p.m./mmHg for, respectively). 4 The data indicate that the resetting of baroreceptors in chronic hypotension is stable and is not accompanied by changes in gain sensitivity, as observed in hypertension. This may account for the normal baroreflex control of HR observed in non-anaesthetized rats.

UCP3's exact physiological function in lipid handling in skeletal and cardiac muscle remains unknown. Interestingly, etomoxir, a fat oxidation inhibitor and strong inducer of UCP3, is proposed for treating both diabetes and heart failure. We hypothesize that the upregulation of UCP3 upon etomoxir serves to protect mitochondria against lipotoxicity. To evaluate UCP3's role in skeletal muscle (skm) and heart under lipid-challenged conditions, the effect of UCP3 ablation was examined in a state of dysbalance between fat availability and oxidative capacity. Wild type (WT) and UCP3(-/-) mice were subjected to high-fat feeding for 14 days. From day 6 onwards, they were given either saline or etomoxir. Etomoxir treatment induced an increase in markers of lipotoxicity in skm compared to saline. This increase upon etomoxir was similar for both, WT and UCP3(-/-) mice, suggesting that UCP3 does not play a role in protection against lipotoxicity. Interestingly, we observed 25 % mortality in UCP3(-/-)s upon etomoxir administration vs. 11 % in WTs. This increased mortality in UCP3(-/-) compared to WT mice could not be explained by differences in cardiac lipotoxicity, apoptosis, fibrosis (histology, immunohistochemistry), oxidative capacity (respirometry) or function (echocardiography). Electrophysiology demonstrated, however, prolonged QRS and QTc intervals and greater susceptibility to ventricular tachycardia upon programmed electrical stimulation in etomoxir-treated UCP3(-/-)s versus WTs. Isoproterenol administration after pacing resulted in 75 % mortality in UCP3(-/-)s vs. 14 % in WTs. Our results argue against a protective role for UCP3 on skm metabolism under lipid overload, but suggest UCP3 to be crucial in prevention of arrhythmias upon lipid-challenged conditions.

Here we present the application of our adeno-associated virus (AAV2) vector where transgene expression is driven by a synthetic, p53-responsive promoter, termed PG, used to supply human vascular endothelial growth factor-A165 (VEGF-A). Thus, p53 is harnessed to promote the beneficial expression of VEGF-A encoded by the AAVPG vector, bypassing the negative effect of p53 on HIF-1α which occurs during cardiac hypertrophy. Wistar rats were submitted to pressure overload induced by thoracic aorta coarctation (TAC) with or without concomitant gene therapy (intramuscular delivery in the left ventricle). After 12 weeks, rats receiving AAVPG-VEGF gene therapy were compared to those that did not, revealing significantly improved cardiac function under hemodynamic stress, lack of fibrosis and reversal of capillary rarefaction. With these functional assays, we have demonstrated that application of the AAVPG-VEGF vector under physiologic conditions known to stimulate p53 resulted in the preservation of cardiac performance.

Peripheral muscle dysfunction is a key mechanism contributing to exercise intolerance (i.e. breathlessness and fatigue) in heart failure patients with preserved ejection fraction (HFpEF); however, the underlying molecular and cellular mechanisms remain unknown. We therefore used an animal model to elucidate potential molecular, mitochondrial, histological, and functional alterations induced by HFpEF in the diaphragm and soleus, while also determining the possible benefits associated with exercise training.Female Dahl salt-sensitive rats were fed a low (CON; n = 10) or high salt (HFpEF; n = 11) diet of 0.3% or 8% NaCl, respectively, or a high salt diet in combination with treadmill exercise training (n = 11). Compared with low-salt rats, high-salt rats developed (P0.05) HFpEF. Compared with CON, the diaphragm of HFpEF rats demonstrated (P0.05): a fibre type shift from fast-to-slow twitch; fibre atrophy; a decreased pro-oxidative but increased anti-oxidant capacity; reduced proteasome activation; impaired in situ mitochondrial respiration; and in vitro muscle weakness and increased fatigability. The soleus also demonstrated numerous alterations (P0.05), including fibre atrophy, decreased anti-oxidant capacity, reduced mitochondrial density, and increased fatigability. Exercise training, however, prevented mitochondrial and functional impairments in both the diaphragm and soleus (P0.05).Our findings are the first to demonstrate that HFpEF induces significant molecular, mitochondrial, histological, and functional alterations in the diaphragm and soleus, which were attenuated by exercise training. These data therefore reveal novel mechanisms and potential therapeutic treatments of exercise intolerance in HFpEF.

