[Show abstract][Hide abstract] ABSTRACT: Despite optimal therapy, the morbidity and mortality of patients presenting with an acute myocardial infarction (MI) remain significant, and the initial mechanistic trigger of myocardial "ischaemia/reperfusion (I/R) injury" remains greatly unexplained. Here we show that factors released from the damaged cardiac tissue itself, in particular extracellular RNA (eRNA) and tumour-necrosis-factor α (TNF-α), may dictate I/R injury. In an experimental in vivo mouse model of myocardial I/R as well as in the isolated I/R Langendorff-perfused rat heart, cardiomyocyte death was induced by eRNA and TNF-α. Moreover, TNF-α promoted further eRNA release especially under hypoxia, feeding a vicious cell damaging cycle during I/R with the massive production of oxygen radicals, mitochondrial obstruction, decrease in antioxidant enzymes and decline of cardiomyocyte functions. The administration of RNase1 significantly decreased myocardial infarction in both experimental models. This regimen allowed the reduction in cytokine release, normalisation of antioxidant enzymes as well as preservation of cardiac tissue. Thus, RNase1 administration provides a novel therapeutic regimen to interfere with the adverse eRNA-TNF-α interplay and significantly reduces or prevents the pathological outcome of ischaemic heart disease.
Thrombosis and Haemostasis 10/2014; 112(6). · 5.76 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Ischaemic heart disease (IHD) remains the leading cause of death and disability worldwide. As a result, novel therapies are still needed to protect the heart from the detrimental effects of acute ischaemia-reperfusion injury, in order to improve clinical outcomes in IHD patients. In this regard, although a large number of novel cardioprotective therapies discovered in the research laboratory have been investigated in the clinical setting, only a few of these have been demonstrated to improve clinical outcomes. One potential reason for this lack of success may have been the failure to thoroughly assess the cardioprotective efficacy of these novel therapies in suitably designed preclinical experimental animal models. Therefore, the aim of this Position Paper by the European Society of Cardiology Working Group Cellular Biology of the Heart is to provide recommendations for improving the preclinical assessment of novel cardioprotective therapies discovered in the research laboratory, with the aim of increasing the likelihood of success in translating these new treatments into improved clinical outcomes.
[Show abstract][Hide abstract] ABSTRACT: Pre-, post-, and remote conditioning of the myocardium are well described adaptive responses that markedly enhance the ability of the heart to withstand a prolonged ischemia/reperfusion insult and provide therapeutic paradigms for cardioprotection. Nevertheless, more than 25 years after the discovery of ischemic preconditioning, we still do not have established cardioprotective drugs on the market. Most experimental studies on cardioprotection are still undertaken in animal models, in which ischemia/reperfusion is imposed in the absence of cardiovascular risk factors. However, ischemic heart disease in humans is a complex disorder caused by, or associated with, cardiovascular risk factors and comorbidities, including hypertension, hyperlipidemia, diabetes, insulin resistance, heart failure, altered coronary circulation, and aging. These risk factors induce fundamental alterations in cellular signaling cascades that affect the development of ischemia/reperfusion injury per se and responses to cardioprotective interventions. Moreover, some of the medications used to treat these risk factors, including statins, nitrates, and antidiabetic drugs, may impact cardioprotection by modifying cellular signaling. The aim of this article is to review the recent evidence that cardiovascular risk factors and their medication may modify the response to cardioprotective interventions. We emphasize the critical need to take into account the presence of cardiovascular risk factors and concomitant medications when designing preclinical studies for the identification and validation of cardioprotective drug targets and clinical studies. This will hopefully maximize the success rate of developing rational approaches to effective cardioprotective therapies for the majority of patients with multiple risk factors.
