Emiliano Medei

Chronic treatment with anabolic steroids induces ventricular repolarization disturbances: cellular, ionic and molecular mechanism.

The illicit use of supraphysiological doses of androgenic steroids (AAS) has been suggested as a cause of arrhythmia in athletes. The objectives of the present study were to investigate the time-course and the cellular, ionic and molecular processes underlying ventricular repolarization in rats chronically treated with AAS. Male Wistar rats were treated weekly for 8 weeks with 10mg/kg of nandrolone decanoate (DECA n=21) or vehicle (control n=20). ECG was recorded weekly. Action potential (AP) and transient outward potassium current (I(to)) were recorded in rat hearts. Expression of KChIP2, Kv1.4, Kv4.2, and Kv4.3 was assessed by real-time PCR. Hematoxylin/eosin and Picrosirius red staining were used for histological analysis. QTc was greater in the DECA group. After DECA treatment the left, but not right, ventricle showed a longer AP duration than did the control. I(to) current densities were 47.5% lower in the left but not in the right ventricle after DECA. In the right ventricle the I(to) inactivation time-course was slower than in the control group. After DECA the left ventricle showed lower KChIP2 ( approximately 26%), Kv1.4 ( approximately 23%) and 4.3 ( approximately 70%) expression while the Kv 4.2 increased in 4 ( approximately 250%) and diminished in 3 ( approximately 30%) animals of this group. In the right ventricle the expression of I(to) subunits was similar between the treatment and control groups. DECA-treated hearts had 25% fewer nuclei and greater nuclei diameters in both ventricles. Our results strongly suggest that supraphysiological doses of AAS induce morphological remodeling in both ventricles. However, the electrical remodeling was mainly observed in the left ventricle.

Macrophage-dependent IL-1β production induces cardiac arrhythmias in diabetic mice

Diabetes mellitus (DM) encompasses a multitude of secondary disorders, including heart disease. One of the most frequent and potentially life threatening disorders of DM-induced heart disease is ventricular tachycardia (VT). Here we show that toll-like receptor 2 (TLR2) and NLRP3 inflammasome activation in cardiac macrophages mediate the production of IL-1β in DM mice. IL-1β causes prolongation of the action potential duration, induces a decrease in potassium current and an increase in calcium sparks in cardiomyocytes, which are changes that underlie arrhythmia propensity. IL-1β-induced spontaneous contractile events are associated with CaMKII oxidation and phosphorylation. We further show that DM-induced arrhythmias can be successfully treated by inhibiting the IL-1β axis with either IL-1 receptor antagonist or by inhibiting the NLRP3 inflammasome. Our results establish IL-1β as an inflammatory connection between metabolic dysfunction and arrhythmias in DM.

Antibodies with beta-adrenergic activity from chronic chagasic patients modulate the QT interval and M cell action potential duration.

AIMS The aim of this study was to investigate whether the sera from chronic chagasic patients (CChPs) with beta-1 adrenergic activity (Ab-beta) can modulate ventricular repolarization. Beta-adrenergic activity has been described in CChP. It increases the L-type calcium current and heart rate in isolated hearts, but its effects on ventricular repolarization has not been described. METHODS AND RESULTS In isolated rabbit hearts, under pacing condition, QT interval was measured under Ab-beta perfusion. Beta-adrenergic activity was also tested in guinea pig ventricular M cells. Furthermore, the immunoglobulin fraction (IgG-beta) of the Ab-beta was tested on Ito, ICa, and Iks currents in rat, rabbit, and guinea pig myocytes, respectively. Beta-adrenergic activity shortened the QT interval. This effect was abolished in the presence of propranolol. In addition, sera from CChP without beta-adrenergic activity (Ab-beta) did not modulate QT interval. The M cell action potential duration (APD) was reversibly shortened by Ab-beta. Atenolol inhibited this effect of Ab-beta, and Ab- did not modulate the AP of M cells. Ito was not modulated by isoproterenol nor by IgG-beta. However, IgG-beta increased ICa and IKs. CONCLUSION The shortening of the QT interval and APD in M cells and the increase of IKs and ICa induced by IgG-beta contribute to repolarization changes that may trigger malignant ventricular arrhythmias observed in patients with chronic chagasic or idiopathic cardiomyopathy.

