RESUMO
BACKGROUND: Ischemic cardiovascular diseases, particularly acute myocardial infarction (MI), is one of the leading causes of mortality worldwide. Indoleamine 2, 3-dioxygenase 1 (IDO) catalyzes 1 rate-limiting step of L-tryptophan metabolism, and emerges as an important regulator of many pathological conditions. We hypothesized that IDO could play a key role to locally regulate cardiac homeostasis after MI. METHODS: Cardiac repair was analyzed in mice harboring specific endothelial or smooth muscle cells or cardiomyocyte or myeloid cell deficiency of IDO and challenged with acute myocardial infarction. RESULTS: We show that kynurenine generation through IDO is markedly induced after MI in mice. Total genetic deletion or pharmacological inhibition of IDO limits cardiac injury and cardiac dysfunction after MI. Distinct loss of function of IDO in smooth muscle cells, inflammatory cells, or cardiomyocytes does not affect cardiac function and remodeling in infarcted mice. In sharp contrast, mice harboring endothelial cell-specific deletion of IDO show an improvement of cardiac function as well as cardiomyocyte contractility and reduction in adverse ventricular remodeling. In vivo kynurenine supplementation in IDO-deficient mice abrogates the protective effects of IDO deletion. Kynurenine precipitates cardiomyocyte apoptosis through reactive oxygen species production in an aryl hydrocarbon receptor-dependent mechanism. CONCLUSIONS: These data suggest that IDO could constitute a new therapeutic target during acute MI.
Assuntos
Células Endoteliais/metabolismo , Indolamina-Pirrol 2,3,-Dioxigenase/uso terapêutico , Cinurenina/uso terapêutico , Infarto do Miocárdio/tratamento farmacológico , Animais , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/farmacologia , Cinurenina/farmacologia , Camundongos , Infarto do Miocárdio/fisiopatologiaRESUMO
Recessive forms of catecholaminergic polymorphic ventricular tachycardia (CPVT) are induced by mutations in genes encoding triadin or calsequestrin, two proteins that belong to the Ca2+ release complex, responsible for intracellular Ca2+ release triggering cardiac contractions. To better understand the mechanisms of triadin-induced CPVT and to assay multiple therapeutic interventions, we used a triadin knockout mouse model presenting a CPVT-like phenotype associated with a decrease in calsequestrin protein level. We assessed different approaches to rescue protein expression and to correct intracellular Ca2+ release and cardiac function: pharmacological treatment with kifunensine or a viral gene transfer-based approach, using adeno-associated virus serotype 2/9 (AAV2/9) encoding the triadin or calsequestrin. We observed that the levels of triadin and calsequestrin are intimately linked, and that reduction of both proteins contributes to the CPVT phenotype. Different combinations of triadin and calsequestrin expression level were obtained using these therapeutic approaches. A full expression of each is not necessary to correct the phenotype; a fine-tuning of the relative re-expression of both triadin and calsequestrin is required to correct the CPVT phenotype and rescue the cardiac function. AAV-mediated gene delivery of calsequestrin or triadin and treatment with kifunensine are potential treatments for recessive forms of CPVT due to triadin mutations.
Assuntos
Calsequestrina/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Musculares/metabolismo , Taquicardia Ventricular/metabolismo , Alcaloides/uso terapêutico , Animais , Arritmias Cardíacas/tratamento farmacológico , Cálcio/metabolismo , Sinalização do Cálcio/genética , Calsequestrina/genética , Dependovirus , Modelos Animais de Doenças , Terapia Genética/métodos , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Camundongos , Camundongos Knockout , Proteínas Musculares/genética , Contração Miocárdica/efeitos dos fármacos , Contração Miocárdica/genética , Miócitos Cardíacos/metabolismo , Parvovirinae/genética , Fenótipo , Ratos , Taquicardia Ventricular/tratamento farmacológico , Taquicardia Ventricular/patologia , Transdução Genética , TransfecçãoRESUMO
RATIONALE: Although the second messenger cyclic AMP (cAMP) is physiologically beneficial in the heart, it largely contributes to cardiac disease progression when dysregulated. Current evidence suggests that cAMP is produced within mitochondria. However, mitochondrial cAMP signaling and its involvement in cardiac pathophysiology are far from being understood. OBJECTIVE: To investigate the role of MitEpac1 (mitochondrial exchange protein directly activated by cAMP 1) in ischemia/reperfusion injury. METHODS AND RESULTS: We show that Epac1 (exchange protein directly activated by cAMP 1) genetic ablation (Epac1-/-) protects against experimental myocardial ischemia/reperfusion injury with reduced infarct size and cardiomyocyte apoptosis. As observed in vivo, Epac1 inhibition prevents hypoxia/reoxygenation-induced adult cardiomyocyte apoptosis. Interestingly, a deleted form of Epac1 in its mitochondrial-targeting sequence protects against hypoxia/reoxygenation-induced cell death. Mechanistically, Epac1 favors Ca2+ exchange between the endoplasmic reticulum and the mitochondrion, by increasing interaction with a macromolecular complex composed of the VDAC1 (voltage-dependent anion channel 1), the GRP75 (chaperone glucose-regulated protein 75), and the IP3R1 (inositol-1,4,5-triphosphate receptor 1), leading to mitochondrial Ca2+ overload and opening of the mitochondrial permeability transition pore. In addition, our findings demonstrate that MitEpac1 inhibits isocitrate dehydrogenase 2 via the mitochondrial recruitment of CaMKII (Ca2+/calmodulin-dependent protein kinase II), which decreases nicotinamide adenine dinucleotide phosphate hydrogen synthesis, thereby, reducing the antioxidant capabilities of the cardiomyocyte. CONCLUSIONS: Our results reveal the existence, within mitochondria, of different cAMP-Epac1 microdomains that control myocardial cell death. In addition, our findings suggest Epac1 as a promising target for the treatment of ischemia-induced myocardial damage.