Heart failure with preserved ejection fraction (HFpEF) is a condition that accounts for approximately 50 % of heart failure cases with the prevalence increased with advancing age. As of now, no effective treatment is available for HFpEF, which calls for continued efforts towards novel therapies. Dahl salt-sensitive (Dahl SS) rats have recently been reported as an experimental model of HFpEF, although a specific diagnostic criteria for HFpEF is still unclear in rodents. We aimed to provide clear criteria to identify HFpEF in Dahl SS rats. After a follow-up of 28 weeks, adult female Dahl SS rats receiving high salt (HS, 8 % NaCl) diet developed chronic hypertension (209 ± 80 vs. 147 ± 55 mm Hg; P 2 , respectively; P < 0.05) and EF > 50 % (67.7 ± 1.5 %). Evidence that HS rats have developed HFpEF was observed only in rats with left atrial dimension (LAD)/body weight (BW), E/A, and E/E’ ratios above the 75 th percentile of the LS group (17.50 mm/kg, 1.53, and 14.25, respectively). In addition, HS rats diagnosed with HFpEF had increased LV end-diastolic pressure and plasma NT-proBNP compared to LS rats (12.8 ± 3.4 vs. 5.8 ± 0.8 mm Hg, and 78.7 ± 18.0 vs. 17.7 ± 3.5 pg/mL, respectively; P < 0.05), while no significant changes in LAD/BW, E/A, E/E’, and plasma NT-proBNP were demonstrated in HS rats not matching the suggested criteria for HFpEF. Distance run was not different between HS and LS groups. Survival rate was 39.9 % in HS compared to 94.7 % in LS rats ( P = 0.0001), with stroke as the main cause of death (69.6 % incidence in HS rats). These results provide the first clear criteria for diagnosis of HFpEF in Dahl SS rats. Our findings have important implications for future preclinical studies aiming to develop novel therapeutic strategies targeting diastolic dysfunction in HFpEF.

Abstract —Exercise training attenuates arterial hypertension and increases baroreflex sensitivity in spontaneous hypertension. However, no information exists regarding the portion of the baroreflex arch in which this attenuation takes place. We tested the hypothesis that exercise training increases the afferent pathway sensitivity of baroreflex control in both normotensive and spontaneously hypertensive rats (SHR). Arterial pressure and whole-nerve activity of the aortic baroreceptor (multifiber preparation) were evaluated in 30 male rats assigned to 4 groups: sedentary and exercise-trained normotensive rats and sedentary and exercise-trained SHR. Exercise training was performed on a motor treadmill, 5 d/wk for 60 minutes, gradually progressing toward a speed of 26.8 m/min. Exercise training reduced mean arterial pressure in conscious exercise-trained SHR (183±4 versus 165±7 mm Hg). The relation between changes in aortic baroreceptor discharge and changes in systolic arterial pressure increased significantly in exercise-trained normotensive rats (2.09±0.1 versus 1.44±0.1%/mm Hg) and exercise-trained SHR (0.92±0.1 versus 0.71±0.1%/mm Hg) compared with their respective sedentary rats. Likewise, the average aortic baroreceptor gain sensitivity (calculated by logistic equation) was significantly higher in exercise-trained normotensive rats (2.25±0.19 versus 1.77±0.03%/mm Hg) and exercise-trained SHR (1.07±0.04 versus 0.82±0.05%/mm Hg) compared with their respective sedentary control rats. In conclusion, exercise training increases aortic baroreceptor gain sensitivity in normotensive and SHR, thus improving baroreceptor sensitivity, which may result in a more efficient arterial pressure regulation by the baroreflexes.

