Evidence of anti-cardiac antibodies and complement activation was found in a large number of patients with end-stage cardiomyopathy regardless of the aetiology. Adenosine triphosphate synthase appears to be a new prominent antigenic stimulus; but more interestingly, the simultaneous co-existence of activated complement components suggests that this humoral mechanism may participate in disease progression.

Stress-induced cardiomyopathy, triggered by acute catecholamine discharge, is a syndrome characterized by transient, apical ballooning linked to acute heart failure and ventricular arrhythmias. Rats receiving an acute isoproterenol (ISO) overdose (OV) suffer cardiac apex ischemia-reperfusion damage and arrhythmia, and then undergo cardiac remodeling and dysfunction. Nevertheless, the subcellular mechanisms underlying cardiac dysfunction after acute damage subsides are not thoroughly understood. To address this question, Wistar rats received a single ISO injection (67 mg/kg). We found in vivo moderate systolic and diastolic dysfunction at 2 wk post-ISO-OV; however, systolic dysfunction recovered after 4 wk, while diastolic dysfunction worsened. At 2 wk post-ISO-OV, cardiac function was assessed ex vivo, while mitochondrial oxidative metabolism and stress were assessed in vitro, and Ca(2+) handling in ventricular myocytes. These were complemented with sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA), phospholamban (PLB), and RyR2 expression studies. Ex vivo, basal mechanical performance index (MPI) and oxygen consumption rate (MVO2) were unchanged. Nevertheless, upon increase of metabolic demand, by β-adrenergic stimulation (1-100 nM ISO), the MPI versus MVO2 relation decreased and shifted to the right, suggesting MPI and mitochondrial energy production uncoupling. Mitochondria showed decreased oxidative metabolism, membrane fragility, and enhanced oxidative stress. Myocytes presented systolic and diastolic Ca(2+) mishandling, and blunted response to ISO (100 nM), and all these without apparent changes in SERCA, PLB, or RyR2 expression. We suggest that post-ISO-OV mitochondrial dysfunction may underlie decreased cardiac contractility, mainly by depletion of ATP needed for myofilaments and Ca(2+) transport by SERCA, while exacerbated oxidative stress may enhance diastolic RyR2 activity.

Background and Purpose Despite the importance of mitochondrial Ca2+ to metabolic regulation and cell physiology, little is known about the mechanisms that regulate Ca2+ entry into the mitochondria. Accordingly, we established a system to determine the role of the mitochondrial Ca2+ uniporter in an isolated heart model, at baseline and during increased workload following β‐adrenoceptor stimulation. Experimental Approach Cardiac contractility, oxygen consumption and intracellular Ca2+ transients were measured in ex vivo perfused murine hearts. Ru360 and spermine were used to modify mitochondrial Ca2+ uniporter activity. Changes in mitochondrial Ca2+ content and energetic phosphate metabolite levels were determined. Key Results The addition of Ru360, a selective inhibitor of the mitochondrial Ca2+ uniporter, induced progressively and sustained negative inotropic effects that were dose‐dependent with an EC50 of 7 μM. Treatment with spermine, a uniporter agonist, showed a positive inotropic effect that was blocked by Ru360. Inotropic stimulation with isoprenaline elevated oxygen consumption (2.7‐fold), Ca2+‐dependent activation of pyruvate dehydrogenase (5‐fold) and mitochondrial Ca2+ content (2.5‐fold). However, in Ru360‐treated hearts, this parameter was attenuated. In addition, β‐adrenoceptor stimulation in the presence of Ru360 did not affect intracellular Ca2+ handling, PKA or Ca2+/calmodulin‐dependent PK signalling. Conclusions and Implications Inhibition of the mitochondrial Ca2+ uniporter decreases β‐adrenoceptor response, uncoupling between workload and production of energetic metabolites. Our results support the hypothesis that the coupling of workload and energy supply is partly dependent on mitochondrial Ca2+ uniporter activity.

Metabolic syndrome (MS) increases cardiovascular risk and is associated with cardiac dysfunction and arrhythmias, although the precise mechanisms are still under study. Chronic inflammation in MS has emerged as a possible cause of adverse cardiac events. Male Wistar rats fed with 30% sucrose in drinking water and standard chow for 25–27 weeks were compared to a control group. The MS group showed increased weight, visceral fat, blood pressure, and serum triglycerides. The most important increases in serum cytokines included IL-1β (7-fold), TNF-α (84%), IL-6 (41%), and leptin (2-fold), the latter also showing increased gene expression in heart tissue (35-fold). Heart function ex vivo in MS group showed a decreased mechanical performance response to isoproterenol challenge (ISO). Importantly, MS hearts under ISO showed nearly twofold the incidence of ventricular fibrillation. Healthy rat cardiomyocytes exposed to MS group serum displayed impaired contractile function and Ca2+ handling during ISO treatment, showing slightly decreased cell shortening and Ca2+ transient amplitude (23%), slower cytosolic calcium removal (17%), and more frequent spontaneous Ca2+ release events (7.5-fold). As spontaneous Ca2+ releases provide a substrate for ventricular arrhythmias, our study highlights the possible role of serum proinflammatory mediators in the development of arrhythmic events during MS.