Assuntos
Fatores de Troca do Nucleotídeo Guanina/biossíntese , Mitocôndrias Cardíacas/metabolismo , Miócitos Cardíacos/metabolismo , Animais , Animais Recém-Nascidos , Morte Celular/fisiologia , Células Cultivadas , Insuficiência Cardíaca/metabolismo , Insuficiência Cardíaca/patologia , Humanos , Masculino , Microdomínios da Membrana/metabolismo , Microdomínios da Membrana/patologia , Camundongos , Camundongos Knockout , Mitocôndrias Cardíacas/patologia , Miócitos Cardíacos/patologia , RatosRESUMO
Besides its role in calcium (Ca2+) homeostasis, the sarco-endoplamic reticulum (SR/ER) controls protein folding and is tethered to mitochondria. Under pathophysiological conditions the unfolded protein response (UPR) is associated with disturbance in SR/ER-mitochondria crosstalk. Here, we investigated whether ER stress altered SR/ER-mitochondria links, Ca2+ handling and muscle damage in WT (Wild Type) and mdx mice, the murine model of Duchenne Muscular Dystrophy (DMD). In WT mice, the SR/ER-mitochondria links were decreased in isolated FDB muscle fibers after injection of ER stress activator tunicamycin (TM). Ca2+ imaging revealed an increase of cytosolic Ca2+ transient and a decrease of mitochondrial Ca2+ uptake. The force generating capacity of muscle dropped after TM. This impaired contractile function was accompanied by an increase in autophagy markers and calpain-1 activation. Conversely, ER stress inhibitors restored SR/ER-mitochondria links, mitochondrial Ca2+ uptake and improved diaphragm contractility in mdx mice. Our findings demonstrated that ER stress-altered SR/ER-mitochondria links, disturbed Ca2+ handling and muscle function in WT and mdx mice. Thus, ER stress may open up a prospect of new therapeutic targets in DMD.
Assuntos
Sinalização do Cálcio , Cálcio/metabolismo , Estresse do Retículo Endoplasmático , Mitocôndrias Musculares/metabolismo , Distrofia Muscular de Duchenne/metabolismo , Retículo Sarcoplasmático/metabolismo , Animais , Autofagia/genética , Calpaína/genética , Calpaína/metabolismo , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos mdx , Mitocôndrias Musculares/genética , Mitocôndrias Musculares/patologia , Contração Muscular/genética , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/patologia , Retículo Sarcoplasmático/genética , Retículo Sarcoplasmático/patologiaRESUMO
AIMS/HYPOTHESIS: Mitochondria-associated endoplasmic reticulum membranes (MAMs) are regions of the endoplasmic reticulum (ER) tethered to mitochondria and controlling calcium (Ca(2+)) transfer between both organelles through the complex formed between the voltage-dependent anion channel, glucose-regulated protein 75 and inositol 1,4,5-triphosphate receptor (IP3R). We recently identified cyclophilin D (CYPD) as a new partner of this complex and demonstrated a new role for MAMs in the control of insulin's action in the liver. Here, we report on the mechanisms by which disruption of MAM integrity induces hepatic insulin resistance in CypD (also known as Ppif)-knockout (KO) mice. METHODS: We used either in vitro pharmacological and genetic inhibition of CYPD in HuH7 cells or in vivo loss of CYPD in mice to investigate ER-mitochondria interactions, inter-organelle Ca(2+) exchange, organelle homeostasis and insulin action. RESULTS: Pharmacological and genetic inhibition of CYPD concomitantly reduced ER-mitochondria interactions, inhibited inter-organelle Ca(2+) exchange, induced ER stress and altered insulin signalling in HuH7 cells. In addition, histamine-stimulated Ca(2+) transfer from ER to mitochondria was blunted in isolated hepatocytes of CypD-KO mice and this was associated with an increase in ER calcium store. Interestingly, disruption of inter-organelle Ca(2+) transfer was associated with ER stress, mitochondrial dysfunction, lipid accumulation, activation of c-Jun N-terminal kinase (JNK) and protein kinase C (PKC)ε and insulin resistance in liver of CypD-KO mice. Finally, CYPD-related alterations of insulin signalling were mediated by activation of PKCε rather than JNK in HuH7 cells. CONCLUSIONS/INTERPRETATION: Disruption of IP3R-mediated Ca(2+) signalling in the liver of CypD-KO mice leads to hepatic insulin resistance through disruption of organelle interaction and function, increase in lipid accumulation and activation of PKCε. Modulation of ER-mitochondria Ca(2+) exchange may thus provide an exciting new avenue for treating hepatic insulin resistance.