Abstract We studied the effects of acute and chronic exercise on the arterial baroreflex and chemosensitive cardiopulmonary baroreflex (CCB) in spontaneously hypertensive rats (SHR). Arterial baroreflex and CCB were evaluated in normotensive rats (NR, n=11) and SHR (n=5) at rest and after 30 minutes of an acute bout of exercise (45 minutes at 50% of V o 2 max). In addition, these baroreflexes were evaluated in sedentary (n=5) and exercise-trained (n=9) SHR. Exercise training was performed on a motor treadmill, 5 days/week, during 60 minutes, at 50% of V o 2 max. Baroreflex bradycardia and tachycardia, analyzed by baroreflex sensitivity index (Δ heart rate/Δ mean arterial pressure), were significantly lower in SHR than in NR (0.7±0.1 versus 2.0±0.1 and 1.8±0.2 versus 3.4±0.1 beats per minute [bpm]/mm Hg, respectively). During the recovery period from acute exercise, baroreflex bradycardia was significantly higher than at rest only in SHR (1.7±0.1 versus 0.7±0.1 bpm/mm Hg). Hypotension and bradycardia induced by CCB stimulation (5-hydroxytryptamine, IV) were similar between SHR and NR, and an acute exercise bout did not change these responses. Exercise training markedly improved baroreflex bradycardia and tachycardia in SHR (1.9±0.1 versus 0.7±0.1 and 2.9±0.1 versus 1.8±0.2 bpm/mm Hg, respectively). Exercise-trained rats had greater bradycardiac (118±26 versus 14±2 and 209±30 versus 19±5 bpm to 1 and 2 μg/kg 5-HT, respectively) and hypotensive (30±6 versus 15±3 and 45±7 versus 17±2 mm Hg to 1 and 2 μg/kg 5-hydroxytryptamine, respectively) responses to CCB stimulation. In conclusion, an acute bout of exercise increases baroreflex bradycardia in SHR, and exercise training attenuates hypertension concomitant with improved arterial baroreflex and CCB sensitivity in SHR.

MicroRNAs (miRNAs) are a class of non-coding RNAs of similar to 22 nucleotides in length, and constitute a novel class of gene regulators by imperfect base-pairing to the 3'UTR of protein encoding messenger RNAs. Growing evidence indicates that miRNAs are implicated in several pathological processes in myocardial disease. The past years, we have witnessed several profiling attempts using high-density oligonucleotide array-based approaches to identify the complete miRNA content (miRNOME) in the healthy and diseased mammalian heart. These efforts have demonstrated that the failing heart displays differential expression of several dozens of miRNAs. While the total number of experimentally validated human miRNAs is roughly two thousand, the number of expressed miRNAs in the human myocardium remains elusive. Our objective was to perform an unbiased assay to identify the miRNOME of the human heart, both under physiological and pathophysiological conditions. We used deep sequencing and bioinformatics to annotate and quantify microRNA expression in healthy and diseased human heart (heart failure secondary to hypertrophic or dilated cardiomyopathy). Our results indicate that the human heart expresses >800 miRNAs, the majority of which not being annotated nor described so far and some of which being unique to primate species. Furthermore, >250 miRNAs show differential and etiology-dependent expression in human dilated cardiomyopathy (DCM) or hypertrophic cardiomyopathy (HCM). The human cardiac miRNOME still possesses a large number of miRNAs that remain virtually unexplored. The current study provides a starting point for a more comprehensive understanding of the role of miRNAs in regulating human heart disease.

Traditional pulmonary artery banding (PAB) is not always suitable for mature subpulmonary ventricle retraining. We sought to assess in detail the myocardial morphologic adaptations of two different protocols for inducing right ventricular (RV) hypertrophy in an adult animal model.Eighteen adult goats were distributed into three groups: sham (no systolic overload), traditional (continuous systolic overload), and intermittent (daily 12-hour systolic overload). Systolic overload was adjusted to achieve a 0.7 RV-to-aortic pressure ratio. All animals underwent weekly echocardiographic studies, and hemodynamic evaluations were performed 3 times a week. After 4 weeks, the animals were humanely killed for morphologic assessment.A 37.2% increase was observed in the RV wall thickness of the intermittent group (p0.05), but no significant echocardiographic changes were observed in the other two groups. The intermittent and traditional groups had a 55.7% and 36.7% increase in RV mass, respectively, compared with the sham group (p0.05). No differences were observed in myocardial water content of the three groups (p=0.27). RV myocardial fiber and nuclei diameters were increased in the intermittent group compared with the sham group (p0.05). The area of collagen deposition in the RV interstitium was increased 98% in traditional group compared with the sham group (p0.05). No significant cellular proliferation occurred in any group.This study suggests that a more effective and harmless hypertrophy can be achieved in adult animals using intermittent PAB compared with the traditional approach.