[Show abstract][Hide abstract] ABSTRACT: The mechanism through which the protein kinase Akt (also called PKB), protects the heart against acute ischaemia-reperfusion injury (IRI) is not clear. Here, we investigate whether Akt mediates its cardioprotective effect by modulating mitochondrial morphology. Transfection of HL-1 cardiac cells with constitutively active Akt (caAkt) changed mitochondrial morphology as evidenced by an increase in the proportion of cells displaying predominantly elongated mitochondria (73 ± 5.0 % caAkt vs 49 ± 5.8 % control: N=80 cells/group; p< 0.05). This effect was associated with delayed time taken to induce mitochondrial permeability transition pore (MPTP) opening (by 2.4 ± 0.5 fold; N=80 cells/group: p< 0.05); and reduced cell death following simulated IRI (32.8 ± 1.2 % caAkt vs 63.8 ± 5.6 % control: N=320 cells/group: p< 0.05). Similar effects on mitochondrial morphology, MPTP opening, and cell survival post-IRI, were demonstrated with pharmacological activation of Akt using the known cardioprotective cytokine, erythropoietin (EPO). The effect of Akt on inducing mitochondrial elongation was found to be dependent on the mitochondrial fusion protein, Mitofusin-1 (Mfn1), as ablation of Mfn1 in mouse embryonic fibroblasts (MEFs) abrogated Akt-mediated mitochondrial elongation. Finally, in vivo pre-treatment with EPO reduced myocardial infarct size (as a % of the area at risk) in adult mice subjected to IRI (26.2 ± 2.6 % with EPO vs 46.1 ± 6.5 % in control; N=7/group: p< 0.05), and reduced the proportion of cells displaying myofibrillar disarray and mitochondrial fragmentation observed by electron microscopy in adult murine hearts subjected to ischaemia from 5.8 ± 1.0 % to 2.2 ± 1.0 % (N=5 hearts/group; p< 0.05). In conclusion, we found that either genetic or pharmacological activation of Akt protected the heart against acute ischaemia-reperfusion injury by modulating mitochondrial morphology.
[Show abstract][Hide abstract] ABSTRACT: Remote ischaemic preconditioning (RIPC), using brief cycles of limb ischaemia/reperfusion, is a non-invasive, low-cost intervention that may reduce perioperative myocardial injury (PMI) in patients undergoing cardiac surgery. We investigated whether RIPC can also improve short-term clinical outcomes.
[Show abstract][Hide abstract] ABSTRACT: Hypoxia inducible factor-1 (HIF-1) has been reported to promote tolerance against acute myocardial ischemia-reperfusion injury (IRI). However, the mechanism through which HIF-1 stabilization actually confers this cardioprotection is not clear. We investigated whether HIF-1¦Á stabilization protects the heart against acute IRI by preventing the opening of the mitochondrial permeability transition pore (MPTP) and the potential mechanisms involved.
[Show abstract][Hide abstract] ABSTRACT: Fetal cardiomyocyte adaptation to low levels of oxygen in utero is incompletely understood, and is of interest as hypoxia tolerance is lost after birth, leading to vulnerability of adult cardiomyocytes. It is known that cardiac mitochondrial morphology, number and function change significantly following birth, although the underlying molecular mechanisms and physiological stimuli are undefined. Here we show that the decrease in cardiomyocyte HIF-signaling in cardiomyocytes immediately after birth acts as a physiological switch driving mitochondrial fusion and increased postnatal mitochondrial biogenesis. We also investigated mechanisms of ATP generation in embryonic cardiac mitochondria. We found that embryonic cardiac cardiomyocytes rely on both glycolysis and the tricarboxylic acid cycle to generate ATP, and that the balance between these two metabolic pathways in the heart is controlled around birth by the reduction in HIF signaling. We therefore propose that the increase in ambient oxygen encountered by the neonate at birth acts as a key physiological stimulus to cardiac mitochondrial adaptation.
Journal of Molecular and Cellular Cardiology 06/2014; · 5.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A number of 'proof-of-concept' trials suggest that remote ischaemic preconditioning (RIPC) reduces surrogate markers of end-organ injury in patients undergoing major cardiovascular surgery. To date, few studies have involved hard clinical outcomes as primary end-points.
[Show abstract][Hide abstract] ABSTRACT: The morbidity and mortality from ischemic heart disease (IHD) remain significant worldwide. The treatment for acute myocardial infarction has improved over the past decades, including early reperfusion of occluded coronary arteries. Although it is essential to re-open the artery as soon as possible, paradoxically this leads to additional myocardial injury, called acute ischemia-reperfusion injury (IRI), for which currently no effective therapy is available. Therefore, novel therapeutic strategies are required to protect the heart from acute IRI in order to reduce myocardial infarction size, preserve cardiac function and improve clinical outcomes in patients with IHD. In this review article, we will first outline the pathophysiology of acute IRI and review promising therapeutic strategies for cardioprotection. These include novel aspects of mitochondrial function, epigenetics, circadian clocks, the immune system, microvesicles, growth factors, stem cell therapy and gene therapy. We discuss the therapeutic potential of these novel cardioprotective strategies in terms of pharmacological targeting and clinical application.