Assuntos
Cálcio/metabolismo , Retículo Endoplasmático/metabolismo , Resistência à Insulina/fisiologia , Mitocôndrias/metabolismo , Animais , Linhagem Celular , Peptidil-Prolil Isomerase F , Ciclofilinas/genética , Ciclofilinas/metabolismo , Hepatócitos/metabolismo , Humanos , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Fígado/metabolismo , Masculino , Camundongos , Camundongos KnockoutRESUMO
BACKGROUND: The mitochondrial permeability transition pore (PTP) has been established as an important mediator of ischemia-reperfusion-induced cell death. The matrix protein cyclophilin D (CypD) is the best known regulator of PTP opening. Therefore, the authors hypothesized that isoflurane, by inhibiting the respiratory chain complex I, another regulator of PTP, might reinforce the myocardial protection afforded by CypD inhibition. METHODS: Adult mouse or isolated cardiomyocytes from wild-type or CypD knockout (CypD-KO) mice were subjected to ischemia or hypoxia followed by reperfusion or reoxygenation. Infarct size was assessed in vivo. Mitochondrial membrane potential and PTP opening were assessed using tetramethylrhodamine methyl ester perchlorate and calcein-cobalt fluorescence, respectively. Fluo-4 AM and rhod-2 AM staining allowed the measurement, by confocal microscopy, of Ca transient and Ca transfer from sarcoplasmic reticulum (SR) to mitochondria after caffeine stimulation. RESULTS: Both inhibition of CypD and isoflurane significantly reduced infarct size (-50 and -37%, respectively) and delayed PTP opening (+63% each). Their combination had no additive effect (n = 6/group). CypD-KO mice displayed endogenous protection against ischemia-reperfusion. Isoflurane depolarized the mitochondrial membrane (-28%, n = 5), decreased oxidative phosphorylation (-59%, n = 5), and blunted the caffeine-induced Ca transfer from SR to mitochondria (-22%, n = 7) in the cardiomyocytes of wild-type mice. Importantly, this transfer was spontaneously decreased in the cardiomyocytes of CypD-KO mice (-25%, n = 4 to 5). CONCLUSIONS: The results suggest that the partial inhibitory effect of isoflurane on respiratory complex I is insufficient to afford a synergy to CypD-induced protection. Isoflurane attenuates the Ca transfer from SR to mitochondria, which is also the prominent role of CypD, and finally prevents PTP opening.
Assuntos
Cálcio/metabolismo , Ciclofilinas/metabolismo , Precondicionamento Isquêmico Miocárdico , Isoflurano/administração & dosagem , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Anestésicos Inalatórios/administração & dosagem , Animais , Peptidil-Prolil Isomerase F , Complexo I de Transporte de Elétrons/metabolismo , Masculino , Camundongos , Camundongos Knockout , Mitocôndrias Cardíacas/efeitos dos fármacos , Mitocôndrias Cardíacas/metabolismoRESUMO
Loss of T-tubules (TT), sarcolemmal invaginations of cardiomyocytes (CMs), was recently identified as a general heart failure (HF) hallmark. However, whether TT per se or the overall sarcolemma is altered during HF process is still unknown. In this study, we directly examined sarcolemmal surface topography and physical properties using Atomic Force Microscopy (AFM) in living CMs from healthy and failing mice hearts. We confirmed the presence of highly organized crests and hollows along myofilaments in isolated healthy CMs. Sarcolemma topography was tightly correlated with elasticity, with crests stiffer than hollows and related to the presence of few packed subsarcolemmal mitochondria (SSM) as evidenced by electron microscopy. Three days after myocardial infarction (MI), CMs already exhibit an overall sarcolemma disorganization with general loss of crests topography thus becoming smooth and correlating with a decreased elasticity while interfibrillar mitochondria (IFM), myofilaments alignment and TT network were unaltered. End-stage post-ischemic condition (15days post-MI) exacerbates overall sarcolemma disorganization with, in addition to general loss of crest/hollow periodicity, a significant increase of cell surface stiffness. Strikingly, electron microscopy revealed the total depletion of SSM while some IFM heaps could be visualized beneath the membrane. Accordingly, mitochondrial Ca(2+) studies showed a heterogeneous pattern between SSM and IFM in healthy CMs which disappeared in HF. In vitro, formamide-induced sarcolemmal stress on healthy CMs phenocopied post-ischemic kinetics abnormalities and revealed initial SSM death and crest/hollow disorganization followed by IFM later disarray which moved toward the cell surface and structured heaps correlating with TT loss. This study demonstrates that the loss of crest/hollow organization of CM surface in HF occurs early and precedes disruption of the TT network. It also highlights a general stiffness increased of the CM surface most likely related to atypical IFM heaps while SSM died during HF process. Overall, these results indicate that initial sarcolemmal stress leading to SSM death could underlie subsequent TT disarray and HF setting.