The name of the fifth author was spelled incorrectly. The correct name is: Serve Olieslagers.

BackgroundVentricle retraining with abrupt systolic overload can cause myocardial edema and necrosis, followed by late ventricular failure. Intermittent systolic overload could minimize the inadequacy of conventional pulmonary artery banding. The present study compared ventricle function under dobutamine stress in 2 protocols of systolic overload in young goats.MethodsNineteen young goats were divided into 3 groups: sham (n = 7; no systolic pressure overload), continuous (n = 6; systolic overload maintained for 96 hours), and intermittent (n = 6; 4 periods of 12-hour systolic overload, paired with a 12-hour resting period). Echocardiographic and hemodynamic evaluations were performed daily. The myocardial performance index and ejection fraction were evaluated at rest and during dobutamine stress. The goats were then killed for morphologic evaluation.ResultsThe intermittent group underwent less systolic overload than the continuous group (P

We tested the hypothesis that small changes in angiotensin I-converting enzyme (ACE) expression can alter the vascular response to injury. Male mice containing one, two, three, and four copies of the Ace gene with no detectable vascular abnormality or changes in blood pressure were submitted to cuff-induced femoral artery injury. Femoral thickening was higher in 3- and 4-copy mice (42.4 ± 4.3% and 45.7 ± 6.5%, respectively) compared with 1- and 2-copy mice (8.3 ± 1.3% and 8.5 ± 0.9%, respectively). Femoral ACE levels from control and injured vessels were assessed in 1- and 3-copy Ace mice, which represent the extremes of the observed response. ACE vascular activity was higher in 3- vs. 1-copy Ace mice (2.4-fold, P < 0.05) in the control uninjured vessel. Upon injury, ACE activity significantly increased in both groups [2.41-fold and 2.14-fold ( P < 0.05) for 1- and 3-copy groups, respectively] but reached higher levels in 3- vs. 1-copy Ace mice ( P < 0.05). Pharmacological interventions were then used as a counterproof and to indirectly assess the role of angiotensin II (ANG II) on this response. Interestingly, ACE inhibition (enalapril) and ANG II AT1 receptor blocker (losartan) reduced intima thickening in 3-copy mice to 1-copy mouse values ( P < 0.05) while ANG II treatment significantly increased intima thickening in 1-copy mice to 3-copy mouse levels ( P < 0.05). Together, these data indicate that small physiologically relevant changes in ACE, not associated with basal vascular abnormalities or blood pressure levels, do influence the magnitude of cuff-induced neointima thickening in mice.

Adjustable pulmonary trunk (PT) banding device may induce a more physiologic ventricle retraining for the two-stage Jatene operation. This experimental study evaluates the acute hypertrophy (96 hours) of the right ventricle (RV) submitted to an intermittent pressure overload.Five groups of seven young goats were distributed according to RV intermittent systolic overload duration (0, 24, 48, 72 and 96 hours). The zero-hour group served as a control group. Echocardiographic and hemodynamic evaluations were performed daily. After completing the training program for each group, the animals were sacrificed for water content and cardiac masses evaluation.There was a significant increase in RV free wall thickness starting with the 48-hour group (p0.05). However, a decreased RV ejection fraction, associated with an important RV dilation and a significant increase in the RV volume to mass ratio was observed at 24-hour training period, when compared to 96-hour period (p=0.003), with subsequent recovery throughout the protocol. A 104.7% increase in RV mass was observed in the 96-hour group, as compared to the control group, with no differences in water content between these two groups. The daily mean increase in RV mass during the study period was 21.6% +/- 26.8%. The rate of RV mass acquisition for the overall study period of intermittent systolic overload was 0.084 g/h +/- 0.035 g/h.Intermittent PT banding has allowed a significant RV mass acquisition in the 96-hour trained group. No myocardial water content changes were observed in this group, suggesting an increased myocardial protein synthesis.