[Show abstract][Hide abstract] ABSTRACT: Mitochondria are critical for sustaining life, not only as the essential powerhouses of cells but as critical mediators of cell survival and death. Mitochondrial dysfunction has been identified as a key perturbation underlying numerous pathologies including myocardial ischemia-reperfusion injury and the subsequent development of impaired left ventricular systolic function and compensatory cardiac hypertrophy. This article outlines the role of mitochondrial dysfunction in these important cardiac pathologies and highlights current cardioprotective strategies and their clinical efficacy in acute myocardial infarction and heart failure patients. Finally, we explore novel mitochondrial targets and evaluate their potential future translation for clinical cardioprotection.
[Show abstract][Hide abstract] ABSTRACT: Novel therapeutic targets are required to protect the heart against cell death from acute ischemia-reperfusion injury (IRI). Mutations in the DJ-1 (PARK7) gene in dopaminergic neurons induce mitochondrial dysfunction and a genetic form of Parkinson's disease. Genetic ablation of DJ-1 renders the brain more susceptible to cell death following ischemia-reperfusion in a model of stroke. Although DJ-1 is present in the heart, its role there is currently unclear. We sought to investigate whether mitochondrial DJ-1 may protect the heart against cell death from acute IRI by preventing mitochondrial dysfunction. Overexpression of DJ-1 in HL-1 cardiac cells conferred the following beneficial effects: reduced cell death following simulated IRI (30.4±4.7% with DJ-1 versus 52.9±4.7% in control; n=5, P<0.05); delayed mitochondrial permeability transition pore (MPTP) opening (a critical mediator of cell death) (260±33 s with DJ-1 versus 121±12 s in control; n=6, P<0.05); and induction of mitochondrial elongation (81.3±2.5% with DJ-1 versus 62.0±2.8% in control; n=6 cells, P<0.05). These beneficial effects of DJ-1 were absent in cells expressing the non-functional DJ-1(L166P) and DJ-1(Cys106A) mutants. Adult mice devoid of DJ-1 (KO) were found to be more susceptible to cell death from in vivo IRI with larger myocardial infarct sizes (50.9±3.5% DJ-1 KO versus 41.1±2.5% in DJ-1 WT; n≥7, P<0.05) and resistant to cardioprotection by ischemic preconditioning. DJ-1 KO hearts showed increased mitochondrial fragmentation on electron microscopy, although there were no differences in calcium-induced MPTP opening, mitochondrial respiratory function or myocardial ATP levels. We demonstrate that loss of DJ-1 protects the heart from acute IRI cell death by preventing mitochondrial dysfunction. We propose that DJ-1 may represent a novel therapeutic target for cardioprotection.
Cell Death & Disease 02/2014; 5:e1082. · 6.04 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In adult cardiomyocytes there are known to be 3 different sub-populations of mitochondria: Subsarcolemmal (SS), Interfibrillar (IF), and Perinuclear (PN), which appear to have distinctive properties and shapes. Here, we investigate the mitochondrial morphology of these 3 different mitochondrial subgroups and examine the effect of ischaemia on these parameters.
We investigated mitochondrial morphology in adult rodent hearts at baseline and in response to ischemia using electron microscopy (x3000 magnification). In six randomly selected electron micrographs of longitudinally-arranged cardiomyocytes, the following characteristics were determined (approx. 500 mitochondria in total): (1) Surface area (SA); (2) Fret's diameter (the longest distance between any points in a given mitochondrion); (3) Perimeter; (4) Aspect ratio: the length-to-width ratio; (5) Roundness: 4. SA/π.major axis2; (6) Circularity: 4π. SA/perimeter2 (1.0 indicates a perfect circle and as the value approaches zero it indicates an elongated shape).
In adult rat cardiomyocytes (n = 3 animals), IF mitochondria have a significantly greater SA [μm(2)] (IF 0.37 ± 1.08 vs. SS 0.30 ± 1.07, PN 0.27 ± 1.03; P < 0.0001), Fret's diameter (IF 0.93 ± 0.02 vs. SS 0.83 ± 0.02, PN 0.79 ± 0.18, P < 0.0001), and Perimeter (IF 2.48 ± 1.08 vs. SS 2.21 ± 1.07 PN 2.11 ± 1.03, P < 0.0001), when compared to SSM and PN mitochondria. However, there were no major differences in Aspect ratio, Roundness or Circularity. IF mitochondria in the ischaemic adult murine heart had a smaller Area, Perimeter, Fret's diameter, Aspect ratio, and a greater degree of Roundness and Circularity.
Interfibrillar mitochondria are generally larger and longer than SSM and PN mitochondria. In response to ischaemia, the IF mitochondria undergo fragmentation, generating smaller more rounded mitochondria.