Assuntos
Insuficiência Cardíaca/patologia , Mitocôndrias Cardíacas/ultraestrutura , Miócitos Cardíacos/ultraestrutura , Miofibrilas/ultraestrutura , Sarcolema/ultraestrutura , Animais , Elasticidade , Camundongos , Microscopia de Força Atômica , Microscopia EletrônicaRESUMO
BACKGROUND: Under physiological conditions, Ca(2+) transfer from the endoplasmic reticulum (ER) to mitochondria might occur at least in part at contact points between the 2 organelles and involves the VDAC1/Grp75/IP3R1 complex. Accumulation of Ca(2+) into the mitochondrial matrix may activate the mitochondrial chaperone cyclophilin D (CypD) and trigger permeability transition pore opening, whose role in ischemia/reperfusion injury is well recognized. We questioned here whether the transfer of Ca(2+) from ER to mitochondria might play a role in cardiomyocyte death after hypoxia-reoxygenation. METHODS AND RESULTS: We report that CypD interacts with the VDAC1/Grp75/IP3R1 complex in cardiomyocytes. Genetic or pharmacological inhibition of CypD in both H9c2 cardiomyoblasts and adult cardiomyocytes decreased the Ca(2+) transfer from ER to mitochondria through IP3R under normoxic conditions. During hypoxia-reoxygenation, the interaction between CypD and the IP3R1 Ca(2+) channeling complex increased concomitantly with mitochondrial Ca(2+) content. Inhibition of either CypD, IP3R1, or Grp75 decreased protein interaction within the complex, attenuated mitochondrial Ca(2+) overload, and protected cells from hypoxia-reoxygenation. Genetic or pharmacological inhibition of CypD provided a similar effect in adult mice cardiomyocytes. Disruption of ER-mitochondria interaction via the downregulation of Mfn2 similarly reduced the interaction between CypD and the IP3R1 complex and protected against hypoxia-reoxygenation injury. CONCLUSIONS: Our data (1) point to a new role of CypD at the ER-mitochondria interface and (2) suggest that decreasing ER-mitochondria interaction at reperfusion can protect cardiomyocytes against lethal reperfusion injury through the reduction of mitochondrial Ca(2+) overload via the CypD/VDAC1/Grp75/IP3R1 complex.
Assuntos
Sinalização do Cálcio/fisiologia , Hipóxia Celular/fisiologia , Retículo Endoplasmático/fisiologia , Mitocôndrias Cardíacas/fisiologia , Miócitos Cardíacos/patologia , Oxigênio/toxicidade , Animais , Linhagem Celular , Células Cultivadas/metabolismo , Peptidil-Prolil Isomerase F , Ciclofilinas/deficiência , Ciclofilinas/genética , Ciclofilinas/fisiologia , Proteínas de Choque Térmico HSP70/fisiologia , Técnicas In Vitro , Receptores de Inositol 1,4,5-Trifosfato/fisiologia , Membranas Intracelulares/fisiologia , Masculino , Proteínas de Membrana/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Complexos Multiproteicos , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/ultraestrutura , Técnicas de Patch-Clamp , Distribuição Aleatória , Ratos , Canal de Ânion 1 Dependente de Voltagem/fisiologiaRESUMO
Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited arrhythmogenic disease so far related to mutations in the cardiac ryanodine receptor (RYR2) or the cardiac calsequestrin (CASQ2) genes. Because mutations in RYR2 or in CASQ2 are not retrieved in all CPVT cases, we searched for mutations in the physiological protein partners of RyR2 and CSQ2 in a large cohort of CPVT patients with no detected mutation in these two genes. Based on a candidate gene approach, we focused our investigations on triadin and junctin, two proteins that link RyR2 and CSQ2. Mutations in the triadin (TRDN) and in the junctin (ASPH) genes were searched in a cohort of 97 CPVT patients. We identified three mutations in triadin which cosegregated with the disease on a recessive mode of transmission in two families, but no mutation was found in junctin. Two TRDN mutations, a 4 bp deletion and a nonsense mutation, resulted in premature stop codons; the third mutation, a p.T59R missense mutation, was further studied. Expression of the p.T59R mutant in COS-7 cells resulted in intracellular retention and degradation of the mutant protein. This was confirmed after in vivo expression of the mutant triadin in triadin knock-out mice by viral transduction. In this work, we identified TRDN as a new gene responsible for an autosomal recessive form of CPVT. The mutations identified in the two families lead to the absence of the protein, thereby demonstrating the importance of triadin for the normal function of the cardiac calcium release complex in humans.