Toll-like receptor 4 activation promotes cardiac arrhythmias by decreasing the transient outward potassium current (Ito) through an IRF3-dependent and MyD88-independent pathway

Cardiac arrhythmias are one of the main causes of death worldwide. Several studies have shown that inflammation plays a key role in different cardiac diseases and Toll-like receptors (TLRs) seem to be involved in cardiac complications. In the present study, we investigated whether the activation of TLR4 induces cardiac electrical remodeling and arrhythmias, and the signaling pathway involved in these effects. Membrane potential was recorded in Wistar rat ventricle. Ca(2+) transients, as well as the L-type Ca(2+) current (ICaL) and the transient outward K(+) current (Ito), were recorded in isolated myocytes after 24 h exposure to the TLR4 agonist, lipopolysaccharide (LPS, 1 μg/ml). TLR4 stimulation in vitro promoted a cardiac electrical remodeling that leads to action potential prolongation associated with arrhythmic events, such as delayed afterdepolarization and triggered activity. After 24 h LPS incubation, Ito amplitude, as well as Kv4.3 and KChIP2 mRNA levels were reduced. The Ito decrease by LPS was prevented by inhibition of interferon regulatory factor 3 (IRF3), but not by inhibition of interleukin-1 receptor-associated kinase 4 (IRAK4) or nuclear factor kappa B (NF-κB). Extrasystolic activity was present in 25% of the cells, but apart from that, Ca(2+) transients and ICaL were not affected by LPS; however, Na(+)/Ca(2+) exchanger (NCX) activity was apparently increased. We conclude that TLR4 activation decreased Ito, which increased AP duration via a MyD88-independent, IRF3-dependent pathway. The longer action potential, associated with enhanced Ca(2+) efflux via NCX, could explain the presence of arrhythmias in the LPS group.

In vitro effects of acute amiodarone and dronedarone on epicardial, endocardial, and M cells of the canine ventricle

Amiodarone (AM) is an antiarrhythmic agent widely used in the treatment of ventricular and supraventricular arrhythmias. Dronedarone (DR) is a new compound with a pharmacological profile similar to that of AM, but iodine free. We previously demonstrated that chronic AM treatment reduces transmural dispersion of repolarization (TDR) in the canine heart. We used standard microelectrode technique to evaluate the effects of acute AM (100 µM) and DR (30 µM) on epicardial (EPI), endocardial (ENDO), and M region tissues obtained from the left ventricular wall of the canine heart. Amiodarone (100 µM, 120 min of exposure) produced little change in the action potential duration of ENDO and EPI tissues, but it shortened the action potential of M cells, especially at slow rates, leading to a decrease in TDR. Similar results were observed with DR. Acute AM (100 µM) and DR (30 µM) eliminated d-sotalol—induced early afterdepolarizations (EADs) and triggered activity in 3 of 3 and 2 of 6 M cell preparations, respectively. The reduction of TDR and the elimination of EAD-induced triggered activity differentiates AM and DR from other class III agents. These effects may explain the efficacy and low arrhythmogenicity of acute AM and suggest a potential safe use of DR as an antiarrhythmic agent.

Noninvasive method for electrocardiogram recording in conscious rats: feasibility for heart rate variability analysis.

Heart rate variability (HRV) analysis consists in a well-established tool for the assessment of cardiac autonomic control, both in humans and in animal models. Conventional methods for HRV analysis in rats rely on conscious state electrocardiogram (ECG) recording based on prior invasive surgical procedures for electrodes/transmitters implants. The aim of the present study was to test a noninvasive and inexpensive method for ECG recording in conscious rats, assessing its feasibility for HRV analysis. A custom-made elastic cotton jacket was developed to fit the rats mean thoracic circumference, with two pieces of platinum electrodes attached on its inner surface, allowing ECG to be recorded noninvasively in conscious, restrained rats (n=6). Time- and frequency-domain HRV analyses were conducted, under basal and autonomic blockade conditions. High-quality ECG signals were obtained, being feasible for HRV analysis. As expected, mean RR interval was significantly decreased in the presence of atropine (p <0.05) and increased in the presence of propranolol (p<0.001). Also, reinforcing the reliability of the method, low- and high-frequency HRV spectral powers were significantly decreased in the presence of propranolol (p <0.05) and atropine (p< 0.001), respectively. In summary, the present work describes a novel, inexpensive and noninvasive method for surface ECG recording in conscious rats.

Acute In Vitro Effects of Dronedarone, an Iodine-Free Derivative, and Amiodarone, on the Rabbit Sinoatrial Node Automaticity: A Comparative Study

Amiodarone is a potent antiarrhythmic drug commonly used in the treatment of supraventricular and ventricular arrhythmias. Dronedarone is a recently developed iodine-free compound (Sanofi Recherche), structurally related to amiodarone. Amiodarone and dronedarone have shown similar long-term effects on sinoatrial node automaticity in vivo and in vitro in the rabbit heart. In the present study, we used a microelectrode technique to compare the acute in vitro electrophysiologic effects of amiodarone (100 µM) and dronedarone (100 µM) on the rabbit sinus node. Like amiodarone, dronedarone induces a marked reduction in sinus node automaticity, evidenced by decreases in spontaneous beating rate, action potential amplitude, and slope of phase 4 depolarization. Isoproterenol dose-dependently increases sinus node automaticity in the presence of either amiodarone or dronedarone. The data suggest that dronedarone may be a useful antiarrhythmic alternative to amiodarone in the treatment of supraventricular arrhythmias.

