Rebecca Parodi-Rullan

Direct Renin Inhibition Exerts an Anti-hypertrophic Effect Associated with Improved Mitochondrial Function in Post-infarction Heart Failure in Diabetic Rats

Background: In addition to hypertension control, direct renin inhibition has been shown to exert direct beneficial effects on the heart in post-infarction cardiac remodeling. This study elucidates the possible contribution of mitochondria to the anti-hypertrophic effects of the direct renin inhibitor aliskiren in post-infarction heart failure complicated with diabetes in rats. Methods: Diabetes was induced in male Sprague-Dawley rats by a single injection of streptozotocin (IP, 65 mg/kg body weight). After 7 days, the animals were randomly assigned to 4 groups: sham, heart failure, sham+aliskiren, and heart failure+aliskiren. Post-infarction HF was induced by coronary artery ligation for 4 weeks. Results: showed that heart failure reduced ejection fraction and cardiac output by 41% (P<0.01) and 42% (P<0.05), respectively, compared to sham-operated hearts. Cardiac dysfunction was associated with suppressed state 3 respiration rates and respiratory control index in mitochondria, and increased mitochondrial permeability transition pore (PTP) opening. In addition, heart failure reduced expression of the major mitochondrial sirtuin, SIRT3 and increased acetylation of cyclophilin D, a regulatory component of the PTP. Aliskiren significantly improved cardiac function and abrogated mitochondrial perturbations. Conclusion: Our results demonstrate that aliskiren attenuates post-infarction remodeling which is associated with its beneficial effects on mitochondria.

Mitochondria-targeted ROS scavenger improves post-ischemic recovery of cardiac function and attenuates mitochondrial abnormalities in aged rats

Mitochondria-generated reactive oxygen species (ROS) play a crucial role in the pathogenesis of aging and age-associated diseases. In this study, we evaluated the effects of XJB-5-131 (XJB), a mitochondria-targeted ROS and electron scavenger, on cardiac resistance to ischemia-reperfusion (IR)-induced oxidative stress in aged rats. Male adult (5-month old, n=17) and aged (29-month old, n=19) Fischer Brown Norway (F344/BN) rats were randomly assigned to the following groups: adult (A), adult+XJB (AX), aged (O), and aged+XJB (OX). XJB was administered 3 times per week (3mg/kg body weight, IP) for four weeks. At the end of the treatment period, cardiac function was continuously monitored in excised hearts using the Langendorff technique for 30 min, followed by 20 min of global ischemia, and 60-min reperfusion. XJB improved post-ischemic recovery of aged hearts, as evidenced by greater left ventricular developed-pressures and rate-pressure products than the untreated, aged-matched group. The state 3 respiration rates at complexes I, II and IV of mitochondria isolated from XJB-treated aged hearts were 57% (P<0.05), 25% (P<0.05) and 28% (P<0.05), respectively, higher than controls. Ca(2+)-induced swelling, an indicator of permeability transition pore opening, was reduced in the mitochondria of XJB-treated aged rats. In addition, XJB significantly attenuated the H2O2-induced depolarization of the mitochondrial inner membrane as well as the total and mitochondrial ROS levels in cultured cardiomyocytes. This study underlines the importance of mitochondrial ROS in aging-induced cardiac dysfunction and suggests that targeting mitochondrial ROS may be an effective therapeutic approach to protect the aged heart against IR injury.

The Role of PPARα in Metformin-Induced Attenuation of Mitochondrial Dysfunction in Acute Cardiac Ischemia/Reperfusion in Rats

Metformin, an anti-diabetic drug, exerts cardioprotection against ischemia-reperfusion (IR) through the activation of AMPK. However, the molecular mechanisms underlying these beneficial effects remain elusive. In this study, we examined the role of PPARα in mediating cardioprotective effects of metformin on mitochondria. Hearts of male Sprague-Dawley rats perfused by Langendorff were subjected to IR in the presence or absence of metformin and the PPARβ inhibitor, GW6471. IR reduced cardiac function and compromised the structural integrity of cardiac cells evidenced by increased LDH release from the hearts. In addition, IR induced mitochondrial dysfunction as evidenced by reduced respiration and increased mitochondrial permeability transition pore (PTP) opening. However, metformin-treated hearts demonstrated improved post-ischemic recovery of cardiac function and reduced cell death that were associated with increased state 3 respiration at complexes I and II (by 27% and 32%, respectively, both p < 0.05) and decreased PTP opening (by 27%, p < 0.05) compared to untreated hearts. The protective effects of metformin on cardiac function and mitochondria were blocked by GW6471. Thus, our results demonstrate that inhibition of PPARα attenuates the beneficial effects of metformin on mitochondria in acute IR.