Assuntos
Arritmias Cardíacas/genética , Proteínas de Transporte/genética , Morte Súbita Cardíaca , Proteínas Musculares/genética , Taquicardia Ventricular/genética , Animais , Arritmias Cardíacas/metabolismo , Western Blotting , Células COS , Cálcio/metabolismo , Proteínas de Transporte/metabolismo , Membrana Celular/metabolismo , Chlorocebus aethiops , Retículo Endoplasmático/metabolismo , Saúde da Família , Feminino , Genes Recessivos , Predisposição Genética para Doença/genética , Humanos , Masculino , Camundongos , Camundongos Knockout , Proteínas Musculares/metabolismo , Mutação , Miócitos Cardíacos/metabolismo , Linhagem , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Taquicardia Ventricular/metabolismo , Taquicardia Ventricular/patologiaRESUMO
In vitro human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) can differentiate into functional cardiomyocytes (CMs). Protocols for cardiac differentiation of hESCs and hiPSCs include formation of the three-dimensional cell aggregates called embryoid bodies (EBs). The traditional suspension method for EB formation from clumps of cells results in an EB population heterogeneous in size and shape. In this study we show that forced aggregation of a defined number of single cells on AggreWell plates gives a high number of homogeneous EBs that can be efficiently differentiated into functional CMs by application of defined growth factors in the media. For cardiac differentiation, we used three hESC lines and one hiPSC line. Our contracting EBs and the resulting CMs express cardiac markers, namely myosin heavy chain α and ß, cardiac ryanodine receptor/calcium release channel, and cardiac troponin T, shown by real-time polymerase chain reaction and immunocytochemistry. Using Ca(2+) imaging and atomic force microscopy, we demonstrate the functionality of RyR2 to release Ca(2+) from the sarcoplasmic reticulum as well as reliability in contractile and beating properties of hESC-EBs and hiPSC-EBs upon the stimulation or inhibition of the ß-adrenergic pathway.
Assuntos
Corpos Embrioides/fisiologia , Células-Tronco Pluripotentes Induzidas/fisiologia , Miócitos Cardíacos , Retículo Sarcoplasmático/metabolismo , Cálcio/metabolismo , Diferenciação Celular , Linhagem Celular , Forma Celular , Tamanho Celular , Humanos , Miócitos Cardíacos/citologia , Miócitos Cardíacos/fisiologia , Cadeias Pesadas de Miosina/metabolismo , Reprodutibilidade dos Testes , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Troponina T/metabolismoRESUMO
Myocardial ischemic disease is the major cause of death worldwide. After myocardial infarction, reperfusion of infracted heart has been an important objective of strategies to improve outcomes. However, cardiac ischemia/reperfusion (I/R) is characterized by inflammation, arrhythmias, cardiomyocyte damage, and, at the cellular level, disturbance in Ca(2+) and redox homeostasis. In this study, we sought to determine how acute inflammatory response contributes to reperfusion injury and Ca(2+) homeostasis disturbance after acute ischemia. Using a rat model of I/R, we show that circulating levels of TNF-α and cardiac caspase-8 activity were increased within 6 h of reperfusion, leading to myocardial nitric oxide and mitochondrial ROS production. At 1 and 15 d after reperfusion, caspase-8 activation resulted in S-nitrosylation of the RyR2 and depletion of calstabin2 from the RyR2 complex, resulting in diastolic sarcoplasmic reticulum (SR) Ca(2+) leak. Pharmacological inhibition of caspase-8 before reperfusion with Q-LETD-OPh or prevention of calstabin2 depletion from the RyR2 complex with the Ca(2+) channel stabilizer S107 ("rycal") inhibited the SR Ca(2+) leak, reduced ventricular arrhythmias, infarct size, and left ventricular remodeling after 15 d of reperfusion. TNF-α-induced caspase-8 activation leads to leaky RyR2 channels that contribute to myocardial remodeling after I/R. Thus, early prevention of SR Ca(2+) leak trough normalization of RyR2 function is cardioprotective.