[Show abstract][Hide abstract] ABSTRACT: Despite state-of-the-art reperfusion therapy, morbidity and mortality remains significant in patients with an acute myocardial infarction. Therefore, novel strategies to limit myocardial ischemia-reperfusion injury are urgently needed. Mineralocorticoid receptor (MR) antagonists are attractive candidates for this purpose, since several clinical trials in patients with heart failure have reported a survival benefit with MR antagonist treatment. MRs are expressed by several cells of the cardiovascular system, including cardiomyocytes, cardiac fibroblasts, vascular smooth muscle cells, and endothelial cells. Experiments in animal models of myocardial infarction have demonstrated that acute administration of MR antagonists, either before ischemia or immediately at the moment of coronary reperfusion, limit infarct size. This action appears to be independent of the presence of aldosterone and cortisol, which are the endogenous ligands for the MR. The cardioprotective effect is mediated by a nongenomic intracellular signaling pathway, including adenosine receptor stimulation, and activation of several components of the Reperfusion Injury Salvage Kinase (RISK) pathway. In addition to limiting infarct size, MR antagonists can improve scar healing when administered shortly after reperfusion and can reduce cardiac remodeling post myocardial infarction when initiated after initial infarct healing. Clinical trials are currently being performed studying whether early administration of MR antagonists can indeed improve prognosis in patients with an acute myocardial infarction, independent of the presence of heart failure.
[Show abstract][Hide abstract] ABSTRACT: The dipeptidyl peptidase-4 (DPP-4) inhibitors Sitagliptin and Vildagliptin lower blood glucose by augmenting endogenous levels of glucagon-like peptide-1 (GLP-1), an incretin which also confers cardioprotection. As such, we hypothesized that treatment with DPP-4 inhibitors are also cardioprotective.
In ex vivo experiments: Male Sprague--Dawley rats were randomized to receive by oral gavage either Vildagliptin (20 mg/kg/day), Sitagliptin (100 mg/kg/day), or water for 2 weeks. Excised hearts were Langendorff-perfused with buffer containing either 5 mmol/L or 11 mmol/L glucose and subjected to 35 minutes ischaemia/120 minutes reperfusion. In in vivo experiments: Male young Wistar and Sprague--Dawley rats, middle aged Wistar and Goto-Kakizaki diabetic rats were randomized to receive by oral gavage either Sitagliptin (100 mg/kg/day), or water for 2 weeks. Rats were then subjected to 30 minutes ischaemia/120 minutes reperfusion and infarct size ascertained.
Two weeks pre-treatment with either Vildagliptin or Sitagliptin reduced ex vivo myocardial infarction (MI) size in hearts perfused with buffer containing 11 mmol/L glucose but not 5 mmol/L glucose. This effect was abolished by Exendin 9--39 (GLP-1 receptor antagonist) and H-89 (PKA antagonist). Treatment of perfused hearts with native GLP-1 was also glucose-sensitive, reducing MI size, at glucose concentrations 7, 9, and 11 mmol/L but not at 5 mmol/L. Finally, Sitagliptin reduced in vivo MI size in middle aged Wistar (7-8 mmol/L glucose) and Goto-Kakizaki (9-10 mmol/L glucose) rats where blood glucose was elevated, but not in young Wistar (5 mmol/L glucose) or Sprague--Dawley (5 mmol/L glucose) rats, where blood glucose was normal.
We find that chronic treatment with DPP-4 inhibitors reduced MI size, via the GLP-1 receptor-PKA pathway, in a glucose-dependent manner. Glucose-sensitive cardioprotection of endogenous GLP-1 in diabetic patients may in part explain why intensive control of serum glucose levels has been associated with increased cardiovascular risk.
[Show abstract][Hide abstract] ABSTRACT: Mutations in PTEN inducible kinase-1 (PINK1) induce mitochondrial dysfunction in dopaminergic neurons resulting in an inherited form of Parkinson's disease. Although PINK1 is present in the heart its exact role there is unclear. We hypothesized that PINK1 protects the heart against acute ischemia reperfusion injury (IRI) by preventing mitochondrial dysfunction.