Targeting the mitochondrial permeability transition: cardiac ischemia-reperfusion versus carcinogenesis.

Cardiovascular diseases and cancer continue to be major causes of death worldwide, and despite intensive research only modest progress has been reached in reducing the morbidity and mortality of these awful diseases. Mitochondria are broadly accepted as the key organelles that play a crucial role in cell life and death. They provide cells with ATP produced via oxidative phosphorylation under physiological conditions, and initiate cell death through both apoptosis and necrosis in response to severe stress. Oxidative stress accompanied by calcium overload and ATP depletion induces the mitochondrial permeability transition (mPT) with formation of pathological, non-specific mPT pores (mPTP) in the mitochondrial inner membrane. Opening of the mPTP with a high conductance results in matrix swelling ultimately inducing rupture of the mitochondrial outer membrane and releasing pro-apoptotic proteins into the cytoplasm. The ATP level is the determining factor in deciding whether cells die through apoptosis or necrosis. Cardiac cells undergoing ischemia followed by reperfusion (IR) possess exactly the same conditions mentioned above to induce mPTP opening. Due to its critical role in cell death, inhibition of mPTP opening has been accepted as a major therapeutic approach to protect the heart against IR. In contrast to cardiac IR, cancer cells exhibit less sensitivity to pore opening which can be in part explained by increased expression of mPTP compounds/modulators and metabolic remodeling. Since the main goal of chemotherapy is to provoke apoptosis, mPT induction may represent an attractive approach for the development of new cancer therapeutics to induce mitochondria-mediated cell death and prevent cell differentiation in carcinogenesis. This review focuses on the role of the mPTP in cardiac IR and cancer, and pharmacological agents to prevent or initiate mPT-mediated cell death, respectively in these diseases.

High Sensitivity of SIRT3 Deficient Hearts to Ischemia-Reperfusion Is Associated with Mitochondrial Abnormalities

Aim: Sirtuins are NAD+-dependent deacetylases that regulate cell metabolism through protein acetylation/deacetylation, and SIRT3 is the major deacetylase among mitochondrial isoforms. Here, we elucidated the possible role of acetylation of cyclophilin D, a key regulator of the mitochondrial permeability transition pore (mPTP), in mitochondria-mediated cardiac dysfunction induced by ischemia-reperfusion (IR) in wild type (WT) and SIRT3 knockout (SIRT3-/-) mice. Materials and Methods: Isolated and Langendorff-mode perfused hearts of WT and SIRT3-/- mice were subjected to 25-min global ischemia followed by 60-min of reperfusion in the presence or absence of the mPTP inhibitor, sanglifehrin A (SfA). Results: Analysis of mitochondrial sirtuins demonstrated that SIRT3 deficiency upregulated SIRT4 with no effect on SIRT5 expression. Hearts of SIRT3-/- mice exhibited significantly less recovery of cardiac function at the end of IR compared to WT mice. Intact (non-perfused) SIRT3-/- hearts exhibited an increased rate of Ca2+-induced swelling in mitochondria as an indicator of mPTP opening. However, there was no difference in mPTP opening and cyclophilin D acetylation between WT and SIRT3-/- hearts subjected to IR injury. Ca2+-stimulated H2O2 production was significantly higher in SIRT3-/- mitochondria that was prevented by SfA. Superoxide dismutase activity was lower in SIRT3-/- heart mitochondria subjected to IR which correlated with an increase in protein carbonylation. However, mitochondrial DNA integrity was not affected in SIRT3-/- hearts after IR. Conclusion: SIRT3 deficiency exacerbates cardiac dysfunction during post-ischemic recovery, and increases mPTP opening and ROS generation without oxidative damage to mitochondrial proteins and DNA.

Mitochondrial permeability transition in cardiac ischemia–reperfusion: whether cyclophilin D is a viable target for cardioprotection?