Assuntos
Caspase 8/metabolismo , Ventrículos do Coração/patologia , Traumatismo por Reperfusão Miocárdica/enzimologia , Traumatismo por Reperfusão Miocárdica/patologia , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Animais , Ativação Enzimática , Fluorescência , Traumatismo por Reperfusão Miocárdica/sangue , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Miocárdio/metabolismo , Miocárdio/patologia , Fenantridinas/metabolismo , Ratos , Ratos Endogâmicos WKY , Fator de Necrose Tumoral alfa/sangue , Remodelação VentricularRESUMO
Atrial cardiomyopathy is defined as any complex of structural, architectural, contractile or electrophysiological changes affecting atria, with the potential to produce clinically relevant manifestations. Most of our knowledge about the mechanistic aspects of atrial cardiomyopathy is derived from studies investigating animal models of atrial fibrillation and atrial tissue samples obtained from individuals who have a history of atrial fibrillation. Several noninvasive tools have been reported to characterize atrial cardiomyopathy in patients, which may be relevant for predicting the risk of incident atrial fibrillation and its related outcomes, such as stroke. Here, we provide an overview of the pathophysiological mechanisms involved in atrial cardiomyopathy, and discuss the complex interplay of these mechanisms, including aging, left atrial pressure overload, metabolic disorders and genetic factors. We discuss clinical tools currently available to characterize atrial cardiomyopathy, including electrocardiograms, cardiac imaging and serum biomarkers. Finally, we discuss the clinical impact of atrial cardiomyopathy, and its potential role for predicting atrial fibrillation, stroke, heart failure and dementia. Overall, this review aims to highlight the critical need for a clinically relevant definition of atrial cardiomyopathy to improve treatment strategies.
Assuntos
Apêndice Atrial , Fibrilação Atrial , Cardiomiopatias , Acidente Vascular Cerebral , Animais , Humanos , Fibrilação Atrial/diagnóstico , Átrios do Coração , Cardiomiopatias/diagnóstico , Cardiomiopatias/terapiaRESUMO
Duchenne muscular dystrophy (DMD) is characterized by myofiber death from apoptosis or necrosis, leading in many patients to fatal respiratory muscle weakness. Among other pathological features, DMD muscles show severely deranged metabolic gene regulation and mitochondrial dysfunction. Defective mitochondria not only cause energetic deficiency, but also play roles in promoting myofiber atrophy and injury via opening of the mitochondrial permeability transition pore. Autophagy is a bulk degradative mechanism that serves to augment energy production and eliminate defective mitochondria (mitophagy). We hypothesized that pharmacological activation of AMP-activated protein kinase (AMPK), a master metabolic sensor in cells and on-switch for the autophagy-mitophagy pathway, would be beneficial in the mdx mouse model of DMD. Treatment of mdx mice for 4 weeks with an established AMPK agonist, AICAR (5-aminoimidazole-4-carboxamide-1-ß-d-ribofuranoside), potently triggered autophagy in the mdx diaphragm without inducing muscle fiber atrophy. In AICAR-treated mdx mice, the exaggerated sensitivity of mdx diaphragm mitochondria to calcium-induced permeability transition pore opening was restored to normal levels. There were associated improvements in mdx diaphragm histopathology and in maximal force-generating capacity, which were not linked to increased mitochondrial biogenesis or up-regulated utrophin expression. These findings suggest that agonists of AMPK and other inducers of the autophagy-mitophagy pathway can help to promote the elimination of defective mitochondria and may thus serve as useful therapeutic agents in DMD.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Autofagia , Diafragma/enzimologia , Diafragma/patologia , Distrofia Muscular Animal/enzimologia , Distrofia Muscular Animal/patologia , Aminoimidazol Carboxamida/análogos & derivados , Aminoimidazol Carboxamida/farmacologia , Animais , Diafragma/fisiopatologia , Diafragma/ultraestrutura , Metabolismo Energético/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Técnicas In Vitro , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Poro de Transição de Permeabilidade Mitocondrial , Complexos Multiproteicos , Contração Muscular/efeitos dos fármacos , Distrofia Muscular Animal/fisiopatologia , Oxirredução/efeitos dos fármacos , Proteínas/metabolismo , Ribonucleotídeos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TORRESUMO
Recent studies indicate that members of the multidrug-resistance protein (MRP) family belonging to ATP binding cassette type C (ABCC) membrane proteins extrude cyclic nucleotides from various cell types. This study aimed to determine whether MRP proteins regulate cardiac cAMP homeostasis. Here, we demonstrate that MRP4 is the predominant isoform present at the plasma membrane of cardiacmyocytes and that it mediates the efflux of cAMP in these cells. MRP4-deficient mice displayed enhanced cardiac myocyte cAMP formation, contractility, and cardiac hypertrophy at 9 mo of age, an effect that was compensated transiently by increased phosphodiesterase expression at young age. These findings suggest that cAMP extrusion via MRP4 acts together with phosphodiesterases to control cAMP levels in cardiac myocytes.