Over-expressing PINK1 in HL-1 cardiac cells reduced cell death following simulated IRI (29.2±5.2% PINK1 versus 49.0±2.4% control; N = 320 cells/group P<0.05), and delayed the onset of mitochondrial permeability transition pore (MPTP) opening (by 1.3 fold; P<0.05). Hearts excised from PINK1+/+, PINK1+/- and PINK1-/- mice were subjected to 35 minutes regional ischemia followed by 30 minutes reperfusion. Interestingly, myocardial infarct size was increased in PINK1-/- hearts compared to PINK1+/+ hearts with an intermediate infarct size in PINK1+/- hearts (25.1±2.0% PINK1+/+, 38.9±3.4% PINK1+/- versus 51.5±4.3% PINK1-/- hearts; N>5 animals/group; P<0.05). Cardiomyocytes isolated from PINK1-/- hearts had a lower resting mitochondrial membrane potential, had inhibited mitochondrial respiration, generated more oxidative stress during simulated IRI, and underwent rigor contracture more rapidly in response to an uncoupler when compared to PINK1+/+ cells suggesting mitochondrial dysfunction in hearts deficient in PINK1.
We show that the loss of PINK1 increases the heart's vulnerability to ischemia-reperfusion injury. This may be due, in part, to increased mitochondrial dysfunction. These findings implicate PINK1 as a novel target for cardioprotection.
PLoS ONE 06/2013; 8(4):e62400. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: AIMS: Old age and diabetes are risk factors which often coexist increasing the vulnerability of the heart to the lethal effects of ischemia-reperfusion injury (IRI). However, to our knowledge, no investigations have examined IRI and cardioprotective signalling in animal models bearing these comorbidies concomitantly. The ability of the heart to recover following IRI is greatly dependent on its innate cardioprotective potential, in which a central role is played by Akt. We aimed to investigate in an aging diabetic rat model, the susceptibility of the heart to IRI, the achievability of ischemic preconditioning (IPC) against this lethal event and the changes in Akt signalling, as the main prosurvival intracellular pathway.Methods and ResultsOur data showed that the isolated hearts of aged, diabetic Goto-Kakizaki rats were more susceptible to sub lethal injury and not amenable to cardioprotection via IPC, compared to younger diabetic rat hearts. Western blot analysis of the heart tissue suggested a chronic up regulation of Akt phosphorylation, and reduced expression of the mitochondrial regulator PGC-1α and of the antioxidant enzyme catalase, potentially due to the Akt up regulation. Moreover, no further activation of Akt could be achieved following IPC. CONCLUSIONS: An increased susceptibility to IRI in the aged, diabetic heart could be a consequence of impaired Akt signalling due to chronic Akt phosphorylation. Additional Akt phosphorylation required for IPC protection may therefore not be possible in the aged diabetic rat heart and may explain why this cardioprotective maneuver cannot be achieved in these hearts.
Cardiovascular Research 05/2013; · 5.81 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: OBJECTIVES: The aim of this study was to determine the accuracy of the contrast "bolus only" T1 mapping cardiac magnetic resonance (CMR) technique for measuring myocardial extracellular volume fraction (ECV). BACKGROUND: Myocardial ECV can be measured with T1 mapping before and after contrast agent if the contrast agent distribution between blood/myocardium is at equilibrium. Equilibrium distribution can be achieved with a primed contrast infusion (equilibrium contrast-CMR [EQ-CMR]) or might be approximated by the dynamic equilibration achieved by delayed post-bolus measurement. This bolus only approach is highly attractive, but currently limited data support its use. We compared the bolus only technique with 2 independent standards: collagen volume fraction (CVF) from myocardial biopsy in aortic stenosis (AS); and the infusion technique in 5 representative conditions. METHODS: One hundred forty-seven subjects were studied: healthy volunteers (n = 50); hypertrophic cardiomyopathy (n = 25); severe AS (n = 22); amyloid (n = 20); and chronic myocardial infarction (n = 30). Bolus only (at 15 min) and infusion ECV measurements were performed and compared. In 18 subjects with severe AS the results were compared with histological CVF. RESULTS: The ECV by both techniques correlated with histological CVF (n = 18, r(2) = 0.69, p < 0.01 vs. r(2) = 0.71, p < 0.01, p = 0.42 for comparison). Across health and disease, there was strong correlation between the techniques (r(2) = 0.97). However, in diseases of high ECV (amyloid, hypertrophic cardiomyopathy late gadolinium enhancement, and infarction), Bland-Altman analysis indicates the bolus only technique has a consistent and increasing offset, giving a higher value for ECVs above 0.4 (mean difference ± limit of agreement for ECV <0.4 = -0.004 ± 0.037 vs. ECV >0.4 = 0.040 ± 0.075, p < 0.001). CONCLUSIONS: Bolus only, T1 mapping-derived ECV measurement is sufficient for ECV measurement across a range of cardiac diseases, and this approach is histologically validated in AS. However, when ECV is >0.4, the bolus only technique consistently measures ECV higher compared with infusion.