Background Tachycardia is commonly observed in hypertensive patients, predominantly mediated by regulatory mechanisms integrated within the autonomic nervous system. The genetic loci and genes associated with increased heart rate in hypertension, however, have not yet been identified. Methods An F2 intercross of Spontaneously Hypertensive Rats (SHR) × Brown Norway (BN) linkage analysis of quantitative trait loci mapping was utilized to identify candidate genes associated with an increased heart rate in arterial hypertension. Results Basal heart rate in SHR was higher compared to that of normotensive BN rats (365 ± 3 vs. 314 ± 6 bpm, p < 0.05 for SHR and BN, respectively). A total genome scan identified one quantitative trait locus in a 6.78 cM interval on rat chromosome 8 (8q22–q24) that was responsible for elevated heart rate. This interval contained 241 genes, of which 65 are known genes. Conclusion Our data suggest that an influential genetic region located on the rat chromosome 8 contributes to the regulation of heart rate. Candidate genes that have previously been associated with tachycardia and/or hypertension were found within this QTL, strengthening our hypothesis that these genes are, potentially, associated with the increase in heart rate in a hypertension rat model.

The time-course of changes in renal sympathetic nerve activity (RSNA), arterial and cardiopulmonary baroreflexes sensitivities was evaluated in conscious rats eight hours (8 h) and ten days (10 day) after myocardial infarction (MI), induced by coronary artery ligation. RSNA was recorded by a platinum electrode implanted in left renal nerve. Arterial and cardiopulmonary baroreflexes sensitivities were evaluated by changes in blood pressure and serotonin administration, respectively. Both 8 h and 10 day groups presented hypotension (103 ± 4 vs. 102 ± 2 vs. 115 ± 4 mm Hg), but only 8 h showed tachycardia (422 ± 22 vs. 378 ± 11 vs. 384 ± 9 bpm) when compared to Control rats. RSNA was depressed 8 h after MI and increased in 10 day group (12 ± 2 vs. 39 ± 8 vs. 22 ± 2 mV/cycle). Although arterial baroreflex control of heart rate was similar in all groups, the arterial baroreflex control of RSNA in 8 h group was impaired during reductions (− 0.35 ± 0.10 vs. − 1.66 ± 0.23 vs. − 0.09 ± 0.14 mV/cycle/mm Hg) or increases (− 0.77 ± 0.17 vs. − 1.63 ± 0.58 vs. − 1.66 ± 0.17 mV/cycle/mm Hg) in blood pressure when compared to Control animals. Moreover, cardiopulmonary baroreflex bradycardic response was increased in 8 h rats and normalized in 10 day group. The results suggest that the increased cardiopulmonary baroreflex sensitivity in 8 h may contribute to the reduction in the tonic level of RSNA as well as in the impairment of the baroreflex control of RSNA in the presence of hypotension.

Dynamic exercise training has been recommended as an antihypertensive therapy and as a way to modify the effects of many cardiovascular risk factors (Arakawa, 1993; Arroll and Beaglehole, 1992; Kelly and McClellan, 1994: see references 1-3 in the paper). However, the mechanisms underlying the blood-pressure lowering effect of chronic exercise are still poorly understood. It has been suggested that a decrease in sympathetic tone is one of the major effects elicited by chronic exercise on the cardiovascular system. The importance of the sympathetic component is confirmed in this review, since it was found that in spontaneously hypertensive rats (SHR) a marked decrease in sympathetic activity occurred after exercise training. Moreover, our findings suggest that this effect is mediated by improving the depressed baroreceptor function, which is, in part, responsible for the attenuation of the baroreflex sensitivity observed in the sedentary SHR (Krieger et al., 1998, 1999; see references 4 and 5 in the paper).