Assuntos
AMP Cíclico/metabolismo , Homeostase , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Miócitos Cardíacos/metabolismo , 1-Metil-3-Isobutilxantina/farmacologia , Animais , Western Blotting , Cardiomegalia/diagnóstico por imagem , Cardiomegalia/genética , Cardiomegalia/metabolismo , Membrana Celular/metabolismo , Células Cultivadas , Ecocardiografia , Feminino , Regulação Enzimológica da Expressão Gênica , Isoenzimas/genética , Isoenzimas/metabolismo , Masculino , Camundongos , Camundongos Knockout , Microscopia Confocal , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Contração Miocárdica/genética , Contração Miocárdica/fisiologia , Miócitos Cardíacos/citologia , Miócitos Cardíacos/efeitos dos fármacos , Inibidores de Fosfodiesterase/farmacologia , Diester Fosfórico Hidrolases/genética , Diester Fosfórico Hidrolases/metabolismo , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de TempoRESUMO
Patients with Duchenne muscular dystrophy (DMD) have a progressive dilated cardiomyopathy associated with fatal cardiac arrhythmias. Electrical and functional abnormalities have been attributed to cardiac fibrosis; however, electrical abnormalities may occur in the absence of overt cardiac histopathology. Here we show that structural and functional remodeling of the cardiac sarcoplasmic reticulum (SR) Ca(2+) release channel/ryanodine receptor (RyR2) occurs in the mdx mouse model of DMD. RyR2 from mdx hearts were S-nitrosylated and depleted of calstabin2 (FKBP12.6), resulting in "leaky" RyR2 channels and a diastolic SR Ca(2+) leak. Inhibiting the depletion of calstabin2 from the RyR2 complex with the Ca(2+) channel stabilizer S107 ("rycal") inhibited the SR Ca(2+) leak, inhibited aberrant depolarization in isolated cardiomyocytes, and prevented arrhythmias in vivo. This suggests that diastolic SR Ca(2+) leak via RyR2 due to S-nitrosylation of the channel and calstabin2 depletion from the channel complex likely triggers cardiac arrhythmias. Normalization of the RyR2-mediated diastolic SR Ca(2+) leak prevents fatal sudden cardiac arrhythmias in DMD.
Assuntos
Arritmias Cardíacas/metabolismo , Ventrículos do Coração/metabolismo , Distrofia Muscular de Duchenne/metabolismo , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Animais , Arritmias Cardíacas/etiologia , Arritmias Cardíacas/genética , Cálcio/metabolismo , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Distrofia Muscular de Duchenne/complicações , Distrofia Muscular de Duchenne/genética , Canal de Liberação de Cálcio do Receptor de Rianodina/genética , Retículo Sarcoplasmático/metabolismo , Proteínas de Ligação a Tacrolimo/metabolismoRESUMO
BACKGROUND: The use of mesenchymal stem cells (MSCs) appears to be a promising therapeutic approach for cardiac repair after myocardial infarction. However, clinical trials have revealed the need to improve their therapeutic efficacy. Recent evidence demonstrated that mitochondria undergo spontaneous transfer from damaged cells to MSCs, resulting in the activation of the cytoprotective and pro-angiogenic functions of recipient MSCs. Based on these observations, we investigated whether the preconditioning of MSCs with mitochondria could optimize their therapeutic potential for ischemic heart disease. METHODS: Human MSCs were exposed to mitochondria isolated from human fetal cardiomyocytes. After 24 h, the effects of mitochondria preconditioning on the MSCs' function were analyzed both in vitro and in vivo. RESULTS: We found that cardiac mitochondria-preconditioning improved the proliferation and repair properties of MSCs in vitro. Mechanistically, cardiac mitochondria mediate their stimulatory effects through the production of reactive oxygen species, which trigger their own degradation in recipient MSCs. These effects were further confirmed in vivo, as the mitochondria preconditioning of MSCs potentiated their therapeutic efficacy on cardiac function following their engraftment into infarcted mouse hearts. CONCLUSIONS: The preconditioning of MSCs with the artificial transfer of cardiac mitochondria appears to be promising strategy to improve the efficacy of MSC-based cell therapy in ischemic heart disease.