Growing number of studies provide strong evidence that the mitochondrial permeability transition pore (PTP), a non-selective channel in the inner mitochondrial membrane, is involved in the pathogenesis of cardiac ischemia–reperfusion and can be targeted to attenuate reperfusion-induced damage to the myocardium. The molecular identity of the PTP remains unknown and cyclophilin D is the only protein commonly accepted as a major regulator of the PTP opening. Therefore, cyclophilin D is an attractive target for pharmacological or genetic therapies to reduce ischemia–reperfusion injury in various animal models and humans. Most animal studies demonstrated cardioprotective effects of PTP inhibition; however, a recent large clinical trial conducted by international groups demonstrated that cyclosporine A, a cyclophilin D inhibitor, failed to protect the heart in patients with myocardial infarction. These studies, among others, raise the question of whether cyclophilin D, which plays an important physiological role in the regulation of cell metabolism and mitochondrial bioenergetics, is a viable target for cardioprotection. This review discusses previous studies to provide comprehensive information on the physiological role of cyclophilin D as well as PTP opening in the cell that can be taken into consideration for the development of new PTP inhibitors.

Acetylation of Mitochondrial Proteins in the Heart: The Role of SIRT3

A growing number of studies have demonstrated the role of post-translational modifications of proteins, particularly acetylation, in human diseases including neurodegenerative and cardiovascular diseases, diabetes, cancer, and in aging. Acetylation of mitochondrial proteins has been shown to be involved in the pathogenesis of cardiac diseases such as myocardial infarction (ischemia-reperfusion) and heart failure. Indeed, over 60% of mitochondrial proteins contain acetylation sites, and most of these proteins are involved in mitochondrial bioenergetics. Mitochondrial non-enzymatic acetylation is enabled by acetyl-coenzyme A abundance and serves as the primary pathway of acetylation in mitochondria. Hence, regulation of enzymatic deacetylation becomes the most important mechanism to control acetylation/deacetylation of mitochondrial proteins. Acetylation/deacetylation of mitochondrial proteins has been regarded as a key regulator of mitochondrial metabolism and function. Proteins are deacetylated by NAD+-dependent deacetylases known as sirtuins (SIRTs). Among seven sirtuin isoforms, only SIRT3, SIRT4, and SIRT5 are localized in the mitochondria. SIRT3 is the main mitochondrial sirtuin which plays a key role in maintaining metabolic and redox balance in the mitochondria under physiological and pathological conditions. SIRT3 regulates the enzymatic activity of proteins involved in fatty acid oxidation, tricarboxylic acid cycle, electron transport chain, and oxidative phosphorylation. Although many enzymes have been identified as targets for SIRT3, cardiac-specific SIRT3 effects and regulations could differ from those in non-cardiac tissues. Therefore, it is important to elucidate the contribution of SIRT3 and mitochondrial protein acetylation/deacetylation in mitochondrial metabolism and cardiac dysfunction. Here, we summarize previous studies and provide a comprehensive analysis of the role of SIRT3 in mitochondria metabolism and bioenergetics under physiological conditions and in cardiac diseases. In addition, the review discusses mitochondrial protein acetylation as a potential target for cardioprotection.

Divergent Effects of Cyclophilin-D Inhibition on the Female Rat Heart: Acute Versus Chronic Post-Myocardial Infarction

Background/Aims: The mitochondrial permeability transition pore opening plays a critical role in the pathogenesis of myocardial infarction. Inhibition of cyclophilin-D (CyP-D), a key regulator of the mitochondrial permeability transition pore, has been shown to exert cardioprotective effects against ischemia-reperfusion injury on various animal models, mostly in males. However, failure of recent clinical trials requires a detailed elucidation of the cardioprotective efficacy of CyP-D inhibition. The aim of this study was to examine whether cardioprotective effects of sanglifehrin A, a potent inhibitor of CyP-D, on post-infarcted hearts depends on reperfusion. Methods: Acute or chronic myocardial infarction was induced by coronary artery ligation with/without subsequent reperfusion for 2 and 28 days in female Sprague-Dawley rats. Cardiac function was estimated by echocardiography. Oxygen consumption rates, ROS production, permeability transition pore opening, protein carbonylation and respiratory supercomplexes were analyzed in isolated cardiac mitochondria. Results: Sanglifehrin A significantly improved cardiac function of reperfused hearts at 2 days but failed to protect after 28 days. No protection was observed in non-reperfused post-infarcted hearts. The respiratory control index of mitochondria was significantly reduced in reperfused infarcted hearts at 2-days with no effect at 28-days post-infarction on reperfused and non-reperfused hearts. Likewise, only a minor increase in reactive oxygen species production was observed at 2-days in non-reperfused post-infarcted hearts. Conclusion: This study demonstrates that CyP-D inhibition exerts cardioprotective effects in reperfused but not in non-reperfused infarcted hearts of female rats, and the effects are observed only during acute post-infarction injury.