P43 We have previously demonstrated that low-intensity exercise traning (ET) diminishes blood pressure and partially restores the sensitivity of the baroreflex bradycardia and tachycardia that are depressed in spontaneously hypertensive rats (SHR). Presently, the influence of the exercise trainig (ET) on the afferent part of the baroreflex (baroreceptor function curve) and its implication on the baroreflex was analysed in SHR and in normal control rats (NCR). NCR and SHR were subdivided in sedentary (S) and ET groups: SHR-S (n=8) and SHR-ET (n=6), and NCR-S (n=8) and NCR-ET (n=8). ET was performed on treadmill, during 60 min, 5 days/wk, at 50% of VO 2 max, for 12 wk. Arterial baroreflex sensitivity was evaluated by bradycardiac responses to phenylephrine (0.5;1;2;4;8 and 16 μg/ml, i.v.) and tachycardiac responses to sodium nitroprusside (0.5;1;2;4;8 and 16 μg/ml, i.v.). Aortic baroreceptor function curve was evaluated under pentobarbital anesthesia (40 mg/kg) during rapid variations of arterial pressure (AT/CODAS, 3kHz per channel). The relationship between changes in baroreceptor discharge (0-100%) and systolic arterial pressure was analysed using a sigmoidal regression. Mean arterial pressure was reduced in SHR-ET compared to SHR-S group (165±7 vs. 183±4 mmHg) but remained inaltared in NCR-ET compared to NCR-S (112±3 vs. 115±3 mmHg). In SHR, ET increased the sensitivity of baroreflex bradycardia (1.9±0.1 vs. 0.7±0.1 bpm.mmHg -1 ) and tachycardia (2.9±0.1 vs. 1.8±0.2 bpm.mmHg -1 ) which were depressed. In NCR-ET baroreflex bradycardia was decreased (1.4±0.1 vs. 1.7±0.1 bpm.mmHg -1 ) but baroreflex tachycardia was increased (4.6±0.5 vs. 3.0±0.2 bpm.mmHg -1 ). ET increased the aortic baroreceptor gaing-sensitivity in both groups: SHR (0.9±0.1 vs. 0.7±0.1 %.mmHg -1 ) and NCR (2.1±0.1 vs. 1.4±0.1 %.mm Hg -1 ). Conclusion:1. ET increases aortic baroreceptor gain-sensitivity in NCR as well in SHR; 2. The improvement of the baroreflex produced by ET in SHR is partially explainedd by the recovery of the baroreceptor sensitivity, which may also participate in the reduction of high blood pressure.

The autonomic nervous system plays an important role in physiological and pathological conditions, and has been extensively evaluated by parametric and non-parametric spectral analysis. To compare the results obtained with fast Fourier transform (FFT) and the autoregressive (AR) method, we performed a comprehensive comparative study using data from humans and rats during pharmacological blockade (in rats), a postural test (in humans), and in the hypertensive state (in both humans and rats). Although postural hypotension in humans induced an increase in normalized low-frequency (LFnu) of systolic blood pressure, the increase in the ratio was detected only by AR. In rats, AR and FFT analysis did not agree for LFnu and high frequency (HFnu) under basal conditions and after vagal blockade. The increase in the LF/HF ratio of the pulse interval, induced by methylatropine, was detected only by FFT. In hypertensive patients, changes in LF and HF for systolic blood pressure were observed only by AR; FFT was able to detect the reduction in both blood pressure variance and total power. In hypertensive rats, AR presented different values of variance and total power for systolic blood pressure. Moreover, AR and FFT presented discordant results for LF, LFnu, HF, LF/HF ratio, and total power for pulse interval. We provide evidence for disagreement in 23% of the indices of blood pressure and heart rate variability in humans and 67% discordance in rats when these variables are evaluated by AR and FFT under physiological and pathological conditions. The overall disagreement between AR and FFT in this study was 43%.

ObjectiveIncreased myocardial glucose-6-phosphate dehydrogenase (G6PD) activity occurs in heart failure. This study compared G6PD activity in 2 protocols of right ventricle (RV) systolic overload in young goats.MethodsTwenty-seven goats were separated into 3 groups: sham (no overload), continuous (continuous systolic overload), and intermittent (four 12-hour periods of systolic overload paired with a 12-hour resting period). During a 96-hour protocol, systolic overload was adjusted to achieve a 0.7 RV/aortic pressure ratio. Echocardiographic and hemodynamic evaluations were performed before and after systolic overload every day postoperatively. After the study period, the animals were humanely killed for morphologic and G6PD tissue activity assessment.ResultsA 92.1% and 46.5% increase occurred in RV and septal mass, respectively, in the intermittent group compared with the sham group; continuous systolic overload resulted in a 37.2% increase in septal mass. A worsening RV myocardial performance index occurred in the continuous group at 72 hours and 96 hours, compared with the sham (P