Assuntos
Células-Tronco Mesenquimais , Infarto do Miocárdio , Isquemia Miocárdica , Camundongos , Animais , Humanos , Isquemia Miocárdica/metabolismo , Infarto do Miocárdio/metabolismo , Miócitos Cardíacos/metabolismo , Mitocôndrias Cardíacas/metabolismo , Células-Tronco Mesenquimais/metabolismoRESUMO
BACKGROUND: Cardiomyocytes use Ca2+ not only in excitation-contraction coupling but also as a signaling molecule promoting, for example, cardiac hypertrophy. It is largely unclear how Ca2+ triggers signaling in cardiomyocytes in the presence of the rapid and large Ca2+ fluctuations that occur during excitation-contraction coupling. A potential route is store-operated Ca2+ entry, a drug-inducible mechanism for Ca2+ signaling that requires stromal interaction molecule 1 (STIM1). Store-operated Ca2+ entry can also be induced in cardiomyocytes, which prompted us to study STIM1-dependent Ca2+ entry with respect to cardiac hypertrophy in vitro and in vivo. METHODS AND RESULTS: Consistent with earlier reports, we found drug-inducible store-operated Ca2+ entry in neonatal rat cardiomyocytes, which was dependent on STIM1. Although this STIM1-dependent, drug-inducible store-operated Ca2+ entry was only marginal in adult cardiomyocytes isolated from control hearts, it increased significantly in cardiomyocytes isolated from adult rats that had developed compensated cardiac hypertrophy after abdominal aortic banding. Moreover, we detected an inwardly rectifying current in hypertrophic cardiomyocytes that occurs under native conditions (i.e., in the absence of drug-induced store depletion) and is dependent on STIM1. By manipulating its expression, we found STIM1 to be both sufficient and necessary for cardiomyocyte hypertrophy in vitro and in the adult heart in vivo. Stim1 silencing by adeno-associated viruses of serotype 9-mediated gene transfer protected rats from pressure overload-induced cardiac hypertrophy. CONCLUSION: By controlling a previously unrecognized sarcolemmal current, STIM1 promotes cardiac hypertrophy.
Assuntos
Sinalização do Cálcio/fisiologia , Cardiomegalia/fisiopatologia , Glicoproteínas de Membrana/fisiologia , Miócitos Cardíacos/fisiologia , Adenoviridae/genética , Fatores Etários , Animais , Animais Recém-Nascidos , Cafeína/farmacologia , Cálcio/metabolismo , Canais de Cálcio , Sinalização do Cálcio/efeitos dos fármacos , Cardiomegalia/metabolismo , Cardiomegalia/patologia , Células Cultivadas , Inibidores Enzimáticos/farmacologia , Inativação Gênica , Técnicas de Transferência de Genes , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Transgênicos , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Técnicas de Patch-Clamp , Inibidores de Fosfodiesterase/farmacologia , Ratos , Sarcolema/metabolismo , Molécula 1 de Interação Estromal , Tapsigargina/farmacologiaRESUMO
BACKGROUND: Patients with metabolic syndrome (MetS) have an increased risk of atrial fibrillation (AF). Impaired Ca2+ homeostasis and mitochondrial dysfunction have emerged as an arrhythmogenic substrate in both patients and animal models of MetS. Whether impaired mitochondrial Ca2+ handling underlies AF associated with MetS remains poorly explored. OBJECTIVES: The aim of this study was to determine the initial mechanisms related to AF susceptibility and mitochondrial dysfunction encountered in metabolic cardiomyopathy. METHODS: A total of 161 mice and 34 patients were studied. Mitochondrial Ca2+ and mitochondrial Ca2+ uniporter complex (MCUC) were investigated in right atrial tissue of patients with (n = 18) or without (n = 16) MetS and of C57Bl/6J mice fed with a high-fat sucrose diet (HFS) for 2 (n = 42) or 12 (n = 39) weeks. Susceptibility to AF was evaluated in isolated sinoatrial tissue and in vivo in mice. RESULTS: Increased expression of the MICUs subunits of the MCUC (1.00 ± 0.33 AU vs 1.29 ± 0.23 AU; P = 0.034) was associated with impaired mitochondrial Ca2+ uptake in patients (168.7 ± 31.3 nmol/min/mg vs 127.3 ± 18.4 nmol/min/mg; P = 0.026) and HFS mice (0.10 ± 0.04 ΔF/F0 × ms-1 vs 0.06 ± 0.03 ΔF/F0 × ms-1; P = 0.0086, and 0.15 ± 0.07 ΔF/F0 × ms-1 vs 0.046 ± 0.03 ΔF/F0 × ms-1; P = 0.0076 in 2- and 12-week HFS mice, respectively). HFS mice elicited a 70% increased susceptibility to AF. The MCUC agonist kaempferol restored MCUC activity in vitro and abolished the occurrence of AF in HFS mice. CONCLUSIONS: Impaired MCUC activity and mitochondrial Ca2+ homeostasis from the early stage of metabolic cardiomyopathy in mice lead to AF. Given that similar defects in cardiac mitochondrial Ca2+ handling are present in MetS patients, the modulation of the MCUC activity represents an attractive antiarrhythmic strategy.