Monitoring of the Parasite Load in the Digestive Tract of Rhodnius prolixus by Combined qPCR Analysis and Imaging Techniques Provides New Insights into the Trypanosome Life Cycle

Background Here we report the monitoring of the digestive tract colonization of Rhodnius prolixus by Trypanosoma cruzi using an accurate determination of the parasite load by qPCR coupled with fluorescence and bioluminescence imaging (BLI). These complementary methods revealed critical steps necessary for the parasite population to colonize the insect gut and establish vector infection. Methodology/Principal Findings qPCR analysis of the parasite load in the insect gut showed several limitations due mainly to the presence of digestive-derived products that are thought to degrade DNA and inhibit further the PCR reaction. We developed a real-time PCR strategy targeting the T. cruzi repetitive satellite DNA sequence using as internal standard for normalization, an exogenous heterologous DNA spiked into insect samples extract, to precisely quantify the parasite load in each segment of the insect gut (anterior midgut, AM, posterior midgut, PM, and hindgut, H). Using combined fluorescence microscopy and BLI imaging as well as qPCR analysis, we showed that during their journey through the insect digestive tract, most of the parasites are lysed in the AM during the first 24 hours independently of the gut microbiota. During this short period, live parasites move through the PM to establish the onset of infection. At days 3–4 post-infection (p.i.), the parasite population begins to colonize the H to reach a climax at day 7 p.i., which is maintained during the next two weeks. Remarkably, the fluctuation of the parasite number in H remains relatively stable over the two weeks after refeeding, while the populations residing in the AM and PM increases slightly and probably constitutes the reservoirs of dividing epimastigotes. Conclusions/Significance These data show that a tuned dynamic control of the population operates in the insect gut to maintain an equilibrium between non-dividing infective trypomastigote forms and dividing epimastigote forms of the parasite, which is crucial for vector competence.

Activation of angiotensin-converting enzyme 2 improves cardiac electrical changes in ventricular repolarization in streptozotocin-induced hyperglycaemic rats

AIMS Diabetic patients present a high level of cardiac arrhythmias and risk of cardiac sudden death. The renin-angiotensin system (RAS) plays a key role in diabetes and cardiac diseases. The present study aimed to evaluate whether an angiotensin-converting enzyme 2 (ACE2) activator, diminazene aceturate (DIZE), could improve the streptozotocin (STZ)-induced electrical changes in ventricular repolarization in hyperglycaemic rats. METHODS AND RESULTS Hyperglycaemia was induced in Wistar male rats with STZ (60 mg/kg/iv). After 4 weeks of STZ injection, rats were daily treated with saline (control) or DIZE (1 mg/kg/gavage) for four consecutive weeks. The cardiac electrical function was evaluated in vivo by electrocardiogram and in vitro by cardiac action potential records in different pacing frequencies. Treatment with DIZE was not able to reverse hyperglycaemia nor body weight loss. However, DIZE reversed hyperglycaemia-induced cardiac electrical changes in ventricular repolarization. Specifically, animals treated with DIZE showed shorter QT and QTc intervals. In addition, ACE2 activation was capable to shorten the cardiac action potential and also reverse the arrhythmic markers. Diminazene aceturate treatment did not induce arrhythmic events in normal, as well as in hyperglycaemic animals. CONCLUSION Our data indicate that activation of ACE2 has a beneficial effect in hyperglycaemic rats, improving the cardiac electrical function. Thus, DIZE represents a promising new therapeutic agent to treat hyperglycaemia-induced cardiac electrical changes in ventricular repolarization.

Could a high-fat diet rich in unsaturated fatty acids impair the cardiovascular system?

BACKGROUND Dyslipidemia results from consumption of a diet rich in saturated fatty acids and is usually associated with cardiovascular disease. A diet rich in unsaturated fatty acids is usually associated with improved cardiovascular condition. OBJECTIVE To investigate whether a high-fat diet rich in unsaturated fatty acids (U-HFD) - in which fatty acid represents approximately 45% of the total calories - impairs the cardiovascular system. METHODS Male, 30-day-old Wistar rats were fed a standard (control) diet or a U-HFD containing 83% unsaturated fatty acid for 19 weeks. The in vivo electrocardiogram, the spectral analysis of heart rate variability, and the vascular reactivity responses to phenylephrine, acetylcholine, noradrenaline and prazosin in aortic ring preparations were analyzed to assess the cardiovascular parameters. RESULTS After 19 weeks, the U-HFD rats had increased total body fat, baseline glucose levels and feed efficiency compared with control rats. However, the final body weight, systolic blood pressure, area under the curve for glucose, calorie intake and heart weight⁄final body weight ratio were similar between the groups. In addition, both groups demonstrated no alteration in the electrocardiogram or cardiac sympathetic parameters. There was no difference in the responses to acetylcholine or the maximal contractile response of the thoracic aorta to phenylephrine between groups, but the concentration necessary to produce 50% of maximal response showed a decrease in the sensitivity to phenylephrine in U-HFD rats. The cumulative concentration- effect curve for noradrenaline in the presence of prazosin was shifted similarly in both groups. CONCLUSIONS The present work shows that U-HFD did not impair the cardiovascular parameters analyzed.