The signaling pathway for aldosterone-induced mitochondrial production of superoxide anion in the myocardium.

Mineralocorticoid receptor (MR) antagonists decrease morbidity and mortality in heart failure patients for whom oxidative stress is usual; however, the underlying mechanism for this protection is unclear. Since aldosterone stimulates reactive oxygen species (ROS) production in several tissues, we explored its effect and the intracellular pathway involved in the rat myocardium. Aldosterone dose-dependently increased O2(-) production in myocardial slices. At 10 nmol/L, aldosterone increased O2(-) to 165 ± 8.8% of control, an effect prevented not only by the MR antagonists eplerenone and spironolactone (107 ± 7.8 and 103 ± 5.3%, respectively) but also by AG1478 (105 ± 8.0%), antagonist of the EGF receptor (EGFR). Similar results were obtained by silencing MR expression through the direct intramyocardial injection of a lentivirus coding for a siRNA against the MR. The aldosterone effect on O2(-) production was mimicked by the mKATP channel opener diazoxide and blocked by preventing its opening with 5-HD and glibenclamide, implicating the mitochondria as the source of O2(-). Inhibiting the respiratory chain with rotenone or mitochondrial permeability transition (MPT) with cyclosporine A or bongkrekic acid also canceled aldosterone-induced O2(-) production. In addition, aldosterone effect depended on NADPH oxidase and phosphoinositide 3-kinase activation, as apocynin and wortmannin, respectively, inhibited it. EGF (0.1 µg/mL) similarly increased O2(-), although in this case MR antagonists had no effect, suggesting that EGFR transactivation occurred downstream from MR activation. Inhibition of mKATP channels, the respiratory chain, or MPT did not prevent Akt phosphorylation, supporting that it happened upstream of the mitochondria. Importantly, cardiomyocytes were confirmed as a source of aldosterone induced mitochondrial ROS production in experiments performed in isolated cardiac myocytes. These results allow us to speculate that the beneficial effects of MR antagonists in heart failure may be related to a decrease in oxidative stress.

Na+/H+ exchanger and cardiac hypertrophy.

Reactive cardiac hypertrophy (CH) is an increase in heart mass in response to hemodynamic overload. Exercise-induced CH emerges as an adaptive response with improved cardiac function, in contrast to pathological CH that represents a risk factor for cardiovascular health. The Na+/H+ exchanger (NHE-1) is a membrane transporter that not only regulates intracellular pH but also intracellular Na+ concentration. In the scenario of cardiovascular diseases, myocardial NHE-1 is activated by a variety of stimuli, such as neurohumoral factors and mechanical stress, leading to intracellular Na+ overload and activation of prohypertrophic cascades. NHE-1 hyperactivity is intimately linked to heart diseases, including ischemia-reperfusion injury, maladaptive CH and heart failure. In this review, we will present evidence to support that the NHE-1 hyperactivity constitutes a "switch on/off" for the pathological phenotype during CH development. We will also discuss some classical and novel strategies to avoid NHE-1 hyperactivity, and that are therefore worthwhile to improve cardiovascular health.

The positive inotropic effect of endothelin-1 is mediated by mitochondrial reactive oxygen species.

We have previously demonstrated the participation of reactive oxygen species (ROS) in the positive inotropic effect of a physiological concentration of Angiotensin II (Ang II, 1 nM). The objective of the present work was to evaluate the role and source of ROS generation in the positive inotropic effect produced by an equipotent concentration of endothelin-1 (ET-1, 0.4 nM). Isolated cat ventricular myocytes were used to measure sarcomere shortening with a video-camera, superoxide anion (()O(2)(-)) with chemiluminescence, and ROS production and intracellular pH (pH(i)) with epifluorescence. The ET-1-induced positive inotropic effect (40.4+/-3.1%, n=10, p<0.05) was associated to an increase in ROS production (105+/-29 fluorescence units above control, n=6, p<0.05). ET-1 also induced an increase in ()O(2)(-) production that was inhibited by the NADPH oxidase blocker, apocynin, and by the blockers of mitochondrial ATP-sensitive K(+) channels (mK(ATP)), glibenclamide and 5 hydroxydecanoic acid. The ET-1-induced positive inotropic effect was inhibited by apocynin (0.3 mM; 6.3+/-6.6%, n=13), glibenclamide (50 microM; 8.8+/-3.5%, n=6), 5 hydroxydecanoic acid (500 microM; 14.1+/-8.1, n=9), and by scavenging ROS with MPG (2 mM; 0.92+/-5.6%, n=8). ET-1 enhanced proton efflux (J(H)) carried by the Na(+)/H(+) exchanger (NHE) after an acid load, effect that was blocked by MPG. Consistently, the ET-induced positive inotropic effect was also inhibited by the NHE selective blocker HOE642 (5 microM; 9.37+/-6.07%, n=7). The data show that the effect of a concentration of ET-1 that induces an increase in contractility of about 40% is totally mediated by an intracellular pathway triggered by mitochondrial ROS formation and stimulation of the NHE.

Cardioprotective role of IGF-1 in the hypertrophied myocardium of the spontaneously hypertensive rats: A key effect on NHE-1 activity.

AIM: Myocardial Na+/H+ exchanger-1 (NHE-1) hyperactivity and oxidative stress are interrelated phenomena playing pivotal roles in the development of pathological cardiac hypertrophy and heart failure. Exercise training is effective to convert pathological into physiological hypertrophy in the spontaneously hypertensive rats (SHR), and IGF-1-key humoral mediator of exercise training-inhibits myocardial NHE-1, at least in normotensive rats. Therefore, we hypothesize that IGF-1 by hampering NHE-1 hyperactivity and oxidative stress should exert a cardioprotective effect in the SHR. METHODS: NHE-1 activity [proton efflux ( JH+ ) mmol L-1 min-1], expression and phosphorylation; H2O2 production; superoxide dismutase (SOD) activity; contractility and calcium transients were measured in SHR hearts in the presence/absence of IGF-1. RESULTS: IGF-1 significantly decreased NHE-1 activity ( JH+ at pHi 6.95: 1.39 ± 0.32, n = 9 vs C 3.27 ± 0.3, n = 20, P < .05); effect prevented by AG1024, an antagonist of IGF-1 receptor (2.7 ± 0.4, n = 7); by the PI3K inhibitor wortmannin (3.14 ± 0.41, n = 7); and the AKT inhibitor MK2206 (3.37 ± 0.43, n = 14). Moreover, IGF-1 exerted an antioxidant effect revealed by a significant reduction in H2O2 production accompanied by an increase in SOD activity. In addition, IGF-1 improved cardiomyocyte contractility as evidenced by an increase in sarcomere shortening and a decrease in the relaxation constant, underlined by an increase in the amplitude and rate of decay of the calcium transients. CONCLUSION: IGF-1 exerts a cardioprotective role on the hypertrophied hearts of the SHR, in which the inhibition of NHE-1 hyperactivity, as well as the positive inotropic and antioxidant effects, emerges as key players.