Acetogenins are cell-membrane permeable, naturally occurring secondary metabolites of plants such as Annonaceae, Lauraceae and other related phylogenic families. They belong to the chemical derivatives of polyketides, which are synthesized from fatty acid precursors. Although acetogenins have displayed diverse biological activities, the anti-proliferative effect on human cancer cells has been widely reported. Acetogenins are inhibitors of complex I in the electron transport chain therefore they interrupt ATP synthesis in mitochondria. We tested a new acetogenins-enriched extract from the seed of Persea americana in order to investigate if any toxicity was induced on cardiac tissue and determine the involved mechanism. In isolated perfused heart we found that contractility was completely inhibited at an accumulative dose of 77 μg/ml. In isolated cardiomyocytes, the acetogenins-enriched extract induced apoptosis through the activation of the intrinsic pathway at 43 μg/ml. In isolated mitochondria, it inhibited complex I activity on NADH-linked respiration, as would be expected, but also induced permeability transition on succinate-linked respiration. Cyclosporine A, a known blocker of permeability transition, significantly prevented the permeability transition triggered by the acetogenins-enriched extract. In addition, our acetogenins-enriched extract inhibited ADP/ATP exchange, suggesting that an important element in phosphate or adenylate transport was affected. In this manner we suggest that acetogenins-enriched extract from Persea americana could directly modulate permeability transition, an entity not yet associated with the acetogenins' direct effects, resulting in cardiotoxicity.

Casiopeinas are a group of copper-based antineoplastic molecules designed as a less toxic and more therapeutic alternative to cisplatin or Doxorubicin; however, there is scarce evidence about their toxic effects on the whole heart and cardiomyocytes. Given this, rat hearts were perfused with Casiopeinas or Doxorubicin and the effects on mechanical performance, energetics, and mitochondrial function were measured. As well, the effects of Casiopeinas-triggered cell death were explored in isolated cardiomyocytes. Casiopeinas III-Ea, II-gly, and III-ia induced a progressive and sustained inhibition of heart contractile function that was dose- and time-dependent with an IC50 of 1.3 ± 0.2, 5.5 ± 0.5, and 10 ± 0.7 μM, correspondingly. Myocardial oxygen consumption was not modified at their respective IC50, although ATP levels were significantly reduced, indicating energy impairment. Isolated mitochondria from Casiopeinas-treated hearts showed a significant loss of membrane potential and reduction of mitochondrial Ca2+ retention capacity. Interestingly, Cyclosporine A inhibited Casiopeinas-induced mitochondrial Ca2+ release, which suggests the involvement of the mitochondrial permeability transition pore opening. In addition, Casiopeinas reduced the viability of cardiomyocytes and stimulated the activation of caspases 3, 7, and 9, demonstrating a cell death mitochondrial-dependent mechanism. Finally, the early perfusion of Cyclosporine A in isolated hearts decreased Casiopeinas-induced dysfunction with reduction of their toxic effect. Our results suggest that heart cardiotoxicity of Casiopeinas is similar to that of Doxorubicin, involving heart mitochondrial dysfunction, loss of membrane potential, changes in energetic metabolites, and apoptosis triggered by mitochondrial permeability.

Background: While the etiology remains largely unspecified, increasing evidence has highlighted the contribution of autoimmune mechanisms to the pathophysiology of idiopathic cardiomyopathy (iDCM). Elevated cardiac Troponin I (cTnI) has been known to be elevated in patients at risk for or post-diagnosis of adverse cardiac events. Although cTnI stimulation of peripheral blood mononuclear cells (PBMCs) from iDCM patients evoke a heightened autoimmune IL-10 response, the cell types involved in this contribution remain unknown. We hypothesize that regulatory T cells are responsible for the elevated IL-10 production, and that IL-10 and TNFα levels will serve as predictors for changes in cardiac function after diagnoses of iDCM or myocardial infarction. Method: Whole blood samples were collected from patients presenting with left ventricular ejection fraction <40% as part of the Autoimmune Responses to Cardiac Troponin I in Cardiomyopathy (ARCTIC) Study. PBMCs were isolated through density gradient centrifugation and used for ELISpot assays and FACS analysis. For ELISpot assays, MultiScreen plates (Millipore) coated with either anti-IL10 or anti-IFNγ were plated with PBMCs cultured at 300,000 cells/well in either media, human CD3, or cTnI. Development was carried out through sequential incubations of a biotinylated anticytokine antibody and streptavidin-HRP. Spot color was developed by adding aminoethyl-carbazole substrate in H2O2-acetate buffer, and the reaction was stopped with diH2O washing. Images and analysis were performed by an ImmunoSpot Series 1 ELISpot analyzer. For FACS analysis, PBMCs were stimulated by incubation in media or cTnI. Cells were labeled for with a live/dead stain, as well as antibodies for various immunospecific antibodies. Labeled cells were analyzed on a Fortessa SORP cytometer with Diva v6.0 software, and analyzed with FlowJo v9.0 software. Results: ELISpot fluorescence as a measure of IL-10 was found to be increased 5.4-fold in PBMCs stimulated by cTnI as compared to control (P < .001). No significant changes were found in TNFα after cTnI stimulation in ELISpot analysis. FACS analysis found IL-10 to be increased with cTnI stimulation by 8.5% in CD14 + and 8.5% in CD16 + monocytes, as compared to decreases of 1.9% in T-regulatory cells and 1.9% in T-Lymphocytes (P < .05). While TNFα was found to increase among these cell types with stimulation, only the 12.4% increase in CD16 + monocytes was statistically significant (P < .05). T-helper cell populations were found to be unaffected by cTnI stimulation. Conclusions: Our findings suggest that the increased the IL-10 levels found after cTnI stimulation are primarily attributed to production in CD14 + and CD16 + monocytes, rather than contributions by adaptive T-regulatory cells.